Cancer Metastasis through the Lymphovascular System: Biology and Treatment (cancermetastasis2017)
Holiday Inn Golden Gateway, San Francisco, USA, April 20-22, 2017

Thursday, April 20, 2017 - Gold Rush Ballroom - 07:30 - 08:30

Industry Sponsored Breakfast Session (Non-CME) - Genomic Profiling, Sponsored by Foundation Medicine, Inc.

00-01

The value of Comprehensive Genomic Profiling, including immunotherapy & liquid biopsy options

Allison Welsh

Foundation Medicine Inc., Cambridge, MA, United States

This talk will give an overview of the new advancements in Comprehensive Genomic Profiling of cancer.  It will include some of the latest data on the power and value of comprehensive profiling versus smaller panels, as well as the opportunity to use biomarkers such as Tumor Mutational Burden to build towards a more personalized approach to immunotherapy.  Lastly, it will touch on the challenges of genomic profiling from liquid biopsies, and the potential scenarios where ctDNA assays may have the most clinical utility at present.

Keywords: comprehensive genomic profiling, immunotherapy, liquid biopsy, NSCLC

Thursday, April 20, 2017 - Emerald Ballroom - 08:35 - 08:45

Plenary Session (CME) - Welcome & Introductions

00-01

Welcome to Cancer Metastasis through the Lymphovascular System: Biology & Treatment, 7th International Symposium.

Warren Browner

California Pacific Medical Center, San Francisco, United States

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Thursday, April 20, 2017 - Emerald Ballroom - 08:45 - 09:15

Keynote Session (CME) - Donald Morton Surgical Oncology Memorial Lecture

00-01

Clinical utility of sentinel lymph nodes

Charles Balch

University of Texas MD Anderson Cancer Center, Houston, United States

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Thursday, April 20, 2017 - Emerald Ballroom - 09:15 - 10:25

Plenary Session (CME) - Tumor Microenvironment in Cancer Initiation & Progression

00-01

Interactions between cancer and stromal cells: selection of the fittest clones to metastasize

Stanley Leong

California Pacific Medical Ctr, San Francisco, United States

Primary cancer grows within the stromal cells which constitute the tumor microenvironment consisting of normal cells such as fibroblasts, lipocytes, immune cells, lymphatic and vascular vessels and other parenchymal cells. Cancer development is associated with uncontrolled proliferation, suppression of apoptosis, stabilization of telomeres, increase in mutation rates, evasion of the immune system, induction of lymphangiogenesis and acquisition of metastatic properties. Each of these characteristics is genetically dependent. Similar to Darwin’s theory of evolution, the tumor microenvironment exerts either a positive or negative force for tumor growth, thus, acting like a natural selection over the genetic diversity of the tumor population. The greater the genetic diversity within the tumor, the more likely it is for the emergence of the fittest clones being selected by the tumor microenvironment. Using techniques such as multiplexed microscopy, DNA/mRNA profiling, microRNA analysis and gene exon sequencing, we are learning more and more about this intricate relationship between tumor microenvironment and the emergence of more aggressive metastatic clones to systemic sites via the lymphatic and vascular vessels. Understanding such intricate molecular mechanisms, rational therapy may be directed to control or block the process of metastasis.

Keywords: Keywords


00-02

Social evolution and natural selection in cancer progression

Athena Aktipis

Arizona State University,, Tempe, United States

Our bodies are composed of approximately 30 trillion cells that engage in a multitude of social behaviors that make us viable multicellular organisms including cells sharing resources, engaging in division of labor and working together to create and maintain the intracellular environment. Natural selection acts on these and other behaviors of cells in the body, including controlled cell death and proliferation controls, in ways that can lead to cancer initiation and progression. All of this occurs within a complex and dynamic ecological environment that shapes resource availability and hence the selection pressures that cells face during neoplastic progression. In this talk I describe how cancer can be understood as cheating in multicellular cooperation and show results from computational models that suggest that cellular cheating through resource monopolization may be an important contributor to the evolution of invasion and metastasis.   

Keywords: cooperation, evolution, multicellularity, ecology, computational modeling


00-03

Classifying the evolution and ecology of tumors

Carlo Maley

Arizona State University, Tempe, United States

Neoplasms change over time, metastasize and ultimately lead to patient death, through a process of cell-level evolution. This process is driven by genetic and epigenetic alterations. However, the ecology of a neoplastic cell’s microenvironment determines which changes provide adaptive benefits. There is widespread recognition of the importance of these evolutionary and ecological processes in cancer, but to date no system has been proposed for drawing clinically relevant distinctions between how different tumors are evolving. We lack a language with which to communicate and coordinate about the dynamcis and trajectories of cancers. We propose a framework for classifying tumors that is based on four relevant components: (1) diversity of neoplastic cells (intra-tumor heterogeneity) and (2) changes over time in that diversity, which make up an evolutionary index (Evo-index), as well as the (3) hazards to neoplastic cell survival, and (4) resources available to neoplastic cells, which make up an ecological index (Eco-index). Development of this classification system holds promise for enabling clinicians to personalize optimal interventions based on the evolvability of the patient’s tumor. The Evo- and Eco-indices provide a common lexicon for communicating about how neoplasms change in response to interventions, with potential implications for clinical trials, personalized medicine and basic cancer research.

Keywords: evolution, ecology, biomarkers, tumor classification

Thursday, April 20, 2017 - Emerald Ballroom - 11:00 - 12:35

Plenary Session (CME) - Tumor Growth and Metastatic Spread

00-01

Cancer metastasis and the lymphatic system

Marlys Witte

University of Arizona, Tucson, United States

Overview

Historically, prior to the mid-1800's and Virchow's establishment of the "cell theory" of cancer, the "lymph theory" and later the "endothelial theory" of the origin and spread of cancer prevailed; it was also recognized that cancer spread to lymph nodes. Currently, the central importance of the lymphatic system (lymphatics, lymph, lymphocytes, and lymph nodes and its study - lymphology) is increasingly recognized as the environment in which cancer develops, is immersed in and surrounded by, and provides the key route for dissemination to regional lymph nodes and likely also to the blood circulation and distant organs. This brief overview will review fundamental concepts of the lymphatic system as a vasculature, key component of the blood-lymph loop circulation of extracellular fluid, route of entry and transport of large molecules, particles and cells, and also encompassing the immune system. How these concepts bear on cancer development and spread will then be highlighted and specifically related to ECM alterations, epithelial-mesenchymal and mesenchymal-epithelial transition, cancer stem cells, lymphangiogenesis, sentinel lymph nodes, lymphogenous dissemination, benign metastasizing tumors, lymphedema, and tumor immunotherapy.

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00-02

Oncolytic virus mpJX-594 induces widespread cell death in pancreatic neuroendocrine tumors

Donald McDonald

University of California, San Francisco, San Francisco, United States

Oncolytic viruses are promising new treatments for cancer. We examined the anti-tumor effects of a replication-competent, oncolytic vaccinia virus mpJX-594 (mouse-prototype JX-594, a mouse-adapted version of Pexa-Vec) administered intravenously to RIP-Tag2 transgenic mice that spontaneously develop pancreatic neuroendocrine tumors. We found that mpJX-594 infected the endothelial cells of tumor blood vessels within hours of iv injection, before spreading to tumor cells. Endothelial cell infection by mpJX-594 was followed by leakiness and pruning of the tumor blood vessels. Non-tumor vasculature was not infected or detectably altered. Subsequently, the virus infected focal regions of tumor cells in the neuroendocrine tumors. Despite the sites of tumor infection being focal, infiltration by activated CD8 T-lymphocytes and apoptosis of tumor cells were widespread within tumors, and were followed by reduction in tumor size, invasion, and metastasis. The magnitude of mpJX-594 infection and anti-tumor actions were increased when the oncolytic virus was administered with the multi-targeted receptor tyrosine kinase inhibitor sunitinib. Together, these findings revealed that intravenous injection of oncolytic virus mpJX-594 results initially in selective infection of tumor blood vessels followed by focal regions of tumor cell infection and widespread CD8 T-cell influx and cell killing in tumors. Anti-tumor actions of mpJX-594 were stronger when the virus was given together with sunitinib. Even though the oncolytic viral infection of tumor cells was focal, the widespread influx of activated CD8 T-cells and tumor-cell apoptosis contributed to reduced tumor burden, invasion, and metastasis.

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00-03

Angiogenesis & lymphangiogenesis in cancer metastasis: concordant or independent

Michael Dellinger

UT Southwestern Medical Center, Dallas, United States

It is well established that lung tumors induce the formation of lymphatic vessels. However, the molecular mechanisms controlling tumor lymphangiogenesis in lung cancer have not been fully delineated. In the present study, we identify a panel of non-small cell lung cancer (NSCLC) cell lines that induce lymphangiogenesis and use genome-wide mRNA expression to characterize the molecular mechanisms regulating tumor lymphangiogenesis. We show that Calu-1, H1993, HCC461, HCC827, and H2122 NSCLC cell lines form tumors that induce lymphangiogenesis whereas Calu-3, H1155, H1975, and H2073 NSCLC cell lines form tumors that do not induce lymphangiogenesis. By analyzing genome-wide mRNA expression data, we identify a 17-gene expression signature that distinguishes lymphangiogenic from non-lymphangiogenic NSCLC cell lines. Importantly, VEGF-C is the only lymphatic growth factor in this expression signature and is approximately 50-fold higher in the lymphangiogenic group than in the non-lymphangiogenic group. We show that forced expression of VEGF-C by H1975 cells induces lymphangiogenesis and that knockdown of VEGF-C in H1993 cells inhibits lymphangiogenesis. Additionally, we demonstrate that the triple angiokinase inhibitor, nintedanib (small molecule that blocks all FGFRs, PDGFRs, and VEGFRs), suppresses tumor lymphangiogenesis in H1993 tumors. Together, our data suggest that VEGF-C is the dominant driver of tumor lymphangiogenesis in NSCLC and that nintedanib could potentially block tumor lymphangiogenesis in NSCLC patients.

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00-04

Molecular biomarkers for cancer: proteins, genes and exosomes

Mohammed Kashani-Sabet

California Pacific Medical Ctr, San Francisco, United States

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Thursday, April 20, 2017 - Gold Rush Ballroom - 12:35 - 13:45

Industry Sponsored Lunch Session (Non-CME) - Prognostic Biomarker, Sponsored by Castle Biosciences

00-01

Tissue Based Biomarkers of Prognosis in Malignant Melanoma

Jane Messina

Moffitt Cancer Center, Tampa, United States

Despite the explosion of knowledge concerning the genomics of melanoma, accurate prediction of prognosis at the time of diagnosis remains a challenge for practitioners. Conventional staging systems such as the AJCC classification, which use pathologic features of the primary tumor and regional nodal status, are most widely used but there is still significant heterogeneity among patients of the same stage. Thus, there is opportunity for improved predictive tools, and in fact a number of other models have been developed for melanoma. These include web-based prediction tools, mostly incorporating primary and nodal tumor pathology characteristics, and models using other data such as serum LDH. An assay of primary melanoma tumor tissue, DecisionDx-Melanoma, has recently been developed and externally validated. Validation studies have indicated that it can significantly predict metastasis and survival independent of AJCC stage. This session will discuss the ideal development of a prognostic tool for melanoma, compare the current tools, and briefly discuss additional emerging prognostic factors that may represent improvements of our current methods.

Keywords: melanoma prognosis, prognostic assays


00-02

Surgical management implications of prognostic biomarkers in melanoma

Eddy Hsueh

St. Louis University School of Medicine, St. Louis, United States

Recent advancement in molecular prognostication for melanoma may have significant implications for future surgical management of primary melanoma. Results of recent molecular prognositic studies including 31-gene gene expression profiling will be discussed. Implications for surgical margin, sentinel node dissection, and completion lymphadenectomy will be discussed. 

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00-03

Prognostic Biomarkers in the Era of Melanoma Immunotherapy

Jason Luke

The University of Chicago Medicine, Chicago, United States

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Thursday, April 20, 2017 - Emerald Ballroom - 13:50 - 15:10

Plenary Session (CME) - Biomarkers for Cancer Metastasis

00-01

Genomics for cancer metastasis

Marcus Bosenberg

Yale School of Medicine, Guilford, United States

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00-02

Utility of Circulating Cell-Free DNA In Assessing Cutaneous Melanoma Progression

Dave Hoon

John Wayne Cancer Institute, Santa Monica, United States

Melanoma patient blood-based biopsies have limited sensitivity in detecting early-stage regional lymph node metastasis and specific distant organ sites (i.e. melanoma brain metastasis; MBM). We investigated the clinical utility of a sensitive single-molecule next generation sequencing (smNGS) approach in patients’ plasma of early-stage regional lymph node melanoma metastasis and MBMs. We have evaluated bloods and matched tumor(s) when available from melanoma patients using smNGS. This study included a dozen patients with serial bleed analysis of  follow-up from regional metastatic disease to distant organ metastasis recurrence including  MBM patients. cfDNA genomic aberrations including therapeutic targets were detected in pre-operation of stage III/IV patients, including MBM patients. cfDNA mutation detection significantly correlated with tumor burden. Analysis demonstrated more efficiency of cfDNA compare to serum lactate dehydrogenase (LDH) a known blood prognostic marker for stage III/IV melanoma patients. cfDNA mutation analysis captured tumor heterogeneity and arising subclonal mutations upon recurrence. Patients with EGFR and/or MET cfDNA amplification were detect in plasma and were significantly associated with a shorter disease-free survival and overall survival compared. This study demonstrates the clinical utility of diagnosis and monitoring cutaneous melanoma from regional disease to brain metastasis. The studies demonstrate blood biopsy can detect subclinical disease well before onset of clinical disease progression. The approach also allows monitoring of tumor evolution/heterogeneity during distant metastasis progression during follow up.   

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00-03

Tumor associated and microenvironmental drivers of melanoma brain metastasis

Isaac P. Witz

Tel Aviv University, Ramat Aviv, Israel

It is well established that interactions between cancer cells with and non-cancerous cells in the tumor microenvironment and/or with soluble factors released from these cells, drive tumor progression towards metastasis.

Drivers of metastasis are cells or molecules whose activity is required for the targeted migration of tumor cells to the secondary target organ and for their survival and propagation within this organ.

Metastasis drivers may be grouped into 2 major groups: Drivers that are intrinsic to tumor cells and drivers that originate in the microenvironment of the secondary target organ. Both types of drivers are candidates to serve as targets for therapy.

Here we report on the identification and function of CCR4, a tumor-associated driver of    melanoma brain metastasis.                                                                                                             

A key initial event in brain metastasis is the rapid activation of microglial cells. Molecular signals that activate microglia cells induce the release of cytokines which take part in shaping the malignancy phenotype of brain metastasizing tumor cells. Microglial cells function thus as microenvironmental drivers of melanoma brain metastasis.

This study was supported by The Dr. Miriam and Sheldon G. Adelson Medical Research Foundation (Needham, MA).

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Thursday, April 20, 2017 - Emerald Ballroom - 15:45 - 16:15

Keynote Session (CME) - Super Resolution Microscopy to study Molecules within a Cell

00-01

Super-resolution microscopy to study molecules within a cell

William Moerner

Stanford University, Stanford, United States

Conventional optical microscopy at visible wavelengths suffers from severe resolution limitations in that features on scales smaller than about 200 nm cannot be resolved. Since cell receptor proteins, amyloid structures, and other important cellular actors are much smaller, there has been a need for higher resolution. The relatively new method of “super-resolution” fluorescence microscopy has circumvented this resolution limit allowing visualization of detail down to the 20-40 nm range. Super-resolution microscopy can be achieved by stimulated emission depletion, structured illumination, or single-molecule localization microscopy methods.  These approaches can now be used to explore structures that could not be observed previously, in both normal and diseased cells. Specific applications to bacterial protein superstructures, huntingtin aggregates, and other cellular targets will be described, along with potential applications to cancer-related structures.

 

 

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Thursday, April 20, 2017 - Emerald Ballroom - 16:15 - 17:50

Plenary Session (CME) - SLN: Incubator or Marker for Cancer Metastasis?

00-01

Sentinel lymph nodes represent an incubator for metastasis

Mark Faries

John Wayne Cancer Institute, Santa Monica, United States

The role of lymph nodes in the metastatic process has long been controversial. In my view, there is ample evidence to demonstrate that in many cases, lymph nodes represent an incubator for subsequent metastases. Early removal of metastatic nodes, therefore, will decrease the frequency of such spread and improve survival.

Available trial data supports this assertion in many (though not all) cases.

The key question for clinical use is determining in which situations intervention with lymphadenectomy is beneficial. The answer to this question will come from clinical trial data and improved assessments of the metastatic potential of different primary tumors.

Keywords: Elective node dissection, incubator hypothesis, sentinel lymph node.


00-02

Sentinel lymph nodes represent a marker for metastasis

Dale Han

Yale School of Medicine, New Haven, United States

The biology of the primary tumor controls the development of distant metastasis in patients with melanoma.  Because of this, the detection of sentinel lymph node metastasis simply represents a biologic marker of the aggressiveness of the primary tumor.  The incubator hypothesis implies that treatment of nodal metastasis has the potential to halt further disease spread and to improve survival.  The incubator hypothesis also implies that treatment of lower volumes of nodal metastases through completion lymphadenectomy should be associated with the greatest benefit due to decreased disease incubation time.  However, current data questions the validity of the incubator hypothesis since studies have shown that control of nodal micrometastases through completion lymphadenectomy is associated with no survival benefit.  These results show that sentinel lymph node disease simply represents a marker for metastasis and that sentinel lymph node biopsy is a crucial staging tool which provides information on the biology of the primary tumor.

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00-03

Nodal staging, survival and the need for CLND with the SLN procedure

Douglas Reintgen

University of South Florida, Tampa, United States

The status of the regional lymphatic basin in patients with malignant melanoma remains the most powerful predictor of recurrence and survival.   Lymphatic mapping and sentinel lymph node (SLN) biopsy has become the standard of the nodal staging due to lower patient morbidities and more accurate staging.    Investigations have determined the following:

1.  The nodal staging system for melanoma depends on the number of nodes positive so that patients with 1 node positive do better than patients with 2-3 nodes positive, do better than patients with 4 or more nodes positive.  Examining the distribution of nodal metastases in the regional basin has determined that patients with metastatic disease confined to the SLNs have a much better survival compared to patients with SLN and Non-SLN metastases.

2.  The SLN status plays a major role in conditional survival estimates for patients with melanoma.  This analysis shows that if patients live 4 years from their diagnosis without a recurrence, then survival becomes equal no matter what stage of disease the patients were assigned at the time of diagnosis.  However at no time do they achieve the same survival as the normal population.

3.  Gene expression assays have been developed for the early stage melanoma population.  A 31-gene panel can divide patients into 2 groups, either at high risk or low risk for recurrence.  This data has been used in combination with the SLN status to determine prognosis and influence surgical care.  Patients have been identified who are SLN negative but have the Class II signature for high risk for recurrence.  These patients have similar survivals as the microscopic Stage III population staged with SLN biopsy and may in turn be candidates for CLND or adjuvant therapies.

4.  Recent data from Germany has suggested that in the positive SLN population, patients may not need a complete regional node dissection.   This finding needs to be confirmed before guidelines are changed.

Nodal staging for patients with melanoma using the sentinel lymph node procedure remains the standard of care for patients with melanoma.   More recent studies have highlighted the importance of non-SLN metastases, more meaningful condition survival prognostic information, a gene profile assay that compliments the lymphatic mapping procedure, and an observation alternative for patients with a positive SLN.

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00-04

Role of Sentinel Lymph Node Staging in Melanoma Clinical Trials

Corrado Caracò

Istituto Nazionale Tumori Fondazione Pascale Napoli Italy, Naples, Italy

Since the introduction of this procedure in 1992, many trials were conducted to assessed its staging and therapeutic role. Initially  the procedure was analyzed to test the safety and efficacy in the short and long term outcome, if it is at least as effective and/or safe as existing proven techniques and to verify the prognostic role and the prognostic factors correlated to the outcome. The first Multicenter Selective Lymphadenectomy Trial (MSLT-I), comparing the procedure followed by immediate complete lymph node dissection in positive patients with observation only. For intermediate-thickness or thick primary melanomas provided important prognostic information and identified patients with nodal metastases who may benefit from immediate complete lymphadenectomy.Sunbelt Melanoma Trial showed that adjuvant high dose Interferon did not improve survival in patients with a single tumor positive sentinel node and was not adequately powered to detect small differences in disease-free and overall survival. The trial offered important information about technical aspect of the sentinel procedure and prognostic factors correlated with the disease outcome. Sentinel lymph node (SLN) biopsy is now a standard staging procedure for melanoma but the value of completion lymph node dissection (CLND) for patients with SLN metastases is not clear and needs to be clarified. More recently DeCog trial, from German groups, despite a very short follow-up leading to the trial being underpowered, showed no difference in overall survival in patients with positive sentinel node treated with complete lymph node dissection compared  with observation only. In this trial some limitations about the prognostic factors of the primary and the sentinel node reducing “a priori” the usefulness of CLND must be considered. Data from MSLT II trial are not yet available, but the time from study design and final results may represents a bias to lead the clinical interpretation. The changing in staging system and prognostic factors analyzed may lead again to underpowered the trial to identify those subgroups of patients that may benefit of an early nodal dissection. The sentinel procedure provides an accurate and important information and in absence of  MSLT II trial result, for positive sentinel node cases CLND remains the best opportunity to discuss with the patients.

Keywords: cutaneous melanoma, sentinel node, lymph node dissection

Thursday, April 20, 2017 - Gold Rush Ballroom - 17:50 - 19:30

Poster Session (Non-CME) - Poster Session

00-01

Substance p (sp) increases early onset adhesion of leukaemia (Jurkat) and oral squamous cell carcinoma cell lines (OSCC) to endothelial cells (EC) via adhesion molecules: implication for metastasis.

Moustafa Elhousiny1, anura Ariyawardana1, 2, Kate Miller1, Alan Nimmo1

1James cook University, SMITHFIELD, Australia
2Griffith Health Institute, smithfield, Australia
3James cook University, SMITHFIELD, Australia
4James cook University, SMITHFIELD, Australia
5James cook University, SMITHFIELD, Australia
6James cook University, SMITHFIELD, Australia
7James cook University, SMITHFIELD, Australia
8James cook University, SMITHFIELD, Australia
9James cook University, SMITHFIELD, Australia
10James cook University, SMITHFIELD, Australia
11James cook University, SMITHFIELD, Australia

Background: Metastasis is the leading cause of fatality in 90 % of cancer and approximately 60 % of patients will have metastasis at initial diagnosis. The striking similarity which exists between out-flux of tumor mass from the circulation and attraction of the circulating leucocytes to the injured site in inflammation points out to the possibility of same inflammatory mediators might be utilized for both purposes. . In this aspect, SP is a primary regulator of inflammatory extravasation and leukocyte adhesion to vascular endothelial cells. SP is highly expressed in several neoplasms, particularly metastatic lesions. Understanding the adhesion of cancer cells as rate limiting steps in metastasis, will improve outcome of cancer patients. We hypothesized that cancer utilizes the inflammatory mediator SP to adhere to EC, in acute phase, through inducing adhesion molecules expression in blood and solid tumor cell lines. The study was done with the following objectives; (1) to examine the adhesion of different tumor cell lines to endothelial cells and determine the effect of SP on this adhesion in acute phase (1-6). (2)To examine the expression of binding legends of endothelial adhesion molecules in those cell lines. (3) To examine the effect of SP treatment on the expression of these molecules. (4) To examine the effect of neurokinin receptor-1 antagonist (NK-1R) on the adhesion and adhesion molecule expression.
 

