E.I. Antonova1, A.B. Achilov2, N.A. Lengesova3, N.V. Firsova4, E.V. Balacuk5, D.A. Saifutdinova6

1–6 Scientific Research Center for Fundamental and Applied Problems of Bioecology and Biotechnology Ulyanovsk State Pedagogical University named after I.N. Ulyanov (Ulyanovsk, Russia)

1 antonov_67@mail.ru, 2 a.achilow@inbox.ru, 3 lengesova@yandex.ru, 4 n-firsova@mail.ru, 5 balacyxa@mail.ru, 6 dinasajfutdinova17@gmail.com

This article delineates the outcomes of developing an indigenous multiplex test system protocol, leveraging xMAP-ASPE-xTAG technology, for the detection of gene mutations, exemplified by BRAF, MITF-M, and CDKN2A, in cutaneous lesion samples with a preliminary histopathological diagnosis of dysplastic nevus with a pigment component. The study was conducted to assess the probability of malignant transformation of dysplastic nevus with a pigment component into melanoma. The multiplex assay simultaneously identified mutations in the BRAF (rs113488022; p.V600E; c.1799T>A), MITF-M (rs149617956; p.E318K; c.952G>A), and CDKN2A (rs121913386, p.P114L, c.341C>T; rs121913387, p.R58*, c.172C>T; rs121913388, p.R80*, c.238C>T; rs121913389, p.W110*, c.330G>A; rs1057519852, p.W110*, c.329G>A) genes within a single analyte. The resultant protocol parameters were defined according to lesion area, total and target DNA (gene) concentrations, optimal reagent concentrations, and microsphere quantity. Varied reagent concentrations and incubation times facilitated the standardization and optimization of experimental conditions, thereby achieving acceptable sensitivity and robust performance in the detection of the analyzed gene mutations. Validation of the obtained results was performed, and specificity was determined. Mutational analysis revealed histological criteria characteristic of dysplastic nevi (DN). BRAF gene mutations (rs113488022; p.V600E; c.1799T>A) were identified in three out of four samples. One sample exhibited mutations in three genes—BRAF (rs113488022; p.V600E; c.1799T>A), MITF-M (rs149617956; p.E318K; c.952G>A), and CDKN2A (rs121913386; p.P114L; c.341C>T)—the presence of which constitutes an initial stage of melanocytic neoplasia, reflects hereditary predisposition to melanoma development, and indicates dysregulation of the cell cycle and proliferation. Furthermore, the identified germline mutation in the MITF-M gene (p.E318K) signifies a high risk of developing cutaneous malignant melanoma. This work is conducted within the framework of developing an indigenous diagnostic test system utilizing xMAP-ASPE-xTAG technology for the early diagnosis and identification of predisposition to melanoma development.

Antonova E.I., Achilov A.B., Lengesova N.A., Firsova N.V., Balacuk E.V., Saifutdinova D.A. Molecular profiling of melanocytes (displastic nevus with a pigment component). Technologies of Living Systems. 2025. V. 22. № 4. Р. 32-41. DOI: https://doi.org/10.18127/ j20700997-202504-04 (In Russian).

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