FMS-like tyrosine kinase 3 (FLT3) mutations in acute myeloid leukemia (AML) are associated with adverse prognosis. FLT3 inhibitors (FLT3i) improve therapeutic response; however, diverse resistance mechanisms, such as adaptations in lipid metabolism, have been identified. We hypothesized that a lipid-rich ketogenic diet (KD) might alter both host and tumoral lipid metabolism, enhancing responses to FLT3i. In FLT3-mutated AML mouse models, 3 weeks of lard- or plant-based KD improved the efficacy of FLT3i by 2-fold reduction of engraftment and tumor burden. KD increased ketone bodies and lipid accumulation in plasma, liver, and AML cells and also induced a polyunsaturated fatty acid:monounsaturated fatty acid (PUFA:MUFA) imbalance. KD impacted pentoses, hexoses, and amino acid metabolism, enhancing sugar phosphates and vitamins in the host. Mechanistically, KD rewired anabolism toward fatty acid oxidation and glycine-utilizing pathways, modulated the expression of FLT3 signaling pathways and lipid biosynthesis, and promoted tumor cell differentiation. In conclusion, this study shows that KD reduces FLT3i resistance, offering a promising therapeutic solution.

Background Right ventricular (RV) adaptation to chronic thromboembolic pulmonary hypertension (CTEPH) remains incompletely understood. Precise phenotyping of RV reverse remodeling after pulmonary endarterectomy (PEA) may help identify biomarkers and generate hypotheses for future mechanistic and therapeutic studies on RV. We aimed to assess longitudinal changes in RV imaging, histology, and circulating metabolites following PEA. Methods We conducted a prospective study including 10 patients with CTEPH who were referred for PEA and 10 controls with normal RV function who were referred for aortic surgery. All patients underwent preoperative echocardiography, cardiac magnetic resonance, 18-fluoro-deoxy-glucose positron emission tomography, perioperative RV myocardial biopsies, and mass-spectrometry-based untargeted metabolomic analyses of plasma samples. Imaging and metabolomic studies were repeated at 1 and 6 months after surgery in patients with CTEPH to assess their longitudinal evolution. Results Compared to controls, RV histology of patients with CTEPH demonstrated capillary rarefaction, cardiomyocyte hypertrophy, marked fibrosis, and macrophage infiltration, all of which were significantly associated with RV dysfunction, RV-pulmonary arterial (RV-PA) uncoupling, adverse remodeling, and increased myocardial glucose uptake. Circulating metabolic profiles of patients with CTEPH exhibited pronounced alterations in arginine, lysine, purine, and fatty acid metabolic pathways compared to controls, as assessed both cross-sectionally and longitudinally. These metabolic changes were correlated with both histological and imaging features. Conclusions Multimodality imaging, histology, and metabolomics highlighted longitudinal changes associated with RV reverse remodeling following PEA. Capillary rarefaction, cardiomyocyte hypertrophy, fibrosis, and inflammation were observed in CTEPH patients, consistent with previous experimental studies. Alterations in arginine, lysine, purine, and fatty acid metabolic pathways were associated with RV remodeling and recovery, generating hypotheses regarding potential metabolic biomarkers and mechanisms involved in RV adaptation in CTEPH.

Medulloblastoma, the most common malignant brain tumor of childhood, exhibits significant biological complexity that demands deeper exploration. Here, we present a large multiomics dataset integrating data from 384 primary medulloblastoma patient samples across five omic layers: CpG methylome, transcriptome, proteome, phosphoproteome, and metabolome, paired with associated clinical metadata. Data integration revealed intertumoral heterogeneity of lipid metabolism across proteomic subtypes. Notably, while the MYC-FASN-SCD axis drives lipid biosynthesis, pathway inhibition elicits a compensatory escape mechanism in vivo through exogenous fatty acid uptake. Unexpectedly, we demonstrated that MYC triggers lipid storage, creating a unique dependency on lipid droplet-mitochondria communications to sustain tumor maintenance in vivo. Together, this comprehensive analysis reveals a targetable vulnerability downstream of MYC that constitutes a promising therapeutic approach to treat currently untreatable medulloblastoma subtypes.

Table S15: Relative quantification by lipid class

Metabolic steatotic liver disease (MASLD) can progress to hepatocellular carcinoma (HCC). 25

Table S12: gene signatures used to analyze RNA-seq data from patients with AML before/after GILT

Table S2: EnrichR analysis of FLT3 gene signatures

Supplementary materials and methods

Table S4: GSEA results in FLT3-ITD PDX AML cells ex vivo-treated with QUIZ or Veh.

