Investigating the role of hypoxia in CML metabolism

Sarnello, Daniele (2024) Investigating the role of hypoxia in CML metabolism. PhD thesis, University of Glasgow.

Abstract

Chronic myeloid leukaemia (CML) is a blood cancer arising from a haematopoietic stem cell (HSC) carrying the aberrant chromosomal translocation t (9;22) (q34; q11). The resulting “Philadelphia Chromosome” hosts the fusion oncogene BCR::ABL1, which encodes for a constitutive active non-receptor tyrosine kinase: BCR::ABL1. The gold-standard treatment of CML are tyrosine kinase inhibitors (TKIs), such as Imatinib mesylate. However, TKI treatment does not lead to complete eradication of CML cells as shown by relapse upon treatment suspension. Indeed, the main limitation of TKIs regimen is related to the presence of drug-resistant leukaemic stem cells (LSCs).

The bone marrow (BM) hypoxic microenvironment is an essential niche for the self-renewal and survival of primitive HSCs. Oxygen levels in the BM range from <1% to 6% in the sinusoidal cavity. Hematopoietic cells cultured in vitro under hypoxic conditions are mostly in the G0 phase of the cell cycle and, hence, slow proliferating.

Hypoxia (0.5% O2) enhances CML LSCs stem cell properties, clonogenicity and engraftment in immunocompromised mice despite effective TKI-mediated inhibition of downstream BCR::ABL1 signaling. Low O2 levels reduce the pro-apoptotic effect of Imatinib, resulting in the selection of resistant progenitor cells. However, current research suggests that the transcriptional activity of HIF-1α (master regulator of the hypoxia response) is reduced by Imatinib treatment, implying that persistence of CML LSCs in the hypoxic BM microenvironment could be induced by other mechanisms, whose regulation might be independent of this transcription factor.

Despite numerous studies investigating the metabolic profile of CML LSCs, details on how CML LSCs reprogram their metabolic activity once exposed to hypoxia are lacking. Knowing this could help to better understand how this resistant cellular reservoir can properly function in the BM microenvironment.

In this work we initially show that both short-term and long-term hypoxia promotes transcriptional changes in metabolic genes in CML LSCs. Moreover, via metabolomics studies, we demonstrate that CML cells rewire their mitochondrial metabolic activity when exposed to low oxygen levels. Indeed, while glucose is mostly used for lactate production and secretion, hypoxic CML cells increase glutamine uptake and use it to fuel the tricarboxylic acid cycle via both oxidative and reductive metabolism. Interestingly, we demonstrate that pharmacological inhibition of glutaminase 1 significantly kills CML cells when exposed to hypoxia.

With the attempt to understand deeper the molecular mechanisms involved in this metabolic shift, we observed that the receptor-mediated mitophagy genes BNIP3 and BNIP3L (NIX) are upregulated in CML LSCs compared to non-CML cells. Moreover, we show that BNIP3 is required for proper mitochondrial turnover even under normoxia and erythroid differentiation under hypoxia in CML cells.

Furthermore, via extensive metabolic analysis of CML cells, we suggest a novel role of BNIP3-dependent mitophagy in promoting glutamine anaplerosis and reductive carboxylation in CML cells during their adaption to hypoxia. Of note, mice transplantation with BNIP3 depleted CML cells we prove the importance of BNIP3-mediated mitophagy in survival and tumour formation in an in vivo setting.

Together our results indicate a novel role of BNIP3-mediated mitophagy in metabolic reprogramming and sustaining the survival of CML cells in the hypoxic microenvironment by promoting glutaminolysis and reductive carboxylation.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: Q Science > QH Natural history > QH345 Biochemistry; R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cancer Sciences
• Funder's Name: Cancer Research UK (CRUK)
• Supervisor's Name: Helgason, Professor Vignir
• Date of Award: 2024
• Depositing User: Theses Team
• Unique ID: glathesis:2024-84795
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 13 Jan 2025 08:37
• Last Modified: 13 Jan 2025 08:40
• Thesis DOI: 10.5525/gla.thesis.84795
• URI: https://theses.gla.ac.uk/id/eprint/84795