2813 Comprehensive genomic profiling of clinically advanced medullary thyroid carcinoma

2813 Comprehensive genomic profiling of clinically advanced medullary thyroid carcinoma

S562 cells was assessed and compared with the parental cells. Furthermore their sensitivity to carboplatin and 5 fluorouracil (5-FU) was assessed using...

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S562 cells was assessed and compared with the parental cells. Furthermore their sensitivity to carboplatin and 5 fluorouracil (5-FU) was assessed using flow cytometry (annexin V/PI staining) and caspase-3 enzyme assay. Results: Immunohistochemical analysis revealed increased cytoplasmic S100A2 expression in OSCCs (%) as compared to normal mucosa. Kaplan Meier survival analysis revealed that 70.3% OSCC patients which over express S100A2 have significantly reduced disease free survival (p = 0.001). Cox multivariate regression analysis validated the association of S100A2 with tumor recurrence. Down regulation of S100A2 expression in oral cancer cells inhibited their proliferation and induced apoptosis. Notably, annexin V/PI assays exhibited increased apoptosis in SCC4S100A2S1 cells on treatment with carboplatin and 5-FU as compared with SCC4 control cells. SCC4S100A2S1 cells demonstrated enhanced sensitivity to carboplatin and 5-FU which was associated with an increased Caspase 3 activity. Conclusion: Our study demonstrate the association of cytoplasmic S100A2 overexpression with poor prognosis in OSCC patients. Depletion of S100A2 induced sensitivity of oral cancer cells to chemotherapeutic agents, suggesting it to be a potential therapeutic target. No conflict of interest.

2813 POSTER Comprehensive genomic profiling of clinically advanced medullary thyroid carcinoma A. Heilmann1 , V. Subbiah2 , K. Wang3 , J. Elvin4 , J. Chmielecki5 , R. Yelensky3 , J.A. Vergilio4 , R. Erlich1 , D. Lipson3 , J. Ross4 , V. Miller6 , S. Ali7 , P. Stephens3 . 1 Foundation Medicine, Inc, Biomedical Informatics, Cambridge, USA; 2 MD Anderson Cancer Center, Investigational Cancer Therapeutics, Houston, USA; 3 Foundation Medicine, Inc, Research and Development, Cambridge, USA; 4 Foundation Medicine, Inc, Pathology, Cambridge, USA; 5 Foundation Medicine, Inc., Cancer Genomics, Cambridge, USA; 6 Foundation Medicine, Inc, Medical Group, Cambridge, USA; 7 Foundation Medicine, Inc, Clinical Development, Cambridge, USA Background: Medullary thyroid carcinoma (MTC) accounts for 3−5% of all thyroid cancers and metastatic MTC carries a poor prognosis. The targeted therapies vandetanib and cabozantinib have improved the outcomes for patients with metastatic MTC. However, it is critical to understand predictive markers of response to these agents and to develop strategies to overcome resistance to these therapies. Material and Methods: DNA was extracted from 40 microns of FFPE sections from 34 MTC. The median age was 52 years (range 20−84). Nextgeneration sequencing (NGS) based comprehensive genomic profiling (CGP) was performed on hybridization-captured, adaptor ligation based libraries to high, uniform coverage for all coding exons of 182–315 cancerrelated genes plus 14−28 genes frequently rearranged in cancer. The results were evaluated for all classes of genomic alterations (GA), including base substitutions, small INDELs, copy number alterations, and gene fusions. Clinically relevant GA (CRGA) were defined as GA linked to drugs on the market or under evaluation in mechanism driven clinical trials. Results: All 34 MTC (100%) harbored at least one GA, with 82 total alterations identified for an average of 2.4 alterations per tumor (range 1−11). 68% (56/82) of the genomic alterations were considered clinically relevant (CRGA), with an average of 1.6 CRGA per tumor (range 1−7). RET was mutated in 88% (30/34) of cases, with RET M918T responsible for 70% (21/30) of the RET alterations. The other RET alterations were a small INDEL or missense mutations in the extracellular domain, including RET E632_L633del, C634R, C620R, and C618G/R/S, a known vandetinib resistance mutation in the kinase domain (V804M), and RET amplification. Two of the four RET wild-type patients harbored mutations in KRAS or HRAS (3%, 1/34 each). The next most frequent genomic alterations were amplifications of CCND1, FGF3, and FGF19 as well as alterations in CDKN2A (9%, 3/34 each). RAD50, RBM10, TP53, and VHL were each altered in 6% (2/34). One MTC with a RET M918T mutation experienced a durable response to progressively dose-escalated vandetinib, but then developed resistance to this therapy. The addition of the mTOR inhibitor everolimus to continued vandetanib treatment achieved a 25% reduction of tumor volume (RECIST 1.1) in this patient for 8 months, which is ongoing. Other outcomes for MTC cases will be presented. Conclusion: Comprehensive genomic profiling identified the full-breadth of RET mutations in metastatic MTC and possible co-operating oncogenic driver alterations potentially important for the selection of therapy. This approach may refine the use of RET-targeted therapy for these patients. Finally, co-targeting the mTOR pathway is a promising new strategy with