Methodology: We used Leukaemia cell line Jurkat (clone E6-1) and oral squamous cell carcinoma cell lines in different stages of the tumor (DOK, SCC25, CAL27, H157, and BICR22). Endothelial adhesion assay and fluorescence microscopy were used to quantify the adhesion of those cell lines to endothelial cells (HUV-EC-C) when treated with SP in acute phase.  Flow cytometry was used to measure the adhesion molecules (CD49, CD11, and CD15s) in those cell lines. Statistical analysis was done with one-way ANOVA with P<0.05 considered statistically significant.

Results: Treatment of Jurkat, H157 cell lines with SP significantly increased adhesion to EC in a time dependent manner with a peak at 3 hours. Additionally, SP treatment increased the adhesion of CAL27 and DOK cell lines significantly at 3 hours. SP treatment significantly increased the CD49 (VLA-4) in both Jurkat and CAL27 in similar manner with a maximum threshold at 3 hours. Using the NK-1R inhibitor and V-CAM monoclonal antibody significantly inhibited the adhesion to EC as well as the adhesion molecule expression in both time and dose dependent manner compared with the TNF as positive control. Untreated oral cell lines showed increasing expression of their adhesion molecules in proportion to their malignant stages.

Conclusion: Adhesion of tumor cells and endothelial cells represents a critical step in tumor metastasis and extravasation4. This adhesion is vital for survival of tumor cells, therefore, it must occur within few hours of tumor cells entering the blood vessels (i.e. acute phase like inflammation).  Our results, albeit requiring further in-vivo validation, highlights potential role for SP in acute adhesion and extravasation of tumor cells through EC. The study has important implications for the development of novel strategies for “Metastasis Prevention”.

Keywords: Substance P, Extravasation, inflammation, Metastasis, Adhesion, endothelial cells, adhesion molecules, metastasis prevention.


00-02

Beyond Breslow thickness: Do other clinicopathologic factors predict for sentinel node metastasis in patients with melanoma?

Timothy Murtha1, Gang Han2, Daniel Thomas1, Dale Han1

1Section of Surgical Oncology, Department of Surgery, Yale University School of Medicine, New Haven, United States
2Department of Epidemiology and Biostatistics, School of Public Health, Texas A&M University, College Station, United States

Background. Sentinel lymph node biopsy (SLNB) is generally recommended for staging patients with melanomas ≥ 1 mm thick. However, other clinicopathologic factors in addition to thickness may also predict for nodal metastasis. We analyzed the Surveillance, Epidemiology, and End Results (SEER) database to determine the prognostic value of nodal staging and to assess which factors are correlated with SLN disease in patients with melanoma.

Materials and Methods. From 2010 to 2012, 10,013 patients with cutaneous melanoma who were treated with SLNB were identified in the SEER database. Clinicopathologic factors were compared with SLN metastasis and outcome.

Results. Overall median thickness was 1.25 mm; 3269 (32.6%), 5707 (57.0%), and 1019 (10.2%) cases were thin, intermediate thickness, and thick, respectively. Thickness was unknown in eighteen cases (0.2%). SLN metastases were found in 448/10,013 cases (4.5%); 1.5%, 4.8%, and 12.1% of thin, intermediate thickness, and thick cases had a positive SLNB, respectively. Significant predictors of SLN metastasis on multivariable analysis included thickness, mitotic rate, tumor site, race, marital status, and high school completion (P < 0.05). Overall survival was significantly worse in patients with a positive SLNB (P < 0.05). SLN status, age, presence of ulceration, mitotic rate, thickness, and poverty level were significant predictors on multivariable analysis for overall survival (P < 0.05).

Conclusions. Melanoma thickness, used in combination with unique factors such as race and marital status, may improve identification of patients at risk for SLN metastasis. Additionally, SLN status provides powerful staging information and is predictive of survival in patients with thin, intermediate thickness, and thick melanomas.

Keywords: Melanoma, Sentinel lymph node biopsy


00-03

WITHDRAWN

Joy Odili1, Christian Kunte2, Pietro Quaglino3

1St George's Hospital Tooting London, London, United Kingdom
2Department of Dermatology and Allergology, Munich, Germany
3Department of Medical Sciences, Dermatology Clinic, Torino, Italy

ABSTRACT WITHDRAWN

Keywords: Electrochemotherapy, Melanoma, metastasis, Bleomycin, disease control, Palliation


00-04

Identification of Semaphorin 7a as a driver of lymphovascular dissemination in breast cancer patients

Traci Lyons

University of Colorado Anschutz Medical Campus, Aurora, United States

Postpartum breast cancers, defined as breast cancers diagnosed within 5-10 years of last childbirth, are nearly twice as likely to become metastatic; this devastating diagnosis affects thousands of young women annually. In pre-clinical models of postpartum breast cancer we have shown that tumor growth, invasion, lymph vessel density (LVD), lymphovascular invasion (LVI), and lymph node (LN) and lung metastasis are increased when breast tumor cells are implanted into postpartum mouse mammary glands compared to those in nulliparous mice. In addition, we revealed increased LVD and LN metastasis in postpartum patients (Lyons, TR et al., Nat Med 2011 and JCI 2014). We identified Sem7a protein as increased in postpartum breast cancer patient samples. In addition, SEMA7A mRNA expression is 3-fold higher in breast tumor tissue compared to normal and SEMA7A expression is associated with decreased overall and distant metastasis free survival, as well as with increased LVI in large patient datasets (Black, S…Lyons, TR, Oncogene 2016).  Using silencing and overexpression approaches we observe that growth, invasion, LVD, LVI, as well as LN and lung metastasis are dependent on Sem7a. In addition, we observe that Sem7a is important for adhering to lymphatic endothelial mono-layers and survival in non-adherent conditions. Cumulatively, our results suggest that Sem7a supports multiple steps in the metastatic cascade.  As such, we propose that Sem7a is a driver of metastasis for postpartum patients and a risk factor for metastasis in non-postpartum patients and therefore should be explored as a biomarker and therapeutic target.

Keywords: breast cancer, lymphangiogenesis, lymph vessel invasion, lymph node metastasis


00-05

Ablation of Phospholipase D1 & D2 Suppresses Pulmonary Metastasis in MMTV-PyMT Driven Breast Cancer

Eric Roth, Julie-Ann Cavallo, Michael Frohman

Center for Developmental Genetics and the Department of Pharmacological Sciences, Port Jefferson, United States

Background:

For breast cancer patients, metastasis is the primary cause of treatment failure and results in 90% of cancer-related deaths. Thus far, therapeutic approaches have largely focused on treatments that inhibit growth and proliferation of primary tumors, but few have achieved prevention of metastasis. Understanding the critical factors that drive cells through the complex metastatic cascade will allow for new targeted therapies.

The phospholipase D (PLD) isoforms 1 and 2 generate the lipid signaling molecule phosphatidic acid through the hydrolysis of phosphatidylcholine.  This pleotropic second messenger is involved in multiple tumorigenic pathways. PLD1 and PLD2 have been shown to promote several early metastatic steps including migration and invasion from the primary tumor, neoangiogenesis, and survival in circulation. Together, they have also been linked to 5 of the 6 hallmarks of cancer. Although PLD1 and PLD2 are highly related in amino acid sequence and enzymatic function, their cellular roles are not always redundant or additive, but can also be contradictory. As such, it is necessary to elucidate their separate and combined effects.

Materials and Methods:

Double knockout (DKO) mice lacking PLD1 and PLD2 were crossed with mice containing a mouse mammary tumor virus (MMTV) polyoma middle-T antigen (PyMT) transgene in a C57B6 background. These MMTV-PyMT mice spontaneously develop breast cancer that regularly metastasizes to the lung. The development of tumors is first noted by palpation around 8 weeks of age, and then monitored by caliper measurements when possible, typically around 12 weeks of age. The mice are euthanized at 22 weeks of age to collect mammary fat pads and lungs for measurements of size and weight, histological assessment, and the presence of metastasis.

Results:

PLD1-/- mice exhibit reduced metastasis relative to wild-type mice. We show here a further reduction in DKO mice that results in a dramatic suppression of lung metastasis. Furthermore, DKO mice have a decrease in tumor burden compared to both wild-type and PLD1-/- mice.

Conclusions:

Although PLD1 and PLD2 have been shown to have both complimentary and conflicting roles in cancer cell development, these and other studies offer promising results in support of their therapeutic inhibition. The overtly normal phenotype of DKO mice suggests that PLD inhibitors should have minimal toxicity, allowing them to be added to current-best therapeutic approaches.

Keywords: Phospholipase D, MMTV, breast cancer, animal model


00-06

Novel mechanisms of APE/Ref-1 in metastasis and potential as prognostic biomarker

SUN YANG

Chapman University School of Pharmacy, Irvine, United States

Cutaneous melanoma is the cancer with the fastest rate of increase in incidence, whereas most rates for other cancers are decreasing. Particularly, metastatic melanoma is very lethal and among the most drug resistant malignancies. Understanding the molecular and biochemical basis of melanomagenesis and progression is essential for developing novel and active targeted therapeutics.

Apurinic/apyrimidinic endonuclease/Redox factor-1 (APE/Ref-1) is a multifunctional protein that plays an important role in both DNA damage repair and cellular response to oxidative stress and is remarkably elevated in melanoma compared to normal melanocytes. Our previous studies demonstrated that the knockdown of APE/Ref-1 sensitized melanoma cells to chemo-treatment and significantly reduced metastatic potential. Our studies on JB6 cells showed that in combination with reactive oxygen species (ROS), APE/Ref-1 facilitated malignant transformation as evidence of increased colony formations in soft agar. However, limited progress has been made toward understanding the detailed mechanisms of APE/Ref-1 in melanomagenesis and progression.

E-cadherin/Snail signaling pathway plays an important role in epithelial-mesenchymal transition. Decreased E-cadherin expression has been well-documented to accompany the vertical growth phase and have been associated with elevated invasion potential. Our studies have showed that depletion of APE/Ref-1 resulted in recovery of E-cadherin expression in metastatic melanoma cells. Detected by RT-PCR, the E-cadherin level was increased to ~8 folds of control in APE/Ref-1 depleted melanoma cells. Snail, which represses transcription of E-cadherin, was found overexpressed in human melanoma. Depletion of APE/Ref-1 was also shown to inhibit Snail expression levels. Notably, Immuno-precipitation analysis demonstrated direct association between APE/Ref-1 and SNAIL protein in all tested human melanoma cell lines, which is dynamically changed with distinct stages. The APE/Ref-1/Snail complex was significantly increased in late stage of primary melanoma cells (vertical growth phase) compare to early stage and metastatic melanoma cells, which may contribute to the loss of E-cadherin and allow melanoma cells to escape from keratinocyte growth control.

Current studies are underway to ascertain whether APE/Ref-1 nuclear translocation plus Snail expression serves as a determinant of malignant transformation and melanoma progression, especially in patients with primary melanoma of Breslow depth (1-5mm).

Keywords: Melanoma, APE/Ref-1, E-cadherin, Snail, biomarker


00-07

The effect of human skin lymph on tumor cell proliferation

Marzanna Zaleska1, 2, Waldemar Olszewski2

1Medical Research Center, Warsaw, Poland
2Central Clinical Hospital, Warsaw, Poland

Background. All tissues and organ cells have their own environment made up of extracellular matrix and mobile tissue fluid. Tissue fluid is a mixture of capillary filtrate containing nutrients with metabolic and waste products of cells. It percolates the cells and enters lymphatic to become lymph. Tumor cells, both primary and metastatic, proliferate, at least in the initial stage, in the tissue fluid (lymph) specific for the tissue in which they are located.

Aim. The aim was to examine the effects of human skin and subcutaneous tissue lymph on proliferation of rapidly growing, established, lymphoid and non-lymphoid human tumor cell lines which either grow or metastasize to skin and subcutaneous tissue.

Material and methods. Lymph was collected from cannulated leg superficial lymphatics in volunteers in a study on the effect of muscular contractions on tissue fluid flow (Ethics Committee consent). It was added to tumor cell cultures at various concentrations. B cell lines: Raji. Rael, Balm 1, KM3. T cell lines: Molt 4, CCRF-CEM, CCRF-HSB, Jurkat. Melanoma  cell lines FMEX, LOX. Sarcoma cell lines  OHS, OS3, SaOS-2.Epithelioma carcinoma  HT3 , HeLa. Transformed keratinocytes  Hacat.  Short term tumor cell cultures: 3 days in RPMI 1640 with 5, 20, 40, 80 and 100% of lymph ,3HTdR  was added 18h before harvesting. Long term culture in soft agar colony assay 14 to 21 days with 5,20, 40, 80% of lymph. Preincubation for 1-3 days in 5,20, 40, 80% of  lymph and short term culture.

Results. Different tumor cell lines displayed considerable variations in the proliferation rate  when cultured in media were supplemented with lymph from human skin. Low concentrations had stimulatory effect on some tumor lines. High lymph concentrations (40-100%) had an inhibitory effect on most cultured cell lines , but stimulatory effect was also observed.  Preincubation of tumor cells with lymph demonstrated that the growth regulatory lymph factors might be absorbed by tumor cells.

Conclusions. Results indicate that the in vivo tumor growth  in its tissue-specific humoral environment may have different kinetics than in the in vitro standard culture media.

 

  

Keywords: CC531, vaccine, lymph


00-08

New experimental tumor vaccine- immunization with NaCl  mummified CC531 cells evokes host reaction against live tumor cells

Waldemar Olszewski1, Marzanna Zaleska1, 2

1Central Clinical Hospital, Warsaw, Poland
2Medical Research Center, Warsaw, Poland
3Central Clinical Hospital, Warsaw, Poland
4Medical Research Center, Warsaw, Poland
5Medical Research Center, Warsaw, Poland
6Medical Research Center, Warsaw, Poland
7Medical Research Center, Warsaw, Poland
8Medical Research Center, Warsaw, Poland
9Medical Research Center, Warsaw, Poland
10Medical Research Center, Warsaw, Poland
11Medical Research Center, Warsaw, Poland
12Medical Research Center, Warsaw, Poland
13Medical Research Center, Warsaw, Poland
14Medical Research Center, Warsaw, Poland

Background. The CC531 tumor cells are rat colon adenocarcinoma  cells. They grow in vitro  forming monolayers. Inoculated intra-peritoneally, intraportally or subcutaneously they form nodules within 14 to 21 days. The tissues surrounding tumor nodule become infiltrated by host mononuclear cells. In advances stages fibrous wall forms around the nodules and central parts become necrotic. Since these is a local cellular response to the CC531 line, the idea arouse to locally immunize tissues by inoculation of non-dividing metabolically inactive tumor cells. Immunization raises memory cells and the subsequently inoculated live CC531 cells should be destroyed in a host memory response mechanism. We described previously that cells as lymphocytes  and monocytes or isolated tissue cells preserved in anhydric pulverized sodium chloride (NaCl) become metabolically inactive, however, they retain their molecular structure intact (Transplantation 2006; 81:1583-1588) and can be used as immunogens. Much the same applies to the isolated tumor cells.

Aim. To immunize tissue, prior to implantation of live CC531, with the NaCl mummified (non-diving, metabolically inactive) CC531 cells and observe kinetics of growth of the implanted live CC531

Material and methods. LEW inbred rats were used. In group 1, 107 live CC531 cells were injected subcutaneously into right paw. Follow up period was 14 and 21 days. In group 2, 107 CC531 cells preserved for 7 days in pulverized NaCl were desalinated and implanted subcutaneously as in group 1. Seven days later 107 live CC531 were implanted into the same paw.  In both groups at the end of the 3rd week tissues with implanted tumor cells and regional lymph nodes were harvested. Specimens were stained with H-E and monoclonal antibodies against CC531 cells, MHC class II (OX6), mononuclear cells (ED1) and W3/13' (lymphocytes). Histological evaluation included: central necrosis area (compared to whole CC531 cell occupied area), density of host cells infiltrating tumor periphery (cells/sq.cm), thickness of fibrotic wall around tumor, number of CC531 cells in cortical area of lymph nodes

Results.Topical immunization of rat paw tissues with NaCl preserved CC531 tumor cells prior to inoculation of live CC531 cells brought about: 1. Dense host cellular infiltrates and focal tumor necrosis, 2. Infiltrates composed of host ED1, OX6 and W3/13 cells seen around and within tumor foci. 3. A 50-100 µ tihick fibrous wall formed around tumor foci. 4.Only few metastatic CC531 cells seen in cortical areas of popliteal  lymph nodes.

Conclusions. The NaCl-mummified CC531 tumor cells do not proliferate after implantation, and when implanted into recipients tissues, they evoke host immune reaction resulting in necrosis of subsequently implanted live CC531 cells.

Keywords: CC531, lymph, metastases, vaccine


00-09

Minimally Invasive Groin Dissection: Clinical Series comparing Video Endoscopic and Robotic Inguinal Lymphadenectomy in Melanoma

Jiashou Xu1, Ajay Upadhyay2, Steven Stanten2, Giovanni Begossi2

1Minimally Invasive/Bariatrics Fellowship Alta Bates Medical Center, Oakland, United States
2First Surgical Consultants Alta Bates Medical Center, Oakland, United States

Background

The groin is the site of metastasis in approximately 15-18% of melanoma of the trunk and lower limb. Superficial groin dissection remains standard of care and carries a significant morbidity rate. In order to minimize the morbidity of radical groin dissection, a minimally invasive approach has been described (video endoscopic inguinal lymphadenectomy: VEIL). VEIL has been shown to be safe with adequate oncologic results. More recently, robotic-assisted video endoscopic inguinal lymphadenectomy (RAVEIL) has been successfully described for urologic malignancies. This study is a limited clinical series comparing VEIL and RAVEIL in cutaneous melanoma patients.

Materials and Methods

Our series reviewed six patients that underwent minimally invasive inguinal lymphadenectomy between 1/1/2011 and 1/1/2016.  Patient characteristics and surgical indications were analyzed. Patient charts were reviewed for perioperative adverse events, conversion to unplanned procedure, complications, pathologic outcomes, length of stay, and melanoma recurrence.

 

Results

Three patients had undergone VEIL and three had RAVEIL. The average age of patients was 61.  Indications for VEIL were positive sentinel node in 1 patient and delayed recurrence of limb melanoma of unknown origin in one patient. Indications for RAVEIL were positive sentinel node in 2 patients, and delayed groin recurrence in one patient. There were no intraoperative complications or conversions to open. All patients were discharged on postoperative day 1. Two patients in the VEIL group had a seroma managed with aspiration and one patient in the RAVEIL group had cellulitis with seroma managed conservatively. One patient in the RAVEIL group had a limited skin necrosis managed conservatively. One patient in each group had limited lymphedema.  Median lymph nodes yield from VEIL was 5 and 9 for the RAVEIL. Two patients in the VEIL group had local recurrence: one above the inguinal ligament 6 months after procedure and one above the Scarpa’s fascia 10 months later. No recurrence has been shown in the RAVEIL group.

 

Conclusions

Minimally invasive approach to the groin for melanoma has limited morbidity with very limited impact on patient quality of life. Both VEIL and RAVEIL are safe procedures. While it is too early to draw conclusions in term of oncologic results, RAVEIL appears definitely to be more effective in nodal dissection in terms of removing the Scarpa’s fascia and dissection above the inguinal ligament.

 

Keywords: minimally invasive, inguinal, lymphadenectomy, robot


00-10

Guided Lymphangiogenesis for the Treatment of Lymphedema. Pilot Study Results

Michael Paukshto

Fibralign Corporation, Union City, United States

To address the limitations of current treatments for secondary lymphedema, our study group developed an experimental surgical procedure based on Autologous Lymph Node Fragment (ALNF) transfer supplemented by nanofibrillar collagen scaffold with and without autologous Adipose Derived Stromal Cells (ADSCs). The efficacy of this scaffold was demonstrated before by histological, functional and imaging analyses in a porcine model of secondary lymphedema. The use of ADSCs has major potential demonstrated in preclinical and clinical studies, but their use is often hampered due to difficulties in harvesting and delivery. The cell injections require large dosages and still have limited effect due to poor survival and migration from the target site. We address these challenges by using ADSC-seeded scaffolds to deliver the cells, support cell survival, maintenance and function precisely at the targeted site. The ongoing pilot study has 8 patients currently enrolled. The ALNF transfer was performed in all patients and additionally 6 patients received the collagen scaffolds alone and 2 patients received the ADSC-seeded scaffolds. Lymphoscintigraphy was conducted before the treatment, 6 and 12 months after the surgery. Volume and bioimpedance measurements (L-Dex U400) were conducted for 0, 3, 6, and 12 months after surgery. A portable closed-loop Mini-Stem system was used for extraction of ADSCs from patient lipoaspirate. Also, a 1.7 ml trocar was used for cell seeding on the scaffolds (~2 hours in incubator) and for subcutaneous delivery to bridge the area of impaired lymphatics. While this is a small ongoing study, safety has been demonstrated as there has been no complications reported over the 12 months. The affected/unaffected limb volume ratio was reduced on average 20%. For two patients, the affected/unaffected limb volume ratio was reduced to nearly normal level. More data will be presented at the time of the conference. While vascularized lymph node transfer is considered to be a more advanced technique than ALNF transfer, there is a great interest in developing countries to have a simpler surgery that could help patients with lymphedema. On the other hand, if the current treatment improves lymphedema, the concept of guiding lymphangiogenesis with collagen scaffolds could potentially improve the efficiency of established vascularized lymph node procedures.

Keywords: secondary lymphedema, lymphangiogenesis, stromal cells, scaffold


00-11

High-resolution 3D fluorescence microscopy of immune checkpoints in melanoma tissue sections

Colin Comerci1, Mehdi Nosrati2, Mohammed Kashani-Sabet2, Stanley Leong2, W. E. Moerner3

1Biophysics Program, Stanford University, Stanford, United States
2Center for Melanoma Research and Treatment, California Pacific Medical Center Research Institute, San Francisco, United States
3Department of Chemistry, Stanford University, Stanford, United States

Background: Immune checkpoint inhibitors, such as anti-PD-1 and anti-CTLA-4, have dramatically improved clinical outcomes for various classes of cancer patients.  However, many questions remain as to why only certain patients respond to such therapies and when combination therapies should be employed.  There is growing evidence that the sub-cellular spatial organization of immune receptors and signaling molecules play an important role in regulating immune cell signaling.  We are working to better understand the nano- to microscale localization of PD-1 on various classes of immune cells in human melanoma tissue sections. 

Materials and Methods: We are labeling human melanoma tissue sections using immunofluorescence.  These tissue sections are then imaged using laser scanning confocal microscopy (LSCM) and Stimulated Emission Depletion (STED) microscopy.   The resolution of LSCM is limited by diffraction to ~300 nm, allowing us to study both the distribution of PD-1 expression on specific subpopulations of lymphocytes throughout the tumor, as well as the sub-cellular 3D architecture of PD-1.  On the other hand, our custom-built STED microscope allows us to surpass the diffraction limit, obtaining a lateral resolution of ~65 nm.  This allows us to study the nanoscale localization of PD-1 in single cells.  Importantly, the images from both techniques provide high-resolution information about PD-1 and its interactions with other receptors inside the tumor microenvironment.

