Proteogenomics analysis to identify acquired resistance-specific alterations in melanoma PDXs on MAPKi therapy [WES]

Therapeutic approaches to treat melanoma include small molecule drugs that target activating protein mutations in pro-growth signaling pathways like the MAPK pathway. While beneficial to the approximately 50% of patients with activating BRAFV600 mutation, mono- and combination therapy with MAPK inhibitors is ultimately associated with acquired resistance. To better characterize the mechanisms of MAPK inhibitor resistance in melanoma, we utilize patient-derived xenografts and apply proteogenomic approaches leveraging genomic, transcriptomic, and proteomic technologies that permit the identification of resistance-specific alterations and therapeutic vulnerabilities. A specific challenge for proteogenomic applications comes at the level of data curation to enable multi-omics data integration. Here, we present a proteogenomic approach that uses custom curated databases to identify unique resistance-specific alternations in melanoma PDX models of acquired MAPK inhibitor resistance. We demonstrate this approach with a NRASQ61L melanoma PDX model from which resistant tumors were developed following treatment with a MEK inhibitor. Our multi-omics strategy addresses current challenges in bioinformatics by leveraging development of custom curated proteogenomics databases derived from individual resistant melanoma that evolves following MEK inhibitor treatment and is scalable to comprehensively characterize acquired MAPK inhibitor resistance across patient-specific models and genomic subtypes of melanoma. Overall design: A patient-derived xenograft (PDX) of an NRASQ61L patient melanoma tumor was established as non-dissociated tumor fragments in NOD scid gamma (NSG) mice and serially passaged without dissociation, in vitro growth cycles, or adjunctive in vivo growth supplements such as Matrigel. To derive the acquired-resistant NRASQ61L tumors, we implanted one tumor fragment (passage #2) per NSG mouse. A cohort of tumor-bearing mice with similar tumor volumes were selected for experimentation. When the tumors reached ~500 mm3, a mouse was treated with the vehicle-chow (V), while four mice were treated with trametinib-embedded chow (R1,2,4,5) to achieve 5 mg/kg/day dosing. All trametinib-treated tumors displayed tumor regression at this chosen dosage. Tumor tissues for NRASQ61L vehicle (V) and four treated samples (R1,2,4,5) were harvested at end-points and prepared for whole exome sequencing (WES) and bulk RNA-sequencing (RNA-seq).