Mixed-lineage leukemia cells undergo unique adaptations in the CNS niche.

KMT2A-rearranged (KMT2A-r) infant leukemia can present as a lymphoid, myeloid, or mixed-lineage leukemia and frequently involves the central nervous system (CNS); yet, the impact of this lineage diversity and plasticity on CNS involvement remains poorly understood. Using a fully murine immunocompetent model of KMT2A-AFF1+ mixed-lineage infant leukemia, we investigated how the CNS niche influences the phenotype and function of leukemia propagating cells (LPCs). Previously defined bone marrow (BM)-derived LPCs were transplanted and shown to engraft the CNS, although not equally; lineage-negative cKit+ common lymphoid progenitor cells were consistently underrepresented in the niche. Transplants of CNS-derived LPCs, modelling relapse, demonstrated reduced systemic repopulation capacity, with many recipients exhibiting stable long-term engraftment without developing overt leukemia, a phenomenon not observed in BM-derived transplants. Transcriptomic profiling of matched CNS- and BM-derived LPCs revealed enrichment of pathways involved in hypoxia, lipid and cholesterol homeostasis, and inflammatory signaling in the CNS. Notably, LPC subsets that successfully adapted to the CNS niche upregulated lipid and fatty acid metabolic programs. CNS-derived LPCs showed increased expression of genes involved in T cell immune modulation, suggesting a skew to a more immunosuppressive environment. These findings indicate that the CNS niche imposes selective pressures that cause lasting metabolic and functional reprogramming of leukemic cells, impairing their ability to reestablish systemic disease and potentially affecting immune cell interactions. Furthermore, these findings may be more generally relevant to primary mixed-lineage infant leukemia and, increasingly important, lineage-switched infant leukemia.