Discipline: Biological Sciences
Subcategory: Biochemistry (not Cell and Molecular Biology and Genetics)
Room: Exhibit Hall
Nord Gilaj - Lehman College
Co-Author(s): Gabriel Bedard MD, PhD Candidate, Albert Einstein College of Medicine, Bronx, New York; Vern L. Schramm PhD, Albert Einstein College of Medicine, Bronx, New York
Genetic deletion of 5’ methylthioadenosine phosphorylase (MTAP) is the most common homozygous deletion (~15%) in human cancers due to its close proximity to tumor suppressor gene locus CDKN2A/B. MTAP deletions are synthetically lethal with co-deletions in the gene encoding the metabolic enzymes methionine adenosyltransferase 2 alpha (MAT2a) and protein arginine methyltransferase 5 (PRMT5). MTAP functions in the methionine salvage pathways to recycle methylthioadenosine (MTA) to methionine, which is used by MAT2a to synthesize s-adenosylmethionine (SAM). SAM is a ubiquitous cofactor for polyamination and methylation reactions in the cell. Current inhibitors of MAT2a are in clinical trials for cancers with MTAP deletions, however this strategy only covers a small fraction of all cancers. We propose that the use of MTAP inhibitor methylthio-DADMe-Immucillin A (MTDIA), transition state analogue developed by Schramm laboratory in combination with MAT2a inhibitor AG-270 will phenocopy the therapeutic effects seen with AG-270 in MTAP-/- cancers. We hypothesize that our novel drug combination will perturb methionine metabolism and have specific downstream effects on the activity of oncotarget protein arginine methyltransferase 5, a known regulator of gene splicing and transcription factor localization. To test this hypothesis, two cell lines representing cancers with rare natural MTAP deletions were selected. MTDIA and AG-270 displayed synergistic cell growth inhibition in MTAP positive FaDu (head and neck squamous cell carcinoma) and HT-29 cells (colorectal carcinoma) by colorimetric WST-1 assay. Next, metabolic disturbances were determined through tandem LC-MS/MS analysis of extracts from cells treated over time. Finally, Western blotting of the PRMT5-specific product symmetric dimethylarginine (SDMA) was performed to demonstrate the inhibition of PRMT5. Here we report that MTDIA and AG-270 together, but not alone, have growth-inhibiting properties. MTDIA phenocopies the metabolic signature of MTAP deletion including increased MTA, and combination with AG-270 further increases the ratio of MTA:SAM by depleting the SAM pool. Finally, PRMT5 is inhibited only under high MTA:SAM ratio conditions, such as with combination therapy. Together, these data suggest that MTDIA will expand the anti-cancer efficacy of MAT2a inhibitors through the specific inhibition of PRMT5 and may extend therapeutic potential to the other 85% of MTAP+ cancers. Continued efforts in identification of gene expression alterations, proteomic quantification of global and specific methylation changes, and safety and efficacy in animal models are underway.
Funder Acknowledgement(s): I would like to thank Gabriel Bedard MD/PhD candidate and Dr. Vern L. Schramm at Albert Einstein College of Medicine for the opportunity to work on this project; their support has been tremendous throughout the process. I’d also like to thank the Office of Grant Support at Albert Einstein College of Medicine and the NIGMS for funding this project.
Faculty Advisor: Gabriel Bedard; Vern L. Schramm, firstname.lastname@example.org
Role: Western blotting, IC50 assays (WST-1) were conducted by me. My advisor helped me with the analysis of LC-MS/MS samples which I prepared.