Results: We have successfully imaged fluorescently labeled PD-1 in human melanoma tissue sections.  Preliminary results suggest PD-1 forms microclusters, similar to what has been previously observed for the T cell receptors.  At the same time, with our high spatial resolution, PD-1 expression is highly heterogeneous across the tumor.  Finally, experiments are in progress to test if PD-1 colocalizes with other immune receptors.

Conclusions: Probing the sub-cellular localization and clustering properties of receptors may prove crucial in understanding and improving immune checkpoint inhibitor therapies.  We have successfully imaged PD-1 using 3D fluorescence microscopy in human melanoma tissue sections, showing PD-1 forms microclusters in the tumor microenvironment.

Keywords: Melanoma; fluorescence microscopy; STED microscopy


00-12

Is every thick melanoma equal?

Meng-Ting Peng, John Wen-Cheng Chang

Chang Gung Memorial Hospital, Hematology and oncology department, Taoyuan, Taiwan

Background: Breslow thickness is an important prognostic factor in malignant melanoma. Thick (>4mm) melanoma without nodal metastasis was classified as stage III in AJCC 5th edition and then revised as stage II in AJCC 6th and 7th edition because of anatomic compartmentalization. Tumor thickness greater than 4 mm was common in acral lentiginous melanoma in Taiwan. The prognosis of these “superthick” melanoma in Asia have not been widely discussed. Here we exam the prognosis of tumor thickness beyond 4-mm.

Materials and Methods: We retrospectively reviewed our databank of non-metastatic melanoma between 2001 and 2010 at Chang Gung Memorial Hospital, Linkuo. A total of 209 patients with non-metastatic melanoma were identified. The second cohort with 181 patients, registered in 1990-2000, was combined into analysis later to verify our findings. Data was collected from primary chart review.

Results: The median overall survival (mOS) of node-negative melanoma was better than node-positive melanoma (p=0.064). However, a significantly shorter OS was identified in T4N0 group, compared with T1-3N1 group. (p=0.02) We suggested that a subgroup of T4N0 had a worse outcome and might be classified as stage III, instead of stage IIC in AJCC 7th tumor staging. When 15mm in thickness as the cut-off value, the median OS of thickness >15mm is significantly worse than thickness ≤15mm in T4N0M0 group. (p=0.025) The median OS of T4(>15mm)N0M0 group is significantly worse then T1-3N1M0 group. (p=0.011) When the two cohorts (registered in 1990-2000 and in 2001-2010) were combined into analysis, the median OS of T4N0 (>15mm) was still significantly worse than T4N0 (≤15mm).

Conclusions: A subgroup of thick melanoma, with thickness greater than 15mm and no nodal metastasis, should be classified as stage III. Further validation in a larger cohort followed by amendment of melanoma staging in Asia is necessary. 

Keywords: melanoma, thickness


00-13

Number of Removed Sentinel Lymph Nodes Does Not Differ between Tc-99m Sulfur Colloid and Tc-99m Tilmanocept in Breast Cancer

Jonathan Unkart, James Proudfoot, Anne Wallace

University of California, San Diego, San Diego, United States

Introduction: Dual-agent mapping for sentinel lymph node (SLN) biopsy generally involves a radiotracer and a dye. Tc-99m sulfur colloid (TcSC) and Tc-99m tilmanocept (TcTM) are the two most common radiopharmaceutical agents used in the United States for SLN biopsy in breast cancer. However, limited data exists that compares operative performance characteristics between the two radiotracers. We sought to compare the number of surgical removed nodes between patients undergoing SLN with TcSC or TcTM.

Methods: We retrospectively reviewed the use of TcSC and TcTM in clinically node-negative breast cancer patients at our institution since 2010. Patients receiving TcSC typically received 2-4 intradermal peritumoral injections of 0.5mCi on day of surgery or 2.0mCi on day prior to surgery. Patients receiving TcTM received a single intradermal peritumoral injection of 0.5mCi on day of surgery or 2.0mCi on day prior to surgery. Intraoperatively, patients received 2-3ml of isosulfan blue dye immediately prior to incision. The mean number of removed SLNs between TcSC + blue dye and TcTM + blue dye was compared with a t-test. The proportion of node-positive patients was assessed with a chi-squared test. A multivariate negative binomial count model was constructed to assess patient and surgical covariates on number of removed nodes.

Results: Since 2010, 1062 clinically node-negative patients underwent SLN biopsy with TcSC + blue dye (n=522) or TcTM + blue dye (n= 540). Overall, the mean number of removed SLNs was 3.0 (SD 1.8). Additionally, 162 patients (15.8%) had at least one positive SLN. The mean number of nodes did not differ between TcSC + blue dye (2.97) vs. TcTM + blue dye (2.94), p = 0.80. The proportion of node-positive patients did not differ between groups, TcSC + blue dye (16.1%) vs. TcTM + blue dye (15.6%), p= 0.88. On multivariate modeling, patient age, surgeon and presence of positive nodes were significantly associated with the number of removed SLNs, however, radiotracer agent did not affect the number of removed SLNs.

Discussion: The number of removed SLNs did not differ between radiopharmaceutical agents in clinically node-negative breast cancer patients. The dual technique of radiopharmaceutical plus blue dye with these two pharmaceutical appear statistically equivalent. Further study is needed to compare the efficacy of the radiopharmaceuticals when used as a single agent.

Keywords:


00-14

Endocardial metastasis from hepatocellular carcinoma to the right atrium

Mayank Patel, Natasha Rai, Kim Lam, Vikas Mehta, Malek Massad, Pam Khosla

Mount Sinai Hospital, Chicago, United States

Background

An uncommon finding in otherwise common hepatocellular carcinoma (HCC) is metastasis to the endocardium. Frequent sites of metastasis include invasion into vascular system, lungs, bone, lymphatics and brain. However, endocardial involvement rarely develops in patients with HCC and carries a poor prognosis. Cardiac metastasis may result from hematogenous spread, direct invasion, or tumor spread via inferior vena cava to the right atrium. We report a case of a 73 year old female patient with unexpected finding of isolated endocardial metastasis from HCC to the right atrium due to HCC. Written informed consent was obtained from the patient’s family.

Materials and Methods

A 73 year old female with PMH of atrial fibrillation, CKD hospitalized for light headedness found to have AKI on CKD, UTI and right atrial masses. Physical exam revealed patient hemodynamically stable with no murmurs, jaundice, or tenderness to palpation.

Laboratory results significant for WBC 4.8, Hb 10.2, Plt 68, Cl 115, Cr 2.0, BUN 32. Coagulation shows PT 23.7, INR 2.1 (4.63 on admission), PTT 35.2. LFTs; Total bilirubin 2.2, AST 43, ALT 32, Alb 2.4. Other studies showed reactive HCV, HCV RNA quantitative 415315, gamma globulins 1.8, and normal AFP at 5.56.

Echocardiogram revealed 2-3 mobile atrial masses with greatest diameters of 3.06  and 3.45  cm. These appeared attached to the interatrial septum. Mild-moderate tricuspid valve regurgitation. LVEF normal at 65-70% with No regional wall motion abnormalities. Left and right coronary angiogram were performed and showed no significant coronary artery disease

U/S abdomen showed an ill-defined hypoechogenic liver lesion. CT chest/abdomen revealed nodular contour of the liver (highly suggestive of cirrhosis with moderate ascites). Median sternotomy with cardiopulmonary bypass and right atrial masses excision was performed and samples sent to pathology.

Results

Histopathologic examination showed a tumor arranged in trabecular pattern. The individual tumor cells showed large vesicular nuclei with prominent nucleoli and abundant amount of eosinophilic cytoplasm. Findings were consistent with poorly differentiated neoplasm on frozen section with final pathology results revealing metastatic Hepatocellular Carcinoma.  The tumor cells were positive for Heppar-1 and were negative for CK7, Alpha feto protein, vimentin, actin, desmin, cytokeratin immunohistochemical stains, confirming the diagnosis.

Conclusions

Only 5% of all cardiac metastasis involve the endocardium. Usually these are larger tumors in the liver. The intracavitary lesion can lead to tricuspid obstruction or tumor emboli.. Typically culprits are  renal  carcinoma and hepatocellular carcinoma. Up to 40 percent of small HCCs can have normal AFP in serum.

Our case is unique in that isolated endocardial metastasis with a small primary HCC unilobar seen only on ultrasound and not CT. Our patient is recovering post operatively currently with the support of plasma and platelet transfusions. Overall prognosis is poor because of underlying cirrhosis and vascular metastasis. Targeted systemic antiangiogenic therapy and local therapy will be tried once patient recovers from surgery in addition to hepatitis C treatment may be contemplated. In conclusion, we report a rare finding of isolated metastasis of HCC to the endocardium.

Keywords:


00-15

Optimizing physiological and mechanical function of the local lymphatic system promotes fluid homeostasis and mitigates protracted changes to the solid phase in scondary lymphedema.

Julia Osborne

Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States
Oncology Rehab, Inc., Centennial, United States

Treatment of lymphatic injury with breast cancer lymph node dissection (BrLND) must include territory-based restoration of the physio-mechanical function of the lymphatic system, and restoration of fluid homeostasis, so as to mitigate protracted changes to solid phase lymphedema.

Physio-mechanical function is optimized by superficial AND subfascial lymphatic treatment. Superficial and subfascial treatment allows increased fluid flow into initial lymphatic (ILs) and powerfully reduces both superficial edema back-flow and subfascial overload.

Subfascial treatment reduces lymphostasis in joint/muscle complexes and reduces occlusion pressure on perforating lymph vessels linking subfascial and superficial systems.

Superficial treatment exerts radial tension on anchoring filaments attached to endothelial cells of the initial lymphatics (ILs). Radial tension is proportional to diameter of IL lumen dilation and widening of junctions between endothelial cells through which lymph fluid enters the lymphatic system.

Territory-based fluid homeostasis is assisted by removal nitric oxide, which disables contractility of collector vessels, and by transition of pro- to anti-inflammatory cytokines. Transition failure leads to inflammation, proteolytic activity and deposition of adipose tissue.

Purpose of the study is to demonstrate treatment leverage in using integrated manual therapy skills to affect positive outcomes in physio-mechanics of ILs, fluid homeostasis, fluid volume reduction and mitigation of solid phase lymphedema.

Method: 24 month Retrospective BrLND case study of clinical lymphedema to evaluate subfascial neuro-musculo-skeletal approach, integrated with superficial MLD, on the physio-mechanical functioning of the local lymphatic system.

Subfascial treatment includes myofascial trigger point release work, joint mobilization and neural tension mobilization to reduce lymphostasis and occlusion pressure on perforating lymphatic vessels. Superficial treatment includes muscle bending techniques and manual lymphatic drainage to exert radial tension on anchoring filaments attached to endothelial cells of ILs.

Surveillance of limb volume was performed with circumferential measurements using a tape measure to provide a scale for volume reduction; L-Dex bioImpedence measurements provided surveillance of lymph homeostasis and fluid consistency.

Fluid to solid ratio was superimposed on a graph using these two measurement systems.

Conclusion: Integrated treatment interventions positively affected the functioning of the locally impacted lymphatic system, resulting in volume reduction and fluid homeostasis, reduced inflammation, reduced proteolytic activity, reduced deposition of adipose tissue and optimizing physio-mechanical functioning of the local lymphatic system so as to mitigate protracted solid phase lymphedema.

Keywords:


00-16

Interaction of macrophages with apoptotic cancer cells inhibits EMT, migration and invasion of cancer epithelial cells.

Jihee Kang, Yong-Bae Kim, Young-Ho Ahn, Jin-Hwa Lee

Ewha Womans University, Seoul, Korea, Republic of (South)

Background: Apoptotic cell clearance by phagocytes (efferocytosis) is essential in tissue homeostasis. However, the effects of efferocytosis in the multistep process of cancer cell dissemination, leading to cancer metastasis, have not been studied thus far.

Materials and Methods: To determine whether interaction of macrophages with dying cancer cells inhibits epithelial-mesenchymal transition (EMT) in various epithelial cancer cells, and decreases cancer cell migration and invasiveness, conditioned medium (CM) from murine macrophages (RAW, BMDMs and M2-like cells) or human blood MDMs exposed to apoptotic cancer cells was added to target cancer epithelial cells in the absence or presence of TGF-β1. Western blot and Real-time quantitative PCR were employed to determine changes in epithelial and mesenchymal markers. Cell migration and invasion were tested using Transwell chambers. Standard 3D culture was performed.

Results: Condition medium from RAW cells exposed to apoptotic 344SQcells (ApoSQ-exposed CM) inhibited TGF-β1-induced EMT, based on morphological cellular alterations, and EMT marker mRNA and protein expression profiles. We confirmed the anti-EMT effects of various types of apoptotic cancer cell-exposed CM in the human non-small cell lung cancer (NSCLC) cell line A549 and other human cancer cells lines, including breast (MDA-MB-231), colon (COLO320HSR), and prostate (PC3) cancer cells. In addition to RAW cells, CM from ApoSQ-exposed primary isolated mouse bone marrow-derived macrophages (BMDMs), or their IL-4-derived M2 phenotype, substantially inhibited TGF-β1-induced changes in EMT markers in 344SQ cells. CM from blood monocyte-derived macrophages (MDMs) from healthy humans and NSCLC (adenocarcinoma) patients exposed to apoptotic A549 cells (ApoA) showed anti-EMT effects. ApoSQ-exposed CM partially blocked Smad-independent TGF-β1 signaling, including the p38 MAP kinase and Akt pathways, in 344SQ cells. ApoSQ- -exposed CM prevented TGF-β1-induced cancer cell migration and invasion. In addition, TGF-β1-induced gene-level enhancement of cancer stem-like cell markers was reduced by treatment with ApoSQ-exposed CM. Moreover, 3D Matrigel culture confirmed the anti-invasive effect of ApoSQ-exposed CM, which caused 344SQ cells to recover their lost polarity and acinus-like colonies, and caused invasive projection by TGF-β1.

Conclusions: programming macrophages by apoptotic cancer cells might create a tumor microenvironment antagonizing cancer progression.

 

Keywords: apoptotic cancer cells, macrophages, EMT, migration, invasion, cancer cells


00-17

Treatment of regional and distant metastasis of women breast cancer with lymphotropic subcutaneous injections of CMF as a first-line chemotherapy

Nadezhda Gariaeva1, 2, Igor Zavgorodnii1, 2, Konstantin Garyaev1, 2

1Private research company "International Clinical Lymphology Center", LLC, Perm, Russia
2Oncological clinic "Lymphatech", LLC, Perm, Russia

Background

Cancer spreads through the lymphovascular system using the sentinel lymph nodes as the primary gateway to distant sites. Therefore, it is essential to eliminate cancer cells in the very first checkpoints and prevent from travelling further.

Scientists proved that lymphotropic method of treatment allows achieving significantly higher drug concentration (up to 3 times) in the targeted lymph nodes and targeted tissues with tumor than using regimes with intravenous administration of chemo drugs. This method allows decreasing drug dose and applying the drug locally to the affected organ and its lymphatic region, including sentinel lymph nodes.

The paper illustrates successful treatment cases of breast cancer with metastasis in regional lymph nodes with lymphotropic chemotherapy that is a subcutaneous administration of modified cytostatics’ solution to the lymphatic region of breast and metastasis.

Methods and materials

Three cases with female breast cancer with SNL and distant metastasis received CMF (cyclophosphamide, methotrexate, fluorouracil) alone as a first line chemotherapy.

Lymphotropic chemotherapy is a targeted administration of cytostatics to the organ affected by cancer and to SNL. The method is performed as weekly subcutaneous injections  cytostatics in the lymphatic region.

Targeted administration ensures lowering single doses to the following figures (cyclophosphamide – 250 mg, methotrexate – 50 mg, fluorouracil – 200 mg).

Prior to the procedure drug composition is adjusted in order to be pH – neutral and do not significantly affect skin, tissues, interstice and lymphatic region when injected subcutaneously.

CT and lab control tests were performed.

Results

Case 1. Patient Us, female, 50 yo. Right breast cancer diagnosed in may 2013, mts in axillar LN. Neo-adjuvant CMF (200/50/250 mg) twice a week performed prior to radical resection. Histopathological tests after radical resection revealed no mts in regional and axillar LN.

Case 2. Patient St, female, 54 yo. Left breast cancer diagnosed in 2006. Radical resection and radiotherapy successful. Relapse in 2014, mts in liver, lungs, vertebra. CA 153-231 ME/ml. Lymphotropic CMF (200/50/250 mg) weekly started in october 2014 through march 2015. 20 single injections performed. Steady decrease in CA 153, mediastinal LN sizes, liver and vertebral mts achieved.

Case 3. Patient A, female, 65 yo. Breast cancer diagnosed in 2017, mts in regional LN, mts in ilium and pubic bone (pain syndrome). Lymphotropic CMF (200/50/250 mg) weekly started in February 2017. Achieved after 4 injections: instant decrease in CA 153, regional LN, reduction in pain syndrome dut to weakening of metastatic process in bones.

Injection site remained safe and satisfactory tolerance of treatment and good quality of life achieved in all cases.

Conclusion

Lymphotropic chemotherapy can be considered as a mean to prevent spreading of cancer cells through the lymphatic system via SNL and to eliminate primary tumor itself. The following destinations of breast cancer metastasis can be covered with lymphotropic injections of cytostatics: regional metastasis in breast, axillar LN, inguinal LN, supraclavicular and subclavian LN, parasternal LN; distant metastasis in mediastinal LN, lungs, liver.

Keywords: Lymphotropic, subcutaneous, CMF, chemotherapy, breast cancer, lymph nodes metastasis


00-18

Modelization of the procoagulant properties of cancer cells and the alterations of the effectiveness of the anticoagulant agents.

Aurélie Rousseau, Patrick Van Dreden

Clinical Research Department, Diagnostica Stago, , France, Gennevilliers, France

Background: The pathogenesis of the prothrombotic state in cancer is complex and may alter the efficiency of the antithrombotic agents but not all mechanisms are entirely understood.

Aims: In the present study we dissected the mechanisms responsible for the procoagulant activity of pancreas adenocarcinoma cells (BXPC3) and human breast carcinoma cells (MCF7) by thrombin generation assay (TG) in different relevant conditions, and we studied their influence on the antithrombotic efficiency of apixaban, fondaparinux and enoxaparin in a new experimental model.

Methods: Cells were cultured and adhered in 96-well plates and normal platelet poor or rich plasma (PPP; PRP) spiked or not with apixaban, fondaparinux or enoxaparin was added. TG was done with CAT® assay in different conditions of reagents, with or without any anti-tissue factor antibody (anti-TF), or corn trypsin inhibitor (CTI; inhibitor of the contact phase of coagulation). Alternatively spliced TF (asTF), TF activity (TFa) and cancer procoagulant (CP) were also assessed.

Results: The TFa and asTF were found in abundant amounts in BXCP3 than MCF7 cells. The CP levels were higher in MCF7. The BXPC3 amplified TG more than MCF7 did. The anti-TF inhibited TG triggered by BXCP3 and MCF7. The CTI had more pronounced inhibitory effect on TG triggered by MCF7. TG enhancement by BXPC3 and MCF7 was mediated by FVII. Factor XII was more important for TG enhancement by MCF7. Comparison on the basis of IC50 showed that in the presence of BXPC3 or MCF7 the efficiency of apixaban was preserved or partially reversed. Fondaparinux, was more vulnerable to the presence of cancer cells as compared to apixaban. The effect of BXCP3 or MCF7 cells on the antithrombotic potency of enoxaparin was of similar magnitude as that on apixaban.

Conclusion: The mechanism of activation of blood coagulation by the BXPC3 is dominated by the TF pathway, MCF7 additionally imply also FXII activation. The type of cancer cells is determinant for the antithrombotic efficiency of the specific factor Xa inhibitors. Modeling procoagulant profile of cancer cells provides an understanding of the procoagulant mechanisms and could evaluate the efficiency of antithrombotic treatment.

 

Keywords: Hypercoagulation, thrombin generation, tissue factor


00-19

WITHDRAWN

Yogesh R. Suryawanshi, Tiantian Zhang, Helene S. Woyczesczyk, Karim Essani

Laboratory of Virology, Department of Biological Sciences, Western Michigan University, Kalamazoo, United States

Keywords: Oncolytic viruses, Tanapoxvirus, Thymidine Kinase, Monocyte Chemoattractant Protein-1 and Interleukin-2

Friday, April 21, 2017 - Gold Rush Ballroom - 07:30 - 08:30

Industry Sponsored Breakfast Session (Non-CME) - Advanced Melanoma Treatment, Sponsored by Amgen

00-01

Clinical Considerations for Treatment in Advanced Melanoma

Jonathan Zager

Moffitt Cancer Center, Tampa, United States

The program is meant to educate healthcare providers on an advanced treatment option covering efficacy, dosing, administration, and safety information. The session will also include a video demonstration of the injection technique for healthcare providers.

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 08:35 - 10:35

Mini-Symposium (CME) - Breast Cancer Mini-Symposium

00-01

Introduction

S. David Nathanson

Henry Ford Health System, Detroit, United States

Keywords:


00-02

Biology and treatment of metastatic breast cancer based on positive clinical trials

Hope Rugo

University of California, San Francisco, San Francisco, United States

Keywords:


00-03

Lymph Nodes as Distribution Hubs for Systemic Metastasis of Breast Cancer

Markus Brown

Institute of Science and Technology Austria, Vienna, Austria

M. Brown1,2, F. P. Assen2, A. Leithner2, J. Abe3, H. Schachner1, G. Asfour1, Z. Horvath1, J.V. Stein3, P. Uhrin4, M. Sixt2, D. Kerjaschki1

 

1 Clinical Institiute of Pathology, Medical University of Vienna, 1090 Vienna, Austria;

2 Institute of Science and Technology (IST Austria), 3400 Klosterneuburg, Austria;

3 Theodor Kocher Institute, University of Bern, 3012 Bern, Switzerland.

4 Institute for Vascular Biology, Medical University of Vienna, 1090 Vienna, Austria

 

Abstract

            Death of cancer patients is caused by metastases in lung, brain and other organs remote from the primary tumor site. Initially, mammary carcinoma colonize draining sentinel lymph nodes before they seed distant organ metastases and it is debated whether tumor subclones with metastatic potential enter the blood stream via lymph nodes and/or vessels within the primary tumor. Here we have established a novel mouse model to determine the ability of tumors to disseminate via the draining lymph node. Recapitulating the route of tumor cells into human sentinel lymph nodes, mammary carcinoma cells were micro-infused into the afferent lymphatic vessels of popliteal lymph nodes, which directly guided them into the subcapsular sinus. Within 3 days tumor cells had infiltrated the paracortex, attached to and intravasated into blood vessels and seeded lung metastases without the involvement of the thoracic duct. Remarkably, the lymph node stroma was exceedingly more efficient in seeding distant metastasis than the orthotopic environment of the mammary fat pad, implying a central role in the process of systemic tumor cell dissemination.

Keywords:


00-04

What have we learned from the breast cancer sentinel node clinical trials?

David Krag

University of Vermont, Burlington, United States

Although some of the more prominent trials first come to mind such as such as ACOSOG Z0011, NSABP B-32, and AMAROS, published articles from around the world describe 46 different randomized trials related to sentinel nodes and breast cancer.

