Abstract
BackgroundTumors can evolve and adapt to therapeutic pressure by acquiring genetic and epigenetic alterations that may be transient or stable. A precise understanding of how such events contribute to intratumoral heterogeneity, dynamic subpopulations, and overall tumor fitness will require experimental approaches to prospectively label, track, and characterize resistant or otherwise adaptive populations at the single-cell level. In glioblastoma, poor efficacy of receptor tyrosine kinase (RTK) therapies has been alternatively ascribed to genetic heterogeneity or to epigenetic transitions that circumvent signaling blockade.ResultsWe combine cell lineage barcoding and single-cell transcriptomics to trace the emergence of drug resistance in stem-like glioblastoma cells treated with RTK inhibitors. Whereas a broad variety of barcoded lineages adopt a Notch-dependent persister phenotype that sustains them through early drug exposure, rare subclones acquire genetic changes that enable their rapid outgrowth over time. Single-cell analyses reveal that these genetic subclones gain copy number amplifications of the insulin receptor substrate-1 and substrate-2 (IRS1 or IRS2) loci, which activate insulin and AKT signaling programs. Persister-like cells and genomic amplifications of IRS2 and other loci are evident in primary glioblastomas and may underlie the inefficacy of targeted therapies in this disease.ConclusionsA method for combined lineage tracing and scRNA-seq reveals the interplay between complementary genetic and epigenetic mechanisms of resistance in a heterogeneous glioblastoma tumor model.
Highlights
Tumors can evolve and adapt to therapeutic pressure by acquiring genetic and epigenetic alterations that may be transient or stable
Outgrowth of fit genetic subclones is a well-established mechanism of drug resistance [1] and recent work has highlighted that genetic subclones may rapidly adapt through dynamic alterations involving extrachromosomal DNA [2, 3]
We found that despite enduring dasatinib-induced inhibition of PDGFRA phosphorylation (Supplementary Fig. S4d), e86var clonal isolates cultured in the absence of dasatinib for > 4 weeks retained their drug-resistant phenotype when re-exposed to dasatinib (Fig. 3c)
Summary
Tumors can evolve and adapt to therapeutic pressure by acquiring genetic and epigenetic alterations that may be transient or stable. A precise understanding of how such events contribute to intratumoral heterogeneity, dynamic subpopulations, and overall tumor fitness will require experimental approaches to prospectively label, track, and characterize resistant or otherwise adaptive populations at the single-cell level. Transient epigenetic changes or cell state transitions that allow tumor cells to persist through drug exposure have been described in several cancer models [4,5,6,7]. Outgrowth of preexisting drug-resistant subpopulations has been tracked in experimental tumor models using DNA barcodes [11]. Holistic assessment of preexisting and dynamic epigenetic and genetic drug resistance mechanisms remains an important goal that requires new methods
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