Abstracts potential to overcome resistance to vandetanib in patients with advanced disease. Conflict of interest: Ownership: All authors except VS have equity iinterest in Foundation Medicine Inc. Other Substantive Relationships: All authors except VS are employees of Foundation Medicine Inc. 2814 POSTER Understanding the mechanism of extracellular vesicles release in head and neck cancer F. Giudice1 , B. Rodrigues1 , T. Lacerda1 , R. Cartaxo1 , A. Baptistella1 , M. Dias1 , L. Kowalski1 , V. Martins1 . 1 A.C.Camargo Cancer Center, ˜ Paulo, Brazil International Research Center, Sao Background: Head and Neck Squamous Cell Carcinoma (HNSCC) is considered one of the six most common malignancies worldwide. It represents more than 90% of all cancers that occur throughout the upper aerodigestive tract. Recent evidences have shown that the tumor development and the metastatic progression are supported by extracellular vesicles (EVs), which are released from the tumor cells to the blood or lymphatic system. The cellular secretion of EVs represents a significant system for short and long-range cell-to-cell communication. In this way, it is essential to understand the molecular mechanisms involved in EVs biogenesis and secretion. Herein, we sought to evaluate key proteins related to endo/exocytosis processes that could regulate EVs’ release. In addition, we tried to recognize the function of HNSCC EVs upon dysplastic keratinocytes and microenvironment cells such as endothelial cells. Material and Methods: EVs derived from HNSCC cell lines (SCC-9, Cal27 and FaDu) were isolated by ultracentrifugation and quantified using Nanoparticle Tracking Analysis (NanoSight® ). A screen targeting for the components of EVs’ biogenesis was also performed through western blotting and the lysosomal compartment was followed by immunofluorescence in live cells. Due to previous results from our laboratory using astrocytes, we also investigated the role of prion protein (PrPC ) in EVs secretion. Finally, the invasion and angiogenic potential of dysplastic keratinocytes (DOK cell line) and endothelial cells (HUVEC cell line) were evaluated, respectively, after treatment with EVs secreted by HNSCC. Results: Comparing the three cell lines, FaDu cells secreted less EVs than SCC-9 and Cal27. Remarkably, FaDu cells presented higher expression of Rab7, a small GTPase that directs vesicles to lysosomes, and lower levels of PrPC , a scaffold molecule, which modulates the organization of signaling complexes. Immunofluorescence experiments showed the presence of numerous perinuclear clusters positive for LAMP-1 (lysosome marker) in FaDu and Cal27 cells, while SCC-9 showed LAMP1-positive organelles dispersed throughout its cytoplasm. These findings indicate that the majority of EVs formed by FaDu cells can be driven to lysosome compartment instead of being released to the extracellular milieu. Functional assays showed that Rab7 knockdown or PrPC overexpression increase EV secretion. Interestingly, the treatment of DOK and HUVEC cells with EVs derived from HNSCC cells enhanced the invasion and angiogenic potential of keratinocytes and endothelial cells, respectively. Conclusions: Taken together, our data highlights a role for Rab7 and PrPC proteins in the modulation of EVs secretion by HNSCC. Moreover, the in vitro functional assays suggest the clinical importance of regulating EVs secretion for the controlling of tumor progression and aggressiveness. No conflict of interest. 2815 POSTER Functional polymorphisms in antioxidant genes and occurrence of recurrent disease in patients with Hurthle cell carcinoma G. Pilko1 , B. Krhin2 , N. Besic1 , K. Goricar3 , V. Dolzan3 . 1 Institute of Oncology Ljubljana, Department of Surgical Oncology, Ljubljana, Slovenia; 2 Institute of Oncology, Department of Laboratory Diagnostics, Ljubljana, Slovenia; 3 Institute of Biochemistry, Faculty of Medicine, University of Ljubljana, Pharmacogenetics Laboratory, Ljubljana, Slovenia Background: Hurthle cells of the thyroid gland are very rich in mitochondria and oxidative enzymes. As the high level oxidative metabolism may lead to increased production of reactive oxygen species (ROS), antioxidant defense is of great importance. Common functional polymorphisms in antioxidant genes can decrease individual’s capacity for antioxidant defense. The resulting higher level of oxidative stress has been associated with increased risk for many human cancers, but there are no data on polymorphisms in antioxidant genes in Hurthle cell adenoma (HCA) and Hurthle cell carcinoma (HCC). In the present study, we investigated