This presentation will describe selected aspects of what we have learned from these randomized trials. The most important thing is that the surgical community has not sufficiently come to grips with conducting clinical trials. It is easy to critique the shortcomings of individual trials. The more important question is why there are shortcomings in the first place. The worldwide community should come together to make sure that the important questions are asked and definitively answered. The results from an effective clinical trial are not controversial.

The methods to conduct clinical trials are remarkably simple. This starts with an important clinical question that if answered will lead to better care. The parameters related to answering the clinical question involve basic statistics. The statistics should be sufficiently sophisticated but should also be intuitively clear to all participants. Then there needs to be a commitment by participants to conduct the trial. Our surgical societies need to be much more prominent in setting the standards that inspire the membership to enter patients in clinical trials. Effective tools that describe methods for training and quality control of surgical trials exist and are not that complicated nor expensive. Dissemination of the results should be so clear and transparent that it is understandable after reading the article once. The results from an effective clinical trial are not ambiguous.

The published articles that report randomized clinical trials for sentinel nodes and breast cancer can be found at www.TreeofMedicine.com . If you do not already have access to the database go to the Tree of Medicine website and click on “Topics” located in the top right of the home page. Then click on Sentinel Node and Breast cancer and navigate to and click on “Sign up for free”. Once in the data base look under the first level heading “Randomized trials”. 

Keywords:


00-05

Targeted Axillary Dissection (TAD) for Breast Cancer: Less is More

Henry Kuerer

Univeristy of Texas MD Anderson Cancer Center, Houston, United States

There has been continued interest by several groups in expanding the role of sentinel lymph node dissection (SLND) for biopsy confirmed node-positive patients who receive neoadjuvant chemotherapy as a way to restage the axilla in hopes of sparing women who convert to node-negative status from the morbidity of complete nodal dissection although previous trials have shown higher than desirable false-negative rates (FNR). While sentinel lymph node dissection alone may not accomplish this goal, there are novel techniques, such as TAD which utilizes a 125I radioactive seed localization of known previous marked nodal metastases with concurrent SLND. Placement of a radioiodine seed in known axillary metastatic nodes was originally described by investigators in the Netherlands. Multiple techniques have been described using tattooed nodes and wire-localization techniques.  Caudle et al. described the initial trial results from MD Anderson using targeted axillary dissection in the Journal of Clinical Oncology in 2016 and the technique now allows for reliable nodal staging after chemotherapy.  Of 208 patients enrolled, 191 underwent axillary lymph node dissection (ALND) with residual disease identified in 120 (63%). The clipped node revealed metastases in 115 resulting in a FNR of 4.2% (95% CI 1.4-9.5) for the clipped node.  In patients undergoing SLND and ALND (n=118), the FNR was 10.1% (95% CI 4.2-19.8). Adding evaluation of the clipped node reduced the FNR to 1.4% (95% CI 0.03-7.3) (p=0.03). The clipped node was not retrieved as a SLN in 23% (31/134) including 6 with negative SLNs but metastasis in the clipped node. This explains why it is essential to identify, excise, and examine the specific biopsied initial node with the documented metastases for this technique to an accurate staging method. Updated results at the 2017 SSO Symposium were presented by Caudle et al and TAD followed by ALND has now been performed in 176  patients with a FNR of 2.4% (3/127, 95% CI 0.05-6.8). As a result of these trial results we selectively utilize TAD and perform completion axillary dissection only when metastases is identified in the clipped and or positive SLN.  Selection of appropriate patients for this new technique will be presented.

Keywords:


00-06

Molecular heterogeneity of breast cancer in different racial groups

Lisa Newman

Henry Ford Health System, Detroit, United States

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 11:10 - 12:20

Plenary Session (Non-CME) - Novel Frontiers in Detecting Cancer Metastasis

00-01

Navigating the immunological microenvironment, in situ, to uncover the hidden clues of cancer immunology

Edward Stack

PerkinElmer, Hopkinton, United States

The tumor microenvironment remains a complex and challenging entity supporting tumor growth, immune escape, and potential paths to metastasis. With novel immunotherapies such as ipilimumab and nivolumab demonstrating benefit, even in the metatstatic context, approaches that direct the patient’s own immune system against tumors are proving valuable.  However, further advances will require a detailed understanding of the tumor microenvironment and characterization of the location and status of immune cells and their interaction with tumor.  This will require methods that provide phenotyping of immune and cancer cells combined with the cytoarchitectonics of the tumor. To achieve this, a practical method for simultaneous immunohistochemistry of up to 6 biomarkers in a single tissue section using standard unlabeled primary antibodies will be described. In addition, approaches for multiplexed imaging, single cell quantitative analysis, automated phenotyping and bioinformatics that enable new insights into cancer biology and the tumor microenvironment will be presented. These will then be leveraged in analyses of multiple cancer types, where it is established that the host-tumor interaction is complex and difficult to characterize with standard immunohistochemistry or flow cytometry. But by leveraging multiplexed IHC in situ, we will demonstrate the unique spatial relationships of immune phenotypes in and around both epithelial and non-epithelial tumor types, and show how this data has the potential to form the basis of assays that can guide therapy and monitor response. Further data regarding functional assessment of regulatory immunologic co-factor expression will be discussed.

Keywords:


00-02

Cancer Neoantigens: Targets for Immunotherapy

Mojca Skoberne

Gritstone Oncology, Cambridge, United States

Genetic instability is a hallmark of cancer and, consequently, each cancer patient’s tumor genome is different from their normal cells. The DNA difference leads to the creation of new protein antigens that are specific to tumor cells. These new antigens, known as tumor-specific neoantigens (TSNAs), can be recognized and targeted by the immune system.

Gritstone Oncology is exploiting this inherent vulnerability of tumor cells by identifying a patient’s unique set of tumor antigens and using them in a therapeutic immunization strategy. The company’s approach involves three key steps: 1) Evaluating each patient’s tumor using real-time next-generation sequencing and custom bioinformatics to identify and catalog the individual TSNAs, 2) Using proprietary algorithms to identify the lead candidate TSNAs predicted to activate tumor-specific T cells, 3) Delivering personalized synthetic TSNAs in a vaccine base to patients in combination with immune checkpoint blockade.

Keywords: neoantigens, immunotherapy, vaccine, cancer vaccine, antitumor immunity


00-03

Novel platform for miRNA profiling directly from biofluids using FirePlex® Technology

Daniel Pregibon

Abcam Inc., Cambridge, United States

To address the need for circulating miRNA biomarker discovery and verification, we developed the FirePlex® microRNA assay. This assay enables multiplex detection of up to 65 microRNA targets per sample in 96-well format with readout on standard flow cytometers and analysis with an included bioinformatics software package. This assay combines particle­-based multiplexing, using patented FirePlex® hydrogel particles, with single­ step RT-PCR signal amplification using universal primers. Thus, the FirePlex microRNA assay leverages PCR sensitivity while eliminating the need for separate reverse transcription reactions and mitigating amplification biases introduced by target­-specific qPCR. The assay can reliably detect as few as 1000 microRNA copies per sample with a linear dynamic range of ~5 logs, and without the need of prior RNA purification, making the assay ideally suited for profiling in serum, plasma or urine. Furthermore, the ability to multiplex targets in each well eliminates the need to split valuable samples into multiple reactions. Results from the FirePlex microRNA assay are displayed and interpreted using our included Firefly Analysis Workbench, which allows visualization, normalization, and export of experimental data with only a few mouse clicks.  To aid discovery and verification of biomarkers, we have generated fixed panels for Oncology, Cardiology, Neurology, Immunology, and Liver Toxicology. These carefully curated panels include hemolysis markers to assess sample quality, as well as critical normalization factors.  Here we present the data from several studies investigating circulating and tumor microRNA profiles using the FirePlex microRNA Assay. Together, this novel combination of bioinformatics tools and multiplexed, high­-sensitivity assays enables rapid discovery and validation of microRNA biomarker signatures from fluid specimens.

Keywords:

Friday, April 21, 2017 - Crystal Ballroom - 12:20 - 13:30

Industry Sponsored Lunch Session (Non-CME) - Checkpoint Inhibitors, Sponsored by Bristol-Myers Squibb

00-01

Dual Immune Checkpoint Inhibition for the First Line Treatment of BRAF Wild Type and BRAF Mutant Metastatic Melanoma.

Steven O'Day

John Wayne Cancer Institute, Santa Monica, United States

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 13:35 - 15:05

Plenary Session (CME) - Advances in Cancer Treatment

00-01

Treatment of intransit metastatic melanoma

Jonathan Zager

Moffitt Cancer Center, Tampa, United States

Metastatic melanoma in a locoregional fashion is a difficult clinical scenario to treat. Presentations vary widely. Treatments range from surgical excision to intra-arterial therapy such as isolated limb infusion and isolated limb perfusion, intralesional therapy with oncolytic viruses such as TVEC, systemic therapy's with immunotherapy or targeted therapy also play a role. Decision making is based on age of the patient, medical comorbidities, burden of disease, available treatments, molecular analysis of the tumor for mutations and a strong and close patient and physician relationship. This talk will discuss the available options to treat locoregional metastatic melanoma.

Keywords:


00-02

Latest development in targeted therapy in solid tumors

Kevin Kim

California Pacific Medical Ctr, San Francisco, United States

Since development of trastumumab for HER-2/neu amplified breast cancer and imatininb for chronic myelogenous leukemia harboring a bcr-abl translocation, a number of targeted drugs have been approved by the Food and Drug Administration in the United States because of superior tumor control and/or prolongation of overall survival compared to the conventional cytotoxic chemotherapy drugs. These include EGFR inhibitors for EGFR mutant lung cancer, ALK translated lung cancer, BRAF inhibitors for V600 BRAF mutant melanoma, PARP inhibitors for BRCA-mutant ovarian cancers among others.  The success of targeted therapy depends on identification of relevant functional genetic and epigenetic alterations and development of potent selective drugs against these genetic / molecular targets. Although an increasing number of targeted drugs are being developed against these known and/or newly discovered targets, a majority of patients with solid tumor do not benefit from these drugs due to emergence of tumor resistance to the drugs or a lack of actionable targets.  To overcome these clinical dilemma, close collaborations between basic scientists and clinicians will be absolutely essential.

Keywords:


00-03

Extended survival time correlates to immunological changes in circulating lymphocytes of patients with pancreatic cancer after NK cell-enriched cell therapy: A retrospective study.

Akihiro Shimosaka, Junichi Masuyama

New City Osaki Clinic, Tokyo, Japan

Background

Pancreatic cancer (PC) is one of the malignancies with the worst prognosis because of rapid metastasis as well as invasion to surrounding tissues even with chemotherapy. Cellular immunotherapy is a candidate to improve the difficulty of treatment for PC.  We retrospectively studied the overall survival (OS) of PC patients treated with a combination of chemotherapy and NK cell-enriched immunotherapy (NK-CT) and the relationship between OS and immunological changes in peripheral blood lymphocytes (PBL) before and after the immunotherapy. 

 

Patients and Methods

We studied 77 patients with advanced PC received NK-CT in our clinic since 2004.  In this study, out of all PC patients, 46 patients with ECOG Performance Status 0-1 were retrospectively analyzed to determine the clinical outcome and the immunological changes of PBL after NK-CT.  Median age was 65 years, and 74% of them had distant metastases.  Almost all cases received chemotherapy with gemcitabine and/or S-1.  NK-CT was carried out average 10 times every two weeks with chemotherapy in parallel.  NK cells were expanded by a culture method using CD52-costimulation of peripheral blood mononuclear cells (PBMC) isolated from cancer patients (Cytotherapy 18:80, 2016).  Muromonab and alemtuzumab were used for costimulation of PBMC.  Immunological changes of PBL were assessed using flow cytometry.

 

Results

The estimated OS was 15.5 months and the 1-year survival rate was 63%, and the median survival time after NK-CT was 11.0 months.  NK-CT was well tolerated by all patients without treatment-related toxicities excluding transient fever (~38oC).  We investigated the effect of infused NK cells on clinical outcome and the relationship between immunological changes of PBL and survival time after NK-CT.  The result suggests that the proportion and number of infused NK cells do not correlate to the length of survival. However, when 46 patients were divided into the short-term survivors (<11 months, median 6.1 months, n = 19) and the long-term survivors (≧11 months, median 17.4 months, n = 27) after NK-CT, we found a significant increase in NK cell cytotoxic activity and the numbers of lymphocytes, NK cells, NKG2D+ cells, and a significant decrease in the ratios of CD4/CD8 T cells in PBL in the long-term survivors, while there were no significant changes in any indicators in the short-term survivors. 

 

Conclusions

The findings suggest that sustained enhancement of cytotoxic profiles in circulating lymphocytes after NK-CT may contribute to survival extension observed in this study.

Keywords: cellular immunotherapy, NK cell, immune monitering, pancretic cancer


00-04

Novel treatment for metastatic gynecological cancer based on new biology

John Chan

California Pacific Medical Ctr, San Francisco, United States

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 15:05 - 15:35

Keynote Session (CME) - Molecular Imaging of Cancer Metastasis

00-01

Molecular imaging of cancer metastasis

Sanjiv Gambhir

Stanford University, Stanford, United States

Molecular imaging continues to expand with growing numbers of imaging agents and strategies for the study of cancer in the living subject. A background will be provided on how one can take a molecular target, develop imaging agents against that target, and perform multimodality molecular imaging in both preclinical and clinical models. Applications to pre-clinical models of cancer as well as clinical translation will be discussed.  Strategies for imaging with multiple modalities including radionuclide, optical, ultrasound, and photoacoustic imaging will be discussed. Methods to image cancer cells and immune cells including within the lymphatics will be highlighted. Through continued collaborations across multiple fields it should be possible to improve cancer detection and management.

Keywords: multimodality imaging
molecular imaging
cancer management

Friday, April 21, 2017 - Emerald Ballroom - 16:10 - 17:20

Plenary Session (Non-CME) - Translating the Promise of Academic Research into Reality

00-01

Industry Persepctive: David R. Parkinson, MD, President & CEO, Essa Pharmaceuticals, Inc., Vancouver, Canada

David Parkinson

Essa Pharmaceuticals, Inc., Vancouver, Canada

Keywords:


00-02

Academic Perspective: Sanjiv (Sam) Gambhir, MD, PhD, Stanford University, Stanford, USA

Sanjiv Gambhir

Stanford University, Stanford, United States

Keywords:


00-03

Technology Licensing Perspective: Mona Wan, MS, MBA, Office of Technology Licensing, Stanford University, Stanford, USA

Mona Wan

Office of Technology Licensing, Stanford University, Stanford, United States

Keywords:


00-04

Regulatory Perspective

Libero Marzella

FDA, Office of Drug Evaluation IV, Silver Spring, United States

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 17:20 - 17:50

Plenary Session (Non-CME) - Molecular Med. for Monitoring CTC, Sponsored by Clearbridge

00-01

RealTime Personalized Molecular Medicine For Monitoring CTC In Melanoma Patients Receiving CheckPoint Inhibitors

Dave Hoon

John Wayne Cancer Institute, Santa Monica, United States

Blood biopsy in cancer patients has come into clinical fruition in the last several years. The approach provides realtime precision molecular medicine with clinical utility. Blood biopsy can consist of circulating tumor cells(CTC) in which both RNA and DNA biomarkers can be assessed.  The utilization of modern molecular techniques and instruments has recently allowed better quantification and assessment of CTC. Melanoma is highly metastatic cancers whereby we have shown molecular analysis of CTCs are important prognostic factors in early and late stage disease. With the improvement in therapy efficacy in melanoma patients, blood biopsies become a necessary tool to monitor and make clinical decisions more rapidly and efficiently than traditional approaches to provide effective realtime precision medicine treatments. Studies in blood biopsy are moving fast forward although clinical utility still needs to be developed for specific stages and status of cancer patients.  I will discuss our approach of assessing CTC using the ClearCell FX1 microfluidic system using the CTchip FR (Clearbridge BioMedics).   Will demonstrate our approach of monitoring CTC in melanoma patients receiving CheckPoint inhibitors. Downstream assays include multimarker mRNA melanoma genes and BRAF V600E mutation, and cell culturing.  Our studies demonstrate the utility of microfluidic CTC isolation and downstream molecular assays for monitoring melanoma patients

This session is sponsored by: Clearbridge BioMedics

 

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 17:50 - 18:20

Plenary Session (Non-CME) - Immuno- Oncology: A CRO’s Perspective

00-01

Immuno-Oncology: A CRO’s perspective

Jai Balkissoon

PPD, Berkeley, United States

A contract research organization (CRO) is an organization that provides support to the pharmaceutical, biotechnology, and medical device industries in the form of research services outsourced on a contract basis. One global CRO called PPD has taken the initiative to create a formal immuno-Oncology (IO) program to help sponsors develop their IO agents in the most efficient manner. At PPD, with our siginificant experience in IO drug development, we have developed an IO center of excellence (IOCOE) and created a formal IO training program that ensures all team members at both the CRO and sponsor will be properly trained to manage all IO clinical trials. At PPD we have have an understanding of the development strategy for IO agents which includes regulatory considerations, competitive landscape, enrollment challenges and operational considerations. A collaboration between sponsors and a CRO that has a unique understanding of the IO development area can have beneficial outcomes that can impact the succesful delivery of effective IO drugs to patients.

Keywords:

Friday, April 21, 2017 - Emerald Ballroom - 18:20 - 18:40

Plenary Session (Non-CME) - Lab Testing to Complement Immuno-Oncology Ther. for Melanoma

00-01

The Laboratory Testing to Complement Immuno-Oncology Therapy for Melanoma

Thomas Moss

The MT Group, Inc., Van Nuys, United Kingdom

From Anti-PD1 to Anti-CTLA4 to T-cell therapy, immune-oncology is being used to treat a growing population of melanoma patients. Companion diagnostic testing identifies who is eligible for which therapy (e.g. PDL1). Surrogate marker testing is used to determine efficacy of therapy and identify progression (e.g. CTCs, circulating DNA, etc.). This talk will emphasize the current approaches to both of these types of assays in relation to the immuno-oncology therapy for melanoma patients. The talk will also review the importance of sample integrity and how biobanks can solve many of these issues.

Keywords: Companion Diagnostic, CTCs, Liquid Biopsy

Friday, April 21, 2017 - Oregon Ballroom - 18:40 - 19:15

Rapid Fire Abstract Session (Non-CME) - Rapid Fire: Tumor Microenvironment & Cancer Progression

00-01

TGF-b1-induced EMT promotes dendritic cell properties and lymphatic dissemination of cancer cells

Jonas Fuxe1, Mei-Fong Pang2, Nikolina Giotopoulou1, Mikael Karlsson1, Anna-Maria Georgoudaki1

1Department of Microbiology, Tumor and Cell biology Karolinska Institutet, Stockholm, Sweden
2Chemical and Biological Engineering Princeton University, Princeton, United States

Background: The lymphatic system is a major gateway for tumor cell dissemination in breast cancer and many other forms of cancer. Yet, it is not clear how tumor cells find their way into lymphatic vessels and whether this is a regulated process, or more of a stochastic event. Recently, we found that mammary tumor cells undergoing TGF-b1-induced epithelial-mesenchymal transition (EMT) become activated for targeted migration through the lymphatic system via CCR7/CCL21-mediated chemotaxis. Furthermore, we found that tumor cells undergoing TGF-b1-induced EMT acquire properties of immune cells. Based on these results we hypothesize that EMT is an activation step, which activates tumor cells for chemotactic migration towards inflammatory signals, similar to how dendritic cells (DCs) become activated for targeted migration through the lymphatic system during inflammation.

 

Materials and Methods: To expand on this concept we have performed gene-profiling analysis to analyze immune cell features of tumor EMT cells. We have also developed a 3D-based co-culture assay to study the capacity of EMT cells to migrate towards lymphatic versus vascular endothelial cells. In addition, we have used a footpad model in the mouse to determine how DC properties of EMT cells affect their capacity to home to draining lymph nodes.

 

Results: Gene profiling experiments indicate that tumor cells undergoing EMT express a DC signature. Several genes within this signature have previously been associated with DC migration, but not with EMT or lymphatic spread of cancer cells. Knockdown of these genes affect the capacity of tumor EMT cells to migrate towards lymphatic endothelial cells in 3D cultures, and to disseminate through the lymphatic system, in vivo.

 

Conclusions: We propose that breast cancer cells undergoing TGF-b-induced EMT acquire properties of DCs allowing them to migrate in a targeted fashion through the lymphatic system. 

Keywords: TGF-beta; Cancer; Epithelial-Mesenchymal Transition; Lymphatic dissemination; Immune cells


00-02

miR-93 regulates epithelial-to-mesenchymal transition process in metastatic colorectal cancer by targeting EphA4

Sheng Gao1, 2, Bo Jiang1, 2, Haiyi Liu1, 2, Liming Wu3, 4, 5, Zhe Yang3, 4, 5, Junpeng Shen6, Shu-Sen Zheng3, 4, 5, Wenqi Bai1, 2

1Department of Colorectal Cancer, Shanxi Cancer Hospital and Institute, Taiyuan, China
2Affiliated Cancer Hospital of Shanxi Medical University, Taiyuan, China
3Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health, Hangzhou, China
4Key Laboratory of Organ Transplantation, Hangzhou, China
5Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, Hangzhou, China
6Department of General Surgery,Department of General Surgery,Licheng County People’s Hospital, Licheng, China

miR-93 regulates epithelial-to-mesenchymal transition process in metastatic colorectal cancer by targeting EphA4

Sheng Gao1,2*, Bo jiang1,2,*, Haiyi Liu1,2, Shenghuai Hou1,2,Liming Wu3,4,5, Zhe Yang3,4,5, Junpeng Shen6, Lin Zhou3,4,5, Shu-Sen Zheng3,4,5 and Wenqi Bai1,2

Authors’ Affiliation: 1Department of Colorectal Cancer, Shanxi Cancer Hospital and Institute, Taiyuan 030013, China. 2Affiliated Cancer Hospital of Shanxi Medical University, Taiyuan 030013, China.. 3Key Laboratory of Combined Multi-organ Transplantation, Ministry of Public Health. 4Key Laboratory of Organ Transplantation, Zhejiang Province.5Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China.6Department of General Surgery, Licheng County People’s Hospital, Licheng 647600, China.

Correspondence to: Sheng Gao, Department of Colorectal Cancer, Shanxi Cancer Hospital and Institute, No.3 Xinchun Road, Taiyuan 030013, China,, P.R.China. Phone: 86-351-4651225; Fax: 86-351-4651883. E-mail address: gaoshengmail@163.com.

Abstract

Background: Regulating epithelial-to-mesenchymal transition (EMT) in cancer cells has been widely considered as an approach to combat cancer progression and therapeutic resistance, whereas a limited number of broadly comprehensive investigations of microRNAs involved in metastatic colorectal cancer (mCRC) have been conducted. In this study, we investigated the roles and mechanisms of EMT process in mCRC.

Materials and Methods: We used in situ hybridization and quantitative reverse transcriptase polymerase chain reaction to measure expression of miR-93 in colorectal tissues, nontumor tissues and metastatic liver tissues. CRC cell lines were transduced with lentiviruses that expressed miR-93, its inhibitor sequence targeted miR-93 or a scrambled sequence (control); proliferation, metastasis, invasion and colony formation were analysed. We analysed growth of CRC cells that overexpress miR-93 or its inhibitor in severe combined immune-deficient mice. Western blot, and luciferase reporter assays were used to measure expression and activity of Eph tyrosine kinase receptor (EphA4) and related signalling molecules.

