Lactylation-related LGALS1 promotes immunosuppressive tumor microenvironment and venetoclax resistance in Acute Myeloid Leukemia

Abstract Acute myeloid leukemia (AML) is a highly heterogeneous and immunosuppressive hematologic malignancy. Novel therapies like venetoclax (a BCL-2 inhibitor) combined with hypomethylating agents or low-dose cytarabine have improving overall survival, particularly for older patients, but chemoresistance remains a major clinical challenge. Lactylation is a novel post-translational modification related to tumor progression, immunosuppression, and chemoresistance across various cancers. Lactate, once considered a metabolic waste, is recognized as a key substrate for lactylation. Galectin-1 (LGALS1), a β-galactoside-binding lectin, is often overexpressed in tumors and leads to immune escape, tumor metastasis, and chemoresistance. The role and mechanisms of LGALS1 in AML, particularly concerning lactylation and venetoclax resistance, remain unclear. The aim of this study is to identify the key role of lactylation-related diagnostic/prognostic biomarker LGALS1 in AML immunosuppression and its potential mechanisms leading to venetoclax resistance. Bone marrow samples from AML patients and healthy donors were analyzed for RNA expression. Public whole genome sequencing datasets (TCGA-LAML, GSE37307, GSE71014, GSE116256) were utilized to identify and validate lactylation-related genes (LRGs). Consensus clustering of LRGs clustered AML patients into two clusters. Weighted Gene Co-expression Network Analysis (WGCNA) identified key gene modules related to clinical traits. Five machine-learning algorithms (Random Forest, GBM, XGBoost, SVM-RFE, LASSO) intersected to screen the hub gene LGALS1. Validation included differential expression analysis, survival analysis, diagnostic/prognostic efficacy (ROC/timeROC curves), gene set enrichment (GSEA), and immune infiltration analysis (CIBERSORT, ssGSEA). Analyzing the role of LGALS1 at the cellular level using single-cell RNA sequencing (scRNA-seq) analysis, pseudotime analysis and cell-cell communication analysis. In vitro experiment on AML cell lines (MOLM-13, MV-4-11) involved LGALS1 expression (RT-qPCR and Western blot), knockdown (siRNA transfection), proliferation (CCK-8), venetoclax sensitivity (IC₅₀) and apoptosis (flow cytometry). Multi-omics and machine learning analysis identified LGALS1 as a critical lactylation-related hub gene in AML. LGALS1 was significantly overexpressed in AML patients compared to healthy donors across internal and external datasets, strongly correlated with poor overall survival, and exhibited significant clinical value as both a diagnostic/prognostic biomarker in AML. Immune infiltration analysis revealed that high LGALS1 expression was associated with an immunosuppressive tumor microenvironment (TME), characterized by increased infiltration of M2 macrophages, monocytes, MDSCs, and regulatory T cells (Tregs), alongside decreased anti-tumor CD8⁺T cells and NK cells. ScRNA-seq analysis identified classical dendritic cells (cDCs) as the primary cell type expressing LGALS1 within the AML bone marrow. Pseudotime analysis showed progressing enrichment of LGALS1⁺cDCs during AML progression. Cell-cell communication analysis indicated that LGALS1⁺cDCs exhibited enhanced interaction strength and number with other cell types than LGALS1-cDCs, and as a key role in pathways like ADGRE5 and CD45 promoting an immunosuppressive TME. In vitro experiments demonstrated that knocking down LGALS1 in AML cell lines (MOLM-13, MV-4-11) significantly inhibited cell proliferation and increased sensitivity to venetoclax, reducing IC₅₀ values by approximately 6-fold in MOLM-13 cells and 4-fold in MV-4-11 cells. Mechanistically, LGALS1 knockdown specifically downregulated the anti-apoptotic protein MCL-1, leading to significantly increased apoptosis on venetoclax treatment. We have determined that LGALS1 is a key driver of immunosuppressive TME and venetoclax resistance in AML, highlighting its potential as a diagnostic/prognostic biomarker and therapeutic target. The LGALS1-MCL-1 axis provides a novel strategy to overcome venetoclax resistance.