LKB1 dictates sensitivity to immunotherapy through Skp2-mediated ubiquitination of immune checkpoint proteins in HCC with python analysis
نویسندگان
1 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
2 Department of Pathology, Division of Experimental Pathology, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA
3 School of Pharmacy ,Tehran University of Medical Sciences,Tehran, Iran
4 School of Pharmacy ,Tehran University of Medical Sciences,Tehran, Iran
5 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
6 School of Pharmacy ,Tehran University of Medical Sciences,Tehran, Iran
7 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
8 School of Pharmacy ,Tehran University of Medical Sciences,Tehran, Iran
9 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
10 Immunology Research Center,Tabriz University of Medical Sciences,Tabriz,Iran
11 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
12 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
13 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
14 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
15 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
16 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
17 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
18 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
19 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
20 School of Medicine,Tehran University of Medical Sciences,Tehran, Iran
doi
10.22038/ijbms.2026.91840.19829چکیده
Objective(s): Hepatocellular carcinoma (HCC) is a highly aggressive malignancy with limited treatment options, particularly in advanced stages. Immune checkpoint inhibitors (ICIs) targeting PD-1, PD-L1, and CTLA-4 have shown promise in cancer immunotherapy, but response rates in HCC remain variable. Materials and Methods: Hep3B and HepG2 HCC cells were cultured and genetically manipulated to overexpress or deplete LKB1. Western blotting, real-time PCR, and immunofluorescence were used to assess PD-L1 expression at the protein and mRNA levels. The role of Skp2 in PD-L1 regulation was evaluated through shRNA-mediated knockdown and overexpression. Additionally, kinase-dead LKB1 mutants were expressed to determine the importance of LKB1 kinase activity in PD-L1 stability. ImageJ software and Python-based computational tools were employed for quantitative analysis of immunofluorescence and Western blot data.Results: LKB1 overexpression up-regulated PD-L1 protein levels in HCC cells, while its depletion reduced PD-L1 expression, indicating a post-translational regulatory mechanism. Although Skp2 expression remained unchanged upon LKB1 modulation, Skp2 overexpression in LKB1-deficient cells increased PD-L1 levels, suggesting a context-dependent role for Skp2 in PD-L1 stability. Furthermore, wild-type LKB1, but not the kinase-dead mutant, restored PD-L1 expression, highlighting the essential role of LKB1 kinase activity in PD-L1 regulation.Conclusion: This study identifies LKB1 as a critical regulator of PD-L1 stability in HCC, with implications for tumor immune evasion and immunotherapy response. While Skp2 appears to influence PD-L1 stability in specific contexts, LKB1’s kinase activity is essential for PD-L1 regulation.
