May Elbanna

Genomic profiling is predictive of response to cisplatin treatment but not to PI3K inhibition in bladder cancer patient-derived xenografts

Purpose Effective systemic therapeutic options are limited for bladder cancer. In this preclinical study we tested whether bladder cancer gene alterations may be predictive of treatment response. Experimental design We performed genomic profiling of two bladder cancer patient derived tumor xenografts (PDX). We optimized the exome sequence analysis method to overcome the mouse genome interference. Results We identified a number of somatic mutations, mostly shared by the primary tumors and PDX. In particular, BLCAb001, which is less responsive to cisplatin than BLCAb002, carried non-sense mutations in several genes associated with cisplatin resistance, including MLH1, BRCA2, and CASP8. Furthermore, RNA-Seq analysis revealed the overexpression of cisplatin resistance associated genes such as SLC7A11, TLE4, and IL1A in BLCAb001. Two different PIK3CA mutations, E542K and E545K, were identified in BLCAb001 and BLCAb002, respectively. Thus, we tested whether the genomic profiling was predictive of response to a dual PI3K/mTOR targeting agent, LY3023414. Despite harboring similar PIK3CA mutations, BLCAb001 and BLCAb002 exhibited differential response, both in vitro and in vivo. Sustained target modulation was observed in the sensitive model BLCAb002 but not in BLCAb001, as well as decreased autophagy. Interestingly, computational modelling of mutant structures and affinity binding to PI3K revealed that E542K mutation was associated with weaker drug binding than E545K. Conclusions Our results suggest that the presence of activating PIK3CA mutations may not necessarily predict in vivo treatment response to PI3K targeted therapies, while specific gene alterations may be predictive for cisplatin response in bladder cancer models and, potentially, in patients as well.

EZH2 Modifies Sunitinib Resistance in Renal Cell Carcinoma by Kinome Reprogramming

Acquired and intrinsic resistance to receptor tyrosine kinase inhibitors (RTKi) represents a major hurdle in improving the management of clear cell renal cell carcinoma (ccRCC). Recent reports suggest that drug resistance is driven by tumor adaptation via epigenetic mechanisms that activate alternative survival pathways. The histone methyl transferase EZH2 is frequently altered in many cancers, including ccRCC. To evaluate its role in ccRCC resistance to RTKi, we established and characterized a spontaneously metastatic, patient-derived xenograft model that is intrinsically resistant to the RTKi sunitinib, but not to the VEGF therapeutic antibody bevacizumab. Sunitinib maintained its antiangiogenic and antimetastatic activity but lost its direct antitumor effects due to kinome reprogramming, which resulted in suppression of proapoptotic and cell-cycle-regulatory target genes. Modulating EZH2 expression or activity suppressed phosphorylation of certain RTKs, restoring the antitumor effects of sunitinib in models of acquired or intrinsically resistant ccRCC. Overall, our results highlight EZH2 as a rational target for therapeutic intervention in sunitinib-resistant ccRCC as well as a predictive marker for RTKi response in this disease. Cancer Res; 77(23); 6651-66. ©2017 AACR.

Abstract 955: Inhibition of SEC24D decreases exosome release of the tumor suppressor miR-605 in renal cell carcinoma

Background: MicroRNA 605 (miR-605) has been recently reported as a putative tumor suppressor and its overexpression decreases tumor cell proliferation, migration and clonogenicity. To date, the role of miR-605 in renal cell carcinoma (RCC) has not been investigated. We recently showed the decrease of circulating miR-605 in serum of clear cell renal cell carcinoma (ccRCC) patients who responded to the treatment with the histone deacetylase (HDAC) inhibitor vorinostat, and the VEGF blocker bevacizumab. The current study was designed to investigate the expression of miR-605 and understand the mechanism(s) responsible the extracellular release of miR-605 using RCC cells. Methods: 786-0 cells were treated with and without vorinostat for 24h and condition media were collected, briefly centrifuged to settle the cells and debris, and processed to isolate exosomes using the ExoQuick exosome isolation kit (Systems Biology, CA). Purified exosomes and 786-0 cells were used to isolate RNA and further prepared cDNA to utilize for quantitative RT-PCR analysis. Expression of miR-605 in exosomes and cells was determined by Quantitative RT-PCR using TaqMan MicroRNA Assays with miR-605 primers obtained from Applied Biosystems, NY. To determine the role of secretory protein 24 family member D (SEC24D), a catalytic component of coat protein complex (COPII) involved in the secretory pathway, 786-0 cells treated with vorinostat were used to determine SEC24D expression by QRT-PCR and Western blot analysis. TCGA data was used to determine correlation of SEC24D expression clear cell renal cell carcinoma patients’ survival. Results: Vorinostat treatment resulted in a significant decrease of miR-605 expression in exosomes and increase (100 fold) of intracellular expression. Furthermore, the increased intracellular miR-605 was associated with the inhibition of SEC24D mRNA and protein expression in 786-0 cells treated with vorinostat. Cancer Genomic data analysis of c-Bioportal from MSKCC of TCGA revealed the overall poor survival of ccRCC patients with alteration of SEC24D. Conclusion: Taken together, our preliminary data suggest that the HDAC inhibitor vorinostat inhibits SEC24D and exosome mediated extracellular secretion of miR-605 in RCC cells. These results suggest that vorinostat treatment retained intracellular miR-605 that target genes involved in cell survival and proliferation in RCC. Further studies will evaluate circulating miR-605 as a predictive biomarker to determine the efficacy of vorinostat in ongoing trials with RCC patients. Citation Format: Sreenivasulu Chintala, Remi Adelaiye-Ogala, Ashley Orillion, Sreevani Arisa, May Elbanna, Roberto Pili. Inhibition of SEC24D decreases exosome release of the tumor suppressor miR-605 in renal cell carcinoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 955.

Abstract 1462: Characterization of patient-derived bladder cancer xenografts: role of xCT in response to cisplatin

Background: A major challenge in cancer drug development has been largely attributed to the inability of cell lines to recapitulate the heterogeneity of human tumors. Patient derived xenografts (PDX) represent a major advance as they are more representative of the clinical setting. However, appropriate characterization of PDXs is necessary to guide biomarker driven drug discovery research. Methods: HE 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1462. doi:10.1158/1538-7445.AM2015-1462