Table S8: Multimodal analysis of shCEBPA versus shCTL MOLM14 cell line

Table S14: PUFA/MUFA ratios in phospholipids

BACKGROUND:Immune cell metabolism and metabolic end products influence the nature and magnitude of immune responses. Various autoimmune and inflammatory diseases are associated with dysregulated cellular metabolism. Intravenous immunoglobulin (IVIG), a therapeutic pooled normal IgG, is extensively used for the immunotherapy of a wide range of autoimmune and inflammatory diseases. Although several cellular and molecular mechanisms of action of IVIG have been reported, the role of IVIG in modulating the immune cell metabolism remains unknown. OBJECTIVE:We sought to investigate the influence of IVIG on the metabolic events of human immune cells. METHODS:We performed metabolic flux analyses on inflammatory mediators-stimulated human PBMCs. Furthermore, intracellular metabolites were extracted from activated PBMCs and subjected to liquid chromatography coupled to high-resolution mass spectrometry. Untargeted global metabolic profiling of PBMCs was performed to determine the metabolic landscape of immune cells and metabolic reprogramming by IVIG. Untargeted lipidomics was used for the mechanistic studies on IVIG-induced lipogenesis. RESULTS:IVIG and its Fc and F(ab')2 fragments regulate the Warburg effect in activated PBMCs depending on the glucose availability. Untargeted global metabolic profiling revealed that IVIG alters the overall metabolic landscape of inflammatory mediators-stimulated PBMCs, blocks prenylation of amino acid cysteine, and promotes lipogenesis of well-known anti-inflammatory lipids such as diacylglycerol and triacylglycerol by shuttling acetyl coenzyme A away from the mevalonate pathway. Mechanistically, IVIG-induced lipogenesis was mediated via F(ab')2 fragments and was dependent on the sialylated glycans of IgG. CONCLUSIONS:Our data indicate that IVIG targets immune cell metabolism and also highlight a novel mechanism of action of IVIG in the context of immunotherapy of autoimmune and inflammatory diseases.

Table S7: Transcriptomics analysis of shCEBPA#2 versus shCTL

Table S13: lipidomic results by lipid species in dox-inducible and constitutives shRNAs and siRNA CTL/CEBPA, empty vector/CEBPA-OE and vehicle/QUIZ conditions in MOLM-14 cells

Supplemental figures S1-S22: Supplementary Figure S1 shows lipid metabolism dependency in FLT3-mutant AML, Supplementary Figure S2 shows anti-leukemic activity of GILT across a panel of AML PDX, Supplementary Figure S3 shows mechanisms of post-translational regulation of C/EBPα expression, Supplementary Figure S4 shows inhibition of C/EBPα phosphorylation and protein expression by FLT3i, Supplementary Figure S5 shows the implication of Ser-21 phosphorylation in post-translational regulation of C/EBPα expression, Supplementary Figure S6 shows that FLT3-ITD regulates the expression of C/EBPα and of proteins related to lipid biosynthesis in AML cell lines, Supplementary Figure S7 shows that FLT3-ITD regulates the expression of genes related to lipid biosynthesis in AML cell lines, Supplementary Figure S8 shows that C/EBPα directly regulates the transcription of lipid biosynthesis genes in AML, Supplementary Figure S9 shows the correlation between CEBPA mRNA expression and FLT3-ITD mutations, Supplementary Figure S10 shows scRNA-seq analysis from PDXTUH84, Supplementary Figure S11 shows scRNA-seq analysis from PDXTUH110, Supplementary Figure S12 shows the evolution of CEBPA_UP and FLT3-ITD_UP signatures in patients with AML treated with GILT, Supplementary Figure S13 shows that C/EBPα regulates rate-limiting lipid biosynthetic enzymes downstream of FLT3-ITD, Supplementary Figure S14 shows that C/EBPα controls lipid amount in FLT3-ITD cell lines, Supplementary Figure S15 shows that FLT3 inhibitors induce a lipid switch increasing neutral lipids dependent on C/EBPα, Supplementary Figure S16 shows that FLT3 inhibitor inhibits fatty acid synthesis fueled by glucose and glutamine, Supplementary Figure S17 shows that FLT3 inhibition decreases monounsaturated fatty acid dependent on C/EBPα, Supplementary Figure S18 shows that FLT3 inhibition increases PUFA/MUFA ratio dependent on C/EBPα, Supplementary Figure S19 shows that ferroptotic cell death induced by FLT3i is mediated by inhibition of SCD-dependent mono-unsaturated FA synthesis, Supplementary Figure S20 shows that FLT3 inhibitors unmask a vulnerability of FLT3-mutant leukemic cells to ferroptosis, Supplementary Figure S21 shows that lipid redox stress induction by GPX4 inhibition primed FLT3i activity in FLT3-ITD AML cells and Supplementary Figure S22 shows combined treatment with GILT and APR-246 in preclinical AML models in vivo.

Table S3: Clinical characteristics of samples from patients with AML used in PDX assays

Table S6: Proteomic analysis of FLT3-ITD AML cell lines treated with vehicle or QUIZ

Table S1: Transcriptome analysis of FLT3-ITD cell lines after FLT3 inhbition