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Background: The MSI/MSS status does not fully explain cancer immunotherapy response in colorectal cancer. Thus, we developed a colorectal cancer-specific method that predicts cancer immunotherapy response.Methods: We used gene expression data of 454 samples (MSI = 131, MSI-L = 23, MSS = 284, and Unknown = 16) and developed a TMEPRE method that models signatures of CD8+ T-cell infiltration and CD8+ T-cell exhaustion states in the tumor microenvironment of colorectal cancer. TMEPRE model was validated on three RNAseq datasets of melanoma patients who received pembrolizumab or nivolumab and one RNAseq dataset of purified CD8+ T cells in different exhaustion states.Results: TMEPRE showed predictive power in three datasets of anti-PD1-treated patients (p = 0.056, 0.115, 0.003). CD8+ T-cell exhaustion component of TMEPRE model correlates with anti-PD1 responding progenitor exhausted CD8+ T cells in both tumor and viral infection (p = 0.048, 0.001). The global pattern of TMEPRE on 454 colorectal cancer samples indicated that 10.6% of MSS patients and 67.2% of MSI patients show biological characteristics that can potentially benefit from anti-PD1 treatment. Within MSI nonresponders, approximately 50% showed insufficient tumor-infiltrating CD8+ T cells and 50% showed terminal exhaustion of CD8+ T cells. These terminally exhausted CD8+ T cells coexisted with signatures of myeloid-derived suppressor cells in colorectal cancer.Conclusion: TMEPRE is a colorectal cancer-specific method. It captures characteristics of CD8+ T-cell infiltration and CD8+ T-cell exhaustion state and predicts cancer immunotherapy response. A subset of MSS patients could potentially benefit from anti-PD1 treatment. Anti-PD1 resistance MSI patients with insufficient infiltration of CD8+ T cells or terminal exhaustion of CD8+ T cells need different treatment strategies.

BackgroundFOLFOX is a combination of drugs that is widely used to treat colorectal cancer. The response rate of FOLFOX in colorectal cancer(CRC) is 30-50%. We develop a method that analyzes mechanisms of FOLFOX resistance and predicts whether a patient will benefit from FOLFOX.MethodsGene expression data of 83 stage IV CRC tumor samples (FOLFOX responder n=42, non-responder n=41) were used to develop a supervised learning method IML and analyze subgroups of FOLFOX resistance mechanism. Datasets of 32 FOLFOX treated stage IV CRC patients and 55 FOLFOX treated stage III CRC patients were used as independent validations.ResultsAn iterative supervised learning (IML) method identified two distinct subgroups of CRC patients who resist FOLFOX. Each subgroup relies on different types of DNA damage repair proteins and they are mutually exclusive. Protein-protein networks showed the main mechanism might be the synergistic effect of resisting apoptosis and an altered cell cycle. IML method was validated in two independent validation sets, one FOLFOX treated stage IV CRC patients(HR=2.6, p-value=0.02, 3-years survival rate of the predicted responder group 61.9%, predicted nonresponder group 18.8%) and one FOLFOX treated stage III CRC patients (estimated HR=2.36, p-value=0.02). A subgroup of mesenchymal subtype patients shows the pattern as FOLFOX responders.ConclusionsIML method reflects the underlying biology of FOLFOX resistance and predicts FOLFOX response.

In most colorectal cancer (CRC) patients, outcome cannot be predicted because tumors with similar clinicopathological features can have differences in disease progression and treatment response. Therefore, a better understanding of the CRC biology is required to identify those patients who will benefit from chemotherapy and to find a more tailored therapy plan for other patients. Based on unsupervised classification of whole genome data from 188 stages I–IV CRC patients, a molecular classification was developed that consist of at least three major intrinsic subtypes (A-, B- and C-type). The subtypes were validated in 543 stages II and III patients and were associated with prognosis and benefit from chemotherapy. The heterogeneity of the intrinsic subtypes is largely based on three biological hallmarks of the tumor: epithelial-to-mesenchymal transition, deficiency in mismatch repair genes that result in high mutation frequency associated with microsatellite instability and cellular proliferation. A-type tumors, observed in 22% of the patients, have the best prognosis, have frequent BRAF mutations and a deficient DNA mismatch repair system. C-type patients (16%) have the worst outcome, a mesenchymal gene expression phenotype and show no benefit from adjuvant chemotherapy treatment. Both A-type and B-type tumors have a more proliferative and epithelial phenotype and B-types benefit from adjuvant chemotherapy. B-type tumors (62%) show a low overall mutation frequency consistent with the absence of DNA mismatch repair deficiency. Classification based on molecular subtypes made it possible to expand and improve CRC classification beyond standard molecular and immunohistochemical assessment and might help in the future to guide treatment in CRC patients.

