The acute cellular response to stress generates a subpopulation of reversibly stress-tolerant cells under conditions that are lethal to the majority of the population. Stress tolerance is attributed to heterogeneity of gene expression within the population to ensure survival of a minority. We performed whole transcriptome sequencing analyses of metastatic human breast cancer cells subjected to the chemotherapeutic agent paclitaxel at the single-cell and population levels. Here we show that specific transcriptional programs are enacted within untreated, stressed, and drugtolerant cell groups while generating high heterogeneity between single cells within and between groups. We further demonstrate that drug-tolerant cells contain specific RNA variants residing in genes involved in microtubule organization and stabilization, as well as cell adhesion and cell surface signaling. In addition, the gene expression profile of drug-tolerant cells is similar to that of untreated cells within a few doublings. Thus, single-cell analyses reveal the dynamics of the stress response in terms of cell-specific RNA variants driving heterogeneity, the survival of a minority population through generation of specific RNA variants, and the efficient reconversion of stress-tolerant cells back to normalcy.

EGFR-activating mutations are observed in approximately 15% to 20% of patients with non-small cell lung cancer. Tyrosine kinase inhibitors have provided an illustrative example of the successes in targeting oncogene addiction in cancer and the role of tumor-specific adaptations conferring therapeutic resistance. The compound osimertinib is a thirdgeneration tyrosine kinase inhibitor, which was granted full FDA approval in March 2017 based on targeting EGFR T790M resistance. The compound has received additional FDA approval as first-line therapy with improvement in progression-free survival by suppressing the activating mutation and preventing the rise of the dominant resistance clone. Drug development has been breathtaking in this space with other third-generation compounds at various stages of development: rociletinib (CO-1686), olmutinib (HM61713), nazartinib (EGF816), naquotinib (ASP8273), mavelertinib (PF-0647775), and AC0010. However, therapeutic resistance after the administration of third-generation inhibitors is complex and not fully understood, with significant intertumoral and intratumoral heterogeneity. Repeat tissue and plasma analyses on therapy have revealed insights into multiple mechanisms of resistance, including novel second site EGFR mutations, activated bypass pathways such as MET amplification, HER2 amplification, RAS mutations, BRAF mutations, PIK3CA mutations, and novel fusion events. Strategies to understand and predict patterns of mutagenesis are still in their infancy; however, technologies to understand synthetically lethal dependencies and track cancer evolution through therapy are being explored. The expansion of combinatorial therapies is a direction forward targeting minimal residual disease and bypass pathways early based on projected resistance.

No abstract

Summary Cellular stress results in profound changes in RNA and protein synthesis. How cells integrate this intrinsic, p53-centered program with extracellular signals is largely unknown. We demonstrate that TGFβ1 signaling interferes with the stress response through coordinate transcriptional and translational repression of p53 levels, which reduces p53-activated transcription, and apoptosis in precancerous cells. Mechanistically, E2F4 binds constitutively to the TP53 gene and induces transcription. TGFβ1-activated Smads are recruited to a composite Smad/E2F4 element by an E2F4/p107 complex that switches to a Smad co-repressor, which represses TP53 transcription. TGFβ1 also causes dissociation of ribosomal protein RPL26 and elongation factor eEF1A from p53 mRNA, thereby reducing p53 mRNA association with polyribosomes and p53 translation. TGFβ1-signalling is dominant over stress-induced transcription and translation of p53 and prevents stress-imposed downregulation of Smad proteins. Thus, crosstalk between the TGFβ and p53 pathways defines a major node of regulation in the cellular stress response, enhancing drug resistance.