Cuproptosis-related gene CEP55 as a biomarker of pancreatic adenocarcinoma via multi-omics techniques and experimental validation

Zhang, Riyuan; Xu, Zixia; Zhuang, Yurui; Shi, Yuzhe; Guo, Ziyi; Chen, Chong

doi:10.2478/raon-2025-0042

Abstract

Background

Background. Pancreatic adenocarcinoma (PAAD) is a malignancy with a very poor prognosis. The clinical significance of cuproptosis in PAAD combining single cell data with The Cancer Genome Atlas (TCGA) data is unclear.

Materials and methods

In this study, we first identified gene modules associated with cuproptosis by performing single-cell analysis and weighted co-expression network analysis (WCGNA). According to TCGA data, Cox regression and LASSO regression analysis were used to establish prognostic models, and PAAD patients were divided into high-risk and low-risk groups according to cuproptosis-related risk score. Then 7 algorithms were used to evaluate cancer immune microenvironment, followed by the mutation analysis. The expression levels and prognostic significance of the 8 model genes were analysed using single-gene analysis, Kaplan-Meier survival plots, and quantitative PCR (qPCR) validation. Finally, the biological function of CEP55 in PAAD was verified by in vitro experiments.

Results

We identified cuproptosis-related genes (CRG) in PAAD by performing single-cell analysis and WCGNA, and constructed a cuproptosis-related prognostic model of PAAD by comprehensive bioinformatics analyses. Based on cuproptosis-related risk score, there were significant differences in survival time between two groups. We further constructed a cuproptosis-related risk score-based nomogram to accurately assess PAAD patient prognosis. Immune infiltration analysis revealed that PAAD samples with higher cuproptosis-related scores exhibited significantly lower immune infiltration levels, which may mechanistically underlie their poorer clinical outcomes. Furthermore, the high-risk group had a higher mutation rate of the same mutated gene, which means that they are more likely to benefit from immunotherapy. Finally, we identified that CEP55 was significantly overexpressed in PAAD and correlated with poor patient prognosis. In vitro knockdown of CEP55 effectively suppressed proliferation and invasion capabilities in pancreatic cancer cell lines.

Conclusions

In this study, a novel prognostic model of PAAD was constructed to evaluate the prognosis and immune microenvironment of PAAD patients, and CEP55 was identified as a central gene of PAAD. In vitro studies verified the biological function of CEP55, providing a new potential target for the treatment of PAAD.

