Identification of predictive markers in the cerebrospinal fluid of patients with glioblastoma

Introduction. Glioblastoma  (GB) is not yet curable despite recent advances in the treatment of other malignant solid tumors. The management of GB is based solely on histopathological features, imaging of the tumor and its genomic analysis (somatic mutations in the isocitrate dehydrogenase genes, methylation status of the O⁶-methylguanine-DNA methyltransferase gene promoter). To adapt the treatment to the most recent tumor evolution, molecular information should be received regularly throughout the course of therapy. However, tumor tissue is often not available for diagnosis as the disease progresses. In this regard, the development of less invasive methods, such as analysis of the proteome of biological fluids of patients, is of particular interest. Cerebrospinal fluid (CSF) is an important source disease biomarkers to monitor the presence and progression of the disease.

Aim. To identify proteomic predictive biomarkers in the CSF of patients with GB.

Materials and methods. During the study, samples of patients’ CSF samples, high-resolution proteomic mass spectrometry, modern biochemical methods and bioinformatic technologies were used.

Results. For the first time, the analysis of proteomes of CSF samples of patients with GB obtained before and 7 days after the removal of the primary tumor was carried out. Potential biomarkers of GB have been identified. After their validation using open databases, 11 proteomic predictive markers of GB (S100A9,  S100A8, PLA2G15, PPIB, LTBP2, VIM, LAMB1, STC1, NRP1, COL6A1, HSPA5)  were selected and their role in the molecular mechanisms of gliomagenesis was assessed. Conclusion. The proposed panel of proteomic predictive CSF biomarkers in GB patients can be further used in the development of test systems for assessing the effectiveness of therapy and early detection of disease relapses.

1. Stupp R., Mason W.P., van den Bent M.J. et al. Radiotherapy plus concomitant and adjuvant temozolomide for glioblastoma. N Engl J Med 2005;352:987–96. DOI: 10.1056/NEJMoa043330

2. Yan H., Parsons D.W., Jin G. et al. IDH1 and IDH2 mutations in gliomas. N Engl J Med 2009;360(8):765–73. DOI: 10.1056/nejmoa0808710

3. Sturm D., Witt H., Hovestadt V. et al. Hotspot mutations in H3F3A and IDH1 define distinct epigenetic and biological subgroups of glioblastoma. Cancer Cell 2012;22(4):425–37. DOI: 10.1016/j.ccr.2012.08.024

4. Brennan C.W., Verhaak R.G.W., McKenna A. et al. The somatic genomic landscape of glioblastoma. Cell 2013;155(2):462–77. DOI: 10.1016/j.cell.2013.09.034

5. Wick A., Kessler T., Platten M. et al. Superiority of temozolomide over radiotherapy for elderly patients with RTK II methylation class, MGMT promoter methylated malignant astrocytoma. Neurooncol 2020;22(8):1162–72. DOI: 10.1093/neuonc/noaa033

6. Weller M., van den Bent M., Preusser M. et al. EANO guidelines on the diagnosis and treatment of diffuse gliomas of adulthood. Nat Rev Clin Oncol 2021;18(3):170–86. DOI: 10.1038/s41571-020-00447-z

7. Sørensen M.D., Fosmark S., Hellwege S. et al. Chemoresistance and chemotherapy targeting stem-like cells in malignant glioma. Adv Exp Med Biol 2015;853:111–38. DOI: 10.1007/978-3-319-16537-0

8. Sastry R.A., Shankar G.M., Gerstner E.R. et al. The impact of surgery on survival after progression of glioblastoma: a retrospective cohort analysis of a contemporary patient population. J Clin Neurosci 2018;53:41–7. DOI: 10.1016/j.jocn.2018.04.004

9. Aebersold R., Mann M. Mass-spectrometric exploration of proteome structure and function. Nature 2016;537(7620):347–55. DOI: 10.1038/nature19949

