CD44v6 promotes anti-apoptotic phenotypes in AML via an SPP1-mediated autocrine activation of the MAPK-dusp pathway

Abstract Background:Acute myeloid leukemia (AML) is a highly heterogeneous hematological malignancy originating from the bone marrow. Complex genetic alterations promote clonal diversity among leukemic cells, leading to poor prognosis and high relapse rates. CD44, a widely expressed cell adhesion molecule, plays key roles in tumor biology. Its splice variant CD44v6 is specifically expressed on AML cells but minimally detected in normal hematopoietic stem cells, rendering it a promising immunotherapeutic target. However, the biological functions and regulatory mechanisms of CD44v6 in AML remain poorly understood. Methods: We analyzed clinical AML samples and public datasets (TCGA-LAML) to evaluate the association between CD44v6 expression and patient outcomes. AML cell lines (SKM-1, THP1, MOLM13) with stable overexpression of CD44v6 were constructed to examine its impact on cell proliferation, colony formation, and resistance to standard chemotherapeutic agents (Ara-C and Venetoclax). Bulk RNA sequencing was performed to identify transcriptomic alterations induced by CD44v6 overexpression, and candidate downstream regulatory pathways and ligands were explored via functional analysis and expression validation. Results: CD44v6 expression was significantly elevated in high-risk genetic AML patients and correlated with poor overall survival. Overexpression of CD44v6 enhanced leukemic cell proliferation, clonogenicity, and resistance to Ara-C and Venetoclax-induced apoptosis. Transcriptomic profiling revealed marked activation of the MAPK signaling pathway and upregulation of DUSP1 and DUSP6. Notably, anti-apoptotic proteins MCL-1 and BCL-XL were upregulated, while BCL-2 was downregulated, indicating a BCL-2-independent survival mechanism distinct from that induced by CD44s. SPP1 was identified as a key CD44v6-associated ligand, whose expression was elevated in CD44v6-overexpressing cells. The secreted osteopontin (SPP1) bound to surface CD44v6 in an autocrine loop, further enhancing DUSP6 and MCL-1 expression while suppressing BCL-2. These effects were partially reversed upon CD44v6 monoclonal antibody blockade. Conclusions:Our findings demonstrate that CD44v6 promotes AML progression by driving a chemoresistance-associated transcriptional program through a novel CD44v6-SPP1 autocrine loop. This signaling axis activates MAPK/DUSP pathways and shifts anti-apoptotic dependence toward MCL-1 and BCL-XL.