Abstract P45: Disruption of mTORC2 Activity by Pdcd4–Rictor Binding Attenuates Tumor Growth in NSCLC

Abstract

Programmed cell death 4 (Pdcd4) is a tumor suppressor frequently downregulated in various cancers, including breast, colon, liver, lung, pancreas, prostate, and skin. It inhibits tumor initiation, proliferation, invasion, and metastasis. Biochemically, Pdcd4 suppresses protein translation by inhibiting the helicase activity of eIF4A, a translation initiation factor. This inhibition is thought to reduce the expression of genes critical for tumor progression. Beyond its role in translation, we recently uncovered a translation-independent tumor-suppressive mechanism of Pdcd4. Specifically, Pdcd4 interacts with the rapamycin-insensitive companion of mTOR (Rictor), a key component of the mTORC2 complex. Domain mapping through deletion and mutation analyses identified the Rictor-binding region on Pdcd4. Co-immunoprecipitation and in vitro kinase assays showed that Pdcd4 binding to Rictor disrupts mTORC2 complex formation and inhibits its kinase activity. Reverse phase protein array analysis revealed the elevated levels of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3) in Pdcd4 knockdown cells. This effect was reversed by re-expression of wild-type Pdcd4 but not by a Rictor-binding deficient mutant in Pdcd4-knockdown cells, indicating that Pdcd4 downregulates PFKFB3 protein through its interaction with Rictor. Further analysis demonstrated that Pdcd4 promotes ubiquitin-proteasome-mediated degradation of PFKFB3. Functionally, wild-type Pdcd4, but not the Rictor-binding deficient mutant, significantly suppressed tumor cell proliferation and colony formation in vitro. In vivo, only wild-type Pdcd4 inhibited tumor growth in a xenograft model using nude mice. Taken together, our findings suggest that Pdcd4 suppresses NSCLC cell growth through a novel, translation-independent mechanism involving disruption of mTORC2 complex formation via binding to Rictor, leading to reduced PFKFB3 protein abundance.