Background Classification of breast cancers into molecular subtypes may be important for the proper selection of therapy for patients as tumors with seemingly similar biology can have strikingly different clinical outcomes. We analyzed response to neo-adjuvant chemotherapy as a function of molecular sub-types and show patient survival from the multi-center neo adjuvant trial, I-SPY I (CALGB 150007/150012; ACRIN 6657). Previous analyses had shown that breast cancer subtypes as identified by immunohistochemistry or molecular analyses, have distinct clinical outcome (Esserman, ASCO 2009). Here, we analyze how the 70-gene signature (MammaPrint) together with an 80-gene molecular subtyping profile (BluePrint=Basal-type, Luminal-type, ERBB2-type; de Snoo, ASCO, 2010 and manuscript under review) can sub-classify patients. Materials: This study was carried out on genomics data of core needle biopsies from 149 out of 221 patients enrolled in the I-SPY I trial, a multi-center trial designed to identify predictive markers of pathological complete response (pCR) and disease-free survival (DFS) in women with locally advanced breast cancer (3 cm or greater). All women received neoadjuvant Doxorubicin & Cytoxan followed by paclitaxel chemotherapy. The 70-gene and the subtyping signatures, Basal, Luminal and ERBB2-type were determined on 44K Agilent arrays available through the I-SPY 1 data portal (Esserman, ASCO, 2009 and manuscript under review). Results: The 70-gene signature classified 9% of patients (13/149) as Low Risk, of whom one patient was ERBB2-type, and the other 12 were Luminal-type. None of these patients experienced a pCR. The remaining 136 were classified as 70-gene High Risk (91%), of whom 47% were classified as Luminal-type with a pCR rate of 13%, 41% were Basal-type with a pCR rate of 34%, and 12% were ERBB2-type with a pCR rate of 56%. Patients with BluePrint Basal-type tumors had a 5-year DFS of 61%; ERBB2-type had a 5-year DFS of 78%; 70-gene High Risk/Luminal-type had a 5-year DFS of 73% and 70-gene Low Risk/Luminal-type showed 5-year DFS of 100%. The molecular subtype classification shows significant association to clinically assessed receptor status. However, clinically assessed HER2+ patients were distributed across all molecular subtypes, where ER+HER+ are predominantly classified as Luminal-type. Conclusion: This study was performed with the I-SPY I dataset, which provides a platform to compare, contrast & combine marker signatures to tailor therapy. We show how combining BluePrint, with MammaPrint risk-classification can detect specific groups of patients who are at high risk of recurrence and who would have a higher likelihood to benefit from chemotherapy. Furthermore, MammaPrint Low Risk patients do not benefit from neo-adjuvant therapy, though have excellent survival rates, suggesting that the Low Risk Luminal-type patients could be managed conservatively. Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-06-20.

Background MammaPrint was developed as a diagnostic tool to predict risk of breast cancer metastasis using the expression of 70 genes. To better understand the tumor biology assessed by MammaPrint, we interpreted the biological functions of the 70-genes and showed how the genes reflect the six hallmarks of cancer as defined by Hanahan and Weinberg. Results We used a bottom-up system biology approach to elucidate how the cellular processes reflected by the 70-genes work together to regulate tumor activities and progression. The biological functions of the genes were analyzed using literature research and several bioinformatics tools. Protein-protein interaction network analyses indicated that the 70-genes form highly interconnected networks and that their expression levels are regulated by key tumorigenesis related genes such as TP53, RB1, MYC, JUN and CDKN2A. The biological functions of the genes could be associated with the essential steps necessary for tumor progression and metastasis, and cover the six well-defined hallmarks of cancer, reflecting the acquired malignant characteristics of a cancer cell along with tumor progression and metastasis-related biological activities. Conclusion Genes in the MammaPrint gene signature comprehensively measure the six hallmarks of cancer-related biology. This finding establishes a link between a molecular signature and the underlying molecular mechanisms of tumor cell progression and metastasis.