Results: In this study, we demonstrated that miR-93 regulated the epithelial-mesenchymal transition (EMT) process by targeting EphA4 in metastasis colorectal cancer (mCRC). We examined the fact that CRC tissues and metastatic liver tissues had increased levels of miR-93 compared with the nontumor tissues and cells, by which we identified miR-93 can regulate EMT process. In addition, overexpression of miR-93 increased proliferation of CRC cells, metastasis, invasion and colony formation in vitro, whereas miR-93 depletion reduced these parameters. In severe combined immune-deficient mice, overexpression of miR-93 by CRC cells increased liver metastasis and overexpression of the miR-93 inhibitor reduced it. By further study the role of miR-93 in EMT and tumor metastasis, we identified by microarray analysis that the direct and functional target genes of miR-93 was EphA4. Knockdown of EphA4 phenocopied the effect of miR-93 and ectopic expression of EphA4 restored the effect of miR-93 on proliferation, migration, invasion and liver metastasis in CRC cells.

Conclusions: Our findings for the first time revealed that miR-93 regulates the epithelial-mesenchymal transition (EMT) process by targeting EphA4 to affect liver metastasis in colorectal cancer (CRC). Mechanistically, miR-93 inhibited tumor metastasis by directly targeting EphA4 which is a crucial factor in regulating EMT. Collectively, this study provide new insights into exploring the therapeutic potential of miR-93 which is able to regulate EMT process to affect tumor metastasis, and warrant further study in clinical settings.

Keywords: Metastatic colorectal cancer, microRNA, EMT, tumor metastasis


00-03

Human lymphatic extracellular vesicles: early modulators of regional lymph node metastasis

Rachel Maus1, James Jakub MD2, Trace Christensen3, Wendy Nevala MS4, Svetomir Markovic MD, PhD4

1Mayo Graduate School Mayo Clinic, Rochester, United States
2Department of General Surgery Mayo Clinic, Rochester, United States
3Microscopy and Cell Analysis Core Mayo Clinic, Rochester, United States
4Department of Oncology Mayo Clinic, Rochester, United States

Background: Lymph node metastasis remains a valuable prognostic marker for many solid tumor malignancies, including melanoma. Mediating the linear progression of early lymph node metastasis, the lymphatic network enables critical communication and content exchange between the otherwise remote sites of the primary tumor and draining sentinel lymph node (SLN). Previous studies have demonstrated Th2 polarization and increased immunosuppression amenable to tumor growth and progression define SLNs in patients with melanoma. Importantly, this profile precedes clinical evidence of metastasis, suggesting the lymphatic microenvironment is initially altered independent of tumor cells. In the current study, we evaluated the role of human lymphatic-derived extracellular vesicles (EVs) isolated from lymph fluid in modulating the immune profile of regional lymph nodes.

Materials and Methods: Fresh SLN tissue and proximal afferent lymphatic channels were surgically excised from patients with primary cutaneous melanoma undergoing sentinel lymph node biopsy. Control lymph node tissue was obtained from patients undergoing prophylactic mastectomy. Lymph node immune cell profiles were comprehensively phenotyped by mass cytometry. Lymphatic fluid was perfused from the afferent channel for EV isolation, visualized by electron microscopy (EM) and quantified by nanoparticle tracking analysis. EV protein cargo was characterized by mass spectrometry and validated by EM utilizing immunogold staining.

Results: Lymphatic EVs were identified by EM to be present within the lumen of the afferent lymphatic channel (Figure 1) and isolated EVs were quantified ex vivo by nanoparticle tracking analysis. Profiling of EV cargo by mass spectrometry identified a conserved signature of 266 proteins including enrichment for known immune modulators AHNAK, Galectin-1 and Annexin A1. Co-localization of these proteins with vesicle marker CD63 on isolated lymphatic vesicles was confirmed by EM following double immuno-gold labelling.

Conclusions: To our knowledge, this pilot study provides the first examination of human lymphatic fluid as a viable bio-fluid for isolating mediators of early lymph node metastasis. Characterization of lymphatic EVs and their associated cargo is providing valuable insights regarding the mechanisms responsible for mediating the pre-metastatic immune profile of the sentinel lymph node.

Keywords:


00-04

The prognostic significance of molecular sentinel lymph node staging by one-step nucleic acid amplification (OSNA) in patients with colon cancer - A multicenter cohort study

Benjamin Weixler1, 2, Ragna van der Linden3, Dimitri Rapits4, 5, Leonora Boogerd1, Alexander Vahrmeijer1, Cornelis van de Velde1, Koop Bosscha3, Markus Zuber5

1Department of Surgery, Leiden University Medical Cetern, The Netherlands, Leiden, Netherlands
2Department of Surgery, University Hospital Basel, Basel, Switzerland
3Department of Surgery, Jeroen Bosch Hospital, s'-Hertogenbosch, Netherlands
4Department of Visceral and Transplant Surgery, University Hospital Zurich, Zurich, Switzerland
5Department of Surgery, Cantonal Hospital Olten, Olten, Switzerland

Background: Sentinel lymph node (SLN) mapping with multilevel sectioning and immunohistochemistry (IHC) or one-step nucleic acid amplification (OSNA) were proposed as promising techniques for better lymph node staging in patients with colon cancer. To the best of our knowledge, this is the first study assessing the prognostic significance of OSNA compared to established investigations with hematoxylin & eosin (H&E) or IHC in patients with colon cancer.

Patients and Methods: Standard oncological resection and SLN mapping was performed from 10/2008 to 12/2013 in 177 patients with stage I-III colon cancer operated at three different institutions internationally. In all patients, SLN were examined for the presence of tumor cells with conventional H&E staining, multilevel sectioning with IHC and OSNA. Kappa statistics were used to evaluate the association (agreement) between positive H&E, IHC and OSNA status. Kaplan-Meier survival and Cox regression analysis were performed to determine the prognostic significance of either technique.

Results: There was only moderate agreement between H&E, IHC and OSNA (IHC and H&E [kappa=41%], OSNA and H&E [kappa=47%] and OSNA and IHC [kappa=38%]). Mean disease free survival  (DFS) was 45.9 months for patients with positive H&E status versus 65.1 months for patients with negative H&E status (p<0.001). Mean DFS for positive vs. negative IHC status was 53.5 vs. 64.1 months (p=0.006) and for positive vs. negative OSNA status 49.2 vs. 63.9 months (p<0.001).  Mean overall survival (OS) for positive vs. negative H&E status was 47.4 vs. 63.7 months (p<0.001), for positive vs. negative IHC status 53.4 vs. 63.1 months ((p=0.011) and for positive vs. negative OSNA status 50.2 vs. 62.5 months (p<0.001). In multivariable analysis, positive H&E status was prognostic for DFS (hazard ratio (HR)=2.56, p=0.048) along with positive OSNA status (HR 2.11, p=0.075) and lymph vessel invasion (HR=2.825, p=0.039). The only prognostic factors for OS were  positive H&E status (HR=2.695, p=0.005) and positive OSNA status (HR=2.038, p=0.044).

Conclusions: This study demonstrates that SLN examination with H&E, IHC and OSNA produces discordant findings in most patients. Although all three techniques could identify patients with decreased survival, the combination of H&E staining and OSNA seems to be best to most accurately predict patient outcome. The clinical implications of this combination (H&E and OSNA) have to be evaluated in future oncological trials.

Keywords: colon cancer, sentinel lymph node, SLN, OSNA, immunohistochemistry, prognostic significance


00-05

Tertiary lymphoid structures: a unique intratumoral immune response in liposarcoma with a potential role in the development of high grade disease

William Tseng1, Susan Groshen2, Sophia Hernandez1, Eric Jung1, Shefali Chopra3

1Department of Surgery, Section of Surgical Oncology, University of Southern California, Keck School of Medicine, Los Angeles, United States
2Department of Biostatistics, University of Southern California, Keck School of Medicine, Los Angeles, United States
3Department of Pathology, University of Southern California, Keck School of Medicine, Los Angeles, United States

Background: The immune response plays a critical role in shaping the tumor microenvironment in many cancers. Tertiary lymphoid structures (TLS) are organized aggregates of immune cells that may be sites of antigen presentation within tumors. TLS have been described in several solid tumor types (e.g. melanoma, NSCLC) and we have previously reported their intratumoral presence in well differentiated / dedifferentiated (WD / DD) liposarcoma (Tseng et al., Am J Surg Path 2012; Tseng et al., Sarcoma 2015). This disease is unique in that it frequently has low grade (WD) and high grade (DD) components juxtaposed in the same tumor. We sought to further characterize TLS in WD / DD liposarcoma.

Methods: Clinically-available H&E slides were retrieved for consecutive cases of WD / DD liposarcoma resected at our institution between 2011-2016. Each case was scored for overall TLS density (none, rare, few, some, many, inflammatory) and immunologic maturity (none, aggregate, follicle, germinal center).  Demographic, clinicopathologic and outcome data were also collected. Associations between the data were analyzed for statistical significance using a two sided Cochran-Mantel-Haenszel test (for ordered categorical variables) and Spearman correlation coefficients (for continuous variables). Clinical outcome data were represented with Kaplan-Meier plots and analyzed using the logrank test.

Results: In total, 43 cases were studied, 51% of which had evidence of high grade disease (DD). The presence of TLS was noted in 72% of all cases, more commonly in DD than WD tumors (82% vs. 62%, p = 0.19). There was no significant association with TLS density scores and histology (WD versus DD); however higher TLS maturity scores were seen in DD tumors compared to WD tumors (p = 0.012). Tumors found in deep body locations (retroperitoneum, mediastinum) had both higher TLS density (p = 0.032) and maturity (p = 0.027) scores compared to tumors in superficial locations (extremity, trunk). After stratifying by histology and then presence of TLS, tumors with TLS were more likely to recur, but this pattern was not statistically significant (p > 0.25); however in a post hoc analysis comparing DD tumors with TLS to all others, this subset was particularly likely to recur (p = 0.002).

Conclusions: TLS are commonly found in WD / DD liposarcoma and may play a role in the disease biology. TLS appear to be specifically associated with high grade disease (DD) and seem to confer a greater likelihood of recurrence. These unique aggregates of immune cells may have potential utility as a clinical biomarker. Further investigation of the intratumoral immune response in WD / DD liposarcoma is warranted. This disease may also serve as a "cancer model" to study the dynamic interaction of tumor- and immune cells in the microenvironment of other solid tumors.

Keywords: Tumor microenvironment; immune response; tertiary lymphoid structure


00-06

 WITHDRAWN

Mustafa Ilhan, Cansu Kucukkose, Ozden Yalcin-Ozuysal

Izmir Institute of Technology, Department of Molecular Biology and Genetics, Izmir, Turkey

Keywords:


00-07

Exploring the oncolytic potential of tanapoxvirus for melanoma virotherapy

Tiantian Zhang

Laboratory of Virology Department of Biological Sciences Western Michigan University, Kalamazoo, United States

Background: Melanoma is one of the most common skin cancers with poor prognosis and survival rate. Oncolytic viruses, with the ability of causing replicative oncolysis and expessing toxic or immunostimulatory genes, are an appealing addition to the melanoma therapy. Tanapoxvirus (TPV) is potentially a great candidate for melanoma therapy, as it is a benign virus that has exhibited tumor regression in human colon cancer xenografted in nude mice. TPV-66R gene encodes thymidine kinase (TK) that is more abundant in cancerous cells, and deletion of viral TK gene has widely been used for increasing the virus tumor-selectivity. Previous studies have shown that the TPV-15L protein functionally mimics human neuregulin (NRG) and that NRG promotes the proliferation of melanoma. We hypothesized that TPV-15L protein would enhance melanoma proliferation, and aimed to generate TPV recombinants with either the 15L gene deleted (TPVΔ15L) or with both the 15L and the 66R gene ablated (TPVΔ15LΔ66R). We sought to determine the oncolytic effectiveness of TPVΔ15L and TPVΔ15LΔ66R in human melanoma.

Materials and Methods: Owl monkey kidney (OMK) cells, human lung fibroblasts (WI38) and human melanoma cell line SK-MEL-3 were used. Construction of the recombinant viruses was done by transfecting the virus-infected cells with the plasmids targeted for gene deletion from the viral genome. Western blot was conducted for determining the protein expression. Melanoma tumors were induced by injecting SK-MEL-3 cells subcutaneously in the nude mice. Treatments consisted of intratumoral injection of viruses when the tumor volumes reached 45 ± 4.5 mm3.

Results: The TPV-15L protein exhibits similar effectiveness as NRG in promoting melanoma growth in vitro. The replication kinetics of TPVΔ15L was similar to that of wtTPV. However, TPVΔ15LΔ66R replicated less efficiently than TPVΔ15L and the parental virus in vitro. TPVΔ15L exhibited more robust tumor reduction in the human melanoma-bearing nude mice than other recombinant TPVs in vivo. Our results indicate that the deletion of the 66R gene, but not 15L gene, adversely affected virus replication, and that deletion of 15L (which elevates melanoma proliferation) enhanced virus oncolytic efficacy. Interestingly, an anti-viral activity, which was identified as Interferon-λ1 (IFN-λ1) was secreted in a significantly higher quantity by the cells infected with TPVΔ15L. We show that IFN-λ exhibits a more pronounced antiproliferative effect in melanoma than IFNα and IFNβ in vitro. Further, we demonstrate that anti-IFN-λ1 exerts an inhibitory effect on melanoma cell apoptosis caused by TPVΔ15L infection, which indicates that TPVΔ15L regresses melanoma growth partially through inducing IFN-λ1 release.

Conclusion: Our results indicate that deletion of TPV-15L gene product which facilitates the growth of human melanoma cells can be an effective strategy to enhance the oncolytic potential of TPV for the treatment of melanoma. In addition, we demonstrate the immuno-modulatory activities associated with TPVΔ15L and suggest further exploration of TPVΔ15L as a melanoma virotherapy.

Keywords: Oncolytic virotherapy, Tanapoxvirus, Melanoma, Neuregulin, Thymidine kinase, Interferon, Viral immuno-modulation

Friday, April 21, 2017 - Emerald Ballroom - 18:40 - 19:15

Rapid Fire Abstract Session (Non-CME) - Rapid Fire: Cancer Metastasis & Treatment

00-01

A genome-wide query of somatic copy number alterations to predict lymph node metastases and survival in patients with muscle invasive bladder cancer

Karla Lindquist1, Thomas Sanford1, Terence Friedlander2, Pamela Paris1, 2, Sima Porten1

1Department of Urology, University of California, San Francisco, San Francisco, United States
2Department of Medicine, Division of Hematology & Medical Oncology, University of California, San Francisco, San Francisco, United States

Background: Patients with muscle-invasive bladder cancer (MIBC) have a poor prognosis if the cancer has metastasized to surrounding lymph nodes (LN). Adding tumor-based genomic tests that improve prediction of LN status and prognosis over clinical variables alone would be useful to clinicians in making treatment decisions, potentially improving outcomes for these patients.

Materials and Methods: We performed a genome-wide query of copy number alterations (CNAs) in MIBC tumors from 237 patients in The Cancer Genome Atlas who had radical cystectomy and lymphadenectomy (≥10 nodes) without neoadjuvant treatment. We independently analyzed pathology reports and copy number data to confirm LN status and gene-level CNAs. Using elastic net and logistic regression models, we sought to identify a set of genes with CNAs that predict LN status. We also tested for association between CNAs and survival. 

Results: We identified 26 genes with CNAs that predicted LN status. Those located on chr3p25 and chr11p11 had gains associated with LN positivity after adjusting for age, gender, race, pathological tumor stage, histology, and number of nodes examined (p=0.03). CNAs at these loci were also associated with one-year survival in the cohort overall (p<0.01), as well as in LN-positive patients after adjusting for node stage, LN density, and extracapsular extension (p<0.01). 

Conclusions: We have identified a small set of genes with CNAs in MIBC tumors that robustly predict LN status and one-year survival. A simple copy number-based test based on these genes could potentially improve preoperative LN status determination and help inform adjuvant treatment decisions to improve outcomes in MIBC patients.
 

Keywords: Copy number alterations, lymph node metastases, bladder cancer


00-02

Recurrence risk in patients with melanoma of the external ear: An investigation of surgical approach and sentinel lymph node status

Amanda Truong1, John Hyngstrom1, 2, 3, Robert Andtbacka1, 2, R. Dirk Noyes3, Melissa Wright3, John Snyder3, Kenneth Grossmann1, 4, Douglas Grossman1, 5, Tawnya Bowles1, 2, 3

1Huntsman Cancer Institute, Salt Lake City, UT, United States
2Department of Surgery, University of Utah, Salt Lake City, UT, United States
3Intermountain Healthcare, Salt Lake City, UT, United States
4Department of Medicine, University of Utah, Salt Lake City, UT, United States
5Department of Dermatology, University of Utah, Salt Lake City, UT, United States

Background

Surgical management of external ear melanomas represents a unique challenge due to the ear’s structural complexity and variable lymphatic drainage patterns. Few and conflicting studies of melanomas at this site have resulted in a lack of consensus regarding optimal treatment approach of the primary tumor and regional nodal basin. To address this, a retrospective cohort study was performed to evaluate primary tumor characteristics and treatment decisions, as well as their impact on local, regional, and distant recurrence.

Materials and Methods

Patients diagnosed with invasive external ear melanoma between 2000 and 2011 were identified from the University of Utah and Intermountain Healthcare melanoma databases. Patients older than 18 were included for analysis if they had documented invasive primary melanoma of the external ear, had at least 2 years of documented follow-up, and had no synchronous metastatic disease at time of diagnosis. Patient demographics, tumor site, histologic characteristics, surgical technique (including cartilage sparing vs. cartilage removal procedures), sentinel lymph node location and status, and follow-up data were evaluated using descriptive statistics and Fisher’s exact test for analysis of recurrence risk. 

Results

A total of 156 patients were identified, median age was 62.5 years, 85.9% were male, and mean follow-up was 5.5 years. Primary melanoma location on the external ear included helix (61.5%), lobe (13.5%), and other (25.0%). 75.3% of ear melanomas were diagnosed as stage IA/IB. Lentigo maligna melanoma was the most common histologic subtype (36.6%) and median Breslow depth was 0.86 mm (range: 0.1-7.0 mm). Cartilage sparing surgery was performed in 29 (19.2%) patients, whereas 122 (80.8%) patients had cartilage removal, including 14 (9.5%) patients who underwent a complete auriculectomy with prosthetic reconstruction. 99 (63.5%) patients underwent sentinel lymph node biopsy, with sentinel lymph node(s) identified in 100% of patients and a median number of four nodes. 14.1% of these patients had micrometastatic nodes. Twenty-six (16.7%) patients had a recurrence with a median time to recurrence of 2.3 years (range: 0.4 – 5.5 years). Of the patients with a recurrence, 57.7% recurred initially at a distant site, most commonly to the lung. Increased recurrence risk was associated with primary tumor location (non-helix sites), increased Breslow depth, more advanced initial stage at diagnosis, and micrometastatic sentinel lymph nodes. There was no significant association between recurrence risk and surgical technique, including whether or not cartilage was removed in early stage lesions.

Conclusions

This study comprises one of the largest cohorts of external ear melanomas and sentinel lymph node biopsy. In addition to identifying and reinforcing previously established risk factors for recurrence, our study also highlights the feasibility and importance of sentinel lymph node biopsies in stratifying recurrence risk, even in this area of variable lymphatic drainage. We also report that choice of surgical approach of the primary tumor, including sparing cartilage or not, was not associated with increased local, regional, or distant recurrence risk. Our findings provide more robust data to inform future surgical guidelines of this intricate cutaneous region.

Keywords: auricle, SLNB, surgical technique, cartilage sparing approach,


00-03

PET/CT at the time of diagnosis in patients with melanoma metastatic to sentinel lymph nodes rarely identifies systemic disease.

Edward Miranda1, 2, Beata Jaron1, 2, Mohammed Kashani-Sabet2, Stanley Leong2

11. Center for Complex Reconstruction, San Francisco, United States
2California Pacific Medical Center, San Francisco, United States

Background

Several studies have been published on the appropriate radiographic studies for detecting stage IV disease in patients with melanoma at initial presentation. Of particular interest are those asymptomatic patients who undergo selective lymph node biopsy and are found to have regionally metastatic melanoma to a sentinel lymph node.  Commonly, patients undergo PET/CT and/or brain MRI at the time that metastasis to a sentinel lymph node is found.  The diagnostic yield of these imaging studies is uncertain with false positives being of particular concern.

Materials and Methods

Of 1389 patients identified from a database of individuals who underwent selective sentinel lymphadenectomy for primary melanoma we studied 212 with at least 1 sentinel lymph node positive for metastatic melanoma who had a PET/CT and/or brain MRI. The study was conducted as an IRB-approved, retrospective analysis of a population of patients at a tertiary care referral center.  The main outcome measure was diagnostic yield of the imaging studies.  

Results

The results of 0.6% of the imaging studies were positive for metastatic disease, 92.6% were negative, and 6.7% were indeterminate. Indeterminate results were confirmed to be negative by additional studies ranging from repeated imaging to invasive procedures. The yields are as follows: PET/CT 0.7% and brain MRI 0%; the false positive rate for PET/CT was 7.2%.

Conclusions:

PET/CT and brain MRI rarely reveal systemic metastasis at the time of sentinel lymph node biopsy. In the case of PET/CT, the false positive rate is up to 10 times the true positive rate indicating that the use of PET/CT (and brain MRI) as routine imaging modalities at the time initial detection regional melanoma metastasis should be reconsidered in otherwise asymptomatic patients.

Keywords: Sentinel lymph node, melanoma, PET/CT, MRI, metastasis, staging


00-04

Conditional survival in melanoma in the sentinel lymph node era:  Meaningful prognostic information for patients and physicians

Lauren Kerivan, Michael Reintgen, Steve Shivers, Eric Reintgen, Douglas Reintgen

University of South Florida Morsani School of Medicine

Malignant melanoma is epidemic in the State of Florida and a significant public health problem.   Conditional survival (CS) estimates are more meaningful for patients as they progress in their follow-up from a cancer diagnosis.

Methods:  A retrospective query of a prospective database was used to abstract patients diagnosed with invasive malignant melanoma during the years 1988-2016.   Patients were stratified by Stage of Disease at diagnosis.  Data was generated based on 5 year disease-free survival (DFS) and overall survival (OS)  from the date of diagnosis, as well as 5 year DFS and OS if patients survive 1,2,3 and 4 years without recurrence or death (Conditional Survival).    All deaths in the series were from metastatic melanoma. The best prognosis group, Stage I melanoma, was compared to life tables of survival for the normal US population.

Results:   There were a total of 7531 melanoma patients in the study who received their care at the University of South Florida, Lakeland Regional Cancer Center and Moffitt Cancer Center.  59% of the population was male and the mean age of the group was 57.18 years.  A full 25.1% of the patients were greater than 70 years of age and 6.6% were less than the age of 30.  The distribution of patients according to the Stage of disease at diagnosis was for Stage I, II, III and IV melanoma, 53.4%, 32.5%, 11.1% and 3% respectively.

