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147 An integrated approach for identifying E-cadherin synthetic lethality networks
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Wednesday 19 November 2014
Material and Methods: To evaluate the extent of ITH in colorectal cancer (CRC) with microsatellite instability (MSI), we obtained 4−7 biopsies from 39 CRCs followed by MSI analysis either using the Bethesda MSI evaluation system or Promega system with 5 mononucleotide markers. Results: We found decreased prevalence of MSI (+) by the Promega system compared to the Bethesda system. The overall discordance between the two systems was 54%. In contrast to the previous studies that had shown discordance only in low MSI (MSI-L), our results showed the discordance not only in MSI-L, but also in high MSI (MSI-H) cases. Among the MSI (+) CRCs, ITH of MSI status was identified in 41.7% of CRC by the Bethesda system and 22.2% by the Promega system. In terms of MSI markers, the ITH originated from dinucleotide markers in most cases (69%), but it originated from mononucleotide markers (31%) as well. Pooling of DNA from a regional biopsy with MSI (+) with additional biopsies from stable MSI (MSS) showed that this approach was beneficial to increase the sensitivity of MSI detection. Conclusions: Our results indicate that ITH of MSI phenotype by the Bethesda system is more overestimated than previously identified. However, because there was considerable ITH of MSI subtypes and markers even by the Promega system, our data suggest that analysis of MSI status in multiple regional biopsies is needed for a better evaluation of MSI status in CRC. 144 POSTER Association between interleukin 17/interleukin 17 receptor gene polymorphism and papillary thyroid cancer Y. Eun1 , H. Chung1 , Y.C. Lee1 . 1 Kyung Hee University, Otolaryngology, Seoul, South Korea Background: Interleukin 17 (IL17) is an important cytokine for inflammatory and autoimmune disease. Recently, although numerous studies have been suggested the role of IL17 in tumor development, the mechanism remains to be unknown. The aim of this study is to determine whether single nucleotide polymorphisms (SNPs) in IL17 and IL17R contribute the development of papillary thyroid cancer (PTC) and assess the relationship between IL17 and IL17R SNPs and the clinicopathologic characteristics of PTC. Material and Methods: Eight SNPs located within the three genes of IL17A, IL17RA and IL17RB were genotyped using direct sequencing in 94 patients with PTC and 213 patients without PTC (controls). Genetic data were analyzed using commercially available software. And, the statistical analyses were performed according to clinicopathologic characteristics of PTC. Results: Genotyping analysis demonstrated that the SNP rs4819554 of IL17RA (codominant model 1, OR = 0.39, P = 0.001; and dominant model, OR = 0.45, 95% CI, P = 0.002) and the SNP rs1025689 of IL17RB (dominant model, OR = 0.59, P = 0.043) were significantly associated with the development of PTC. The SNP rs2275913 of IL17A (codominant model 2, OR = 0.19, P = 0.034; dominant model, OR = 0.34, P = 0.033) was significantly associated with multifocality. Furthermore, IL17RA SNP rs4819554 (dominant model, OR = 0.25, P = 0.010) was significantly associated with bilaterality of cancers. Conclusion: In our case–control study of SNPs in the IL17 and IL17R gene in patients with PTC, we demonstrated that IL17RA polymorphism have the possibilities of developing PTC and can influence the bilaterality of PTC. 145 POSTER ARQ 087, a novel pan FGFR-inhibitor crosses the BBB (blood–brain barrier) and distributes to the brain of rats R. Savage1 , T. Hall1 , B. Schwartz1 . 1 ArQule Inc., Woburn, USA Background: ARQ 087 is a potent multi-kinase inhibitor with pan-FGFR activity against FGFR1, FGFR2, FGFR3 and FGFR4 kinases. Preclinical data from human cell lines and xenograft models support the exploration of its anti-tumor activity across a broad range of human solid and hematological malignancies. Material and Methods: QWBA (Quantitative Whole Body Autoradiography) and metabolite profiling studies were conducted with 14 C-ARQ 087 in rats. Free concentrations of 14 C-ARQ 087 related radioactivity were determined in rat brain via microdialysis. The brain regions sampled by microdialysis were the right striatum and left ventricle. The concentrations sampled in the brain were compared to those obtained in peripheral blood sampled from the right jugular vein. Results: From QWBA studies, 14 C-ARQ 087 derived radioactivity was found to distribute to the cerebellum, cerebrum, choroid plexus, corpus callosum, lateral ventricle, meninges and spinal cord (as well as broadly distributing to the rest of the body) after 5-Day repeat dosing in SpragueDawley rats. At 24 hours post dose on Day 5, 14 C-ARQ087 derived
