Pancreatic ductal adenocarcinoma (PDAC), like many KRAS-driven tumors, preferentially loses CDKN2A that encodes an endogenous CDK4/6 inhibitor to bypass the RB-mediated cell cycle suppression. Analysis of a panel of patient-derived cell lines and matched xenografts indicated that many pancreatic cancers have intrinsic resistance to CDK4/6 inhibition that is not due to any established mechanism or published biomarker. Rather, there is a KRAS-dependent rapid adaptive response that leads to the upregulation of cyclin proteins, which participate in functional complexes to mediate resistance. In vivo , the degree of response is associated with the suppression of a gene-expression signature that is strongly prognostic in pancreatic cancer. Resistance is associated with an adaptive gene expression signature which is common to multiple kinase inhibitors, but is attenuated with MTOR inhibitors. Combination treatment with MTOR and CDK4/6 inhibitors had potent activity across a large number of patient derived models of PDAC underscoring the potential clinical efficacy.

Cancer biology is influenced by the tumor microenvironment that impacts on disease prognosis and therapeutic intervention. The inter-relationship of tumor infiltrating lymphocytes, immune response regulators, and a glycolytic tumor environment was evaluated in a cohort of 183 largely consecutive patients with triple negative breast cancer diagnosis. High levels of tumor infiltrating lymphocytes were associated with improved survival of triple negative breast cancer cases. However, elevated levels of PD-L1, CD163, and FOXP3 were individually associated with significantly decreased overall survival. These three determinants were significantly correlated, and could serve to differentiate the prognostic significance of tumor infiltrating lymphocytes. Interestingly, a glycolytic tumor environment, as determined by the expression of MCT4 in the tumor stroma, was associated with the immune evasive environment and poor prognosis. Clustering of all markers defined four distinct triple negative breast cancer subtypes that harbored prognostic significance in multi-variate analysis. Immune and metabolic markers stratify triple negative breast cancer into subtypes that have prognostic significance and implications for therapies targeting immune checkpoints and tumor metabolism.

ObjectiveThis study exploits the intersection between molecular-targeted therapies and immune-checkpoint inhibition to define new means to treat pancreatic cancer.DesignPatient-derived cell lines and xenograft models were used to define the response to CDK4/6 and MEK inhibition in the tumour compartment. Impacts relative to immunotherapy were performed using subcutaneous and orthotopic syngeneic models. Single-cell RNA sequencing and multispectral imaging were employed to delineate effects on the immunological milieu in the tumour microenvironment.ResultsWe found that combination treatment with MEK and CDK4/6 inhibitors was effective across a broad range of PDX models in delaying tumour progression. These effects were associated with stable cell-cycle arrest, as well as the induction of multiple genes associated with interferon response and antigen presentation in an RB-dependent fashion. Using single-cell sequencing and complementary approaches, we found that the combination of CDK4/6 and MEK inhibition had a significant impact on increasing T-cell infiltration and altering myeloid populations, while potently cooperating with immune checkpoint inhibitors.ConclusionsTogether, these data indicate that there are canonical and non-canonical features of CDK4/6 and MEK inhibition that impact on the tumour and immune microenvironment. This combination-targeted treatment can promote robust tumour control in combination with immune checkpoint inhibitor therapy.

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Pancreatic ductal adenocarcinoma (PDAC) is a therapy recalcitrant disease characterized by the aberrations in multiple genes that drive pathogenesis and limit therapeutic response. While CDK4/6 represents a downstream target of both KRAS mutation and loss of the CDKN2A tumor suppressor in PDAC, clinical and preclinical studies indicate that pharmacological CDK4/6 inhibitors are only modestly effective. Since chemotherapy represents the established backbone of PDAC treatment we evaluated the interaction of CDK4/6 inhibitors with gemcitabine and taxanes that are employed in the treatment of PDAC. Herein, we demonstrate that the difference in mechanisms of actions of chemotherapeutic agents elicit distinct effects on the cellular response to CDK4/6 inhibition. Gemcitabine largely ablates the function of CDK4/6 inhibition in S-phase arrested cells when administered contemporaneously; although, when cells recover from S-phase block they exhibit sensitivity to CDK4/6 inhibition. In contrast, pharmacological inhibition of CDK4/6 yields a cooperative cytostatic effect in combination with docetaxel and prevents adaptation and cell cycle re-entry, which is a common basis for resistance to such agents. Importantly, using organoid and PDX models we could confirm the cooperative effects between chemotherapy and CDK4/6 inhibition. These data indicate that the combination of cytotoxic and cytostatic agents could represent an important modality in those tumor types that are relatively resistant to CDK4/6 inhibitors. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: