Lung tumors with distinct p53 mutations respond similarly to p53 targeted therapy but exhibit genotype-specific statin sensitivity.

Lung adenocarcinoma accounts for ∼40% of lung cancers, the leading cause of cancer-related death worldwide, and current therapies provide only limited survival benefit. Approximately half of lung adenocarcinomas harbor mutations in TP53 (p53), making these mutants appealing targets for lung cancer therapy. As mutant p53 remains untargetable, mutant p53-dependent phenotypes represent alternative targeting opportunities, but the prevalence and therapeutic relevance of such effects (gain of function and dominant-negative activity) in lung adenocarcinoma are unclear. Through transcriptional and functional analysis of murine KrasG12D -p53null , -p53R172H (conformational), and -p53R270H (contact) mutant lung tumors, we identified genotype-independent and genotype-dependent therapeutic sensitivities. Unexpectedly, we found that wild-type p53 exerts a dominant tumor-suppressive effect on mutant tumors, as all genotypes were similarly sensitive to its restoration in vivo. These data show that the potential of p53 targeted therapies is comparable across all p53-deficient genotypes and may explain the high incidence of p53 loss of heterozygosity in mutant tumors. In contrast, mutant p53 gain of function and their associated vulnerabilities can vary according to mutation type. Notably, we identified a p53R270H -specific sensitivity to simvastatin in lung tumors, and the transcriptional signature that underlies this sensitivity was also present in human lung tumors, indicating that this therapeutic approach may be clinically relevant.

Mutant Kras copy number defines metabolic reprogramming and therapeutic susceptibilities.

The RAS/MAPK (mitogen-activated protein kinase) signalling pathway is frequently deregulated in non-small-cell lung cancer, often through KRAS activating mutations. A single endogenous mutant Kras allele is sufficient to promote lung tumour formation in mice but malignant progression requires additional genetic alterations. We recently showed that advanced lung tumours from Kras(G12D/+);p53-null mice frequently exhibit Kras(G12D) allelic enrichment (Kras(G12D)/Kras(wild-type) > 1) (ref. 7), implying that mutant Kras copy gains are positively selected during progression. Here we show, through a comprehensive analysis of mutant Kras homozygous and heterozygous mouse embryonic fibroblasts and lung cancer cells, that these genotypes are phenotypically distinct. In particular, Kras(G12D/G12D) cells exhibit a glycolytic switch coupled to increased channelling of glucose-derived metabolites into the tricarboxylic acid cycle and glutathione biosynthesis, resulting in enhanced glutathione-mediated detoxification. This metabolic rewiring is recapitulated in mutant KRAS homozygous non-small-cell lung cancer cells and in vivo, in spontaneous advanced murine lung tumours (which display a high frequency of Kras(G12D) copy gain), but not in the corresponding early tumours (Kras(G12D) heterozygous). Finally, we demonstrate that mutant Kras copy gain creates unique metabolic dependences that can be exploited to selectively target these aggressive mutant Kras tumours. Our data demonstrate that mutant Kras lung tumours are not a single disease but rather a heterogeneous group comprising two classes of tumours with distinct metabolic profiles, prognosis and therapeutic susceptibility, which can be discriminated on the basis of their relative mutant allelic content. We also provide the first, to our knowledge, in vivo evidence of metabolic rewiring during lung cancer malignant progression.

Targeting the activated microenvironment with endosialin (CD248)-directed CAR-T cells ablates perivascular cells to impair tumor growth and metastasis.

BACKGROUND: Targeting of solid cancers with chimeric antigen receptor (CAR)-T cells is limited by the lack of suitable tumor-specific antigens and the immunosuppressive, desmoplastic tumor microenvironment that impedes CAR-T cell infiltration, activity and persistence. We hypothesized that targeting the endosialin (CD248) receptor, strongly expressed by tumor-associated pericytes and perivascular cancer-associated fibroblasts, would circumvent these challenges and offer an exciting antigen for CAR-T cell therapy due to the close proximity of target cells to the tumor vasculature, the limited endosialin expression in normal tissues and the lack of phenotype observed in endosialin knockout mice. METHODS: We generated endosialin-directed E3K CAR-T cells from three immunocompetent mouse strains, BALB/c, FVB/N and C57BL/6. E3K CAR-T cell composition (CD4+/CD8+ ratio), activity in vitro against endosialin+ and endosialin- cells, and expansion and activity in vivo in syngeneic tumor models as well as in tumor-naive healthy and wounded mice and tumor-bearing endosialin knockout mice was assessed. RESULTS: E3K CAR-T cells were active in vitro against both mouse and human endosialin+, but not endosialin-, cells. Adoptively transferred E3K CAR-T cells exhibited no activity in endosialin knockout mice, tumor-naive endosialin wildtype mice or in wound healing models, demonstrating an absence of off-target and on-target/off-tumor activity. By contrast, adoptive transfer of E3K CAR-T cells into BALB/c, FVB/N or C57BL/6 mice bearing syngeneic breast or lung cancer lines depleted target cells in the tumor stroma resulting in increased tumor necrosis, reduced tumor growth and a substantial impairment in metastatic outgrowth. CONCLUSIONS: Together these data highlight endosialin as a viable antigen for CAR-T cell therapy and that targeting stromal cells closely associated with the tumor vasculature avoids CAR-T cells having to navigate the harsh immunosuppressive tumor microenvironment. Further, the ability of E3K CAR-T cells to recognize and target both mouse and human endosialin+ cells makes a humanized and optimized E3K CAR a promising candidate for clinical development applicable to a broad range of solid tumor types.

Age-associated microenvironmental changes highlight the role of PDGF-C in ER+ breast cancer metastatic relapse.

Patients with estrogen receptor (ER)-positive breast cancer are at risk of metastatic relapse for decades after primary tumor resection and treatment, a consequence of dormant disseminated tumor cells (DTCs) reawakening at secondary sites. Here we use syngeneic ER+ mouse models in which DTCs display a dormant phenotype in young mice but accelerated metastatic outgrowth in an aged or fibrotic microenvironment. In young mice, low-level Pdgfc expression by ER+ DTCs is required for their maintenance in secondary sites but is insufficient to support development of macrometastases. By contrast, the platelet-derived growth factor (PDGF)-Chi environment of aging or fibrotic lungs promotes DTC proliferation and upregulates tumor cell Pdgfc expression stimulating further stromal activation, events that can be blocked by pharmacological inhibition of PDGFRα or with a PDGF-C-blocking antibody. These results highlight the role of the changing microenvironment in regulating DTC outgrowth and the opportunity to target PDGF-C signaling to limit metastatic relapse in ER+ breast cancer.