Abstract
Background: Despite comparable survival benefit has been obtained, the drug resistance remarkably reduced lenvatinib clinical efficacy. Here, we aimed to identify the potential mechanism by which cysteine and glycine-rich protein 2 (CSRP2) regulates the development of hepatocellular carcinoma (HCC) and participates in the resistance to lenvatinib. Methods: We harnessed RNA sequencing, multiplex immunofluorescence staining, and hydrodynamic tail vein (HTV) injection HCC model to systematically explore the function of CSRP2 in HCC progression. To precisely delineate how immunosuppressive macrophages, influenced by CSRP2-regulated C-C motif chemokine ligand 28 (CCL28) signaling, respond to lenvatinib-induced cytotoxicity, we established an in vitro co-culture system and conducted functional cytotoxicity assays. Results: Using RNA sequencing, multiplex immunofluorescence staining and HTV injection HCC mouse model, we identified CSRP2 as one of the most significantly upregulated genes in HCC tissues. CSRP2 overexpression drives anti-lenvatinib resistance by inducing high levels of tumor-associated macrophages (TAMs) infiltration and reshaping an immunosuppressive microenvironment. Then flow cytometry, mass spectrometry and chromatin immunoprecipitation were conducted to clarify the underlying mechanism of CSRP2. We showed CSRP2 promotes phosphorylation of activating transcription factor 2 (ATF2) at Thr69/71, leading to the transcriptional activation of CCL28 expression. HCC-derived CCL28 recruits TAMs to drive immunosuppression and anti-lenvatinib tolerance. BI6901, a potent and selective CCR10 antagonist, blocked TAMs recruitment and enhanced T-cell activation. Combining CCR10 inhibition improved the therapeutic benefit of anti-lenvatinib in HCC. Conclusions: These results illustrate that CSRP2 regulates the tumor microenvironment to promote HCC growth and drive lenvatinib tolerance via the CSRP2/ATF2/CCL28 axis. Targeting this pathway could synergize with lenvatinib to treat HCC more effectively.