Disruption of Autophagic Flux and Treatment with the PDPK1 Inhibitor GSK2334470 Synergistically Inhibit Renal Cell Carcinoma Pathogenesis

Background:
Renal cell carcinoma (RCC) often shows mutations that activate the PI3K-Akt-mTOR signaling pathway. The enzyme 3-Phosphoinositide-dependent kinase 1 (PDPK1 or PDK1) is crucial for regulating this pathway. Meanwhile, mTOR serves as the primary factor initiating autophagy. Despite this, our understanding of the interplay between PDPK1 and autophagy in RCC remains limited.

Methods:
We utilized GSK2334470 (GSK470), a novel and highly selective PDPK1 inhibitor, to explore its anticancer effects in two RCC cell lines. Cell proliferation was evaluated through the CCK-8 assay and colony formation tests. The levels of key components in the Akt/mTOR pathway and apoptosis markers were analyzed using Western blotting. Autophagy was assessed by measuring LC3B expression, employing transmission electron microscopy, and utilizing a tandem mRFP-EGFP-LC3 construct. In vivo effects of PDPK1 and the autophagy inhibitor chloroquine were studied in a mouse model with tumors.

Results:
GSK470 significantly reduced cell proliferation and induced apoptosis in both A498 and 786-O RCC cells. It downregulated PDPK1 phosphorylation, leading to the inhibition of downstream Akt1 phosphorylation at Thr308 and Ser473 and a decrease in mTOR complex 1 (mTORC1) activity. Additionally, treatment with insulin-like growth factor-1 (IGF-1) partially reversed the effects induced by GSK470. Notably, treatment of A498 and 786-O cells with GSK470 or siPDPK1 resulted in marked increases in autophagy markers, including autophagosome accumulation, autophagic flux, and LC3B expression. Furthermore, GSK470 and chloroquine exhibited a synergistic effect, inhibiting RCC cell growth both in vitro and in xenograft models, highlighting the protective role of autophagy activation when blocking the PDPK1-Akt-mTOR signaling pathway.

Conclusion:
Our findings provide new insights into the potential of combining PDPK1 inhibition with autophagy suppression as a promising treatment strategy for RCC.