Colorectal cancer patients with the BRAF(p.V600E) mutation have poor prognosis in metastatic setting. Personalized treatment options and companion diagnostics are needed to better treat these patients. Previously, we developed a 58-gene signature to characterize the distinct gene expression pattern of BRAF-mutation-like subtype (accuracy 91.1%). Further experiments repurposed drug Vinorelbine as specifically lethal to this BRAF-mutation-like subtype. The aim of this study is to translate this 58-gene signature from a research setting to a robust companion diagnostic that can use formalin-fixed, paraffin-embedded (FFPE) samples to select patients with the BRAF-mutation-like subtype. BRAF mutation and gene expression data of 302 FFPE samples were measured (mutants = 57, wild-type = 245). The performance of the 58-gene signature in FFPE samples showed a high sensitivity of 89.5%. In the identified BRAF-mutation-like subtype group, 50% of tumours were known BRAF mutants, and 50% were BRAF wild-type. The stability of the 58-gene signature in FFPE samples was evaluated by two control samples over 40 independent experiments. The standard deviations (SD) were within the predefined criteria (control 1: SD = 0.091, SD/Range = 3.0%; control 2: SD = 0.169, SD/Range = 5.5%). The fresh frozen version and translated FFPE version of this 58-gene signature were compared using 170 paired fresh frozen and FFPE samples and the result showed high consistency (agreement = 99.3%). In conclusion, we translated this 58-gene signature to a robust companion diagnostic that can use FFPE samples.

Study design: Cadaveric and clinical studies. Objectives: To investigate the safety and efficiency of modified decancellation posterior closing-wedge osteotomy for traumatic fixed kyphotic deformity of thoracolumbar spine. Methods: Single-level vertebral osteotomy was performed on two groups of fresh-frozen human cadaveric lumbar spines. Group I underwent conventional decancellation posterior closing-wedge osteotomy, and Group II underwent our modified cancellation posterior closing-wedge osteotomy. Sagittal plane angulation as well as anterior height and distance between the most cephalad and caudal endplate were measured before and after osteotomy. Twenty six cases of old thoracolumbar fractures with spinal cord injury were recruited in this study. The mean age was 35.6 years. The mean time from injury to operation was 25 months ranging from 3 months to 11 years. Prior to the index surgery, 9 patients received conservative treatment, and 17 patients underwent surgical treatment. There was complete paraplegia in 10 cases and incomplete paraplegia in 14 (Frankel B in 2 cases, C 10 and D 2). Two patients had no neurological deficit. All patients suffered from low back pain, the average score of Visual Analog Scale (VAS) was 4.5 (range 2.5-6.0). The patients were found to have a mean remaining kyphotic deformity of 35° (range 20°-75°). According to the deformity angles conventional or modified decancellation posterior closing-wedge osteotomy was performed. Results: The mean correction was 36° ± 3.6° for Group I and 49° ± 2.0° for Group II. The mean change in anterior height was only 2–4 mm for Group I and II. All cases were followed up for 10 months to 6 years with a mean of 12.5 months. Successful decompression and satisfied correction of kyphosis was noticed. The post-operatively mean angle of kyphosis deformity was 10.8°, ranging from 0° to 40°. Neurological function recovery was noted in 50% of cases. For complete spinal cord injury, 30% of cases had partial recovery (sensation) whereas recovery was observed in 64.3% of cases with incomplete spinal cord injury. The statistical difference between the groups was p

Microsatellite instability (MSI) occurs in 10–20% of colorectal tumours and is associated with good prognosis. Here we describe the development and validation of a genomic signature that identifies colorectal cancer patients with MSI caused by DNA mismatch repair deficiency with high accuracy. Microsatellite status for 276 stage II and III colorectal tumours has been determined. Full-genome expression data was used to identify genes that correlate with MSI status. A subset of these samples (n = 73) had sequencing data for 615 genes available. An MSI gene signature of 64 genes was developed and validated in two independent validation sets: the first consisting of frozen samples from 132 stage II patients; and the second consisting of FFPE samples from the PETACC-3 trial (n = 625). The 64-gene MSI signature identified MSI patients in the first validation set with a sensitivity of 90.3% and an overall accuracy of 84.8%, with an AUC of 0.942 (95% CI, 0.888–0.975). In the second validation, the signature also showed excellent performance, with a sensitivity 94.3% and an overall accuracy of 90.6%, with an AUC of 0.965 (95% CI, 0.943–0.988). Besides correct identification of MSI patients, the gene signature identified a group of MSI-like patients that were MSS by standard assessment but MSI by signature assessment. The MSI-signature could be linked to a deficient MMR phenotype, as both MSI and MSI-like patients showed a high mutation frequency (8.2% and 6.4% of 615 genes assayed, respectively) as compared to patients classified as MSS (1.6% mutation frequency). The MSI signature showed prognostic power in stage II patients (n = 215) with a hazard ratio of 0.252 (p = 0.0145). Patients with an MSI-like phenotype had also an improved survival when compared to MSS patients. The MSI signature was translated to a diagnostic microarray and technically and clinically validated in FFPE and frozen samples. Copyright © 2012 Pathological Society of Great Britain and Ireland.