References

Siegel RL, Miller KD, Fuchs HE, Jemal A. Cancer Statistics, 2021. CA Cancer J Clin 2021; 71: 7-33. doi: 10.3322/caac.21654
Open DOI Search in Google Scholar Back to article
Moore A, Donahue T. Pancreatic cancer. Jama 2019; 322: 1426. doi: 10.1001/jama.2019.14699
Open DOI Search in Google Scholar Back to article
Rahib L, Smith BD, Aizenberg R, Rosenzweig AB, Fleshman JM, Matrisian LM. Projecting cancer incidence and deaths to 2030: the unexpected burden of thyroid, liver, and pancreas cancers in the United States. Cancer Res 2014; 74: 2913-21. doi: 10.1158/0008-5472.Can-14-0155
Open DOI Search in Google Scholar Back to article
Warshaw AL, del Castillo FC. Pancreatic carcinoma. N Engl J Med 1992; 326: 455-65. doi: 10.1056/nejm199202133260706
Open DOI Search in Google Scholar Back to article
Howlader N, Noone AM, Krapcho M, Garshell J, Miller D, Altekruse SF, et al. SEER cancer statistics review, 1975-2009. [internet]. Bethesda, MD, USA: National Cancer Institute; 2012. [cited 2025 Jan 15]. Available at: http://seer.cancer.gov/csr/1975_2012/
Search in Google Scholar Back to article
Klepsch V, Hermann-Kleiter N, Do-Dinh P, Jakic B, Offermann A, Efremova M, et al. Nuclear receptor NR2F6 inhibition potentiates responses to PD-L1/PD-1 cancer immune checkpoint blockade. Nat Commun 2018; 9: 1538. doi: 10.1038/s41467-018-04004-2
Open DOI Search in Google Scholar Back to article
Li X, Gulati M, Larson AC, Solheim JC, Jain M, Kumar S, et al. Immune checkpoint blockade in pancreatic cancer: trudging through the immune desert. Semin Cancer Biol 2022; 86: 14-27. doi: 10.1016/j.semcancer.2022.08.009
Open DOI Search in Google Scholar Back to article
Heumann T, Azad N. Next-generation immunotherapy for pancreatic ductal adenocarcinoma: navigating pathways of immune resistance. Cancer Metastasis Rev 2021; 40: 837-62. doi: 10.1007/s10555-021-09981-3
Open DOI Search in Google Scholar Back to article
Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell 2011; 144: 646-74. doi: 10.1016/j.cell.2011.02.013
Open DOI Search in Google Scholar Back to article
Fesik SW. Promoting apoptosis as a strategy for cancer drug discovery. Nat Rev Cancer 2005; 5: 876-85. doi: 10.1038/nrc1736
Open DOI Search in Google Scholar Back to article
Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, et al. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell 2012; 149: 1060-72. doi: 10.1016/j.cell.2012.03.042
Open DOI Search in Google Scholar Back to article
Stockwell BR, Friedmann Angeli JP, Bayir H, Bush AI, Conrad M, Dixon SJ, et al. Ferroptosis: a regulated cell death nexus linking metabolism, redox biology, and disease. Cell 2017; 171: 273-85. doi: 10.1016/j.cell.2017.09.021
Open DOI Search in Google Scholar Back to article
Shi J, Gao W, Shao F. Pyroptosis: gasdermin-mediated programmed necrotic cell death. Trends Biochem Sci 2017; 42: 245-54. doi: 10.1016/j. tibs.2016.10.004
Open DOI Search in Google Scholar Back to article
Fatokun AA, Dawson VL, Dawson TM. Parthanatos: mitochondrial-linked mechanisms and therapeutic opportunities. Br J Pharmacol 2014; 171: 2000-16. doi: 10.1111/bph.12416
Open DOI Search in Google Scholar Back to article
Tsvetkov P, Coy S, Petrova B, Dreishpoon M, Verma A, Abdusamad M, et al. Copper induces cell death by targeting lipoylated TCA cycle proteins. Science 2022; 375: 1254-61. doi: 10.1126/science.abf0529
Open DOI Search in Google Scholar Back to article
Li SR, Bu LL, Cai L. Cuproptosis: lipoylated TCA cycle proteins-mediated novel cell death pathway. Signal Transduct Target Ther 2022; 7: 158. doi: 10.1038/s41392-022-01014-x
Open DOI Search in Google Scholar Back to article
Sprooten J, De Wijngaert P, Vanmeerbeerk I, Martin S, Vangheluwe P, Schlenner S, et al. Necroptosis in immuno-oncology and cancer immunotherapy. Cells 2020; 9: 1823. doi: 10.3390/cells9081823
Open DOI Search in Google Scholar Back to article
Zhang Z, Zeng X, Wu Y, Liu Y, Zhang X, Song Z. Cuproptosis-related risk score predicts prognosis and characterizes the tumor microenvironment in hepatocellular carcinoma. Front Immunol 2022; 13: 925618. doi: 10.3389/fimmu.2022.925618
Open DOI Search in Google Scholar Back to article
Li Z, Zhang H, Wang X, Wang Q, Xue J, Shi Y, et al. Identification of cuproptosis-related subtypes, characterization of tumor microenvironment infiltration, and development of a prognosis model in breast cancer. Front Immunol 2022; 13: 996836. doi: 10.3389/fimmu.2022.996836
Open DOI Search in Google Scholar Back to article
