Mechanosensitive ion channel-related genes in hepatocellular carcinoma: Unraveling prognostic genes and their roles in drug resistance and immune modulation

Abstract

Background and aims: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality worldwide, and its etiology involves a complex interplay of genetic and environmental factors. Despite advancements in our understanding of HCC biology and the development of novel therapeutic strategies, the molecular mechanisms underlying its onset, progression, and resistance to therapy remain largely vague. This study aimed to investigate the role of mechanosensitive ion channel-related genes (MICRGs) in HCC, focusing on their potential as prognostic biomarkers and their involvement in immune modulation and drug resistance.

Methods: A comprehensive analysis was conducted using The Cancer Genome Atlas database to identify MICRGs that are upregulated in HCC. Gene expression profiling, bioinformatics tools, and functional experiments were employed to elucidate the role of these channels. In addition, protein-protein interaction (PPI) network analyses and enrichment analyses were performed to explore the biological significance of these genes. An immune cell infiltration analysis was also conducted to understand MICRG-related immune landscape. Single-cell RNA sequencing (scRNA-seq) data were utilized to identify MICRGs in different cell types within the HCC tissue. Deep-learning neural network analysis across patient cohorts was conducted to identify genes associated with sorafenib resistance. Knockdown experiments, cell viability assays, and apoptosis assays on HCC cell lines were performed to examine the role of Piezo-type mechanosensitive ion channel component 1 (PIEZO1) in sorafenib resistance.

Results: The analysis identified a subset of MICRGs, including PIEZO1, that were significantly upregulated in HCC and associated with poor prognosis. The PPI network analysis revealed complex interactions among these genes. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses proposed the involvement of these genes in calcium signaling pathways. Immune cell infiltration analysis demonstrated distinct associations between MICRGs and various immune subpopulations, highlighting their potential roles in immune modulation. scRNA-seq data indicated the upregulation of MICRGs in various cell types in HCC tissues, particularly in endothelial cells and tumor-associated macrophages. Deep-learning neural network analysis across different patient cohorts identified PIEZO1 as a crucial regulator of sorafenib resistance in HCC, which was further validated by functional assays on HCC cell lines.

Conclusions: This study provides evidence that MICRGs, particularly PIEZO1, take on crucial roles in HCC progression and drug resistance. The upregulation of PIEZO1 in HCC cells is associated with poor prognosis and resistance to sorafenib. These findings indicate that PIEZO1 could serve as a potential therapeutic target for overcoming drug resistance and a prognostic biomarker in HCC. Future studies should focus on validating these findings in larger patient cohorts and exploring the functional implications of targeting PIEZO1 in preclinical models.

Keywords

Hepatocellular carcinoma (HCC) / Mechanosensitive ion channel / Piezo-type mechanosensitive ion channel component 1 (PIEZO1) / Drug resistance / Prognosis / Immune infiltration

References

[1] Lee TK, Guan XY, Ma S. Cancer stem cells in hepatocellular carcinoma - from origin to clinical implications. Nat Rev Gastroenterol Hepatol. 2022;19:26-44. https://doi.org/10.1038/s41575-021-00508-3.

[2] Shen P, Jia Y, Zhou W, et al. A biomimetic liver cancer on-a-chip reveals a critical role of LIPOCALIN-2 in promoting hepatocellular carcinoma progres-sion. Acta Pharm Sin B. 2023;13:4621-4637. https://doi.org/10.1016/j.apsb.2023.04.010.

[3] Chen LT, Martinelli E, Cheng AL, et al. Pan-Asian adapted ESMO Clinical Practice Guidelines for the management of patients with intermediate and advanced/relapsed hepatocellular carcinoma: a TOS-ESMO initiative endorsed by CSCO, ISMPO, JSMO, KSMO, MOS and SSO. Ann Oncol. 2020;31:334-351. https://doi.org/10.1016/j.annonc.2019.12.001.

