Metastasis arises owing to tumor cells’ capacity to evade pro-apoptotic signals. Anoikis—the apoptosis of detached cells (from the extracellular matrix (ECM)) is often circumvented by metastatic cells as a result of biochemical and molecular transformations. These facilitate cells’ ability to survive, invade and reattach to secondary sites. Here, we identified deregulated glucose metabolism, oxidative phosphorylation, and proteasome in anchorage-independent cells compared to adherent cells. Metformin an anti-diabetic drug that reduces blood glucose (also known to inhibit mitochondrial Complex I), and proteasome inhibitors were employed to target these changes. Metformin or proteasome inhibitors alone increased misfolded protein accumulation, sensitized tumor cells to anoikis, and impaired pulmonary metastasis in the B16F10 melanoma model. Mechanistically, metformin reduced cellular ATP production, activated AMPK to foster pro-apoptotic unfolded protein response (UPR) through enhanced expression of CHOP in ECM detached cells. Furthermore, AMPK inhibition reduced misfolded protein accumulation, thus highlight relevance of AMPK activation in facilitating metformin-induced stress and UPR cell death. Our findings provide insights into the molecular biology of anoikis resistance and identified metformin and proteasome inhibitors as potential therapeutic options for tumor metastasis.

Despite the remarkable success and efficacy of immune checkpoint blockade (ICB) therapy against the PD-1/PD-L1 axis, it induces sustained responses in a sizeable minority of cancer patients due to the activation of immunosuppressive factors such as myeloid-derived suppressor cells (MDSCs). Inhibiting the immunosuppressive function of MDSCs is critical for successful cancer ICB therapy. Interestingly, lipid metabolism is a crucial factor in modulating MDSCs function. Fatty acid transport protein 2 (FATP2) conferred the function of PMN-MDSCs in cancer via the upregulation of arachidonic acid metabolism. However, whether regulating lipid accumulation in MDSCs by targeting FATP2 could block MDSCs reactive oxygen species (ROS) production and enhance PD-L1 blockade-mediated tumor immunotherapy remains unexplored. Here we report that FATP2 regulated lipid accumulation, ROS, and immunosuppressive function of MDSCs in tumor-bearing mice. Tumor cells-derived granulocyte macrophage-colony stimulating factor (GM-CSF) induced FATP2 expression in MDSCs by activation of STAT3 signaling pathway. Pharmaceutical blockade of FATP2 expression in MDSCs by lipofermata decreased lipid accumulation, reduced ROS, blocked immunosuppressive activity, and consequently inhibited tumor growth. More importantly, lipofermata inhibition of FATP2 in MDSCs enhanced anti-PD-L1 tumor immunotherapy via the upregulation of CD107a and reduced PD-L1 expression on tumor-infiltrating CD8 ⁺ T-cells. Furthermore, the combination therapy blocked MDSC’s suppressive role on T-cells thereby enhanced T-cell’s ability for the production of IFN-γ. These findings indicate that FATP2 plays a key role in modulating lipid-induced ROS in MDSCs and targeting FATP2 in MDSCs provides a novel therapeutic approach to enhance anti-PD-L1 cancer immunotherapy. Research highlights: FATP2 inhibition in MDSCs blocked ROS-mediated immunosuppressive function and promoted MDSCs differentiation to immune-stimulatory phenotype. GM-CSF mediated FATP2 expression via activation of STAT3 signaling leading to lipid accumulation-induced ROS in MDSCs. FATP2 inhibition enhanced anti-PD-L1 tumor immunotherapy by reducing MDSCs immunosuppressive activity in blocking T-cell’s ability to produce IFN-γ. Combination of anti-PD-L1 antibody and FATP2 inhibitor decreased MDSCs accumulation and PD-L1 surface expression in the spleens of LLC-bearing mice. Combined treatment of FATP2 inhibitor and PD-L1 blockade-mediated tumor immunotherapy enhanced tumor-infiltrating CD8 ⁺ T cells activation via increased CD107a and reduced PD-L1 surface expression.