Allergin-1 marks leukemic stem cells and enables immune evasion in AML: A target for therapeutic antibody development

Abstract

Abstract Acute myeloid leukemia (AML) often achieves remission after therapy; however, relapse – driven by chemotherapy-resistant leukemic stem cells (LSCs) – remains a major cause of mortality. LSCs closely resemble hematopoietic stem cells (HSCs) in immunophenotype, making selective targeting difficult and highlighting an unmet need in AML therapy. To address this, we compared gene and protein expression profiles of LSCs and HSCs using public datasets and primary human AML samples across diverse subtypes. MILR1 (encoding Allergin-1) was consistently upregulated in LSC-enriched populations compared to non-LSC populations and HSPCs. In patient-derived xenograft (PDX) models, Allergin-1⁺ AML cells initiated leukemia, whereas Allergin-1⁻ cells did not. Knockdown of Allergin-1 significantly impaired leukemogenesis in PDX mice, establishing Allergin-1 as a functional LSC marker. Using murine models of MLL-AF9-driven AML, Milr1 knockout (KO) delayed leukemia progression and prolonged survival without affecting normal hematopoiesis. Transcriptomic analysis revealed two key effects of Milr1 loss: (1) reduced SHP-1/MAPK signaling, impairing AML stemness, and (2) enhanced natural killer (NK) cell-mediated immune activation. NK cells from Milr1-KO AML mice exhibited increased activation markers and cytotoxicity in co-culture assays, whereas T cell responses remained unchanged. This likely reflects NK cells' reliance on germline-encoded ligands, unlike T cells' dependence on neoantigen presentation. Surface proteomic analysis showed Milr1-KO AML cells exhibited decreased Lgals9 and Cdh2 and increased expression of MHC genes, Stamf7, and Fcgr1, indicating altered immunogenicity likely driven by SHP-1 suppression-induced cellular stress. To therapeutically target Allergin-1, we generated a panel of rabbit-derived monoclonal antibodies (mAbs). From 50 candidates, we identified a high-affinity mAb (Allergin-1 hAb) that antagonizes Allergin-1 activation. Molecular docking predicted that Allergin-1 hAb engages critical residues within the D1 domain of Allergin-1, blocking ligand interaction. In humanized PDX models, Allergin-1 hAb treatment significantly reduced AML burden compared to control IgG, supporting its therapeutic potential. Conclusion: Allergin-1 is a leukemia-specific functional marker that promotes stemness and immune evasion in AML. Targeting Allergin-1 with monoclonal antibodies represents a promising therapeutic strategy for AML treatment.