For all stages of disease as patients with melanoma were followed without recurrence or death, the prognosis improved.  For stage I patients 5 -year DFS calculated from diagnosis increased from 61.7% to 93.2% if patients survive up to 4 years without recurrence.    Similar trends were noted for Stage II -IV disease.   5-year OS from date of diagnosis for Stage I patients also increased from 88.8%  to 98.6% if patients survived 4 years.  5-year OS at any period of time during the recurrence free follow-up period was similar for patients with Stage I (88.8%, 87.2%, 86.2%, 86.4%, 86.2%) and II (66.7%, 62%, 62.8%, 64.2%, 66.1%) disease but improved significantly for Stage III (50.7%, 50.2%, 56.5%, 59.8%, 71.4%) and IV patients (27.6%, 34.9%, 48.3%, 50.0%, 68.7%) as patients survived 1,2,3 and 4 years from diagnosis.  For example, if Stage III patients survive 4 years, the 5-year OS increases from 50.7% to 70.4%.  The best prognostic group, those patients with Stage I melanoma who survive without recurrence for the first 4 years of follow-up continued to have an increased death rate compared to the normal population.   The CS data for patients with Stage I, II and III melanoma do not substantiate any medical/legal causation arguments as long as patients do not have a recurrence in the first 4 years of follow-up.  

Conclusion:  Prognosis improves for melanoma patients if they survive during the follow-up period without recurrence.  This was observed for all stages of disease.  This data provides more meaningful recurrence and survival information for patients and their families, for clinicians, for lawyers in the medical/legal system and for the insurance industry.

Keywords: Conditional Survival, Stage of Melanoma


00-05

Survival of melanoma patients is significantly compromised when the non-sentinel lymph node compartment is found to contain micrometastais following completion lymph node dissection for patients with a positive sentinel lymph node biopsy.

Andrei Rios-Cantu1, Ying Lu2, Servando Cardona-Huerta1, Victor Melendez-Elizondo1, Michael Chen2, Alejandra Gutierrez-Rangel1, Stanley Leong1

1Center for Melanoma Research & Treatment, California Pacific Medical Center, San Francisco, CA, United States
2Departments of Biomedical Data Science, Health Research and Policy, and Radiology, the Stanford Cancer Institute, Stanford University, CA, United States
3Department of Dermatology, University of California, San Francisco, CA, United States

Background/Hypothesis: We hypothesize that the progression of disease from the sentinel lymph node (SLN) to the non-SLN compartment is orderly and is associated with the worsening of the disease status. Thus, the sentinel and non-sentinel lymph node compartments are biologically different in that cancer cells, in general, arrive in the sentinel lymph node compartment before spreading to the non-sentinel lymph node compartment.  To validate this concept, we use a large cohort of melanoma patients from our prospective SLN database.

Design: Retrospective analysis of a prospective melanoma database.

Setting: Academic tertiary medical center.

Patients: Adult cutaneous melanoma (CM) patients (n=291) undergoing CLND after a positive SLN result from 1994 to 2009.

Interventions: Wide local excision and SLN biopsy, followed by CLND.

Main Outcome: Comparison of 5-year disease-free survival and 5-year overall survival between positive and negative CLND groups.

Results: 225 patients had negative CLND and 66 had positive CLND (22.7%). The 5-year disease-free survival rates were 55% (95% CI: 49% - 62%) for patients with no additional LN on CLND versus 14% (95% CI: 8% - 26%) in patients with positive LN on CLND (p<0.0001, log-rank test). The median disease-free survival time was 7.4 years with CLND negative (95% CI: 4.4 to 15+ years) and 1.2 years with CLND positive (95% CI: 1.0 to 1.8 years).

The 5-year overall survival rates were 67% (95% CI: 61% - 74%) for negative CLND versus 38% (95% CI: 28% - 52%) for positive CLND (p<0.0001, log-rank test). The median overall survival time was 12.1 years for CLND negative (95% CI: 9.3 to 15+ years) and 2.5 years for CLND positive (95% CI: 2.2 to 5.7 years).

Conclusion: This study shows that CLND status is a significant prognostic factor for patients with positive SLNs undergoing CLND. In patients, when the non-SLN compartment is not involved with metastasis, their survival is much improved. Thus, information from CLND is important to predict the outcome of melanoma patients with a positive SLN biopsy.

Keywords: Melanoma, Sentinel Lymph Node, Micrometastasis, Survival, SLN, CLND


00-06

Overall survival and outcome of melanoma patients with brain metastasis after the FDA approval of checkpoint inhibitors and BRAF and MEK inhibitors

Elham Vosoughi, Jee Min Lee, James R. Miller, Mehdi Nosrati, David Minor, Roy Abendroth, John W. Lee, Brian T. Andrews, Lewis Z. Leng, Max Wu, Mohammed Kashani-Sabet, Kevin B. Kim

California Pacific Medical Center Research Institute, San Francisco, United States

Background: Patients with melanoma with BM have a poor prognosis with historical series showing a median overall survival (OS) of ~5 months. Novel BRAF/MEK inhibitors and checkpoint inhibitors have dramatically improved OS in patients with advanced melanoma, but demonstrated mostly in those without BM. We evaluated OS and clinical outcomes of patients with BM in the era of these novel therapies.

Materials and Methods: We performed a retrospective review of the medical records of melanoma patients with BM diagnosed between 01/2011 and 06/2015.

Results: We identified 79 patients with BM. The median time from primary melanoma diagnosis to BM was 3.2 yrs (range, 0-29.8 yrs), and the median time from stage IV diagnosis to BM was 2 months (range, 0-103 months). The stage prior to the BM diagnosis was II in 6 (7.6%) IIIA in 1 (1.3%), IIIB in 9 (11.4%), IIIC in 3 (3.8%), IV(M1a) in 6 (7.6%), IV(M1b) in 8 (10.1%) and IV(M1c) in 25 (31.6%). Six (7.6%) patients had brain metastasis as the only site of distant metastasis, and 36 (45.6%) patients had neurological symptoms at BM diagnosis. Twenty-nine (36.7%) had V600 BRAF-mutant melanoma. Thirty-four (43.0%) underwent surgical resection, and 52 (65.8%) were treated with stereotactic radiosurgery. 39 (49.4%), 28 (35.4%) and 24 (30.4%) patients received CTLA-4Ab, PD-1Ab and BRAF(+/-MEK) inhibitors, respectively. Of 55 (69.6%) patients who have died of melanoma progression, 30 (40.0%) died with progressing BM.  The median OS from the diagnosis of BM was 12.8 months (range, 1.1-64.8 months), and the median OS from the time of a stage IV diagnosis was 18.2 months (range, 1.4-158.4 months). The median OS from the time of the first stereotactic radiosurgical treatment was 14 months (range, 1-63 months).

Conclusion: Our finding suggests that OS of patients with BM has been significantly improved in the new era of the novel melanoma therapies.

Keywords: Metastatic Melanoma, Checkpoint Inhibitors, BRAF Inhibitor, MEK Inhibitors


00-07

Utility and feasibility of sentinel lymph node inking for accurate localization of nodal metastases

Alexandra Gangi1, Jeffrey Francis2, Jane Messina3, John Hassani4, Caitlin Porubsky4, Jiannong Li5, Jose Pimiento1, Ann Chen5, Vernon Sondak4, Amod Sarnaik4, Jonathan Zager4

1H. Lee Moffitt Cancer Center Department of Surgical Oncology, Tampa, United States
2Morsani College of Medicine, USF Health, Tampa, United States
3H. Lee Moffitt Cancer Center Department of Anatomic Pathology, Tampa, United States
4H. Lee Moffitt Cancer Center Department of Cutaneous Oncology, Tampa, United States
5H. Lee Moffitt Cancer Center Department of Biostatistics, Tampa, United States

Background:  Sentinel lymph node (SLN) biopsy is standard of care for nodal staging in patients with melanoma. SLN biopsy identifies nodal metastases in approximately 12-16% of patients with clinically node-negative melanoma. However, some literature suggests that ~5% of all tumor-positive nodal basins are incorrectly staged as sentinel node negative. In some cases, retrospective evaluation of the “negative” SLN has shown small tumor deposits that were not in the initial planes of sectioning but were identified by further sectioning of the paraffin-embedded lymph node. This suggests that intraoperative techniques to orient the SLN for histologic sectioning and pathologic analysis could increase the yield of positive SLNs and potentially decrease the rate of false- negative results. We developed a protocol for intraoperative orientation of SLN with sterile ink and studied whether intraoperative inking of the area of maximum blue dye/radioactivity (“hot spot”) within the SLN was feasible and correlated with sites of metastatic disease.  

Materials and Methods:  Patients with primary melanoma undergoing SLN biopsy consented to participate in this IRB-approved trial. Each SLN was evaluated intraoperatively for visual evidence of blue dye and maximum radioactive counts. The area on the SLN with the highest concentration of blue dye and/or radioactivity, if identifiable intraoperatively, was marked with ink from a sterile tissue marking kit (MarginMarker(R)). The histopathology slides were evaluated for the presence of ink and its association to metastases.

Results:  313 patients were enrolled and 309 underwent SLN biopsy. Average age was 61.3 years and most patients were male (58%). Primary melanomas had a median Breslow depth of 1.5 mm and a median of 2 mitoses/mm2 (Table 1). 290 patients had ≥1 inked SLN, while the remainder had no identifiable hot spot. 54 patients had metastatic nodal disease, of whom 40 had the positive node(s) inked for a patient-level sensitivity of 74.1% (95% CI 60.3-85.0%). At the node level, 901 SLNs were removed, of which 688 had ink identified on pathology evaluation. 75 SLNs were found to harbor metastatic disease and 62 of these had been inked and could be evaluated for correlation of the ink with the metastatic site. 48 inked SLNs harbored metastatic disease adjacent to the inked hot spot for node level sensitivity of 77.4% (95% CI 65.0-87.1).

Conclusion:  Surgeon-identified orientation of SLN hot spots is feasible in a majority of cases and correlates with sites of melanoma nodal metastases. SLN inking may focus the pathologist on sections of the SLN at highest risk for metastatic involvement. Further evaluation will be required to determine if the incidence of false negative SLN biopsy results can be decreased by using intraoperative orientation techniques that allow for histopathologic analysis of nonrandom sections of the SLN.

 

Keywords: sentinel lymph node, hot spot


00-08

The Performance of Tc-99m Tilmanocept Allows for Flexible Scheduling and Use in a Variety of Clinical Settings for Sentinel Lymph Node Biopsy in Breast Cancer

Jonathan Unkart, Anne Wallace

Department of Surgery University of California, San Diego, San Diego, United States

Background: No prior studies have investigated the performance of Tc-99m tilmanocept (TcTM) in sentinel lymph node (SLN) biopsy for breast cancer since FDA-approval in March 2013. We sought to investigate the sensitivity and performance in a variety of clinical scenarios.

Methods: Our institution adopted the use of Tc-99m tilmanocept for routine use in May 2013 for breast cancer SLN biopsy in clinically node-negative patients. We assessed the effect of neoadjuvant chemotherapy, use of lymphoscintigraphy and compared one-day versus two-day preoperative injection protocols on sensitivity of detecting a SLN and on total number of removed nodes. Patients received a 0.1ml injection of either 0.5mCi TcTM on day of surgery or 0.1ml injection of 2.0mCi TcTM on day prior to surgery by a nuclear medicine radiologist. Performance and use of lymphoscintigraphy was optional. Intraoperatively, patients received a 2-3ml injection of vital blue dye (VBD) immediately prior to incision. A negative binomial count model was used to assess univariate and multivariate factors influencing the number of removed SLNs with TcTM and VBD. A p-value < 0.05 was used for statistical significance.

Results: Between May 2013 to June 2016, 617 clinically node-negative patients underwent SLN biopsy with TcTM and VBD. A SLN was identified intraoperatively in 100% of patients. On average, 3.0 SLNs (SD 1.8) were removed per patient. Sixty-seven (10.9%) patients were injected with the two-day protocol and 91 (14.7%) patients underwent neoadjuvant chemotherapy. At least one positive-node was detected in 95 (15.4%) patients. Overall, radioactive nodes were identified in 609 (98.7%) patients, blue nodes in 550 (89.1%) patients and both radioactive and blue nodes in 543 (88.0%) patients. There was no statistical difference in the number of removed nodes in patients receiving neoadjuvant chemotherapy, undergoing the “two-day” injection protocol or with the use of lymphoscintigraphy. On multivariate analysis: operating surgeon, age and presence of positive nodes affected the number of removed SLNs.

Conclusion: TcTM reliably identifies SLNs in clinically node-negative breast cancer patients undergoing SLN biopsy. The use of neoadjuvant chemotherapy does not alter the number of removed SLNs. Furthermore, the injection of TcTM can be performed on the day before surgery or the day of surgery without the use of lymphoscintigraphy.

Keywords: tilmanocept, sentinel lymph node biopsy, breast cancer


00-09

Sentinella portable gamma camera detects additional sentinel lymph nodes in melanoma patients

Stanley Leong1, Max Wu1, Ying Lu1, Donald Torre2, Anna von Bakonyi1, Ari Wight1, James Newsom1, William Luckett1, Mohammed Kashani-Sabet1

1California Pacific Medical Ctr, San Francisco, United States
2Stanford University, Palo Alto, United States

Introduction Preoperative imaging combined with intraoperative gamma probe (GP) localization is standard procedure for identifying the sentinel lymph nodes (SLNs) in melanoma patients. The primary aim of this prospective IRB-approved study is to investigate whether an intraoperative portable gamma camera (PGC) improves the intraoperative detection of SLNs when used in conjunction with the GP.

Materials & Methods Planar lymphoscintigraphy and SPECT/CT were performed after injection of a dosage of 600 uCi (1-day protocol) or 2000 uCi (2-day protocol) of [(99m)Tc]Tilmanocept in melanoma patients over 18 years old with a Breslow thickness ≥ 1.0 mm. Preoperatively,  the SLNs were localized and the skin overlying the area was marked by the nuclear medicine physician. Intraoperatively, a strict surgical protocol was followed for each identified draining SLN basin. A GP (Neoprobe 2000) was used to locate and remove the SLNs. The resection bed was then explored by the GP using ‘roaming counts’ at 8 positions of the clock and center to make sure that the entire operative field has a negative GP screening (less than 10% of the hottest node). Finally, the PGC (Sentinella S102, Oncovision S.A., Valencia, Spain) was used after a negative GP screening. The Sentinella® PGC was positioned 5 cm above the resection bed with an image acquisition time of 60 seconds. Any depicted radioactive hotspots after a negative GP screening were considered as additional found SLNs by Sentinella® PGC. The hotspots were further explored using the GP to locate and remove additional detected SLNs (applying the >10% rule).

Results First 50 patients (27 males and 23 females) were included in this report. Preoperative imaging localized 70 SLN basins. Intraoperative GP detected 159 SLNs and Sentinella® PGC found additional 60 SLNs, which were removed with the GP guided by the Sentinella images. One palpable lymph node, not detected by GP was positive for micrometastasis. Thus, the identification rate by Sentinella alone was 27% (60/220). Overall, a SLN positive rate of 8.2% (18/220) and a patient positive rate of 26% (13/50) were found. In 32 patients, Sentinella® PGC imaging detected additional SLNs (55 tumor-negative and 5 tumor-positive SLNs) after a negative gamma probe screening. Two of these 5 tumor-positive SLNs being found by Sentinella® PGC prevented a false-negative procedure. Patients will be enrolled to complete the study with a total anticipated number of 100.

Conclusion Interim analysis of this prospective study to date demonstrates that intraoperative PGC imaging provides additional information when standard technique with GP fails. The use of intraoperative imaging has led to the detection of additional SLNs in a significant number of patients. Thus, intraoperative imaging by PGC holds promise for reducing false negative rate for sentinel lymph node biopsy for melanoma.

Keywords:

Friday, April 21, 2017 - Gold Rush Ballroom - 18:40 - 19:15

Rapid Fire Abstract Session (Non-CME) - Rapid Fire: Late Breaking

00-01

Thromboembolic Risk in Patients with Newly Diagnosed Multiple Myeloma Is Associated with Biomarkers of Hypercoagulability

Fotiou Despina2, Flora Zagouri2, Aurélie Rousseau1, Patrick Vandreden1, Evangelos Terpos2, Meletios Dimopoulos2, Grigorios Gerotziafas3

1Diagnostica Stago France, Genneviliers, France
2National and Kapodistrian University of Athens, Athens, Greece
3Consultation Thrombose – Oncologie Service d'Hématologie Biologique Hôpitaux Universitaires de l'Est Parisien, Paris, France

Background: Multiple myeloma (MM) and other plasma cell dyscrasias (PCD) and the associated immunomodulatory treatments are linked to increased risk of venous thromboembolism (VTE). The identification of patients at VTE risk and the optimization of VTE prevention is an unmet medical need. Elaboration of a risk assessment model (RAM) specific for patients with MM, which includes biomarkers of hypercoagulability, could improve the management of VTE risk.

Aim: We conducted a longitudinal observational study, to explore the relationship of MM with cellular and plasma hypercoagulability aiming to identify the most relevant biomarkers for use in a RAM for VTE in combination with clinical risk factors.

Methods: Newly diagnosed patients with PCD (n=186) were recruited from July 2014 to Dec 2015; including 27 with monoclonal gammopathy of undetermined significance (MGUS), 40 with asymptomatic multiple myeloma (AMM), 79 with multiple myeloma (MM), 30 with AL-amyloidosis, 8 with Waldenstrom’s Macroglobulinemia (WM) and 2 with solitary plasmacytoma. They were compared against 30 healthy age and sex-matched individuals (CG). A systematic compression ultrasound was performed at baseline and at 6-12 months. Blood samples were obtained at diagnosis and at 6-12 months (n=89). Samples of platelet-poor plasma (PPP) were assessed for thrombin generation (TG) with PPP-Reagent® (TF 5pM and 4 μM phospholipids), P-selectin, D-dimers (D-Di), activated FVII (FVIIa), Tissue Factor (TFa), fibrin monomers (FM), and procoagulant phospholipid-dependent clotting time (Procag-PPL). The upper and lower normal limits (UNL and LNL) were calculated by the mean±2SD.

Results: Median age was 67 years (37-89) and 49% of the population was male. Median time to follow up was 7 months (1-12 months). Among MM cases (n=79), symptomatic VTE rate was 10% (n=8) and mortality rate 7.5%. The events included 2 central venous catheter thromboses, 1 Pulmonary Embolism, 2 Deep Vein Thromboses, 2 Superficial Vein Thromboses and 1 Mesenteric Vein Thrombosis. Cases had significantly shorter PPL-ct, higher FTa and DDi, TG (increased Peak), heparanase and P-selectin compared to controls (p<0.001). Along the MGUS- AMM- MM continuum, D-Di concentration (p<0.0001), FM concentration (p<0.05), lag time (p<0.027) and MRI were highest in MM patients. Sixty percent of MM patients had Procoag-PPL: below the LNL, in 63% TFa levels were above the UNL, and 31% had MRI lower than LNL. After 6 months of treatment, FTa, DDi, MRI, peak thrombin and P-selectin levels decreased (p<0.05) and TM and ATIII levels increased (p<0.05).

Conclusion: In patients with PCD, increased procoagulant microparticles of cellular origin is a generalized phenomenon. In addition, patients with MM, are at high risk of VTE and present with significant TF pathway activation and increased TG. A significant fraction, but not all, of the patients present strong signs of plasma hypercoagulability. The finding of high inter-individual variability of TG underlines the heterogeneity of blood coagulation alterations in PCD patients. The data of the prospective part of this study will allow validation of the clinical significance of this finding

Keywords: Multiple Myeloma-Thromboembolic Risk


00-02

Development in ambulatory patients with lung adenocarcinoma on chemotherapy of A clinicobiological Risk Assessment Model for VTE. The ROADMAP study.

Grigoris Gerotziafas1, Aurélie Rousseau2, Ilias Evmorfiadis3, Patrick Vandreden2, Paraskevi Boura4, Andriani Charpidou4, Ioannis Giozos4, Konstatinos Syrigos4

1Unité d’Explorations Fonctionnelles et Génétiques du Risque Vasculaire, Consultation Thrombose – Oncologie Service d'Hématologie Biologique Hôpitaux Universitaires de l'Est Parisien – APHP, Paris, France
2Diagnostica Stago, Gennevilliers, France
3Cancer Biology and Therapeutics Centre de Recherche Saint-Antoine INSERM U938 et Université Pierre et Marie Curie, Paris, France
4Department of Medicine, Athens School of Medicine,, Athens, Greece

Background. In ambulatory patients with advanced or metastatic stage of lung adenocarcinoma (LA) the risk of VTE increases during chemotherapy.  but individual risk factors cannot identify patients at risk. The available RAM are not applicable in patients already on chemotherapy.

Aim. The prospective longitudinal non interventional study ROADMAP was designed to elaborate a RAM for VTE specific for ambulatory patients with LA on chemotherapy.

Methods. Patients with LA on chemotherapy were included and followed up at 3, 6 and 12 months. Documented symptomatic VTE was the end-point of the study. Blood samples were collected at inclusion and assessed for thrombin generation (TG) and procoagulant phospholipids (PPL-ct). Assays and reagents were from Diagnostic Stago (France). Multivariate analysis was performed and the RAM was developed using the logistic regression. Sensitivity, specificity and the predictive value of the RAM were calculated. The ROC Curve was plotted.

Results. The study included 150 patients (mean age 65 years, 73% male). The LA diagnosis was done within 6 months before inclusion in 70% of them and 90% had advanced or metastatic disease at inclusion. In 85% of patients the ECOG performance status was <3. In one year follow up 12 symptomatic VTE episodes occured (8%), 75% of which occurred within the 3 months from inclusion. The RAM includes the following variables: Recent hospitalisation, Time since diagnosis of the cancer, Mean Rate Index of TG and PPL-ct. The ROC analysis gave a AUC value of 0.84. The sensitivity of the RAM was 89% and the specificiy was 70%. The postive predictive value is 16% and the negative predictive value is 98%

Conclusions. The new RAM for VTE is specific for ambulatory patients with LA on chemotherapy and can relibly predict VTE using simple clinical variables and biomarkers of hypercoagulability. This RAM can be used by physitians for the identification of ambulatory lung cancer patients eligible for thromboprophylaxis.

 

Keywords:


00-03

The Futility of Brain MRI for Melanoma Patients with a Positive Sentinel Lymph Node Biopsy.

Edward Miranda1, 2, Beata Jaron1, Stanley Leong2

1Center for Complex Reconstruction, San Francisco, United States
2California Pacific Medical Center, San Francisco, United States

Background

A variety of studies have been published on both the frequency and utility of brain imaging for detecting melanoma brain metastasis.  However, uncertainty remains in the subset of asymptomatic patients who have regional metastasis at the time of selective sentinel lymph node biopsy (SLNB).  In many cases, patients receive brain imaging with magnetic resonance imaging (MRI) when regional metastatic disease is found during SLNB.  These imaging studies have historically been reported as low-yield for distant metastasis.  If the diagnostic yield for MRI tends to be low, based on data, we sought to calculate that maximal incidence melanoma brain metastasis at the time of positive SLNB in our CPMC melanoma database.

Materials and Methods

136 patients with brain MRIs were identified from our prospectively maintained databases of individuals who underwent SLNB for primary melanoma with metastasis to SLNs.  The study was conducted as part of an IRB-approved, retrospective analysis of a population of patients at a tertiary care referral center.  The primary outcome measure was detection of distant metastasis on MRI brain imaging.  A probabilistic model was then applied to the observed data to predict the maximal incidence of melanoma brain metastasis at the time of positive SLNB in asymptomatic melanoma patients.