Poster Session – Preclinical Models radioactivity ranged from 0.540 mg equiv./g (cerebellum) to 4.12 mg equiv./g (choroid plexus) versus 0.756 mg equiv./g in plasma. In brain tissues AUC(0−24 h) ranged from 12.4 (cerebellum) to 408 (choroid plexus) mg equiv.·hr/g versus 34.2 mg equiv.·hr/g in plasma. Metabolite profiling data in rat brain showed that ARQ 087 is present primarily as parent drug. Preliminary microdialysis data in rat brain showed that free unbound ARQ 087 (based on AUC) is present in the striatum (17.45%) and in the left ventricle (16.30%) compared to that in systemic circulation. Conclusions: ARQ 087 distributes to the brain of rats as well as broadly to the rest of the body. Metabolite profiling and preliminary microdialysis studies confirmed that in brain primarily parent ARQ 087 is present and available in part as free ARQ 087. 146 POSTER Establishment of patient-derived xenografts (PDX) models for triple negative breast cancer (TNBC) as a pre-clinical platform for drug development J. Thatte1 , M. Meza1 , J. Ricono1 , T. Broudy1 , C. Mirsaidi1 , P. Nair1 . 1 Molecular Response, San Diego California, USA TNBC comprises 15−20% of breast cancers in the United States. TNBC lacks expression of estrogen receptor, progesterone receptor and receptor tyrosine protein kinase ERBB2 or Her-2/neu oncogene amplification. As a result, this type of breast cancer is difficult to treat as most of the chemotherapies target these 3 receptors. TNBC is an aggressive form of cancer associated with high morbidity, mortality and shorter diseasefree survival. Prognosis and management of TNBC is complicated due to its heterogeneous clinical presentation, histology and response to therapy. PDX models in immune compromised mice have become a valuable tool for preclinical drug development in recent years. Here, we report development and characterization of the TNBC PDX model. We used patient derived TNBC tissues to generate 5 new TNBC models in NOD-SCID mice. Patient tumors were pre-screened for their ER, PR and Her-2/Neu expression by IHC, prior to inoculation in mice. We report efficacy of two standard of care drugs, cisplatin and vinorelbine, in the TNBC PDX model which demonstrates the potential utility of the TNBC model in drug discovery effort in oncology for treatment of TNBC. 147 POSTER An integrated approach for identifying E-cadherin synthetic lethality networks I. Bajrami1 , S.J. Pettitt1 , R. Brough1 , H. Pemberton1 , D. Kastrev1 , Y. Fontebasso1 , J. Frankum1 , J. Campbell1 , A. Ashworth1 , C.J. Lord1 . 1 Institute of Cancer Research, Division of Breast Cancer, London, United Kingdom The E-cadherin (CDH1) tumour suppressor gene encodes a calciumdependent cell–cell adhesion glycoprotein, which has roles in maintaining cell polarity, differentiation, cell migration and survival. E-cadherin dysfunction is a feature common to many epithelial tumours, with the highest incidence occurring in diffuse gastric cancer (50%) and lobular breast cancer (56%) and can occur via CDH1 mutation, deletion or epigenetic silencing. Although E-cadherin dysfunction is relatively common, approaches to target this pathogenic alteration do not as yet exist. We have taken an integrated functional genomics approach to identifying E-cadherin synthetic lethality effects that exploits siRNA. Using a combination of Achilles’ Heel siRNA and small molecule inhibitor screens in histologically and genetically diverse tumour cell line panels, we have identified a compendium of genes whose disruption selectively targets E-cadherin deficient cells. Alongside the Achilles’ Heel screens in tumour cell models, complementary screens in isogenic systems with shRNA and CRISPR-engineered E-Cadherin defects will also be presented. As an orthogonal approach, we have also exploited piggyBac transposonbased mutagenesis in haploid ES cells to generate a diverse mutant library consisting of 100,000 mutants which are now being used in E-cadherin synthetic lethal screens. Together, these systems provide a framework for identifying candidate synthetic lethal effects suitable for mechanistic dissection and subsequent validation. The results from these complementary approaches and subsequent validation of selected effects will be presented.
