Date of Award

2014

Degree Type

Dissertation

Department

Biological, Geological and Environmental Sciences

First Advisor

Almasan, Alexandru

Subject Headings

Lymphocytic leukemia -- Chemotherapy, Cancer -- Molecular aspects, Enzyme inhibitors -- Therapeutic use, Fludarabine, Biochemistry, biology, cellular biology, molecular biology

Abstract

Deregulated mTORC1 contributes to tumorigenesis and chemoresistance. However, mTORC1-specific inhibitors (rapalogs) show modest efficacy in the clinic, as they unleash the feedback inhibition on upstream, prosurvival pathways. An alternative approach is to target downstream functions of mTORC1. We investigated acquired resistance to fludarabine (Flu), a purine analog, active agent for chronic lymphocytic leukemia (CLL). Elevated phospho-p70S6k (T389), an mTORC1 activation marker, predicted Flu resistance in a panel of B-cell tumor lines, and primary CLL cells. However, the rapalogs induced moderate cell death in Flu-resistant (FluR) and primary CLL cells. Activation of oncogenic pathways, including mTORC1, induces profound metabolic reprogramming to provide energy and biosynthetic substrates for tumor growth. Therefore, we investigated the metabolic consequences of mTORC1 activation in FluR compared to Flu-sensitive (FluS) cells, aiming to identify selective vulnerability of FluR cells to interference with specific metabolic pathways (Aim1). Seahorse metabolic analysis revealed mTORC1-mediated increase in glycolysis and mitochondrial respiration, which was uncoupled from ATP synthesis in the FluR cells. Additionally, we observed increased mTORC1-p70S6k dependent phospho-CAD (S1859) and, S-phase population in FluR cells, suggesting accelerated de novo pyrimidine biosynthesis. Consistently, siRNA-mediated knockdown of p70S6K, and the pharmacological inhibition of the enzymes of de novo pyrimidine biosynthesis pathway, or associated mitochondrial respiration were selectively cytotoxic to FluR, but not FluS cells. Such oncogene-mediated metabolic reprogramming confers selective growth and survival advantage to cancer cells. However, the by-products of unrestrained biosynthesis and exhaustive mitochondrial metabolism will cause metabolic stress. Autophagy, by removal and recycling of damaged cellular contents, functions as a prominent cellular metabolic stress adaptation pathway. This suggested autophagy mi

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