10. Jiang Y., Sun A., Zhao Y. et al. Proteomics identifies new therapeutic targets of early-stage hepatocellular carcinoma. Nature 2019;567(7747):257–61. DOI: 10.1038/s41586-019-0987-8

11. Coscia F., Lengyel E., Duraiswamy J. et al. Multi-level proteomics identifies CT45 as a chemosensitivity mediator and immunotherapy target in ovarian cancer. Cell 2014;175(1):159–70. DOI: 10.1016/j.cell.2018.08.065

12. Bryukhovetskiy A., Shevchenko V., Kovalev S. et al. To the novel paradigm of proteome-based cell therapy of tumors: through comparative proteome mapping of tumor stem cells and tissuespecific stem cells of humans. Cell Transplant 2014;23(Suppl. 1):151–70. DOI: 10.3727/096368914X684907

13. Shen F., Zhang Y., Yao Y. et al. Proteomic analysis of cerebrospinal fluid: toward the identification of biomarkers for gliomas. Neurosurg Rev 2014;37(3):367–80. DOI: 10.1007/s10143-014-0539-5

14. Schmid D., Warnken U., Latzer P. et al. Diagnostic biomarkers from proteomic characterization of cerebrospinal fluid in patients with brain malignancies. J Neurochem 2021;158(2):522–38. DOI: 10.1111/jnc.15350

15. Wang H., Mao X., Ye L. et al. The role of the S100 protein family in glioma. J Cancer 2022;13(10):3022–30. DOI: 10.7150/jca.73365

16. Grebhardt S., Veltkamp C., Ströbel P. et al. Hypoxia and HIF‐1 increase S100A8 and S100A9 expression in prostate cancer. Int J Cancer 2012;131(12):2785–94. DOI: 10.1002/ijc.27591

17. Jang J.E., Kim H.P., Han S.W. et al. NFATC3-PLA2G15 fusion transcript identified by RNA sequencing promotes tumor invasion and proliferation in colorectal cancer cell lines. Cancer Res Treat 2019;51(1):391–401. DOI: 10.4143/crt.2018.103

18. Oh Y., Lee E.H., Kang S.G. PPIB is overexpressed in glioblastoma and regulates tumor growth by inducing self-ubiquitination by E3 ligase, Smurf2. Brain Tumor Res Treat 2022;10:S255. DOI: 10.14791/btrt.2022.10.F-1443

19. Zhao J., Liu X., Cong K. et al. The prognostic significance of LTBP2 for malignant tumors: Evidence based on 11 observational studies. Medicine 2022;101(17):e29207. DOI: 10.1097/MD.0000000000029207

20. Wang J., Liang W.J., Min G.T. et al. LTBP2 promotes the migration and invasion of gastric cancer cells and predicts poor outcome of patients with gastric cancer. Int J Oncol 2018;52(6):1886–98. DOI: 10.3892/ijo.2018.4356

21. Wang T.A., Zhou Z., Wang C. et al. LTBP2 knockdown promotes ferroptosis in gastric cancer cells through p62-keap1-Nrf2 pathway. BioMed Res Int 2022;2022:1–15. DOI: 10.1155/2022/6532253

22. Li Q., Aishwarya S., Li J.P. et al. Gene expression profiling of glioblastoma to recognize potential biomarker candidates. Front Genet 2022;13:832742. DOI: 10.3389/fgene.2022.832742

23. Lin H., Hong Y., Huang B. et al. Vimentin overexpressions induced by cell hypoxia promote vasculogenic mimicry by renal cell carcinoma cells. BioMed Res Int 2019;2019:1–13. DOI: 10.1155/2019/7259691

24. Liu T., Guevara O.E., Warburton R.R. et al. Regulation of vimentin intermediate filaments in endothelial cells by hypoxia. Am J Physiol 2010;299(2):363–73. DOI: 10.1152/ajpcell.00057.2010

25. Srivastava C., Irshad K., Dikshit B. et al. FAT1 modulates EMT and stemness genes expression in hypoxic glioblastoma. Int J Cancer 2018;142(4):805–12. DOI: 10.1002/ijc.31092