Results

None (0.0%) of the brain MRIs detected metastatic disease in 136 melanoma patients at the time of positive SLNB.  Using the developed probabilistic model, estimates of the incidence of brain metastasis at the time of diagnosis of melanoma with a positive SLN were calculated.  For an approximated error tolerance of 0.05 the maximal incidence of melanoma brain metastasis at the time of positive SLNB was calculated as less than 2.2%; for a more stringent error tolerance of 0.01, the maximal incidence of was calculated at less than 3.3%.

Conclusions:

Brain MRI rarely reveals distant metastasis at the time of SLNB.  Based on the MRI data, there is a low incidence of melanoma metastasis to the brain at the time of positive SLNB. The use of MRI for patients at the time of positive SLNB should be reserved for symptomatic patients or those who undergo clinical trials that require baseline brain MRI.

Keywords: melanoma, sentinel lymph node, metastasis, staging, MRI, mathematical model


00-04

Use of SLN biopsy for DCIS in patients undergoing mastectomy:  who is at risk for upstaging?

Ahmed Mohsen, Heidi Kosiorek, Bhavika Patel, Matthew Covington, Anamika Basu, Richard Gray, Nabil Wasif, Chee-Chee Stucky, Ann McCullough, Idris Tolgay Ocal, Barbara Pockaj

Mayo Clinic, Phoenix, United States

Background: Ductal carcinoma in situ (DCIS) accounts for over 25% of breast neoplasms diagnosed in the US. This study aims to identify factors influencing the upstaging rate of DCIS in the high risk mastectomy patient subgroup, outline which surgical interventions are optimal and whether all patients require axillary staging.

    

Methods: Patients undergoing mastectomy for newly diagnosed DCIS were retrospectively studied. A univariate analysis of their demographics, methods of diagnosis, complete surgical treatment, pathologic characteristics and outcomes were compiled to determine which variables impacted tumor upstaging to invasive cancer at final pathology. 

 

Results: A total of 212 women were identified. Mean age was 59 years (range 31-88) with 74% postmenopausal. Only two patients had a genetic mutation (BRCA2). Most (95%) were diagnosed mammographically with the findings including calcifications only (77%), calcifications + mass/architectural distortion (9%), and only mass/architectural distortion (9%). Ten patients were diagnosed via MRI and one patient was diagnosed with Paget’s disease on punch biopsy. Size on imaging was stated 69% of the time with a mean of 3.1 cm (range 2-12 cm). When size was unstated, 53% of lesions were unifocal whereas others were extensive/multi-focal.  

 

Most patients were diagnosed by core (84%) and the remainder by excisional biopsy. Thirty-four lesions (16%) were palpable. The majority of DCIS lesions were high grade (55%), with 12% low and 33% intermediate; 34% exhibited comedo-necrosis and 66% were ER-positive. Histologic suspicion of microinvasion was present among 11% of biopsies and the treating surgeon recorded clinical suspicion of invasion among 18%. Most underwent reconstruction (69%), predominantly with implant-based technique (67%) with 41% pursuing bilateral mastectomy. Axillary staging was carried out in 150 patients (71%) with SLN biopsy in all but 1 patient. Thirty-four patients (16%) were upstaged to invasive cancers with a mean tumor size of 1.3 cm. Staging of the invasive cancers was T1mic (n=8), T1a (n=13), T1b (n=5), T1c (n=3), T2 (n=4), and T3 (n=1). Only 1 patient (<1%) had a positive SLNB. Univariate analysis identified that the variables significantly associated with upstaging were a histologic suspicion of microinvasion (50% upstaged, p<0.001), ER- (p=0.026), surgeon suspicion of invasion (36% upstaged, p<0.001), palpable disease (30% upstaged, p=0.02), and pre-operative tumor size >2 cm (p=0.04). Mean tumor size was associated with upstaging with 3.8 vs 3.0 cm, p=0.066 but did not reach statistical significance. All patients that were upstaged had a SLNB.  Factors not found to be associated with upstaging included type of biopsy (core vs excision), grade, comedo-necrosis, mammography findings, or use of MRI.

 

Conclusions: In this high risk population the upstage rate from DCIS to invasive cancer was 16%. Clinical judgement, palpable disease, suspicion of microinvasion, and large tumor size were associated with upstaging. Surgeons can use these factors in deciding to forego SLNB in patients with small, low risk DCIS undergoing mastectomy.

Keywords:


00-05

Melanoma lymphatic drainage patterns identified by tilmanocept

Daniel M Balkin1, Frederick Wang1, Emily M Balkin2, Cristina O’donoghue4, Caitlin Porubsky6, Jaime L Montilla-Soler5, Vernon K Sondak4, Mohammed Kashani-Sabet3, Jonathan S Zager4, Stanley P L Leong3

1Department of Surgery, Division of Plastic and Reconstructive Surgery, University of California San Francisco, San Francisco, United States
2Department of Pediatrics, Division of Pediatric Critical Care, University of California San Francisco, San Francisco, United States
3Center for Melanoma Research and Treatment, California Pacific Medical Center Research Institute, San Francisco, United States
4Department of Cutaneous Oncology, Moffitt Cancer Center, Tampa, United States
5Department of Diagnostic Imaging and Interventional Radiology, Moffitt Cancer Center, Tampa, United States
6Philadelphia College of Osteopathic Medicine/North Fulton Hospital Medical Campus, Roswell, United States

Background:
A variety of methods have been used to identify the SLN in cutaneous melanoma. [99mTc] Tilmanocept, a multivalent mannose-containing radiopharmaceutical designed for SLN detection, is unique in its ability to more precisely determine whether a single SLN or multiple independent SLNs exist based on its biologic intelligent design. The objective of this study is to determine the number of sentinel lymph nodes in melanoma patients using Tilamocept to guide sentinel lymph node (SLN) detection.

Methods:
Multi-center retrospective review of patients with cutaneous melanoma undergoing SLN biopsy using Tilmanocept between January 2008 - August 2014 was conducted. Data analyses included descriptive statistics, t-tests, Fisher’s exact tests and Chi-square tests.

Results:
Of the 573 patients in the cohort, mean age was 60.3 +/- 16.6 years and 62.5% were male. Primary tumor sites included: 33.2% trunk, 27.3% head/neck, 18.5% lower extremity and 20.7% upper extremity. A median of 3 (range 1-28) SLNs were identified and resected by following the rule of 10%. Overall, 83.4% of patients had >1 SLN identified by Tilmanocept. Patients with >1 SLN were more likely to have a positive node (p=0.008), younger (p =0.001), had a higher pathologic AJCC tumor stage (p=0.018) and higher Clark level (p=0.031). Additionally, >1 SLN was associated with tumor location (p<0.001). In a multivariate model, age, tumor location and Breslow depth were significantly associated with >1 SLN. In addition, positive SLN status was significantly associated with younger age, greater Breslow depth, mitosis rate and AJCC tumor stage, and presence of ulceration, microsatellite lesions, lymphatic and vascular invasion.

Conclusions:
Tilmanocept detects multiple independent SLNs in the majority of melanoma patients.

Keywords: Melanoma, Sentinel lymph node, Tilmanocept


00-06

Recurrences in patients with a primary melanoma having a negative sentinel lymph node biopsy

KM Mahuron1, S Cardona-Huerta1, 2, SR Thummala2, JE Cleaver1, M Kashani-Sabet2, SP Leong2

1University of California-San Francisco, San Francisco, United States
2Center for Melanoma Research & Treatment, California Pacific Medical Center, San Francisco, United States

BACKGROUND:

Sentinel lymph node status is considered the most important prognostic indicator for disease-specific survival as well as the best predictor of recurrence in patients with primary cutaneous melanoma. However, up to 20% of patients with an initial negative sentinel lymph node biopsy (SLNB) have recurrence, and distant metastasis is the most common site. This study aims to determine the pattern of recurrences including distant sites for patients with a negative SLNB.

METHODS:

We performed a retrospective review of our melanoma database collected in an academic tertiary-care medical center following IRB approval for the period between 1993 and 2009 which included 2,098 patients who underwent SLNB (16.9% positive vs. 83.1% negative). Patients with primary cutaneous melanoma and an initial negative SLNB were included in this study (n=1,744).  The median follow-up of the patients was 6.9 years. Statistical analysis was performed using Stata v14.1 software.

RESULTS:

The 1,744 patients with an initial negative SLNB were divided into four groups: 1,446 (82.9%) of these patients had no recurrence, 130 (7.4%) had local (within 5 cm of the original lesion) or in-transit (between the primary site and regional LN basin) recurrence, 54 (3.1%) had recurrence that involved regional LN basins, and 114 (6.5%) had distant recurrences with or without local or regional recurrences. The false negative SLNB rate of our study was 6.7% (any nodal recurrences in the SLNB sites).

Of the 114 patients with a negative SLNB who developed distant metastasis, 100 patients (87.7%) presented with distant metastasis without evidence of local, in-transit, or regional LN recurrence.  Nine patients (7.9%) presented with either local or in-transit recurrence in addition to distant metastasis. Five patients (4.4%) presented with recurrence within the regional LN basin along with distant metastasis.

Of the patients that had distant metastasis without local, in-transit, or regional LN recurrence, either with single or multiple location recurrences, the following sites were the most common: 52 patients (61.9%) had lung metastases, 30 patients (35.7%) had brain metastases, and 14 patients (16.6%) had liver metastases. Only 8 of these patients (9.5%) had any evidence of distant LN involvement (outside of regional LN basins).

CONCLUSION:

Only a minority of patients with a negative SLNB (17.1%) developed recurrences. The majority of recurrences in this cohort of patients occurred without evidence of local, in-transit, or regional LN involvement. The most common sites for distant metastases were the lung and brain, and less than 10% of these patients had any evidence of distant LN involvement. These findings suggest that hematogenous rather than lymphatic dissemination may play a role in the cancer progression and development of metastatic disease in some melanoma patients.  Future molecular studies may shed light to the mechanisms of lymphatic versus hematogenous spread of cancer.

Keywords:


00-07

WITHDRAWN

Franklin Huang1, Wen-Fang Tseng2, Frank Johnson3, Shuan Huang4, Jung Huang5

1Dana-Farber Cancer Institute and Harvard Medical School, Boston, United States
2Excelsior Pharmatech Labs., Hsinchu, Taiwan
3Saint Louis University Medical Center, St. Louis, United States
4Auxagen Inc., St. Louis, United States
5St. Louis University School of Medicine, St. Louis, United States

Keywords:


00-08

Long term outcomes of treating stage-II lymphedema patients with free lymph node transfer (LNT)

Dimitrios Dionyssiou1, 2, Efterpi Demiri1, Antonios Tsimponis1, Olga Christina Goula1, Georgia Tatsidou1, Georgios Arsos1

1Department of Plastic Surgery, Medical School, Aristotle University of Thessaloniki, Thessaloniki, Greece
2Fibralign Corporation, Union City, California, United States

Background

Symptomatic lymphedema patients are traditionally managed with physical therapy, but microsurgical methods are used in an increased manner for treating such cases. The purpose of this prospective control study is to evaluate in a long term period of up to six years the effectiveness of free LNT in stage-II lymphedema patients.

Methods

From January 2011 till June 2014 forty-one stage-II upper limb breast cancer related lymphedema patients were included in a prospective study. Clinical examination, MRI and lymphoscintigraphy of the affected limb were performed in all cases. For the initial design of the study, patients were randomly divided in two groups: Group-A patients (n=21, mean age 47 years) underwent microsurgical LNT; Group B patients (n=20, mean age 49 years) were managed by conservative therapy, including manual lymph drainage and compression garments, for six months. Post-operatively, Group A patients followed a similar six-month physiotherapy program. All forty-one patients stopped physiotherapy and compression bandaging at the sixth month and underwent re-examination of the affected extremities at the twelfth month; limb volume was measured, infection episodes were recorded, and subjective information regarding pain, feeling of heaviness and overall functional recovery, was also given by each patient. Patients of Group A continued to follow up yearly and their results are documented.

Results

At the end of the initial study, reduction of the limb volume was observed in both groups; mean reduction was greater in Group-A (57%) than in Group-B (18%). In Group-A, a mean 0,8 infection episodes per patient was documented, while three episodes per patient were recorded in Group-B. All Group-A patients reported painless and feeling of heaviness-free extremities with overall functional improvement. In Group-B, five out of twenty patients were painless, thirteen reported reduction of pain, while nine reported reduction of heaviness; only twelve patients stated subjective functional improvement of the limb, while eight reported no functional changes. 

At the long term follow up with a mean period of 3 years and 11 months (range 30-72 moths), Group A patients recorded a mean volume reduction of 54,4%, mean infection episodes 0,2 per patient per year, and the subjective overall satisfaction remained as high as 100% for all the patients.

Conclusions

LNT represents an effective therapeutic approach for stage-II lymphedema patients; it significantly reduces the limb volume, decreases recurrent infections and improves the overall function of the affected extremity. Future studies are designed for further improvement of the volume reduction, as well as minimization of the infection episodes with the use of new technologies of purified collagen matrix. Our aim is to use the BioBridgeTM purified collagen matrix as a support in accelerating the lymphagiogenesis between the transplanted at the axilla lymph nodes and the upper limb lymphatic vessels.

Keywords: secondary lymphedema, breast cancer, lymph node transfer

Friday, April 21, 2017 - Crystal Ballroom - 19:30 - 22:30

- Gala Dinner in the Crystal Ballroom

Saturday, April 22, 2017 - Gold Rush Ballroom - 07:30 - 08:30

Industry Sponsored Breakfast Session (Non-CME) - Novel Technology for SLN Imaging, Sponsored by Curadel, LLC

00-01

Strategies for Successful SLN Mapping in the Clinic using NIR Fluorescence

John Frangioni

The Curadel Companies, Marlborough, United States

Keywords:


00-02

Lymphatic imaging with light and sound

Russell Witte

University of Arizona, Tucson, United States

The first images of living human lymphatics required the naked eye to visualize intradermally-injected colored dyes. However, imaging the lymphatics with light is typically limited by poor penetration and depth resolution. On the other hand, ultrasound lymphatic imaging suffers from low contrast in soft tissue. This presentation will discuss several advanced, hybrid ultrasound modalities that have the potential to overcome these failings. Photoacoustic imaging (PAI), for example, combines pulses of light with optically-absorbing agents to generate ultrasonic waves that are detected to form an image. At a spatial resolution similar to ultrasound, PAI can detect lymphatic vessels and nodes 1 - 2 cm below the skin surface. In addition, shear wave imaging, now available on cutting-edge clinical ultrasound scanners, provides a quantitative assessment of tissue elasticity, which may be altered in subjects with lymphatic disease. These imaging technologies, along with complementary contrast agents, not only hold promise for better preclinical imaging of the lymphatics, but also for improving the diagnosis, staging, and treatment-decision making in patients with lymphatic disease or advanced cancer.

Keywords: ultrasound, b mode, lymphatic vessels and nodes, stiffness, dyes, ICG, photoacoustic, optoacoustic, shear wave, elasticity, contrast agents


00-03

Enhanced Intraoperative Detection of Sentinel Lymph Nodes, Alessandro Testori,

Alessandro Testori

General Surgeon, Milano, Italy

Keywords:

Saturday, April 22, 2017 - Gold Rush Ballroom - 08:30 - 10:00

Plenary Session (Non-CME) - Negative pressure treatment for LE, Sponsored by PhysioTouch

00-01

PhysioTouch negative pressure treatment supports lymphedema and cancer survivor rehabilitation

Kalle Palomäki

PhysioTouch, Helsinki, Finland

Keywords:

Saturday, April 22, 2017 - Emerald Ballroom - 08:35 - 10:00

Plenary Session (CME) - Gastrointestinal & Genitourinary Cancer Treatment

00-01

Pathologic Evaluation of Sentinel Lymph Nodes in Gastrointestinal Cancers

David Wiese

McLaren Regional Medical Center, Flint, United States

Accurate identification and evaluation of regional lymph nodes is necessary to properly stage cancer patients, which in turn provides prognostic information, and allows selection of appropriate additional therapy.

Sentinel lymph node (SLN) mapping represents a paradigm shift in nodal evaluation.  As the first nodes draining a solid tumor, identification and focused evaluation of the SLNs provides more accurate staging compared to conventional methods.  Based on SLN status, extended lymph-adenectomy in cases of melanoma or breast cancer may be avoided.  In the gastrointestinal (GI) tract, where regional nodes are still harvested as part of standard oncologic resection, SLN mapping provides more accurate nodal staging, and may identify aberrant drainage pathways to assure that the proper nodes are excised.

Topics to be addressed in this discussion will include:  (1) advantages of SLN mapping in GI cancers with discussion of pathologic protocols and evaluation; (2)  comparative focused evaluation of SLNs and non-SLNs; (3) evaluation of non-tumor associated SLN; (4) comparative studies of node sizes between SLNs and non-SLNs; (5) current lines of research and future directions.

Keywords:


00-02

Personalized treatment of upper gastrointestinal cancer based on SLN status

Hiroya Takeuchi

Keio University, Tokyo, Japan

Keywords:


00-03

Selection of Adjuvant Chemotherapy in Early Stage Colon Cancer guided by Sentinel Lymph Node Status

Sukamal Saha

Easton Hospital, Easton, United States

Globally, the incidence of colorectal cancer is about 1.36 million causing about 693, 933 deaths last published in 2012.  Nodal metastases remain pivotal in decision making for adjuvant chemotherapy.  Much research is being done looking at the biology of the primary tumor to prognosticate the tumor.  At this time, patients with stage III disease (node positive) are treated with systemic chemotherapy across the globe.  However, in patients with node negative disease, stage II patients, the role of adjuvant chemotherapy is somewhat controversial.  Some of the biologic markers ie, microsatellite instability, BRAF or K-Ras mutations have been considered for chemotherapeutic decision making.  None of these are being investigated routinely across the globe.  As 15-20% of the so-called node-negative patients still developed metastatic disease within 5 years, many of these patients have been considered for chemotherapy somewhat empirically.  In 1980s and 1990s multiple studies were done to identify various methods to increase the sensitivity of diagnosis of nodal metastases, i.e., increased number of nodes detected, ultrastaging of the nodes with multilevel sections, inmmunohistochemistry, and lately RT-PCR or OSHNA.

More recently, multiple authors have shown the negative prognostic effect of the presence of micrometastases if they remain undiagnosed and untreated.  In 2011, Saad et al showed in a meta-analysis of the prognostic impact of micrometastases papers that presence of these micrometastases reduces the 5-year survival from 82% to 64% and increases the recurrence from 17% to 31%.  However, to diagnose micrometastases in the lymph nodes, all the lymph nodes have to be ultrastaged. This is highly time-consuming and cost prohibitive.  Precisely this reason, sentinel lymph node (SLN) mapping was used in colon cancer as in melanoma and the breast almost 18 years ago.  SLN’s are diagnosed by simple dye injection at the tumor site and doing a resection of the segment of the colon with regional lymphadenectomy along with resection of 1 to 3 SLNs.  The few SLN’s only were then examined by ultra-staging with multilevel sections and IHC.  This resulted in diagnoses of micrometastases in 25% of the node positive patients.  An international data collection was done from major global centers over 3 continents (Table 1).

In this analysis, over 4497 patients were included with average number of lymph nodes per patient was 17.8 and overall success rate of SLNM was 93.7%.  Accuracy, sensitivity and negative predictive value were 90%, 77.3% and 83.5% respectively. Overall nodal positivity was 45%; off these in 36% of the patients, the sentinel lymph node was the only site of metastases and micrometastases were detected in 30% of the patient’s.  Even though skip metastases were found in 22.7% of the patient’s, all patients with true positive sentinel lymph node and false-negative sentinel lymph node (non-sentinel lymph node positive, sentinel lymph node negative) were diagnosed as node positive (stage III) and were treated with systemic therapy.  So unlike in breast cancer and melanoma, patients with skip metastases in colon cancer are neither understaged nor undertreated.  In addition, our data of SLNM in colon cancer showed, it identified aberrant SLN in 20% of the patient’s which changed the extent of the operation compared to conventional surgery. This identified 9.8% of the patient’s with the aberrant SLN as the only site of the positive lymph nodes. These nodes could have been left behind without SLN mapping.

In conclusion, by doing SLN mapping in early stage colon cancer, we can identify 1 to 3 SLN’s in >93% of patients, which then can be ultrastaged by IHC.  This will allow about 25% of the patients to be diagnosed with micrometastases. These patients can then be either treated with systemic therapy or entered into clinical trial with the placebo control arm versus 5-FU/ Leucovorin with or without oxaliplatin.

 

Keywords:


00-04

SLN detection for prostate and bladder cancer using robot-assisted surgery

Timothy Wilson

John Wayne Cancer Center, Santa Monica, United States

Anatomic studies and experience with lymphadenectomy have demonstrated that the lymph drainage for prostate and bladder cancers is wide, bilateral and unpredictable. Clinical trials in muscle invasive bladder cancer patients generally indicate a prognostic and therapeutic benefit of extended bilateral pelvic lymphadenectomy during radical cystectomy compared to patients who undergo a limited pelvic lymphadenectomy during radical cystectomy. The therapeutic benefit of pelvic lymphadenectomy for men undergoing radical prostatectomy is less clear and a heated topic of debate among urological surgeons. In either disease, lymphadenectomy is time consuming and adds to morbidity. The need for sentinel lymph node technique is badly needed. Technology is available now using robot-assisted surgery which open the door for identifying sentinel node metastasis in patients with bladder cancer and prostate cancer.

Keywords:

Saturday, April 22, 2017 - Emerald Ballroom - 10:35 - 12:20

Mini-Symposium (CME) - PATIENT SUMMIT - Lymphedema Following Cancer Therapy

00-01

Introduction of Lymphedema

Saskia Thiadens

Founder National Lymphedema Network, San Francisco, United States

BACKGROUND:  Lymphedema acquired after cancer surgery is a disfiguring condition and the incidence varies widely in  patients following axillary, inguinal and cervical dissection and/or radiation.  Not until the last 3 decades that the lymphatic system has come to the forefront in medicine, and patients are now receiving appropriate treatment.

METHODS:  As a result of the ignorance of lymphedema and lymphatic system disorders the National Lymphedema Network a non-profit organization was established in 1988 with its mission to increase awareness , education, support and expand the knowledge of this disabling condition, not only to patients, but clinicians, researchers and the general public alike.

RESULTS: Today lymphedema is identified as a legitimate disease in the US, and the current standard of treatment “Complete Decongestive Therapy”(CDT) is in place at University based clinics, hospitals, rehab centers: thousands of clinicians have become certified lymphedema therapists: physicians have included the lymphatic system in their existing practice; research is in progress at Academic Institutions; and patients are actively advocating for their rights and convincing their state and federal representatives to get laws in place to support proper treatment and reimbursement.

CONCLUSIONS: Through determination and  collaborative efforts of dedicated clinicians, researchers, patient advocates,  patients at risk or living with lymphedema are now  diagnosed early and receiving appropriate treatment when needed.

