All ETDs from UAB

Advisory Committee Chair

Victor Darley-Usmar

Advisory Committee Members

John Chatham

Anita Hjelmeland

Kasturi Mitra

Hubert Tse

Adam Wende

Document Type

Dissertation

Date of Award

2023

Degree Name by School

Doctor of Philosophy (PhD) School of Medicine

Abstract

Oxidative phosphorylation is an oxygen-dependent metabolic process that provides the majority of ATP used to support essential cellular functions. However, in cancer, limitations in oxygen availability occur during the development and metastasis of tumors. To compensate for the demands of rapidly proliferating cells, many cancers exhibit an increased demand for glycolysis, the TCA cycle, and glutaminolysis. In this thesis, extracellular flux technology and metabolomics were applied in a hypoxia-reoxygenation model to investigate the metabolic adaptations that occur in dynamic changes in oxygen and nutrient availability in cancer cells. We show the significance of glutaminolysis and its substrates in regulating cancer cell bioenergetics and survival in hypoxia-reoxygenation through 1) restricting L-glutamine (L-gln) availability to inhibit oxygen-sensing pathways during hypoxia and damaging mitochondrial Complex I (CI) activity in the electron transport chain (ETC) and, 2) supplementing media with α-ketoglutarate (α-KG) in place of L-gln to partially rescue oxygen-sensing, mitochondrial bioenergetics, CI activity, and viability. To further examine the influence of environmental factors, secondary modifiers of metabolism, nitric oxide (NO) and hydrogen sulfide (H2S), were implemented in our model. These gaseous molecules demonstrated differential effects on mitochondria, with NO showing anti-cancer effects at high doses administered by NO-donor iv compound, DetaNONOate. The relationship between bioenergetics, α-KG availability, and other adjacent L-gln-dependent pathways were assessed such as glutathione (GSH) synthesis. Buthionine sulfoximine, an experimental anti-cancer compound, was used to inhibit GSH synthesis. Although glutaminolysis was not found to be a major contributor to GSH synthesis, α-KG was found to partially protect this process, and GSH aided in protecting mitochondria from reoxygenation damage. Lastly, contributions of CI in regulating electron flux during hypoxia and producing mitochondrially damaging ROS were tested using rotenone, an inhibitor of CI. Our findings revealed α-KG partially reversed pharmacological inhibition of CI and continued to offer partial protection from mitochondrial damage after hypoxia-reoxygenation. Taken together, this work presents novel observations regarding the role of TCA cycle metabolites, L-gln and α-KG, in regulating and protecting mitochondria from oxygen and nutrient limitations in the progression of cancer and may provide a basis for metabolically targeted therapies in treating cancer.

Share

COinS