26. Li Y., Deng G., Qi Y. et al. Bioinformatic profiling of prognosis-related genes in malignant glioma microenvironment. Med Sci Monit 2020;26:e924054-1. DOI: 10.12659/MSM.924054

27. Chen T.Y., Liu Y., Chen L. et al. Identification of the potential biomarkers in patients with glioma: a weighted gene co-expression network analysis. Carcinogenesis 2020;41(6):743–50. DOI: 10.1093/carcin/bgz194

28. Virga J., Bognár L., Hortobágyi T. et al. Prognostic role of the expression of invasion-related molecules in glioblastoma. J Neurol Surg A: Cent Eur Neurosurg 2017;78(1):12–9. DOI: 10.1055/s-0036-1584920

29. Zhang F., Wang X., Bai Y. et al. Development and validation of a hypoxia-related signature for predicting survival outcomes in patients with bladder cancer. Front Genet 2021;12:670384. DOI: 10.3389/fgene.2021.670384

30. Xiong Y., Wang Q. STC1 regulates glioblastoma migration and invasion via the TGF β/SMAD4 signaling pathway. Mol Med Rep 2019;20(4):3055–64. DOI: 10.3892/mmr.2019.10579

31. Yeung H.Y., Lai K.P., Chan H.Y. et al. Hypoxia-inducible factor-1-mediated activation of stanniocalcin-1 in human cancer cells. Endocrinology 2005;146(11):4951–60. DOI: 10.1210/en.2005-0365

32. Sakata J., Sasayama T., Tanaka K. et al. MicroRNA regulating stanniocalcin-1 is a metastasis and dissemination promoting factor in glioblastoma. J Neurooncol 2019;142:241–51. DOI: 10.1007/s11060-019-03113-2

33. Ma X., Gu L., Li H. et al. Hypoxia-induced overexpression of stanniocalcin-1 is associated with the metastasis of early stage clear cell renal cell carcinoma. J Transl Med 2015;13(1):1–14. DOI: 10.1186/s12967-015-0421-4

34. Sun S., Lei Y., Li Q. et al. Neuropilin-1 is a glial cell line-derived neurotrophic factor receptor in glioblastoma. Oncotarget 2017;8(43):74019–35. DOI: 10.18632/oncotarget.18630

35. Liu Y., Liu Y., Gao Y. et al. H19-and hsa-miR-338-3p-mediated NRP1 expression is an independent predictor of poor prognosis in glioblastoma. PloS One 2021;16(11):e0260103. DOI: 10.1371/journal.pone.0260103

36. Fu R., Du W., Ding Z. et al. HIF-1α promoted vasculogenic mimicry formation in lung adenocarcinoma through NRP1 upregulation in the hypoxic tumor microenvironment. Cell Death Dis 2021;12(4):394. DOI: 10.1038/s41419-021-03682-z

37. Han X., Wang Q, Fang S. et al. P4HA1 Regulates CD31 via COL6A1 in the Transition of Glioblastoma Stem-Like Cells to Tumor Endothelioid Cells. Front Oncol 2022;12:836511. DOI: 10.3389/fonc.2022.836511

38. Lin H., Yang Y., Hou C. et al. Identification of COL6A1 as the key gene associated with antivascular endothelial growth factor therapy in glioblastoma multiforme. Genet Test Mol Biomarkers 2021;25(5):334–45. DOI: 10.1089/gtmb.2020.0279

39. Wen X., Chen X., Chen X. Increased expression of GRP78 correlates with adverse outcome in recurrent glioblastoma multiforme patients. Turk Neurosurgery 2020;30(1):11–6. DOI: 10.5137/1019-5149.jtn.21840-17.4

40. Chen Z., Wang H., Zhang Z. et al. Cell surface GRP78 regulates BACE2 via lysosome-dependent manner to maintain mesenchymal phenotype of glioma stem cells. J Exp Clin Cancer Res 2021;40(1):1–17. DOI: 10.1186/s13046-020-01807-4