Branched Chain Amino Acid Catabolism Restricts Alpha-Ketoglutarate Levels Resembling IDH^mut-Driven DNA Hypermethylation in AML Stem Cells

The branched chain amino acid (BCAA) pathway and high levels of BCAA transaminase 1 (BCAT1) have recently been associated with aggressiveness in several cancer entities. However, the mechanistic role of BCAT1 in this process remains largely uncertain. By performing high-resolution proteomic analysis of human acute myeloid leukaemia (AML) stem cell (LSC) and non-LSC populations, we found the BCAA pathway enriched and BCAT1 protein and transcripts overexpressed in LSCs. We show that BCAT1, which transfers α-amino groups from BCAAs to α-ketoglutarate (αKG), is a critical regulator of intracellular αKG homeostasis. Next to its role in the tricarboxylic acid (TCA) cycle, αKG is an essential co-factor for αKG-dependent dioxygenases such as EGLN1 and the TET family of DNA demethylases. Knockdown of BCAT1 in leukaemia cells caused accumulation of αKG leading to EGLN1-mediated HIF1α protein degradation. This resulted in a growth and survival defect and abrogated leukaemia-initiating potential. In contrast, overexpression (OE) of BCAT1 in leukaemia cells decreased intracellular αKG levels and caused DNA hypermethylation via altered TET activity. BCAT1^high AMLs displayed a DNA hypermethylation phenotype similar to cases carrying mutant isocitrate dehydrogenase (IDH^mut), in which TET2 is inhibited by the oncometabolite 2-hydroxyglutarate. High levels of BCAT1 strongly correlate with shorter overall survival in IDH^wtTET2^wt, but not IDH^mut or TET2^mut AMLs. Gene sets characteristic for IDH^mut AMLs were enriched in IDH^wtTET^wtBCAT1^high patient samples. BCAT1^high AMLs showed robust enrichment for LSC signatures, and paired sample analysis revealed a significant increase in BCAT1 levels upon disease relapse. In summary, by limiting intracellular αKG, BCAT1 links BCAA catabolism to HIF1α stability and regulation of the epigenomic landscape mimicking the effects of IDH mutations. In essence, our data strongly suggest that suppression of αKG-dependent dioxygenases constitutes a common feature of AML and, next to mutations in IDH and TET2, we identify the BCAA-BCAT1-αKG axis as alternative, non-genetic mechanism in LSCs to control the activity of these enzymes. Strategies to increase αKG levels, i.e. by inhibition of BCAT1 in order to compromise cancer stem cell function may lead to improved clinical outcomes of patients suffering from AML or other tumour entities expressing high levels of BCAT1 in the absence of IDH or TET mutations.