[4] Tang W, Chen Z, Zhang W, et al. The mechanisms of sorafenib resistance in hepatocellular carcinoma: theoretical basis and therapeutic aspects. Signal Transduct Target Ther. 2020;5:87. https://doi.org/10.1038/s41392-020-0187-x.

[5] Kurebayashi Y, Ojima H, Tsujikawa H, et al. Landscape of immune microenvi-ronment in hepatocellular carcinoma and its additional impact on histological and molecular classification. Hepatology. 2018;68:1025-1041. https://doi.org/10.1002/hep.29904.

[6] Zhou J, Valentini E, Boutros M. Microenvironmental innate immune signaling and cell mechanical responses promote tumor growth. Dev Cell. 2021;56: 1884e 1899 (e5). https://doi.org/10.1016/j.devcel.2021.06.007.

[7] Roy AM, Iyer R, Chakraborty S. The extracellular matrix in hepatocellular car-cinoma: mechanisms and therapeutic vulnerability. Cell Rep Med. 2023;4: 101170. https://doi.org/10.1016/j.xcrm.2023.101170.

[8] Schrader J, Gordon-Walker TT, Aucott RL, et al. Matrix stiffness modulates proliferation, chemotherapeutic response, and dormancy in hepatocellular carcinoma cells. Hepatology. 2011;53:1192-1205. https://doi.org/10.1002/hep.24108.

[9] Karska J, Kowalski S, Saczko J, Moisescu MG, Kulbacka J. Mechanosensitive ion channels and their role in cancer cells. Membranes (Basel). 2023;13:167. https://doi.org/10.3390/membranes13020167.

[10] Cox CD, Poole K, Martinac B. Re-evaluating TRP channel mechanosensitivity. Trends Biochem Sci. 2024;49:693-702. https://doi.org/10.1016/j.tibs.2024.05.004.

[11] Jin P, Jan LY, Jan YN. Mechanosensitive ion channels: structural features rele-vant to mechanotransduction mechanisms. Annu Rev Neurosci. 2020;43: 207-229. https://doi.org/10.1146/annurev-neuro-070918-050509.

[12] Liu S, Xu X, Fang Z, et al. Piezo 1 impairs hepatocellular tumor growth via deregulation of the MAPK-mediated YAP signaling pathway. Cell Calcium. 2021;95:102367. https://doi.org/10.1016/j.ceca.2021.102367.

[13] Li YM, Xu C, Sun B, Zhong FJ, Cao M, Yang LY. Piezo 1 promoted hepatocellular carcinoma progression and EMT through activating TGF-b signaling by recruiting Rab5c. Cancer Cell Int. 2022;22:162. https://doi.org/10.1186/s12935-022-02574-2.

[14] Tan S, Chao R. An exploration of osteosarcoma metastasis diagnostic markers based on tumor-associated neutrophils. Discov Med. 2023;35:300-311. https://doi.org/10.24976/Discov.Med.202335176.31.

[15] Zhao X, Zhang L, Wang J, et al. Identification of key biomarkers and immune infiltration in systemic lupus erythematosus by integrated bioinformatics analysis. J Transl Med. 2021;19:35. https://doi.org/10.1186/s12967-020-02698-x.

[16] Kanehisa M, Goto S. KEGG: kyoto encyclopedia of genes and genomes. Nucleic Acids Res. 2000;28:27-30. https://doi.org/10.1093/nar/28.1.27.

[17] Wang J, Chen X, Wu D, et al. Single-cell and machine learning approaches uncover intrinsic immune-evasion genes in the prognosis of hepatocellular carcinoma. Liver Res. 2024;8:282-294. https://doi.org/10.1016/j.livres.2024.11.001.

[18] Kalbasi A, Ribas A. Tumour-intrinsic resistance to immune checkpoint blockade. Nat Rev Immunol. 2020;20:25-39. https://doi.org/10.1038/s41577-019-0218-4.

