Molecular and genetic analysis of personal cultures of uveal melanoma cells

Relevance. Uveal melanoma (UM) is a subtype of melanoma that represents the majority of adult ocular cancers. The rare frequency of the disease does not allow to conduct large-scale clinical trials of new agents and approaches, which limits progress in its treatment. Half of patients with UM develop metastatic forms of the disease, which generally have unfavorable prognosis. Therefore, personal cell cultures derived from tumor material of patients are relevant for determining molecular and genetic features of tumor melanocytes and for testing new antitumor drugs. Purpose. To evaluate the aggressiveness of choroidal melanoma by molecular and genetic characterization of melanocytes isolated from the tumor. Material and methods. The study was performed using personal cultures of human UM cells. Karyotyping of cells was performed by GTG staining analysis of the metaphase plates, mutational burden was assessed by NGS sequencing, protein products were evaluated by western blotting. Results. In this work, three personal cultures of human UM cells, uMel1, uMel3 and uMel5, cultured for at least 22 passages, were obtained from tumor material of patients diagnosed with UM for the analysis of molecular and genetic characteristics. Since the development of the metastatic form of UM correlates with chromosomal alterations in tumor melanocytes, karyotyping was performed, which showed that in all cultures, most cells had a near-diploid phenotype. Only in uMel1 and uMel3 cells were chromosomal rearrangements specific for UM – in chromosomes 3, 6, and 8. In uMel5 cells, a deletion was found in the region of the long arm of chromosome 11. In addition, the patient with uMel5 tumor had a history of skin melanoma, for which the typical somatic mutation profile differs from that of UM. Somatic mutations in hotspots of ALK, BRAF, EGFR, HER2, IL17RA, KRAS, METand NRASgenes were analyzed by NGS-sequencing method. No mutations were detected at the tested loci in cells from all cultures tested. Analysis of tumor suppressor BAP1 (BRCA-associated protein) showed its low level in uMel3 and uMel5 cell cultures, which may indicate a high risk of metastasis. Conclusion. The obtained personalized cultures of UM cells are models that allow in vitroand in vivoinvestigation of UM with different molecular and genetic signatures. Key words: uveal melanoma, personalized cell cultures, karyotype, BAP1, BRAF, NGS-sequencing