Objective To develop gene expression profiles that characterise KRAS -, BRAF - or PIK3CA -activated- tumours, and to explore whether these profiles might be helpful in predicting the response to the epidermal growth factor receptor (EGFR) pathway inhibitors better than mutation status alone. Design Fresh frozen tumour samples from 381 colorectal cancer (CRC) patients were collected and mutations in KRAS , BRAF and PIK3CA were assessed. Using microarray data, three individual oncogenic and a combined model were developed and validated in an independent set of 80 CRC patients, and in a dataset from metastatic CRC patients treated with cetuximab. Results 175 tumours (45.9%) harboured oncogenic mutations in KRAS (30.2%), BRAF (11.0%) and PIK3CA (11.5%). Activating mutation signatures for KRAS (75 genes), for BRAF (58 genes,) and for PIK3CA (49 genes) were developed. The development of a combined oncogenic pathway signature-classified tumours as ‘activated oncogenic’, or as ‘wildtype-like’ with a sensitivity of 90.3% and a specificity of 61.7%. The identified signature revealed other mechanisms that can activate ERK/MAPK pathway in KRAS , BRAF and PIK3CA wildtype patients. The combined signature is associated with response to cetuximab treatment in patients with metastatic CRC (HR 2.51, p Conclusion A combined oncogenic pathway signature allows the identification of patients with an active EGFR-signalling pathway that could benefit from downstream pathway inhibition.

Classification of breast cancer into molecular subtypes maybe important for the proper selection of therapy, as tumors with seemingly similar histopathological features can have strikingly different clinical outcomes. Herein, we report the development of a molecular subtyping profile (BluePrint), that enables rationalization in patient selection for either chemotherapy or endocrine therapy prescription. An 80-Gene Molecular Subtyping Profile (BluePrint) was developed using 200 breast cancer patient specimens and confirmed on four independent validation cohorts (n = 784). Additionally, the profile was tested as a predictor of chemotherapy response in 133 breast cancer patients, treated with T/FAC neoadjuvant chemotherapy. BluePrint classification of a patient cohort that was treated with neoadjuvant chemotherapy (n = 133) shows improved distribution of pathological Complete Response (pCR), among molecular subgroups compared with local pathology: 56% of the patients had a pCR in the Basal-type subgroup, 3% in the MammaPrint Low-risk, Luminal-type subgroup, 11% in the MammaPrint High-risk, Luminal-type subgroup, and 50% in the HER2-type subgroup. The group of genes identifying Luminal-type breast cancer is highly enriched for genes having an Estrogen Receptor binding site proximal to the promoter-region, suggesting that these genes are direct targets of the Estrogen Receptor. Implementation of this profile may improve the clinical management of breast cancer patients, by enabling the selection of patients who are most likely to benefit from either chemotherapy or from endocrine therapy.

Only a minority of breast cancer patients responds to chemotherapy and we lack predictive biomarkers that help to select a patient-tailored therapy that takes into consideration the molecular heterogeneity of the cancer type. Responsiveness to the clinically important nucleoside analogs gemcitabine and decitabine may be critically determined by Deoxycytidine kinase (DCK) expression as this enzyme is required to convert the inactive prodrugs into their pharmacologically active forms. Here, we examined whether DCK is differentially expressed in breast cancer and evaluated whether DCK expression levels control responsiveness to these nucleoside analogs in vitro by experimentally modulating DCK expression levels. We examined DCK expression in gene expression data sets of breast tumors including the series of 295 consecutive patients that have been classified into low or high risk for recurrence using the MammaPrint 70 gene profile. We found that DCK is expressed at higher levels in patients having poor clinical outcome as judged by the MammaPrint assay. As such, patients that have a poor prognosis may thus be susceptible to treatment with nucleoside analogs. In support of this, we found a causal relationship between DCK levels and sensitivity to these nucleoside analogs in breast cancer cell lines. The data indicate that breast cancers that are at high risk of recurrence express higher levels of DCK, which we find to be strongly correlated to a favorable response to nucleoside analogs. The data suggest that DCK expression in breast cancer could be exploited to select patients that are likely to respond to treatment with nucleoside analogs.