Ji ZH, Ren WZ, Wang HQ, Gao W, Yuan B. Molecular subtyping based on cuproptosis-related genes and characterization of tumor microenvironment infiltration in kidney renal clear cell carcinoma. Front Oncol 2022; 12: 919083. doi: 10.3389/fonc.2022.919083
Open DOI Search in Google Scholar Back to article
Fang Z, Wang W, Liu Y, Hua J, Liang C, Liu J, et al. Cuproptosis-related gene DLAT as a novel biomarker correlated with prognosis, chemoresistance, and immune infiltration in pancreatic adenocarcinoma: a preliminary study based on bioinformatics analysis. Curr Oncol 2023; 30: 2997-3019. doi: 10.3390/curroncol30030228
Open DOI Search in Google Scholar Back to article
Langfelder P, Horvath S. WGCNA: an R package for weighted correlation network analysis. BMC Bioinformatics 2008; 9: 559. doi: 10.1186/14712105-9-559
Open DOI Search in Google Scholar Back to article
Tibshirani R. The lasso method for variable selection in the Cox model. Stat Med 1997; 16: 385-95. doi: 10.1002/(sici)1097-0258(19970228)16:4<;385::aid-sim380>3.0.co;2-3
Open DOI Search in Google Scholar Back to article
Hidalgo M. Pancreatic cancer. N Engl J Med 2010; 362: 1605-17. doi: 10.1056/NEJMra0901557
Open DOI Search in Google Scholar Back to article
Von Hoff DD, Ervin T, Arena FP, Chiorean EG, Infante J, Moore M, et al. Increased survival in pancreatic cancer with nab-paclitaxel plus gemcitabine. N Engl J Med 2013; 369: 1691-703. doi: 10.1056/NEJMoa1304369
Open DOI Search in Google Scholar Back to article
Conroy T, Desseigne F, Ychou M, Bouché O, Guimbaud R, Bécouarn Y, et al. FOLFIRINOX versus gemcitabine for metastatic pancreatic cancer. N Engl J Med 2011; 364: 1817-25. doi: 10.1056/NEJMoa1011923
Open DOI Search in Google Scholar Back to article
Couzin-Frankel J. Breakthrough of the year 2013. Cancer immunotherapy. Science 2013; 342: 1432-3. doi: 10.1126/science.342.6165.1432
Open DOI Search in Google Scholar Back to article
Kumar A, Swain CA, Shevde LA. Informing the new developments and future of cancer immunotherapy: future of cancer immunotherapy. Cancer Metastasis Rev 2021; 40: 549-62. doi: 10.1007/s10555-021-09967-1
Open DOI Search in Google Scholar Back to article
Morrison AH, Byrne KT, Vonderheide RH. Immunotherapy and prevention of pancreatic cancer. Trends Cancer 2018; 4: 418-28. doi: 10.1016/j. trecan.2018.04.001
Open DOI Search in Google Scholar Back to article
Yin C, Alqahtani A, Noel MS. The next frontier in pancreatic cancer: targeting the tumor immune milieu and molecular pathways. Cancers 2022; 14: 2619. doi: 10.3390/cancers14112619
Open DOI Search in Google Scholar Back to article
Li F, He C, Yao H, Liang W, Ye X, Ruan J, et al. GLUT1 regulates the tumor immune microenvironment and promotes tumor metastasis in pancreatic adenocarcinoma via ncRNA-mediated network. J Cancer 2022; 13: 2540-58. doi: 10.7150/jca.72161
Open DOI Search in Google Scholar Back to article
Garrido-Laguna I, Hidalgo M. Pancreatic cancer: from state-of-the-art treatments to promising novel therapies. Nat Rev Clin Oncol 2015; 12: 319-334. doi: 10.1038/nrclinonc.2015.53
Open DOI Search in Google Scholar Back to article
Tong X, Tang R, Xiao M, Xu J, Wang W, Zhang B, et al. Targeting cell death pathways for cancer therapy: recent developments in necroptosis, pyroptosis, ferroptosis, and cuproptosis research. J Hematol Oncol 2022; 15: 174. doi: 10.1186/s13045-022-01392-3
Open DOI Search in Google Scholar Back to article
Qin Y, Liu Y, Xiang X, Long X, Chen Z, Huang X, et al. Cuproptosis correlates with immunosuppressive tumor microenvironment based on pan-cancer multiomics and single-cell sequencing analysis. Mol Cancer 2023; 22: 59. doi: 10.1186/s12943-023-01752-8
Open DOI Search in Google Scholar Back to article
Guo B, Yang F, Zhang L, Zhao Q, Wang W, Yin L, et al. Cuproptosis induced by ROS responsive nanoparticles with elesclomol and copper combined with αPD-L1 for enhanced cancer immunotherapy. Adv Mater 2023; 35: e2212267. doi: 10.1002/adma.202212267
Open DOI Search in Google Scholar Back to article
Liu WQ, Lin WR, Yan L, Xu WH, Yang J. Copper homeostasis and cuproptosis in cancer immunity and therapy. Immunol Rev 2024; 321: 211-27. doi: 10.1111/imr.13276
Open DOI Search in Google Scholar Back to article
Li J, Yin J, Li W, Wang H, Ni B. Molecular subtypes based on cuproptosis-related genes and tumor microenvironment infiltration characteristics in pancreatic adenocarcinoma. Cancer Cell Int 2023; 23: 7. doi: 10.1186/s12935-022-02836-z
Open DOI Search in Google Scholar Back to article
Wang G, Chen B, Su Y, Qu N, Zhou D, Zhou W. CEP55 as a promising immune intervention marker to regulate tumor progression: a pan-cancer analysis with experimental verification. Cells 2023; 12: 2457. doi: 10.3390/cells12202457
