Quantitative characterization of TUBB3 expression parameters in the tissue of ovarian cancer
AbstractIntroduction. The expansion of the spectrum of molecular prognostic and predictive markers of tumors is a modern trend aimed at increasing the effectiveness of drug therapy for tumors of various origins, including ovarian cancer, the results of which remain unsatisfactory. In this regard, the cytoskeleton protein TUBB3, which is expressed in epithelial tumors, has significant potential. Basic research has proven that TUBB3 overexpression increases the metastatic potential of tumor cells and induces resistance to a wide range of drugs. However, in translational studies with a semi-quantitative analysis of TUBB3 expression, estimates of the contribution of this protein to the aggressiveness of ovarian cancer and the effectiveness of chemotherapy are ambiguous.
The aim of the study was a quantitative assessment of TUBB3 expression parameters in ovarian cancer tissue and determination of the correlation of the identified parameters with the clinical characteristics of the neoplasm.
Material and methods. Surgical samples of ovarian cancer tissue were examined (n = 51). Primary monoclonal antibodies to TUBB3 (clone EP1569Y, Abcam, England) and secondary antibodies conjugated with DyLight650 dye (ab98510, Abcam, England) were used. Fluorescence was measured using a Navios flow cytometer (Beckman Coulter, USA). The number of stained cells was determined in the FlowJo 10.0.8 program (USA) by the Kolmogorov–Smirnov method. Three indicators of TUBB3 expression were calculated: level (%), intensity (units), index (units). Statistical analysis of the normality of the distribution of TUBB3 expression indices, sample comparisons and correlation estimates were carried out in the GraphPad Prism 6.0 program (GraphPad Software, USA).
Results. In the studied sample of tumors, TUBB3 was detected in 100 % of cases with significant differences in quantitative indicators (up to 10 times) in different patients, which «coincides» with clinical data on the heterogeneity of the patient’s response to drug therapy and the overall survival time. There was a weak association between the level and intensity of TUBB3 expression and the absence of differences in indicators depending on the stage of ovarian cancer and the degree of tumor differentiation. In a comparative study in the same patients with primary and metastatic ovarian cancer (tumor cells from ascitic fluid in peritoneal carcinomatosis) The stability of the molecular phenotype has been demonstrated in less than 40 % of cases. In the remaining patients, the level of TUBB3 expression in tumor cells increased in half of the cases, decreased in half.
Conclusion. Quantitative immunofluorescence analysis of TUBB3 expression in ovarian cancer tissue revealed this protein in 100 % of cases with a median TUBB3 expression level of 55 % and pronounced heterogeneity of indicators in different patients. There were no differences in the level, intensity and expression index of TUBB3 depending on the stage of the disease and the degree of differentiation of the tumor. Multidirectional changes in the expression level of TUBB3 were noted in the cells of metastatic ovarian cancer compared with the primary tumor of the same patients. The generated database will allow for an accurate analysis of the predictive significance of TUBB3 expression in tumors and to determine quantitative indicators that contribute to drug resistance.
Keywords: ovarian cancer; TUBB3; tumor-associated marker
For citation. Bogush T.A., Osipova A.A., Lee А., Bogush Е.А., Grishanina A.N., Ravcheeva A.B., Kosorukov V.S. Quantitative characterization of TUBB3 expression parameters in the tissue of ovarian cancer. Immunologiya. 2024; 45 (4): 495–504. DOI: https://doi.org/10.33029/1816-2134-2024-45-4-495-504 (in Russian)
Funding. The research was carried out with the support of the Russian Science Foundation (Project No. 24-25-20080, https://rscf.ru/project/24-25-20080/).
Conflict of interests. Authors declare no conflict of interests.
Authors’ contribution. All authors made an equal contribution to the study, have read the final version of the manuscript and agree with its contents.
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