[19] Li H, Guo L, Cai Z. TCN1 is a potential prognostic biomarker and correlates with immune infiltrates in lung adenocarcinoma. World J Surg Oncol. 2022;20:83. https://doi.org/10.1186/s12957-022-02556-8.

[20] Danaher P, Warren S, Lu R, et al. Pan-cancer adaptive immune resistance as defined by the tumor inflammation signature (TIS): results from The Cancer Genome Atlas (TCGA). J Immunother Cancer. 2018;6:63. https://doi.org/10.1186/s40425-018-0367-1.

[21] Zhou X, Du J, Liu C, et al. A pan-cancer analysis of CD161, a potential new immune checkpoint. Front Immunol. 2021;12:688215. https://doi.org/10.3389/fimmu.2021.688215.

[22] Clemente MRC, Felix N, Navalha DDP, et al. Long-term impact of home-based monitoring after an admission for acute decompensated heart failure: a sys-tematic review and meta-analysis of randomised controlled trials. EClini-calMedicine. 2024;71:102541. https://doi.org/10.1016/j.eclinm.2024.102541.

[23] Zheng W, Rawson S, Shen Z, et al. TMEM63 proteins function as monomeric high-threshold mechanosensitive ion channels. Neuron. 2023;111:3195e 3210 (e7). https://doi.org/10.1016/j.neuron.2023.07.006.

[24] Affo S, Yu LX, Schwabe RF. The role of cancer-associated fibroblasts and fibrosis in liver cancer. Annu Rev Pathol. 2017;12:153-186. https://doi.org/10.1146/annurev-pathol-052016-100322.

[25] Hamill OP, Martinac B. Molecular basis of mechanotransduction in living cells. Physiol Rev. 2001;81:685-740. https://doi.org/10.1152/physrev.2001.81.2.685.

[26] Xiao B. Mechanisms of mechanotransduction and physiological roles of PIEZO channels. Nat Rev Mol Cell Biol. 2024;25:886-903. https://doi.org/10.1038/s41580-024-00773-5.

[27] Zhang X, Zhao Y, Li M, et al. A synergistic regulation works in matrix stiffness-driven invadopodia formation in HCC. Cancer Lett. 2024;582:216597. https://doi.org/10.1016/j.canlet.2023.216597.

[28] Guan SH, Hu WJ, Wang XY, Gu YX, Zhou DH. New perspectives in prognosti-cation of hepatocellular carcinoma: the role and clinical implications of transient receptor potential family genes. World J Gastrointest Oncol. 2024;16: 2862-2864. https://doi.org/10.4251/wjgo.v16.i6.2862.

[29] Wei X, Li J, Xie H, et al. Chloride intracellular channel 1 participates in migration and invasion of hepatocellular carcinoma by targeting maspin. J Gastroenterol Hepatol. 2015;30:208-216. https://doi.org/10.1111/jgh.12668.

[30] Liu T, Han X, Zheng S, et al. CALM1 promotes progression and dampens che-mosensitivity to EGFR inhibitor in esophageal squamous cell carcinoma. Cancer Cell Int. 2021;21:121. https://doi.org/10.1186/s12935-021-01801-6.

[31] Rosenberg P. VDAC 2 as a novel target for heart failure: Ca2þ at the sarcomere, mitochondria and SR. Cell Calcium. 2022;104:102586. https://doi.org/10.1016/j.ceca.2022.102586.

[32] Zhang H, Zhang X, Xu W, Wang J. TMC 5 is highly expressed in human cancers and corelates to prognosis and immune cell infiltration: a comprehensive bioinformatics analysis. Front Mol Biosci. 2022;8:810864. https://doi.org/10.3389/fmolb.2021.810864.

[33] Zhang X, Shao J, Wang C, et al. TMC 7 functions as a suppressor of Piezo2 in primary sensory neurons blunting peripheral mechanotransduction. Cell Rep. 2024;43:114014. https://doi.org/10.1016/j.celrep.2024.114014.