Keywords:


00-02

  Cancer-related lymphedema and incidence

Paula Stewart

Trustpoint Rehab, Murfreesboro, United States

Keywords:


00-03

Imaging techniques for the lymphatic system

Michael Bernas

University of Arizona, Tuscon, United States

Presentation will consist of an overview of current imaging techniques used to visualize the lymphatic system including description of machines, image acquisition, techniques, and sample images. Historical imaging methods will be presented followed by current imaging techniques (nuclear medicine, MR, CT, US, ICG fluorescence) with a focus on lymphangioscintigraphy (LAS). Examples of LAS imaging patterns from patients with and without lymphatic abnormalities will be shown and participants will have a chance to test their new knowledge. Participants completing the session should gain an understanding the imaging of lymphatic and lymphatic-related conditions and both discussion and questions will be highly encouraged. 

Keywords:


00-04

Genetics of breast cancer related lymphedema

Betty Smoot

University of California San Francisco, San Francisco, United States

Findings on the specific risk factors for the development of breast cancer-related lymphedema (BCRL) are variable. Axillary lymph node dissection, higher body mass index, more lymph nodes removed, field of radiation, postoperative seroma, infection, and early edema are linked to the development of BCRL. However, it is unclear why, in the presence of similar phenotypic risk factors, some women develop lymphedema after breast cancer treatment, and others do not.

This presentation will provide a brief overview of the preliminary findings from our research group and others, which suggest that variations in lymphatic, angiogenic, pro-inflammatory cytokine, and potassium channel genes are associated with increased risk for the development of lymphedema after breast cancer treatment. These associations do not, however, explain all of the variability in the occurrence of BCRL. Additional research is needed to identify other biological pathways that contribute to the development and progression of BCRL.

Because the etiology of BCRL is multifactorial, information on genetic variations in biological pathways involved in the lymphatic system that are associated with development of BCRL can lead to the identification of high-risk patients. 

Keywords:


00-05

Treatment and advocacy of cancer related LE

Saskia Thiadens

Founder National Lymphedema Network (NLN), San Francisco, United States

BACKGROUND: Lymphedema (LE), a chronic condition is an abnormal accumulation of protein-rich fluid in the interstitium, which leads to limb swelling.  Secondary or cancer related LE can result from damage to the lymphatic circulation following surgery, chemotherapy and/or radiation.  During the last three decades the lymphatic system and appropriate treatment has come to the forefront of medicine and “Complete decongestive Therapy” (CDT) has become standard of care.

METHODS:  In early 1990 a handful of clinicians came to the realization of the ignorance of LE and appropriate treatment and with support of the National Lymphedema Network (NLN) Complete decongestive Therapy was introduced, and the first Training course for therapists was established.  Since that time clinical experts, researchers, educators, patient advocates and industry representatives have worked tireless hours in creating awareness, risk reduction, education, support, and research in lymphedema management.  Protocols for early detection of LE in cancer patients are in place. Randomized studies in clinical, diagnostic and genetic research are ongoing and surgery in selected patients is promising. Patients and patient advocates have and continue to play a major role in educating local and federal representatives to support appropriate treatment and reimbursement.

CONCLUSION:  Appropriate treatment for LE is in place. Thousands of therapists are certified; pre and post-operative protocols in cancer centers are in place, patients are educated in risk reduction ongoing research is seen at Academic institutions, and most important patients are receiving safe and appropriate treatment and enjoying day to day living.

Keywords:


00-06

Surgical treatments for lymphedema

Jay Granzow

UCLA School of Medicine, Manhattan Beach, United States

Keywords:

Saturday, April 22, 2017 - Oregon Ballroom - 12:20 - 13:30

Industry Sponsored Lunch Session (Non-CME) - Update on Lymphedema Clinical Trials, Sponsored by Eiger

00-01

Update on Lymphedema: Current Clinical Trials at Stanford

Stanley Rockson

Stanford University School of Medicine, Stanford, United States

In recent years, there has been extensive progress in the basic and translational investigation of lymphedema mechanisms, pathogenesis and therapeutics. The presentation will focus on the translational and clinical application of our Stanford research into lymphedema pathogenesis and therapeutics

Keywords: lymphedema, inflammation, patient registries, biospecimens, vascularized lymph node transfer, pharmacology

Saturday, April 22, 2017 - Gold Rush Ballroom - 12:20 - 13:30

Industry Sponsored Lunch Session (Non-CME) - Radiotherapy & Total Cancer Care, Sponsored by Varian

00-01

Technology Evolution for Value-Based Healthcare

Baldassarre Stea

University of Arizona, Tuscon, United States

Keywords:

Saturday, April 22, 2017 - Emerald Ballroom - 13:35 - 15:05

Plenary Session (CME) - Radiotherapy of Cancer

00-01

New Frontiers in Cancer Therapy: Combining Radiotherapy with Immunotherapy to Treat Cancer

Baldassarre Stea

University of Arizona, Tuscon, United States

Keywords:


00-02

Radiotherapy of cancer: adjuvant approach

Roy Abendroth

California Pacific Medical Ctr, San Francisco, United States

Radiation therapy is a valuable tool in treating cancer. This talk will focus on the adjuvant use of radiation therapy, meaning using radiation therapy to address subclinical microscopic disease. We will survey different cancer types as examples of this treatment, and focus somewhat more in depth on the adjuvant use of radiation therapy and melanoma.

Keywords:


00-03

Radiotherapy for oligometastatic cancer

Charles Hsu

University of Arizona, Tuscon, United States

Metastatic cancer represents a spectrum of disease with subsets of patients defined as oligometastatic with a limited number of lesions and a typically less aggressive course.  Localized therapy, including stereotactic body radiation therapy (SBRT), has been shown to improve progression free survival.  Current studies are planned to examine the impact of SBRT on survival for oligometastatic patients.  We will review the history, rationale, data, and future directions for the treatment of oligometastatic malignancies with radiation. 

Keywords:


00-04

New Frontiers in Treatment of Lung Cancer

Charles Hsu

University of Arizona, Tuscon, United States

Both small cell (SCLC) and non-small cell lung cancer (NSCLC) have had significant developments in treatment of both locoregional as well as distant disease.  We will review recent attempts to improve delivery, better personalize treatment, minimize toxicity, and incorporate novel modalities (such as immunotherapy and Tumor Treating Fields) for the treatment of the various stages of lung cancer.

Keywords:

Saturday, April 22, 2017 - Emerald Ballroom - 15:05 - 16:05

Plenary Session (CME) - Trainee & Young Investigator Awardee Presentations

00-01

Intravital imaging of lymph node metastasis reveals its ability to contribute to distant metastasis

Ethel Pereira1, Dmitriy Kedrin1, 12, Giorgio Seano1, Olivia Gautier1, 4, Dennis Jones1, Eelco Meijer1, Shan-Min Chin1, Shuji Kitahara1, Jonathan Chang13, Elizabeth Beech1, Han-Sin Jeong9, Michael Carroll14, Alphonse Taghian10, Timothy Padera1

1Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
2Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
3Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
4Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, United States
5Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
6Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
7Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
8Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
9Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea, Republic of (South)
10Department of Radiation Oncology, Massachusetts General Hospital, Boston, United States
11Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, United States
12Division of Gastroenterology Massachusetts General Hospital and Harvard Medical School, Boston, United States
13Graduate Program in Immunology, Division of Medical Sciences, Harvard Medical School and Program in Cellular and Molecular Medicine, Children’s Hospital Boston, Boston, United States
14Department of Pediatrics, Harvard Medical School and Program in Cellular and Molecular Medicine, Children's Hospital Boston, Boston, United States

Background: The presence of lymph node metastasis in patients with solid tumors is associated with tumor aggressiveness, poorer prognosis and the recommendation for systemic therapy. However, whether tumor cells exit the lymph node and contribute to distant metastases remains controversial. Evidence shows that treating disease in the lymph node improves survival in some patients. In this study, we address the controversy of whether tumor cells colonizing the axillary lymph nodes could disseminate to distant sites.

Materials and Methods: We used syngeneic murine cell lines representing breast (4T1), melanoma (B16F10), head and neck (SCCVII) cancer that spontaneously metastasize to the lymph node. We engineered these cells to express Dendra2, a photoconvertable protein, to determine if metastatic tumor cells from the lymph node are able to seed distant organs like the lung. Using high-resolution multiphoton microscopy and a chronic lymph node window we monitored the behavior of tumor cells in the lymph node and their interaction with blood vessels.

Results: By photoconverting cancer cells only in lymph nodes, we show that spontaneous lymph node metastasis from breast cancer and melanoma mouse models can leave the lymph node and enter the systemic blood circulation. Our data show that the circulating tumor cells originating from the lymph node are viable and are able to proliferate in vitro. Using a combination of intravital microscopy and immunofluorescence staining, our data show that 7.4%±1.8% of isolated tumor cells in the lymph node are intraluminal and an additional 16.3%±1.9% were within 5μm of a blood vessel compared to an expected 8.8%±0.9% if they were randomly distributed. Further, we identified lung micrometastases that originated from the lymph node in both mouse models. Analysis of human metastatic lymph node samples from head and neck cancer patients revealed perivascular association of isolated tumor cells as well as the presence of cancer cells inside blood vessels similar to our observations in the mouse models.

Conclusions: Together, our data show for the first time that in spontaneous breast and melanoma mouse models, tumor cells in the lymph node can invade blood vessels, exit the lymph node and colonize distant organs like the lung. These studies exemplify the importance of treating metastatic disease in the lymph node, which could potentially lead to distant metastasis. 

Keywords: Lymph node, metastasis, circulating tumor cells, intravital imaging, distant metastasis


00-02

Nanoparticle to treat established lymph node metastasis and primary tumor

Isaac Adjei1, 2, Vinod Labhasetwar2, 3

1Department of Biomedical Engineering, University of Florida, Gainesville, United States
2Department of Biomedical Engineering, Lerner Research Institute, Cleveland Clinic, Cleveland, United States
3Taussig Cancer Institute, Cleveland Clinic, Cleveland, United States

Background: Lymph node metastasis increases the probability of cancer relapse and spread to other tissues even after resection of the primary tumor. Here we demonstrate nanoparticles (NPs) that efficiently transport chemotherapy to lymph nodes with metastasis and primary tumor for effective cancer treatment.

Materials and Methods: Nanoparticles (NPs) were formulated from poly (glycolide-co-lactide) using a modified single emulsion-solvent evaporation technique. The NPs were characterized for size, charge and surface morphology. Biodistribution of NPs into lymph nodes and tumors was determined after intravenous (IV) and subcutaneous (SQ) injection in a murine model of prostate cancer. Efficacy of paclitaxel (PTX) loaded NPs at inhibiting the progression of both established tumor and lymph node metastases were also evaluated.

Results: The formulated NPs which were 112 nm in size localized into all lymph nodes along a targeted lymphatic network after SQ injection (Fig.1A). Lymph nodes with metastasis showed a 6-fold greater NP accumulation compared to normal lymph nodes. The NPs ultimately entered the blood and accumulated into tumors at 14% injected dose/g of tumor (ID/GT) compared to 1% ID/GT when injected IV. A single SQ injection of PTX-loaded NPs (PTX-NP-SQ) in murine prostate cancer models with established tumors and lymph node metastasis decreased metastasis burden by 500% (Fig.1B) and increased tumor doubling time by 40% compared to IV injection (PTX-NP-IV). This resulted in 20% increase in average survival time compared to those injected intravenously.

Conclusions: These results demonstrate that NPs can effectively deliver therapeutics primary tumors and lymph node metastases for better cancer treatment.

Keywords:


00-03

Cutaneous Immune Factors and Regression of In-Transit/Lymphatic Melanoma After Intralesional Injection of Bacille Calmette-Guerin

Maris Jones1, Junbao Yang2, Leland Foshag4, Peter Sieling2, Delphine Lee2, 3, Mark Faries4

1Division of Surgical Oncology, John Wayne Cancer Institute, Providence Saint John's Health Center, Santa Monica, CA, Santa Monica, United States
2Dirks/Dougherty Laboratory for Cancer Research, Department of Translational Immunology, John Wayne Cancer Institute, Providence Saint John's Health Center, Santa Monica, CA, Santa Monica, United States
3Division of Dermatology, Los Angeles Biomedical Research Institute, Harbor-UCLA Medical Center, Torrance, CA, Torrance, United States
4Melanoma Research Program, John Wayne Cancer Institute, Providence Saint John's Health Center, Santa Monica, CA, Santa Monica, United States

Background:

Mycobacterium bovis Bacille Calmette-Guerin (BCG) is a known intralesional (ILBCG) therapeutic option for stage III in-transit/lymphatic melanoma. The mechanism of action is unknown; however, regressions in up to 50% of injected lesions and 17% of uninjected lesions have been reported. BCG and other mycobacteria express ligands capable of stimulating γ9δ2 T cells. We hypothesize that γ9δ2 T cells play a role in promoting BCG-mediated antitumor immunity in patients treated with ILBCG.

Materials and Methods:

Eight patients were diagnosed with stage III in-transit/lymphatic melanoma and treated with ILBCG per study protocol. BCG injected and uninjected lesions were resected and subjected to immunohistochemistry and RNAseq gene expression analyses.

Results:

BCG injection induced a significant increase of γ9δ2 T cell infiltration in BCG injected lesions as determined by immunohistochemistry and RNAseq gene expression analyses. Furthermore, BCG injection elicited: the expression of chemokines (such as CXCL9, 10, and 11) capable of attracting γδT cells, the expression of antigenic ligands (BTN3A1) capable of activating γ9δ2 T cells in BCG injected lesions, and the secretion of cytokines by activated γδ T cells. Interestingly, we found that γδ T cell infiltration was associated with the regression of BCG uninjected lesions.

Conclusions:

These data strongly suggest that γ9δ2 T cells contribute to in transit/lymphatic melanoma regressions induced by ILBCG. γ9δ2 T cells may serve as a useful target for future immunotherapeutic study.

Keywords:


00-04

Epigenetic alterations of spliceosome factors are associated with organ-specific melanoma metastasis

Diego Marzese1, Nellie Nelson2, Ling Takeshima1, Sandy Hsu2, Dave Hoon1, 2

1Department of Translational Molecular Medicine, John Wayne Cancer Institute at Providence Saint John's Health Center, Santa Monica, United States
2Sequencing Center, John Wayne Cancer Institute at Providence Saint John's Health Center, Santa Monica, United States

Background: Metastatic melanoma cells exhibit an extraordinary phenotypic plasticity, not only in adapting to unfamiliar microenvironments, but also in surviving aggressive treatments and immune response. A major source of phenotypic variability is the alternative splicing of the messenger RNA (mRNA). This process is catalyzed by one of the most complex cellular machineries, the spliceosome, which is composed of ribonucleoproteins and polypeptides denominated spliceosome factors (SF). Changes in the expression level of SFs lead to a splicing reprogramming that has been linked to different types of cancer progression. We have recently demonstrated that the interplay between epigenomic and transcriptomic alterations of SFs generates melanoma cells with an enhanced predisposition for brain metastasis. Material and Methods: Using the Illumina Human 450K BeadChip technology, we generated genome-wide DNA methylation maps of 133 melanoma specimens, including melanocytes and nevi (n=9), primary melanomas (PRM; n=15), lymph node metastases (LNM; n=27), extracranial metastases (n=37), and brain metastases (MBM; n=45). This analysis included 15 patients with PRM or LNM paired with multiple (>2) distant organ metastases (DOM). To ensure highly representative DNA methylation patterns, all the specimens were microdissected. Additionally, DNA methylation and gene expression data for DOM (n=67) generated by the Cancer Genome Atlas (TCGA) were integrated to validate our findings. Results: This epigenomic profiling allowed us to evaluate changes in the DNA methylation levels of 6,370 genomic regions affecting 356 genes encoding splicing and mRNA editing factors. Overall, 58 genomic regions affecting 12 genes were differentially methylated during melanoma progression to metastasis (Absolute [PRM–DOM]>0.3; Wilcoxon test, FDR-p<0.01). In agreement with our previous studies, this analysis identified epigenetic alterations on ESRP1, ESRP2 and PTBP1 genes which, as we recently described, are involved in melanoma metastasis in an independent patient cohort. These regions were primarily located in gene body CpG island regions suggesting a positive influence on gene expression rates. Interestingly, liver metastases presented hypermethylation of 16 genomic regions overlapping the ADARB2 gene which regulates the splicing of pre-micro-RNAs. Conversely, bowel metastases showed hypomethylation of the ADARB2 gene. ESRP1 and ESRP2 genes were hypomethylated in spleen and liver metastases but hypermethylated in lung and brain metastases. Analysis of TCGA RNA-sequencing data showed changes in the splicing program of several targets of these SFs. Conclusions: Aberrant splicing has a significant impact on melanoma progression to metastasis. The characterization of epigenetic regulation of SF genes influenced by metastasis microenvironment opens up a new avenue for understanding the affinity of melanoma cells for specific metastatic niches.

Keywords: Epigenomic Regulation, Alternative Splicing, Melanoma Metastasis, DNA methylation


00-05

Innovations in Lymphatic Microsurgery -  MLVA and FLA-LVSP: The Complete Treatment Package

Corrado Campisi

Department of Plastic, Reconstructive & Aesthetic Surgery ,Unit for Lymphatic Microsurgery, Salus Hospital, GVM Care & Research, Reggio Emilia, Italy

Background: Despite the common assertion that little is understood about the pathophysiology of disorders of the lymphatic system and that satisfactory treatments are lacking, there is a rich history in medicine dating back close to 500 years of research on the lymphatic system. In the past 40 years there has been a burgeoning interest in research on, and clinical application for, diseases involving the lymphatic system. Development of microsurgical techniques has significantly advanced the field of Lymphology and the treatment of lymphedema secondary to oncological surgery.

 

Methods:The author reports on the development of microsurgery for lymphedema and new developments with particular reference to the wealth of experience in treating patients in Genoa, Italy. The “single-site” microsurgical technique is based on the identification, with the use of Patent Blue Dye / Fluorescent Micro-lymphography (PDE Test), of the lymhatics in the axillary or inguinal-crural regions (“single-site”) and the completion of derivative multiple lymphatic–venous anastomoses (MLVA). The lymphatics are anastomosed with telescopic technique to multiple tributary vein, for example, the axillary vein or the saphenous vein, depending on the affected limb.

In cases of advanced lymphedema, patients are treated by a recently developed Fibro-Lipo-Lymph-Aspiration technique to improve this chronic swelling, using a Lymph Vessel Sparing Procedure (FLLA-LVSP). Using microlymphography techniques to highlight the lymphatic pathways, the excess adipose tissue was carefully aspirated with the FLLA-LVSP procedure.

 

Results: With the “single-site” MLVA, 4000 patients obtained significant reductions in excess limb volume of over 84%, with an average follow-up of 15 years or more. Over 86% of patients with earlier stages of disease (stage IB or IIA) progressively stopped using conservative therapies and 42% of patients with later stages (stages IIB and III) decreased the frequency of physical therapies. DLA attacks considerably reduced by over 91%.

For 250 advanced cases, 0.80L on average for the upper limb and 2.42L for the lower limb was removed with the FLLA-LVSP. For the upper limb, there was an average pre-surgery excess volume of 20.19%, which reduced to 2.68% after the FLLA-LVSP (Z-score =-6.90, p<0.001). Similarly, for the lower limb, there was an average pre-surgery excess limb volume of 21.24% and a reduction to 2.64% post-operatively (Z-score=-3.57, p<0.01). No episodes of post-operative infection occurred. See: Figure 1, pre, post-MLVA, and post-FLLA-LVSP.

 

Conclusion: MLVA or techniques when performed at a single-site produce excellent outcomes in the treatment of secondary lymphedemas, giving the possibility of a complete restoration of lymphatic flow in early stages of disease when tissue changes are minimal. In cases of advanced lymphedema, the FLLA-LVSP is efficient. It is possible to complete an entire leg within 90 minutes. Recovery time is short and cosmetic results are immediate. More importantly, the removal of excess tissue is completed without further damage to lymphatic vessels, providing evidence of the efficacy of FLLA-LVSP in limb-reshaping, whilst maintaining the optimal lymphatic flow restored by Lymphatic Microsurgery. With this procedure, in some patients, it is possible to achieve almost 100% volume reduction and to eliminate the use of compressive stockings.

Keywords: "single-site" lymphatic microsurgery, advanced secondary lymphedema, liposuction, lymph vessel sparing, fibroadipose tissue,
indocyanine green microlymphography


00-06

Cryofluorescence Tomography for Single-Cell and Micron-Level of Analysis of Cancer Metastasis and the Lymphovascular System – Bridging the Microscopic/Mesoscopic Divide

Robert Holt1, Mark Bordo2, Sanjana Pannem1, Jacob Hesterman1, John Frangioni2, Jack Hoppin1, Ajay Verma3

1inviCRO, LLC, Boston, United States
2Curadel, LLC, Marlborough, United States
3Biogen, Cambridge, United States

Background: Fluorescence molecular imaging is a valuable imaging modality at both the cellular and the whole-body imaging regimes.  Although valuable for pathological analysis, fluorescence histology has a severely limited field of view.  Similarly, bulk fluorescence imaging (such as with epi-illumination schemes) is either highly surface weighted, or limited to a resolution of a few cubic millimeters at best using tomographic approaches.  Cryofluorescence tomography (CFT) is proposed as an imaging scheme to bridge the gap between microscopic and mesoscopic imaging methods, while still exploiting the vast and growing library of fluorescent tracers.

CFT is comprised of serial cryoslicing and high resolution imaging of a tissue sample block at multiple visible and NIR wavelengths. For each slice, a series of fluorescence images is captured simultaneous with white light images. In this way, valuable molecular fluorescence information is gathered along with anatomical information. Furthermore, imaging is performed on the block face rather than on transferred slices, thus completely eliminating registration artifacts caused by tissue deformation, folding, and tearing. Imaging is performed by interfacing a cryoslicer (Leica 3050s) with a surgical fluorescence imaging system (FLARE model R1, Curadel, Marlborough, MA). This imaging scheme can be used to automatically recover 3D fluorescence molecular information at mesoscopic resolution, with imaging times of under an hour for a whole mouse.

Materials and Methods: To test this imaging scheme, a rat was injected intrathecally with a near-infrared fluorophore having peak emission at 800 nm and intravenously with a near-infrared fluorophore having peak emission at 700 nm. At one hour, the kidneys were resected and imaged using CFT with 25 micron slices to be able to follow tracer clearance from the CSF to the blood and blood to urine using near-infrared fluorescence. Basic structural analysis based on the white light images was also performed.  A color-based segmentation was performed to segment the vasculature of the sample.

Results: A comparison of the different imaging channels (Figure) shows a snapshot comparison of the renal clearance of the two tracers through different injection routes at approximately 25 micron voxel size.  The images show that it is possible to resolve molecular and structural features on the same order of magnitude as lymphatic channels and the vasculature.

Conclusions: Using this new CFT system, two independent biomarkers can be imaged simultaneously over an entire mouse or rat with resolution far superior to whole-body nuclear imaging methods. By utilizing FLARE® lipophilic cell tracking dyes, single cells can be labeled and tracked anywhere in the animal. This should be particularly useful for tracking the migration and homing of metastatic cells. Using NIR fluorescent targeting ligands, virtually any in vivo target can be assessed over large volumes of tissue. Multiple examples of CFT for imaging cancer metastasis, lymphovascular processes, and biomarkers will be presented.

Keywords: fluorescence, histology, cryoimaging, cryofluorescence

Saturday, April 22, 2017 - Emerald Ballroom - 16:05 - 16:15

Plenary Session (Non-CME) - Closing Remarks, Stanley, P.L. Leong, MD, FACS