Non-Canonical Role Of Pdk1 As A Negative Regulator Of Apoptosis Through Macromolecular Complexes Assembly At The Er-Mitochondria Interface In Oncogene-Driven Nsclc

Simple Summary Co-targeting of glucose metabolism and oncogene drivers in patients with non-small cell lung cancer (NSCLC) has been proposed as a potentially effective therapeutic strategy. Here, we demonstrate that downregulation of pyruvate dehydrogenase kinase 1 (PDK1), an enzyme of glycolytic cascade, enhances maximal respiration of cancer cells by upregulating mitochondrial complexes of oxidative phosphorylation (OXPHOS) and improves tumor response to tyrosine kinase inhibitors by promoting apoptosis. Furthermore, we provided consistent evidence that PDK1 drives the formation of macromolecular complexes at the ER-mitochondria interface involving PKM2, Bcl-2 and Bcl-xL and serves as an indirect anchorage of anti-apoptotic proteins to the mitochondrial membrane. Our findings taken together highlighted a non-canonical role of PDK1 as a negative regulator of apoptosis, thus coupling the glycolytic phenotype to drug resistance. The major translational relevance of this study is to provide a rational basis for combined therapeutic strategies targeting PDK1 and oncogene drivers in NSCLC patients. Here, we tested whether co-targeting of glucose metabolism and oncogene drivers may enhance tumor response to tyrosine kinase inhibitors (TKIs) in NSCLC. To this end, pyruvate dehydrogenase kinase 1 (PDK1) was stably downregulated in oncogene-driven NSCLC cell lines exposed or not to TKIs. H1993 and H1975 cells were stably transfected with scrambled (shCTRL) or PDK1-targeted (shPDK1) shRNA and then treated with MET inhibitor crizotinib (1 mu M), double mutant EGFR(L858R/T790M) inhibitor WZ4002 (1 mu M) or vehicle for 48 h. The effects of PDK1 knockdown on glucose metabolism and apoptosis were evaluated in untreated and TKI-treated cells. PDK1 knockdown alone did not cause significant changes in glycolytic cascade, ATP production and glucose consumption, but it enhanced maximal respiration in shPDK1 cells when compared to controls. When combined with TKI treatment, PDK1 downregulation caused a strong enhancement of OXPHOS and a marked reduction in key glycolytic enzymes. Furthermore, increased levels of apoptotic markers were found in shPDK1 cells as compared to shCTRL cells after treatment with TKIs. Co-immunoprecipitation studies showed that PDK1 interacts with PKM2, Bcl-2 and Bcl-xL, forming macromolecular complexes at the ER-mitochondria interface. Our findings showed that downregulation of PDK1 is able to potentiate the effects of TKIs through the disruption of macromolecular complexes involving PKM2, Bcl-2 and Bcl-xL.