PT-005 - INTEGRATED GENOMICS AND TRANSCRIPTOMICS ANALYSES USING HAPMAP CELL LINES REVEAL NOVEL MODULATORS OF CELLULAR RESPONSE TO CYCLOPHOSPHAMIDE.

M. Gbadamosi¹, N. Bhise², T. Mitra Ghosh², E. Molchan¹, O. Olabige³, C. Yang³, D. Mitchell³, B. Fridley⁴, J. Lamba²; ¹Department of Pharmacotherapy and Translational Research, College of Pharmacy, University of Florida, Gainesville, FL, United States, ²Department of Pharmacotherapy and Translational Research, College of Pharmacy, University of Florida, ³Department of Neurosurgery, College of Medicine, University of Florida, ⁴Department of Biostatistics and Bioinformatics, Moffitt Cancer Center.

Background: Cyclophosphamide (CTX) is an alkylating agent used to treat breast cancer (BC) and other cancers with diverse clinical responses. Studies on this diversity have focused on genomic variations affecting CTX metabolism while those that impact CTX cellular sensitivity remain more obscure. The main cytotoxic metabolite produced by the metabolism of CTX is phosphoramide mustard (PM). In this study, we assessed genomic variation associated with variability in PM chemosensitivity.

Methods: Fifty-three HapMap cell lines were treated with varying concentrations of PM and IC50 values were calculated using MTT assays. IC50 values and publicly available genotyping data (N = 2e+6 single-nucleotide polymorphism (SNPs)) were used to conduct a genome-wide association study to identify SNPs associated with cellular CTX sensitivity. Then, using transcriptomic data and external databases and 5-year overall survival (OS) and transcriptomic data from CTX-treated BC patients (CTX-BC; N = 511) we investigated potential functional and clinical consequences, respectively.

Results: 143 SNPs were associated with PM IC50 at a threshold of P < 10e-4 including novel SNPs proximal to tumor suppressors and oncogenes. Top signals arose from the KIAA0586 locus with 6 SNPs highly associated with PM resistance therein. Notably, rs4143897 (P = 6.2e-7) and rs10483698 (P = 2.3e-7) impact KIAA0586 splicing and are known to reduce KIAA0586 expression. In CTX-BC patients, KIAA0586 expression was protective (HR = 0.45, P = 0.03), suggesting these SNPs cause CTX resistance by reducing KIAA0586 expression or activity. Using transcriptomic data, we identified a hotspot on chromosome 6 containing 5 SNPs associated with PM resistance and altered expression of 15 genes, 3 of which were significantly associated with lower OS in CTX-BC patients. One top signal, rs226928 (P = 3e-5), was associated with higher expression of MYLK4, a tumor progression-associated kinase. MYLK4 expression was positively correlated with PM IC50 (P = 0.04) and associated with lower OS in CTX-BC patients (HR = 3.00, P = 0.03).

Conclusion: Together, we report novel SNPs associated with cellular CTX sensitivity with robust evidence for functional and clinical consequences. Pending validation, these results may guide CTX treatment and provide insight into new drug targets for combinatorial treatment strategies.