RT Journal
A1 Ho, Yu-Jui
A1 Anaparthy, Naishitha
A1 Molik, David
A1 Mathew, Grinu
A1 Aicher, Toby
A1 Patel, Ami
A1 Hicks, James
A1 Hammell, Molly Gale
T1 Single-cell RNA-seq analysis identifies markers of resistance to targeted BRAF inhibitors in melanoma cell populations
JF Genome Research 
JO Genome Research 
YR 2018 
FD September 01 
VO 28 
IS 9 
SP 1353 
OP 1363 
DO 10.1101/gr.234062.117 
UL http://genome.cshlp.org/content/28/9/1353.abstract 
AB Single-cell RNA-seq's (scRNA-seq) unprecedented cellular resolution at a genome-wide scale enables us to address questions about cellular heterogeneity that are inaccessible using methods that average over bulk tissue extracts. However, scRNA-seq data sets also present additional challenges such as high transcript dropout rates, stochastic transcription events, and complex population substructures. Here, we present a single-cell RNA-seq analysis and klustering evaluation (SAKE), a robust method for scRNA-seq analysis that provides quantitative statistical metrics at each step of the analysis pipeline. Comparing SAKE to multiple single-cell analysis methods shows that most methods perform similarly across a wide range of cellular contexts, with SAKE outperforming these methods in the case of large complex populations. We next applied the SAKE algorithms to identify drug-resistant cellular populations as human melanoma cells respond to targeted BRAF inhibitors (BRAFi). Single-cell RNA-seq data from both the Fluidigm C1 and 10x Genomics platforms were analyzed with SAKE to dissect this problem at multiple scales. Data from both platforms indicate that BRAF inhibitor-resistant cells can emerge from rare populations already present before drug application, with SAKE identifying both novel and known markers of resistance. These experimentally validated markers of BRAFi resistance share overlap with previous analyses in different melanoma cell lines, demonstrating the generality of these findings and highlighting the utility of single-cell analysis to elucidate mechanisms of BRAFi resistance.