Open DOI Search in Google Scholar Back to article
Neuzillet C, Tijeras-Raballand A, Ragulan C, Cros J, Patil Y, Martinet M, et al. State of the art and future directions of pancreatic ductal adenocarcinoma therapy. Pharmacol Ther 2015; 155: 80-104. doi: 10.1016/j. pharmthera.2015.08.006
Open DOI Search in Google Scholar Back to article
Xiao Y, Yu D. Tumor microenvironment as a therapeutic target in cancer. Pharmacol Ther 2021; 221: 107753. doi: 10.1016/j.pharmthera.2020.107753
Open DOI Search in Google Scholar Back to article
Joyce JA, Fearon DT. T cell exclusion, immune privilege, and the tumor microenvironment. Science 2015; 348: 74-80. doi: 10.1126/science.aaa6204
Open DOI Search in Google Scholar Back to article
Kleeff J, Beckhove P, Esposito I, Herzig S, Huber PE, Lohr JM, et al. Pancreatic cancer microenvironment. Int J Cancer 2007; 121: 699-705. doi: 10.1002/ijc.22871
Open DOI Search in Google Scholar Back to article
Korc M. Pancreatic cancer-associated stroma production. Am J Surg 2007; 194: S84-86. doi: 10.1016/j.amjsurg.2007.05.004
Open DOI Search in Google Scholar Back to article
Danaher P, Warren S, Lu R, Samayoa J, Sullivan A, Pekker I, et al. Pancancer adaptive immune resistance as defined by the Tumor Inflammation Signature (TIS): results from The Cancer Genome Atlas (TCGA). J Immunother Cancer 2018; 6: 63. doi: 10.1186/s40425-018-0370-7
Open DOI Search in Google Scholar Back to article
Lutz ER, Wu AA, Bigelow E, Sharma R, Mo G, Soares K, et al. Immunotherapy converts nonimmunogenic pancreatic tumors into immunogenic foci of immune regulation. Cancer Immunol Res 2014; 2: 616-31. doi: 10.1158/2326-6066.CIR-14-0027
Open DOI Search in Google Scholar Back to article
J Gunderson A, Rajamanickam V, Bui C, Bernard B, Pucilowska J, Ballesteros-Merino C, et al. Germinal center reactions in tertiary lymphoid structures associate with neoantigen burden, humoral immunity and long-term survivorship in pancreatic cancer. Oncoimmunology 2021; 10: 1900635. doi: 10.1080/2162402x.2021.1900635
Open DOI Search in Google Scholar Back to article
Tang R, Zhang Y, Liang C, Xu J, Meng Q, Hua J, et al. The role of m6A-related genes in the prognosis and immune microenvironment of pancreatic adenocarcinoma. PeerJ 2020; 8: e9602. doi: 10.7717/peerj.9602
Open DOI Search in Google Scholar Back to article
Wang L, Zhang S, Li H, Xu Y, Wu Q, Shen J, et al. Quantification of m6A RNA methylation modulators pattern was a potential biomarker for prognosis and associated with tumor immune microenvironment of pancreatic adenocarcinoma. BMC Cancer 2021; 21: 876. doi: 10.1186/s12885-021-08550-9
Open DOI Search in Google Scholar Back to article
Wang G, Yang L, Gao J, Mu H, Song Y, Jiang X, et al. Identification of candidate biomarker ASXL2 and its predictive value in pancreatic carcinoma. Front Oncol 2021; 11: 736694. doi: 10.3389/fonc.2021.736694
Open DOI Search in Google Scholar Back to article
Lestari B, Utomo RY. CEP55 inhibitor: Extensive computational approach defining a new target of cell cycle machinery agent. Adv Pharm Bull 2022; 12: 191-99. doi: 10.34172/apb.2022.021
Open DOI Search in Google Scholar Back to article
Kalimutho M, Sinha D, Jeffery J, Nones K, Srihari S, Fernando WC, et al. CEP55 is a determinant of cell fate during perturbed mitosis in breast cancer. EMBO Mol Med 2018; 10: e8566. doi: 10.15252/emmm.201708566
Open DOI Search in Google Scholar Back to article
Sinha D, Nag P, Nanayakkara D, Duijf PHG, Burgess A, Raninga P, et al. Cep55 overexpression promotes genomic instability and tumorigenesis in mice. Commun Biol 2020; 3: 593. doi: 10.1038/s42003-020-01304-6
Open DOI Search in Google Scholar Back to article
Zhang X, Xu Q, Li E, Shi T, Chen H. CEP55 predicts the poor prognosis and promotes tumorigenesis in endometrial cancer by regulating the Foxo1 signaling. Mol Cell Biochem 2023; 478: 1561-71. doi: 10.1007/s11010-022-04607-w
Open DOI Search in Google Scholar Back to article
Ye Y, Chen Z, Shen Y, Qin Y, Wang H. Development and validation of a fourlipid metabolism gene signature for diagnosis of pancreatic cancer. FEBS Open Bio 2021; 11: 3153-70. doi: 10.1002/2211-5463.13074
Open DOI Search in Google Scholar Back to article
Luo X, Linghu M, Zhou X, Ru Y, Huang Q, Liu D, et al. Merestinib inhibits cuproptosis by targeting NRF2 to alleviate acute liver injury. Free Radic Biol Med 2025; 229: 68-81. doi: 10.1016/j.freeradbiomed.2025.01.029
Open DOI Search in Google Scholar Back to article