Riboflavin Drives Nucleotide Biosynthesis and Iron-Sulfur Metabolism to Promote Acute Myeloid Leukemia

• Published in: Blood 2024-11, Vol.144, p.626-626
• Main Authors: Bjelosevic, Stefan, Fauth, Rahel, Do, Brian T, Alexe, Gabriela, Ryback, Birgitta, Basanthakumar, Allen T, Merickel, Lucy A, Salhotra, Silvi, Taillon, Audrey, Jackson, Thomas D, DiGiovanni, Giulia, Munim, Muhammad Bin, Elbashir, Ryan, Lin, Shan, Puigserver, Pere, Root, David E., Vander Heiden, Matthew G, Stegmaier, Kimberly
• Format: Article
• Language: English
• ISSN: 0006-4971
• DOI: 10.1182/blood-2024-208370

Cofactors, many of which are vitamin-derived, are organic molecules with diverse biological functions. They are required for many metabolic pathways and enzymatic reactions, and thus can influence cell proliferation and differentiation. Though the role of vitamins in non-malignant contexts is relatively well characterized, their role in carcinogenesis is not well defined. Here, we sought to molecularly characterize how vitamins contribute to oncogenesis in acute myeloid leukemia (AML). We performed vitamin depletion screens in AML cells using culture medium designed to mimic human plasma. These screens revealed deprivation of riboflavin (Vitamin B2) as detrimental to AML proliferation. Analysis of the Cancer Dependency Map (DepMap) and in vivo CRISPR-Cas9 screens identified the first, rate-limiting enzyme of riboflavin metabolism, riboflavin kinase (RFK), as a hematological cancer dependency. Genetic knockout (KO) of RFK or exogenous depletion of riboflavin in genetically diverse AML cell lines and patient-derived xenograft (PDX) cells induced AML differentiation and apoptosis. Riboflavin depletion was rescued by addition of flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), the cofactor products of riboflavin metabolism. We validated that the growth of non-hematological cancer cells was largely unperturbed by exogenous deprivation of riboflavin or RFK KO, highlighting the specificity of this vulnerability in AML. To understand the mechanistic basis of riboflavin dependency in AML, we performed mass spectrometry-based metabolomics studies. Whilst RFK loss dramatically reduced FMN and FAD pools, it also selectively depleted pyrimidine nucleotide species. A subsequent loss-of-function screen using riboflavin restricted culture medium and a metabolism-focused CRISPR-Cas9 library showed that loss of multiple genes encoding purine nucleotide biosynthesis enzymes promoted cell growth in riboflavin-restricted conditions, thereby alleviating the nucleotide imbalance induced by RFK loss. Accordingly, uridine supplementation partially restored proliferation in riboflavin deficient media. Nucleotide imbalance was accompanied by strongly impaired assembly of mitochondrial complex I and II which led to concomitant collapse in oxidative phosphorylation; indeed, loss of respiratory complex I and II subunits enhanced the anti-leukemic effects of riboflavin depletion in our screen. Our CRISPR-Cas9 screen also revealed, unexpectedly, that loss of iron-sulfur