Abstract
Acute myeloid leukemia (AML) is a molecularly heterogeneous malignant disease of the bone marrow resulting from arrest of hematopoietic precursors and uncontrolled proliferation of myeloblasts. Inhibition of the BET-family co-activator BRD4 (BETi) shows encouraging pre-clinical activity but limited clinical activity as a single agent. We previously demonstrated that MYC suppression by BETi induces metabolic and mitochondrial changes in AML blasts leading to a collapse in mitochondrial respiration. AML blasts that are resistant to BETi undergo comparable mitochondrial respiration despite impaired glycolysis and glutaminolysis. The metabolic pathway allowing BETi-resistant AML blasts to maintain metabolic integrity is unclear. Therefore, to identify how AML blast cells metabolically bypass BET inhibition, we used two different cell lines: BETi-resistant MOLM-13 and BETi-susceptible MV-4-11 cells. AML cell lines were treated with BETi for 48 hr and global untargeted metabolic mass spectrometry revealed that glycolysis, glutaminolysis, and fatty acid metabolism were not leveraged to maintain tricarboxylic acid (TCA) cycle activity (Fig. 1A). Rather, an increased NADH/NAD+ ratio and intracellular lactate accumulation suggested that lactate was being utilized as an alternative carbon source to sustain mitochondrial respiration. In support, MOLM-13 cells treated with BETi accumulated higher levels of mitochondrial lactate dehydrogenase, which preferentially converts lactate into pyruvate to fuel the TCA cycle. The heightened capacity of MOLM-13 cells to utilize lactate better maintained cellular viability but failed to support cellular replication, suggesting that lactate utilization promotes cellular senescence during BET inhibition. Therefore, we employed three chemical inhibitors of lactate utilization to couple with BETi: AZD3965 to prevent extracellular lactate uptake, UK5099 to block lactate import into the mitochondria, and oxamate to broadly impair lactate dehydrogenase activity. All three chemical inhibitors of lactate utilization were sufficient to render MOLM-13 cells susceptible to BET inhibition, as mitochondrial respiration, ATP production, and cellular viability were decreased to levels comparable to MV-4-11 cells. A similar correlation was observed in patient samples where AML blasts with increased resistance to BETi were predisposed to utilize lactate as an alternative carbon source and rendered susceptible to BET inhibition with lactate utilization inhibitors. Furthermore, these results were translated in vivo as low-dose combination of BETi and oxamate significantly reduced burdens of BETi-resistant cells in murine models (Fig. 1B).
These results demonstrate that some AML blasts metabolically adapt to overcome BET inhibition by consuming lactate but that combinations of BETi and lactate utilization inhibitors can stymie this resistance. Recognizing the metabolic switching to utilize lactate in AML therapy resistance, and studying lactate utilization inhibitors in clinical settings, especially in BETi-based therapies should be considered.
Disclosures
Stubbs:Incyte Corp.: Current Employment. Rathmell:Tempest Therapeutics: Research Funding; Sitryx Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Caribou Biosciences: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Nirogy Therapeutics: Membership on an entity's Board of Directors or advisory committees; Merck: Consultancy; Pfizer: Consultancy; Mitobridge: Consultancy; Incyte Corp.: Research Funding; Calithera Biosciences: Research Funding. Savona:Taiho Pharmaceutical: Consultancy; TG Therapeutics: Consultancy, Other: Travel expenses, Research Funding; Karyopharm Therapeutics: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; ALX Oncology: Research Funding; Astex Pharmaceuticals: Research Funding; Ryvu Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy; Incyte Corporation: Research Funding; Geron: Consultancy; Novartis: Consultancy; Sierra Oncology: Consultancy, Other: travel expenses; Bristol Myers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: travel expenses; Forma: Consultancy; AbbVie: Consultancy, Other: travel expenses.