Wednesday 19 November 2014
Material and Methods: To evaluate the extent of ITH in colorectal cancer (CRC) with microsatellite instability (MSI), we obtained 4−7 biopsies from 39 CRCs followed by MSI analysis either using the Bethesda MSI evaluation system or Promega system with 5 mononucleotide markers. Results: We found decreased prevalence of MSI (+) by the Promega system compared to the Bethesda system. The overall discordance between the two systems was 54%. In contrast to the previous studies that had shown discordance only in low MSI (MSI-L), our results showed the discordance not only in MSI-L, but also in high MSI (MSI-H) cases. Among the MSI (+) CRCs, ITH of MSI status was identified in 41.7% of CRC by the Bethesda system and 22.2% by the Promega system. In terms of MSI markers, the ITH originated from dinucleotide markers in most cases (69%), but it originated from mononucleotide markers (31%) as well. Pooling of DNA from a regional biopsy with MSI (+) with additional biopsies from stable MSI (MSS) showed that this approach was beneficial to increase the sensitivity of MSI detection. Conclusions: Our results indicate that ITH of MSI phenotype by the Bethesda system is more overestimated than previously identified. However, because there was considerable ITH of MSI subtypes and markers even by the Promega system, our data suggest that analysis of MSI status in multiple regional biopsies is needed for a better evaluation of MSI status in CRC. 144 POSTER Association between interleukin 17/interleukin 17 receptor gene polymorphism and papillary thyroid cancer Y. Eun1 , H. Chung1 , Y.C. Lee1 . 1 Kyung Hee University, Otolaryngology, Seoul, South Korea Background: Interleukin 17 (IL17) is an important cytokine for inflammatory and autoimmune disease. Recently, although numerous studies have been suggested the role of IL17 in tumor development, the mechanism remains to be unknown. The aim of this study is to determine whether single nucleotide polymorphisms (SNPs) in IL17 and IL17R contribute the development of papillary thyroid cancer (PTC) and assess the relationship between IL17 and IL17R SNPs and the clinicopathologic characteristics of PTC. Material and Methods: Eight SNPs located within the three genes of IL17A, IL17RA and IL17RB were genotyped using direct sequencing in 94 patients with PTC and 213 patients without PTC (controls). Genetic data were analyzed using commercially available software. And, the statistical analyses were performed according to clinicopathologic characteristics of PTC. Results: Genotyping analysis demonstrated that the SNP rs4819554 of IL17RA (codominant model 1, OR = 0.39, P = 0.001; and dominant model, OR = 0.45, 95% CI, P = 0.002) and the SNP rs1025689 of IL17RB (dominant model, OR = 0.59, P = 0.043) were significantly associated with the development of PTC. The SNP rs2275913 of IL17A (codominant model 2, OR = 0.19, P = 0.034; dominant model, OR = 0.34, P = 0.033) was significantly associated with multifocality. Furthermore, IL17RA SNP rs4819554 (dominant model, OR = 0.25, P = 0.010) was significantly associated with bilaterality of cancers. Conclusion: In our case–control study of SNPs in the IL17 and IL17R gene in patients with PTC, we demonstrated that IL17RA polymorphism have the possibilities of developing PTC and can influence the bilaterality of PTC. 145 POSTER ARQ 087, a novel pan FGFR-inhibitor crosses the BBB (blood–brain barrier) and distributes to the brain of rats R. Savage1 , T. Hall1 , B. Schwartz1 . 1 ArQule Inc., Woburn, USA Background: ARQ 087 is a potent multi-kinase inhibitor with pan-FGFR activity against FGFR1, FGFR2, FGFR3 and FGFR4 kinases. Preclinical data from human cell lines and xenograft models support the exploration of its anti-tumor activity across a broad range of human solid and hematological malignancies. Material and Methods: QWBA (Quantitative Whole Body Autoradiography) and metabolite profiling studies were conducted with 14 C-ARQ 087 in rats. Free concentrations of 14 C-ARQ 087 related radioactivity were determined in rat brain via microdialysis. The brain regions sampled by microdialysis were the right striatum and left ventricle. The concentrations sampled in the brain were compared to those obtained in peripheral blood sampled from the right jugular vein. Results: From QWBA studies, 14 C-ARQ 087 derived radioactivity was found to distribute to the cerebellum, cerebrum, choroid plexus, corpus callosum, lateral ventricle, meninges and spinal cord (as well as broadly distributing to the rest of the body) after 5-Day repeat dosing in SpragueDawley rats. At 24 hours post dose on Day 5, 14 C-ARQ087 derived
