Metabolic Reprogramming in Myelodysplastic Syndromes

Myelodysplastic syndromes (MDS), lie at the interface of normal differentiation and leukemic transformation, being characterised by polymorphic presentation and elusive pathophysiology. Failure to establish a pathogenic relationship with specific genetic aberrations, along with general paucity in innovative and effective treatment strategies, led us to seek the possible role of a dysregulated metabolism in the disease pathobiology.

Plasma and CD34+ progenitors isolated from 19 previously untreated MDS patients (WHO 2022 LB#9, IB1#8, IB2#2 - CD34+ pellet purity 97-99%) and 10 age-sex matched controls (CTRLs) were subjected to untargeted and targeted mass spectrometry-based metabolomics. Data analysis took place using Proteowizard and MetaboAnalyst 5.0 coupled to our biostatistics pipeline (R language). Statistical analysis involved one-factor exploration, data filtering, normalisation, and univariate assessments. For chemometrics, we performed PCA and OPLS-DA for two-group comparisons and analogous methods like PLS-DA for multiple-group evaluations. Clustering techniques were employed along with classification methods. Pathway-related outcomes were obtained after functional analysis. Visual analytics included scatter plots, heatmaps, and pathway exploration, all contributing to a thorough examination of the mass spectrometry data.

A commercially available myeloid NGS panel showed no correlation between metabolomic status and CD34+ mutational burden of both MDS and CTRL. Mitochondrial metabolism was maintained in MDS CD34+ cells. Tricarboxylic acid (TCA) cycle integrity was established by calculating the absolute values of related metabolites with no statistically significant difference from the CD34+ in the CTRL group. Mitochondrial metabolic reliance was also confirmed by extreme upregulation of beta-alanine and histidine metabolism which was attributed to optimisation of organellar function, as well as by calculated NADH/NAD+ and FADH2/FAD ratios. Based on statistical importance and enrichment factor, MDS CD34+ progenitors were found to rely on glutamine (GLN) and secondarily on glucose (GLUC) as primary carbon sources. Carbon cycling through normal anaplerosis and reductive carboxylation led to increased citrate production, that was reflected upon the uniformly increased citrate/a Ketoglutarate ratio. Increased citrate usage occurred via biosynthetic fatty acid reactions, highly featured in MDS progenitors. Complete GLN mitochondrial oxidation was also documented, producing energy via coupling of the GLN-fuelled TCA to the electron transport chain. Downstream nitrogen detoxification with urea cycle and aspartate-aspartic acid metabolism was as uniformly increased in MDS. Massive upregulation of the L-2-hydroxyglutarate (L-2HG) enantiomer in all MDS CD34+ progenitors, was also documented, despite the absence of IDH 1,2 mutations.

GLUC was suggested to feed several essential anabolic and redox pathways, namely the PPP or the serine-threonine-glycine metabolism and the one-carbon cycle, all of which were highly enriched in our samples. Absolute value of LAC did not significantly differ between controls and MDS, neither in CD34+ cells nor in BM plasma, although a trend for increased values was observed in IB MDS. Discrepancies between CD34+ progenitors from LBs and IBs did exist regarding mostly their redox state through the GSH/GSSG and NADP/NADPH ratios, showing a decreased potential to deal with oxidative stress in the lower risk samples; the finding may as well explain the increased apoptotic yield described in LB MDS.

Overall, a mitochondrially orchestrated metabolism with primary GLN catabolic utilisation and non fermenting cytoplasmic GLUC driven biosynthesis, with massive production of L2HG describes the bioenergetic state of CD34+ MDS progenitors. The resulting continuous epigenetic and oxidative stress, via GLN and fatty acid metabolic reprograming explains the apoptotic features of LB MDS, while driving the malignant nature of IB MDS.

No relevant conflicts of interest to declare.