Oncogenic (mutant) Ras protein Kirsten rat sarcoma viral oncogene homolog (KRAS) promotes uncontrolled proliferation, altered metabolism, and loss of genome integrity in a cell-intrinsic manner. Here, we demonstrate that CD4⁺ T cells when incubated with tumor-derived exosomes from mutant (MT) KRAS non-small-cell lung cancer (NSCLC) cells, patient sera, or a mouse xenograft model, induce phenotypic conversion to FOXP3⁺ Treg-like cells that are immune-suppressive. Furthermore, transfecting T cells with MT KRAS cDNA alone induced phenotypic switching and mathematical modeling supported this conclusion. Single-cell sequencing identified the interferon pathway as the mechanism underlying the phenotypic switch. These observations highlight a novel cytokine-independent, cell-extrinsic role for KRAS in T cell phenotypic switching. Thus, targeting this new class of Tregs represents a unique therapeutic approach for NSCLC. Since KRAS is the most frequently mutated oncogene in a wide variety of cancers, the findings of this investigation are likely to be of broad interest and have a large scientific impact.

Drug resistance, a major challenge in cancer therapy, is typically attributed to mutations and genetic heterogeneity. Emerging evidence suggests that dynamic cellular interactions and group behavior also contribute to drug resistance. However, the underlying mechanisms remain poorly understood. Here, we present a new mathematical approach with game theoretical underpinnings that we developed to model real-time growth data of non-small cell lung cancer (NSCLC) cells and discern patterns in response to treatment with cisplatin. We show that the cisplatin-sensitive and cisplatin-tolerant NSCLC cells, when co-cultured in the absence or presence of the drug, display dynamic group behavior strategies. Tolerant cells exhibit a ‘persister-like’ behavior and are attenuated by sensitive cells; they also appear to ‘educate’ sensitive cells to evade chemotherapy. Further, tolerant cells can switch phenotypes to become sensitive, especially at low cisplatin concentrations. Finally, switching treatment from continuous to an intermittent regimen can attenuate the emergence of tolerant cells, suggesting that intermittent chemotherapy may improve outcomes in lung cancer.

Mitochondria are dynamic organelles that constantly fuse and divide, forming dynamic tubular networks. Abnormalities in mitochondrial dynamics and morphology are linked to diverse pathological states, including cancer. Thus, alterations in mitochondrial parameters could indicate early events of disease manifestation or progression. However, finding reliable and quantitative tools for monitoring mitochondria and determining the network parameters, particularly in live cells, has proven challenging. Here, we present a 2D confocal imaging-based approach that combines automatic mitochondrial morphology and dynamics analysis with fractal analysis in live small cell lung cancer (SCLC) cells. We chose SCLC cells as a test case since they typically have very little cytoplasm, but an abundance of smaller mitochondria compared to many of the commonly used cell types. The 2D confocal images provide a robust approach to quantitatively measure mitochondrial dynamics and morphology in live cells. Furthermore, we performed 3D reconstruction of electron microscopic images and show that the 3D reconstruction of the electron microscopic images complements this approach to yield better resolution. The data also suggest that the parameters of mitochondrial dynamics and fractal dimensions are sensitive indicators of cellular response to subtle perturbations, and hence, may serve as potential markers of drug response in lung cancer.