Co-delineating genomic and transcriptomic modes of resistance to MEK inhibition in individual triple negative breast cancer cells.

D.M. Arvapalli1, I. Salas-Gonzalez1, T.V. Morozova1, Dan R. Goulet2, G.L. Johnson2, V.J. Weigman1, J.A.A. West1, G.L. Harton1, J.S. Zawistowski1

1BioSkryb Genomics, Inc., Durham, NC, USA

2Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, NC, USA

Triple negative breast cancer (TNBC) tumors are frequently driven by MAPK signaling and are initially susceptible to MEK inhibition, yet resistance invariably develops. This resistance occurs, in part, by transcriptional adaptation where signaling is bypassed to reactivate MAPK signaling through a different node, yet resistance can also develop or co-develop through genomic modification. We therefore exploited single-cell ResolveOME™ chemistry to simultaneously capture whole genome single nucleotide variation (SNV), copy number variation (CNV) and full-transcript RNAseq from the same individual cell to define multifaceted contributions to drug resistance and to probe single-cell heterogeneity of these contributions for lineage identification. We employed two TNBC models of resistance to the MEK inhibitor trametinib whereby transcriptional and epigenetic contributions to the resistance have been extensively characterized, yet in which comprehensive assessment of genomic contributions to resistance has not yet been performed. In an epithelial subpopulation of SUM-229PE cells, previous work demonstrated resistance to trametinib coincided with upregulated KRAS expression relative to treatment-naive parental cells and manifested as one of the most statistically significant differentially-expressed genes. The genomic arm of ResolveOME and BaseJumper™ analysis software unveiled a genomic lesion, a ~650 kilobase block of differential allelic identity between parental and trametinib-resistant cells encompassing the KRAS gene—indicating structural variation at the locus and a candidate genomic mechanism underlying KRAS transcript upregulation and consequential MEK inhibitor resistance. The transcriptomic arm of the ResolveOME workflow concurrently validated the upregulation of KRAS and previously known gene sets while defining single-cell heterogeneity within. We also subjected parental and trametinib-resistant SUM-159PT cells, a mesenchymal model of the claudin-low molecular subtype of TNBC, to ResolveOME profiling. We observed expression modulation of CEBP family of enhancer factors and SMARC family chromatin remodeling factors, and current efforts are focused on linking resistant cell-specific SNVs proximal to these and other upregulated factors in both models as candidate regulatory variants in the paradigm of epithelial mesenchymal transition (EMT). The unification of genomic and transcriptomic data here uncovered DNA variation underlying the observed transcriptional modulation and provides a framework for ultimately seeking targetable variation in longitudinal primary patient samples.