Metabolic Rewiring of AML-Experienced Hematopoietic Stem and Progenitor Cells

Relapse is the leading cause of mortality in acute myeloid leukemia (AML). Inflammation is linked to disease prognosis in AML, but its role in remission and relapse are not well understood. Inflammation is also a known regulator of hematopoietic stem and progenitor cells (HSPCs), and recent studies identified an unexpected role in their long term innate immune reprogramming.

We recently reported in a murine model of AML that residual healthy HSPCs contribute to the inflammatory bone marrow (BM) microenvironment. In additional interim pilot studies, we adoptively transferred “AML-experienced” HSPCs (HSPC^AML) to naïve recipients, followed 16 weeks later by a lipopolysaccharide (LPS) challenge. Bulk RNA-Seq from these HSPC^AML revealed significant transcriptional changes compared to a control cohort. To further test whether HSPC^AML develop a long-term innate immune memory of sterile inflammation in the AML BM niche, we turned to a clinically relevant inducible model of KMT2A AML (MLL-AF9). We generated chimera by co-transplanting BM from inducible (i)MLL-AF9 transgenic mice (CD45.1; rtTA^+/+ hMLL-AF9-IRES-NGFR^+/+) and WT mice (CD45.1/2) into WT recipients (CD45.2). This provides a dynamic fully tractable model where leukemia exposure of healthy HSPC and remission can be dynamically controlled in vivo via doxycycline (DOX).

Recent studies in experimental infection have defined altered immunometabolism as a key component of inflammatory HSPC immune reprogramming. We induced AML for 5 weeks (on DOX), followed by 4 weeks off DOX, and harvested FACS-purified HSPC^AML (CD45.1/2⁺ NGFR^- Lin^- cKit⁺ Sca1⁺) to perform a real-time metabolic function assay (Seahorse). Results showed elevated basal extracellular acidification rate (ECAR) indicative of heightened basal glycolysis in HSPC^AML. Consistent with this observation, RNA-Seq analysis of HSPC^AML from identically prepared cohorts revealed 3974 differentially expressed genes (DEGs), with gene set enrichment for glycolysis pathways. Models of trained immunity (TI) indicate that memory generation is epigenetically imprinted, and ATAC-Seq analysis of HSPC^AML revealed a significant increase in differentially accessible regions in HSPC^AML. Through GSEA analysis of differentially accessible promoters and enhancer regions, we observed enrichment for inflammation and metabolic related pathways in HSPC^AML. Lastly, by subjecting chimeric animals off DOX to LPS challenge, we found enrichment of tumor necrosis factor (TNF) associated gene sets in HSPC^AML via RNA Seq analysis, demonstrating a heighten inflammatory response consistent with trained cells.

Altogether, our data suggest that sterile inflammation in the AML BM niche trains healthy HSPC through a process of immune reprogramming, consistent with a model whereby leukemia may durably alter hematopoietic function.