Abstract 5561: Single-cell analysis reveals an adaptive, transiently heritable, slowly-dividing, drug-resistant state inhibitable by drug combinations
Authors: Fallahi-Sichani Mohammad , Becker Verena , Izar Benjamin , Baker Gregory J. , Lin Jia-Ren , Boswell Sarah A. , Garraway Levi A. , Sorger Peter K. 
Published: July 2017

Adaptation and fractional response of tumor cells to targeted inhibitors of oncogenic pathways creates a population of viable tumor cells from which fully resistant clones can ultimately arise. Thus, understanding transient drug adaptation is key for both improving the effectiveness of treatment and delaying/controlling acquired resistance. Despite the wealth of information available about feedback mechanisms associated with adaptive resistance, most of our knowledge in this area comes from studying drug response in bulk tumor cell populations. Furthermore, the phenotypic consequences of drug adaptation have been often studied at a few fixed time-points, when drug-adapted cells exhibit a high population-average activity in multiple pro-growth signaling cascades. It therefore remains unclear how the initial responses to drug relate to subsequent phenotypes such as cell death or adaptation. This is likely a key point for designing novel approaches to overcome fractional drug response in tumor cells and to achieve durable therapy.

We use real-time live-cell imaging, single-cell analysis and molecular profiling to show that exposure of BRAFV600E melanoma cells to RAF/MEK inhibitors elicits a time-variable and heterogeneous response in which some cells die, some arrest and the remainder adapt to drug. Drug-adapted cells up-regulate markers of the neural crest (e.g. NGFR), a melanocyte precursor, and grow slowly. The drug-induced slowly-cycling NFGRHigh state is only transiently stable, reverting to the drug-naïve state within two weeks of drug withdrawal as measured by the restoration of RAF/MEK inhibitor sensitivity, accelerated rate of cell division and reduced expression of NGFR. Transcriptional and biochemical profiling of cell lines and human tumors implicates a role for the c-Jun/ECM/FAK/Src cascade in driving the de-differentiated resistance program. We identify multiple drugs targeting this cascade as well as BET bromodomain inhibitors that block this resistance program in cell lines and in a BRAFV600E melanoma xenograft model and increase sensitivity and maximal effect (Emax) of RAF/MEK inhibitors. Our study reveals directly how drug adaptation happens in individual tumor cells leading to emergence of heterogeneous cell sub-populations with reduced drug-sensitivity that may be targeted by drug combinations.

Citation Format: Mohammad Fallahi-Sichani, Verena Becker, Benjamin Izar, Gregory J. Baker, Jia-Ren Lin, Sarah A. Boswell, Levi A. Garraway, Peter K. Sorger. Single-cell analysis reveals an adaptive, transiently heritable, slowly-dividing, drug-resistant state inhibitable by drug combinations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5561. doi:10.1158/1538-7445.AM2017-5561