Introduction Adjuvant therapy for stage II patients is recommended for patients with high risk features, especially with T4 tumors, and recommended for all stage III patients. Adjuvant therapy is not indicated for patients with MSI-H status who are considered of being at low risk of disease relapse. However, this leaves the majority of stage II patients with an undetermined risk. ColoPrint is an 18-gene expression classifier that identifies patients at higher risk of disease relapse. Together with a genomic classifier to determine MSI-status, ColoPrint can help making treatment decisions for early stage colon cancer patients. Methods ColoPrint was developed using whole genome expression data and was validated in public datasets (n = 322) and independent patient cohorts from 5 European hospitals (n = 341). Tissue specimen, clinical parameters, MSI-status and follow-up data (median follow-up 70 months) for patients were available and the ColoPrint index was determined using validated diagnostic arrays. The 64-gene MSI-signature (MSI-Print) was developed using full genome expression data of 276 primary stage II and III colorectal tumors with known MSI-status (n = 29 MSI-H, n = 247 MSS). Uni-and multivariate analysis was performed on the pooled stage II and stage IIIA patient set and in the subsets of patients who were stage IIIA or stage II T3/ MSS. Results In the analysis of all stage II and IIIA patients, ColoPrint classified two-third of stage II patients as being at lower risk. The 3-year Relapse-Free-Survival (RFS) was 91% for Low Risk and 74% for patients at higher risk with a HR of 2.9 (p = 0.001). Clinicopathological parameters from the 2012 NCCN guidelines (T4, perforation, Conclusion The diagnostic MSI signature identifies patients with MSI-H status with high accuracy. Combined with ColoPrint, the prognostic accuracy in stage II and stage IIIA patients can be significantly improved, thereby facilitating the identification of patients at higher risk who might be considered for additional treatment.

Introduction Previous analysis of the gene expression data of mutational subgroups in the PETACC3 trial, revealed a very homogenous gene expression pattern in BRAF mutant CRC patients; while KRAS mutant CRC contain at least three different patient groups in terms of gene expression. One of these KRAS mutant groups together with 10% of KRAS/BRAF WT (WT2) patients, resembled the gene expression found in the BRAF mutants. This BRAF-like subgroup is characterized by poor prognosis. In this article we investigate, if heterogeneity in CRC patients can be partly explained by diverse signaling downstream of KRAS, to which the DUSPs take part. Material and methods The expression of DUSP1, DUSP4 and DUSP6, DUSPs previously shown to be induced by mutations in genes of the MAPK pathway, was analyzed in 4 different CRC patient microarray datasets (PETACC3, cetuximab, TCGA and Agendia dataset). We investigated, if patients with low and high DUSP expression behave differently in terms of response, OS, PFS and differ in clinical characteristics both in PETACC3 and TCGA dataset. Findings Unlike DUSP1, DUSP4 and DUSP6 were differentially overexpressed between BRAF mutants and KRAS mutants on one side and between BRAF mutants and WT2 patients on the other side. When we subdivided the patients into low and high DUSP expression within KRAS mutant and WT2 patients, we found a significant difference in OS in both mutational subgroups with high DUSP4 expression in KRAS mutants associated with a better OS and in WT2 associated with a worse OS. Multivariate analysis in the BRAF WT patients in both PETACC3 and cetuximab dataset showed that high DUSP4 expression was still associated with a worse OS (HR = 1.23; p Conclusions DUSP4 is predictive for OS in CRC patients with KRAS mutations. Interestingly, within KRAS mutant and WT2 patients high DUSP4 expression correlates with clinical characteristics that are typical for BRAF mutant CRC patients. If DUSP4 may represent a novel biomarker for a subset of CRC patients needs further research. Download : Download full-size image Fig. 1 . Box plots with DUSP4 expression according to different mutational groups in PETACC3 dataset. Download : Download full-size image Fig. 2 . Kaplan-Meier curve for overall survival according to DUSP4 expression (low and high) in KRAS WT patients.