[34] Li M, Zhang X, Wang M, et al. Activation of Piezo 1 contributes to matrix stiffness-induced angiogenesis in hepatocellular carcinoma. Cancer Commun (Lond). 2022;42:1162-1184. https://doi.org/10.1002/cac2.12364.

[35] Yuan Z, Li Y, Zhang S, et al. Extracellular matrix remodeling in tumor pro-gression and immune escape: from mechanisms to treatments. Mol Cancer. 2023;22:48. https://doi.org/10.1186/s12943-023-01744-8.

[36] Liu Y, Yao X, Zhao Y, et al. Mechanotransduction in response to ECM stiffening impairs cGAS immune signaling in tumor cells. Cell Rep. 2023;42:113213. https://doi.org/10.1016/j.celrep.2023.113213.

[37] Nicolas-Boluda A, Vaquero J, Vimeux L, et al. Tumor stiffening reversion through collagen crosslinking inhibition improves T cell migration and anti-PD-1 treatment. Elife. 2021;10:e58688. https://doi.org/10.7554/eLife.58688.

[38] Zhang J, Li J, Hou Y, et al. Osr 2 functions as a biomechanical checkpoint to aggravate CD8þ T cell exhaustion in tumor. Cell. 2024;187:3409e 3426 (e24). https://doi.org/10.1016/j.cell.2024.04.023.

[39] Yu KX, Yuan WJ, Wang HZ, Li YX. Extracellular matrix stiffness and tumor-associated macrophage polarization: new fields affecting immune exclusion. Cancer Immunol Immunother. 2024;73:115. https://doi.org/10.1007/s00262-024-03675-9.

[40] Batlle E,Massagué J. Transforming growth factor-b signaling in immunity and cancer. Immunity. 2019;50:924-940. https://doi.org/10.1016/j.immuni.2019.03.024.

[41] Bordeleau F, Califano JP, Negron Abril YL, et al. Tissue stiffness regulates serine/arginine-rich protein-mediated splicing of the extra domain B-fibronectin isoform in tumors. Proc Natl Acad Sci U S A. 2015;112:8314-8319. https://doi.org/10.1073/pnas.1505421112.

[42] Safina A, Vandette E, Bakin AV. ALK 5 promotes tumor angiogenesis by upre-gulating matrix metalloproteinase-9 in tumor cells. Oncogene. 2007;26: 2407-2422. https://doi.org/10.1038/sj.onc.1210046.

[43] Jiang Y, Zhang H, Wang J, Liu Y, Luo T, Hua H. Targeting extracellular matrix stiffness and mechanotransducers to improve cancer therapy. J Hematol Oncol. 2022;15:34. https://doi.org/10.1186/s13045-022-01252-0.

[44] Cambria E, Coughlin MF, Floryan MA, Offeddu GS, Shelton SE, Kamm RD. Linking cell mechanical memory and cancer metastasis. Nat Rev Cancer. 2024;24:216-228. https://doi.org/10.1038/s41568-023-00656-5.

[45] Hu Y, Tian H, Chen W, et al. The critical role of the Piezo1/b-catenin/ATF 4 axis on the stemness of Gli1þ BMSCs during simulated microgravity-induced bone loss. Adv Sci (Weinh). 2023;10:e2303375. https://doi.org/10.1002/advs.202303375.

[46] Bo H, Wu Q, Zhu C, Zheng Y, Cheng G, Cui L. PIEZO 1 acts as a cancer suppressor by regulating the ROS/Wnt/b-catenin axis. Thorac Cancer. 2024;15:1007-1016. https://doi.org/10.1111/1759-7714.15278.

[47] Lei ZN, Tian Q, Teng QX, et al. Understanding and targeting resistance mech-anisms in cancer. MedComm (2020). 2023; 4:e265. https://doi.org/10.1002/mco2.265.