2002 – PERTURBATION OF LEUKEMIC STEM CELL STATES REVEALS A TGFβ-DRIVEN NICHE REMODELING PROGRAM IN AML

Abstract

The bone marrow (BM) niche regulates hematopoietic stem cell function and contributes to therapy resistance and relapse in acute myeloid leukemia (AML). However, how defined leukemic stem cell (LSC) states remodel their direct niche remains incompletely understood.

Here, we combined genetically controlled in vivo perturbation models with scRNA-seq, 3D imaging, and functional assays to dissect LSC-driven niche remodeling. We used NPM1/FLT3-ITD/DNMT3A AML xenografts with wild-type or knockout of LSC regulator HLF to model LSChigh and LSClow conditions. Human leukemic and mouse stromal compartments were profiled at single-cell resolution, followed by cell–cell communication and gene regulatory network analyses.

AML induced niche remodeling, with expansion of Cxcl12⁺ adipogenic progenitors and reduction of osteo-lineage populations, particularly in LSChigh AML. This was accompanied by expansion of Fmod⁺ and Cd34⁺ fibroblasts.

Single-cell analysis revealed heterogeneity within AML, including quiescent, stress-adapted, and proliferative LSC states enriched in LSChigh AML, whereas LSClow showed increased myeloid and erythroid blast states. Distinct LSC subsets expressed TGFB1, IL1B, and other niche-modulating factors. Ligand prioritization identified TGFβ as a dominant LSC-derived signal. Fmod⁺ fibroblasts showed elevated TGFβ activity, and Creb3l1 was associated with fibroblast expansion, defining a pro-fibrotic state. In BM MSC–AML cocultures, LSChigh AML recapitulated this in vivo phenotype by inducing an Fmod⁺ fibroblast-like program with increased TGFβ activity and Creb3l1 regulon upregulation. Perturbations showed that TGFβ signaling and CREB3L1 modulation alter stromal states and leukemic cell retention.

Together, this work links LSC heterogeneity to microenvironmental reprogramming and identifies an LSC–TGFβ–CREB3L1 axis as a potential therapeutic target in AML.

The bone marrow (BM) niche regulates hematopoietic stem cell function and contributes to therapy resistance and relapse in acute myeloid leukemia (AML). However, how defined leukemic stem cell (LSC) states remodel their direct niche remains incompletely understood.

Here, we combined genetically controlled in vivo perturbation models with scRNA-seq, 3D imaging, and functional assays to dissect LSC-driven niche remodeling. We used NPM1/FLT3-ITD/DNMT3A AML xenografts with wild-type or knockout of LSC regulator HLF to model LSChigh and LSClow conditions. Human leukemic and mouse stromal compartments were profiled at single-cell resolution, followed by cell–cell communication and gene regulatory network analyses.

AML induced niche remodeling, with expansion of Cxcl12⁺ adipogenic progenitors and reduction of osteo-lineage populations, particularly in LSChigh AML. This was accompanied by expansion of Fmod⁺ and Cd34⁺ fibroblasts.

Single-cell analysis revealed heterogeneity within AML, including quiescent, stress-adapted, and proliferative LSC states enriched in LSChigh AML, whereas LSClow showed increased myeloid and erythroid blast states. Distinct LSC subsets expressed TGFB1, IL1B, and other niche-modulating factors. Ligand prioritization identified TGFβ as a dominant LSC-derived signal. Fmod⁺ fibroblasts showed elevated TGFβ activity, and Creb3l1 was associated with fibroblast expansion, defining a pro-fibrotic state. In BM MSC–AML cocultures, LSChigh AML recapitulated this in vivo phenotype by inducing an Fmod⁺ fibroblast-like program with increased TGFβ activity and Creb3l1 regulon upregulation. Perturbations showed that TGFβ signaling and CREB3L1 modulation alter stromal states and leukemic cell retention.

Together, this work links LSC heterogeneity to microenvironmental reprogramming and identifies an LSC–TGFβ–CREB3L1 axis as a potential therapeutic target in AML.