Poster Session – Preclinical Models radioactivity ranged from 0.540 mg equiv./g (cerebellum) to 4.12 mg equiv./g (choroid plexus) versus 0.756 mg equiv./g in plasma. In brain tissues AUC(0−24 h) ranged from 12.4 (cerebellum) to 408 (choroid plexus) mg equiv.·hr/g versus 34.2 mg equiv.·hr/g in plasma. Metabolite profiling data in rat brain showed that ARQ 087 is present primarily as parent drug. Preliminary microdialysis data in rat brain showed that free unbound ARQ 087 (based on AUC) is present in the striatum (17.45%) and in the left ventricle (16.30%) compared to that in systemic circulation. Conclusions: ARQ 087 distributes to the brain of rats as well as broadly to the rest of the body. Metabolite profiling and preliminary microdialysis studies confirmed that in brain primarily parent ARQ 087 is present and available in part as free ARQ 087. 146 POSTER Establishment of patient-derived xenografts (PDX) models for triple negative breast cancer (TNBC) as a pre-clinical platform for drug development J. Thatte1 , M. Meza1 , J. Ricono1 , T. Broudy1 , C. Mirsaidi1 , P. Nair1 . 1 Molecular Response, San Diego California, USA TNBC comprises 15−20% of breast cancers in the United States. TNBC lacks expression of estrogen receptor, progesterone receptor and receptor tyrosine protein kinase ERBB2 or Her-2/neu oncogene amplification. As a result, this type of breast cancer is difficult to treat as most of the chemotherapies target these 3 receptors. TNBC is an aggressive form of cancer associated with high morbidity, mortality and shorter diseasefree survival. Prognosis and management of TNBC is complicated due to its heterogeneous clinical presentation, histology and response to therapy. PDX models in immune compromised mice have become a valuable tool for preclinical drug development in recent years. Here, we report development and characterization of the TNBC PDX model. We used patient derived TNBC tissues to generate 5 new TNBC models in NOD-SCID mice. Patient tumors were pre-screened for their ER, PR and Her-2/Neu expression by IHC, prior to inoculation in mice. We report efficacy of two standard of care drugs, cisplatin and vinorelbine, in the TNBC PDX model which demonstrates the potential utility of the TNBC model in drug discovery effort in oncology for treatment of TNBC. 147 POSTER An integrated approach for identifying E-cadherin synthetic lethality networks I. Bajrami1 , S.J. Pettitt1 , R. Brough1 , H. Pemberton1 , D. Kastrev1 , Y. Fontebasso1 , J. Frankum1 , J. Campbell1 , A. Ashworth1 , C.J. Lord1 . 1 Institute of Cancer Research, Division of Breast Cancer, London, United Kingdom The E-cadherin (CDH1) tumour suppressor gene encodes a calciumdependent cell–cell adhesion glycoprotein, which has roles in maintaining cell polarity, differentiation, cell migration and survival. E-cadherin dysfunction is a feature common to many epithelial tumours, with the highest incidence occurring in diffuse gastric cancer (50%) and lobular breast cancer (56%) and can occur via CDH1 mutation, deletion or epigenetic silencing. Although E-cadherin dysfunction is relatively common, approaches to target this pathogenic alteration do not as yet exist. We have taken an integrated functional genomics approach to identifying E-cadherin synthetic lethality effects that exploits siRNA. Using a combination of Achilles’ Heel siRNA and small molecule inhibitor screens in histologically and genetically diverse tumour cell line panels, we have identified a compendium of genes whose disruption selectively targets E-cadherin deficient cells. Alongside the Achilles’ Heel screens in tumour cell models, complementary screens in isogenic systems with shRNA and CRISPR-engineered E-Cadherin defects will also be presented. As an orthogonal approach, we have also exploited piggyBac transposonbased mutagenesis in haploid ES cells to generate a diverse mutant library consisting of 100,000 mutants which are now being used in E-cadherin synthetic lethal screens. Together, these systems provide a framework for identifying candidate synthetic lethal effects suitable for mechanistic dissection and subsequent validation. The results from these complementary approaches and subsequent validation of selected effects will be presented.