466 Background: Microsatellite Instability (MSI-H) occurs in 10-20% of colon tumors and has been attributed predominantly to gene silencing of DNA mismatch repairs (MMR) genes by mutation or methylation, including MSH2, PMS2 and in particular MLH1. Methods: MSI status for 276 primary stage II and III colorectal tumors have been determined (n=29 MSI-H, n=247 MSS and MSI-L) by immunohistochemically staining of MLH1 and PMS2 (for 90 patients) or by PRC amplification of six microsatellite DNA regions from paired normal and tumor tissues (for 186 patients). Full genome gene expression data for the same 276 tumors was available and used to identify genes that correlate with MSI-H status. All samples had full clinical information including 5-year follow-up and ColoPrint results (as described in Salazar et al. JCO 2011). Results: Most MSI-H patients were classified as ColoPrint low risk (26 of 29 patients; 90%) however this is only as subgroup of the 195 patients identified by ColoPrint as low risk. While ColoPrint had great prognostic power in this dataset (HR 2.37, p=0.0006), it does not identify MSI-H patients with high specificity. We therefore developed a MSI-gene signature of 64 genes using a 10-fold cross validation procedure to complement ColoPrint. The MSI-signature identifies MSI-H patients with high sensitivity (93.1% (27/29)) and high specificity (87.9% (217/247)) and an overall accuracy of 88.4% (244/276). The MSI-signature has prognostic power in the subset of stage II patients (n=215) with HR = 2.38 (95% CI 1.15-4.93, p=0.06. However, in multivariate analysis of MSI status and all clinical factors, ColoPrint was the only significant prognostic factor (HR 3.2; p=0.0014). The MSI-signature is currently validated in an independent patients set of 132 stage II patients (31 MSI-H, 101 MSS). Conclusions: The diagnostic MSI signature identifies patients with MSI-H status with high accuracy. In combination with the prognostic power of ColoPrint, the identification of MSI-H patients might add additional information for treatment of stage II patients.

537 Background: Classification of breast cancer into molecular subtypes may be important for the proper selection of therapy, as tumors with seemingly similar biology can have strikingly different clinical outcomes. We have previously developed an 80-gene molecular subtyping profile (BluePrint) for the classification of breast cancer into three molecular subtypes: triple-negative (“Basal-type”); hormone receptor-positive (“Luminal-type”) and ERBB2-positive (“ERBB2-type”). In this study, we sought to determine if Luminal-type tumors are characterized by active ER signaling. Methods: ERα ChIP-seq data from proliferating MCF-7 breast cancer cells was used from a publically available dataset [Ross-Innes, 2010]. The transcription start sites of the 58 genes in the gene set that identifies the Luminal-type subgroup were determined after which the presence of Estrogen Receptor binding sites was analyzed within a window of −/+ 20 kb from the transcription start site. The sequence track was visualized using the UC...

403 Background: Approximately 8-15% of colorectal (CRC) patients carry an activating mutation in BRAF. This CRC subtype is associated with poor outcome and with resistance, both to chemotherapeutic treatments and to tailored drugs. We recently showed that BRAF (V600E) colon cancers (CCs) have a characteristic gene expression signature (1, 2) which is found also in subsets of KRAS mutant and KRAS-BRAF wild type (WT2) tumors. Tumors having this gene signature, referred as “BRAF-like”, have a similar poor prognosis irrespective of the presence of the BRAF (V600E) mutation. By using a shRNA-based genetic screen in BRAF mutant CC cell lines we aimed to identify genes and pathways necessary for survival and growth of BRAFmutant CC. Such studies may reveal additional targets for therapy and potentially provide new biomarkers for patient stratification Methods: We identified 363 genes that are selectively overexpressed in BRAF mutant tumors as compared to WT2 type tumors, based on gene expression profiles of the PETACC3 (1) and Agendia (2) datasets. The TRC human genome-wide shRNA collection (TRC-Hs1.0) was used to generate a 1815 hairpins sub-library targeting those identified genes (BRAF library). BRAF(V600E) CC cell lines were infected with the BRAF library and screened for shRNAs that cause lethality. LIM1215 CC cell line (WT2) was used as a control. Cells stably expressing the shRNA library were cultured for 13 days, after which shRNAs were recovered by PCR. Deep sequencing was applied to determine the specific depletion of shRNA in BRAF(V600E) cells as compared to LIM1215 cells Results: Candidate genes were identified by using following filtering criteria: depletion in BRAF(V600E) cells by at least 50% and depletion in BRAF(V600E) cells 1, 5-fold higher than in control cells with the corresponding p-value to be ≤ 0.1. A total of 34 genes met our criteria of which 6 genes were presented with more than one hairpin and were concordant across the cell lines selected for validation. Conclusions: We identified candidate synthetic lethal genes in BRAF mutant CC cell lines. Functional analysis is ongoing. Data will be presented. References 1. J Clin Oncol 2012 Apr 20;30(12):1288-9 2. Gut (2012). doi:10.1136/gutjnl-2012-302423