The retinoblastoma protein mediates metabolic reprogramming in lung cancer.

Abstract

Lung cancer is among the most frequently diagnosed cancers and is the leading cause of cancer-related deaths worldwide. One of the hallmark events in lung cancer pathogenesis is deregulation of the cell cycle. The retinoblastoma protein (pRb) is a tumor suppressor that is deleted, mutated, or inactivated in most lung cancer cases. Canonically, pRb functions to regulate cell cycle progression by repressing the transcriptional activity of the E2F family of transcription factors, inhibiting S phase entry. Although the cell cycle functions of pRb have been well established, recent studies have highlighted a functional role for pRb in controlling cellular metabolism. This body of work describes the role of pRb in mediating metabolic reprogramming in both in vitro and in vivo models of lung cancer. To define metabolic pathways regulated by pRb in vitro, I performed several metabolic assays and stable-isotope labeled metabolomics studies in A549 lung adenocarcinoma cells. pRb activity was manipulated via treatment with palbociclib, a CDK4/6 inhibitor which inhibits the phosphorylation and subsequent inactivation of pRb. Palbociclib decreases nucleotide biosynthesis by reducing activity of glucose 6-phosphate dehydrogenase, the rate limiting enzyme in the pentose phosphate pathway (PPP), without altering glycolysis. Additionally, palbociclib increases glutamine dependency for mitochondrial function and sensitizes A549 cells to the glutaminase inhibitor, CB-839. The effects of palbociclib on both the PPP and glutamine utilization occur in an pRb-dependent manner. Separately, using a combination of steady-state and stable-isotope labeled metabolomics, I assessed global changes in metabolism resulting from the loss of the gene encoding pRb (Rb1) in Kras-driven lung tumors in vivo. Depletion of Rb1 in a mutant Kras-driven mouse model of lung cancer enhances glucose metabolism, via upregulation of key genes within glycolysis, without altering mitochondrial pyruvate oxidation. Moreover, pRb-deficient tumors do not exhibit alterations in glutamine anaplerosis or utilization of alternative nutrient sources (e.g. circulating lactate) compared to lung tumors with intact pRb. Together the work described in this dissertation expands our knowledge of the metabolic phenotype resulting from pRb dysfunction in lung cancer, and highlights specific metabolic perturbations that may be exploited therapeutically based on pRb status in patient lung tumors.