[TERTp mutation screening using digital droplet PCR of collection of 52 paired DNA samples from blood plasma and tumor tissue in patients with glioblastoma].

Objective: To test the possibility of creating a non-invasive system for differential diagnosis and molecular-genetic characterization of tumor lesions of the CNS based on the determination of mutations of promoter of telomerase (TERTp) C228T and C250T catalyst subunit through the example of glioblastoma.

Material and methods: The collection of blood plasma samples and tissues of freshly frozen tumor samples was obtained in pairwise correspondence from 52 patients, who were treated in NMRC of Neurosurgery named after Academician N.N. Burdenko in 2022-2023. The digital droplet PCR (ddPCR) method using fluorescently labeled TaqMan probes was used for absolute quantification of C228T and C250T TERTp mutations in DNA samples extracted from the collection by commercial kits. The analysis of ddPCR data was performed with a baseline depending on the level of control reaction signal without adding a DNA matrix.

Results: Analysis of the sample of 52 patients' paired DNA (plasma-tissue) has shown the presence of C228T and C250T mutations in tumor samples: C228T was found in 33 cases (63.5%), and C250T in 14 cases (26.9%). In samples of cell-free DNA (CFDNA), 5 samples with C228T mutation and 3 samples with C250T mutation were found. C228T mutation was found in 15.1% and C250T mutation - in 21.4%.

Conclusion: TERTp mutations, specific for tumor DNA of glioblastomas, are determined by ddPCR in both the tumor sample and the CFDNA of the blood plasma, which indicates the transition of tumor DNA into the circulating (ctDNA) and that the ctDNA passes through the blood-brain barrier (BBB) of patients. The possibility of non-invasive molecular genetic characterization of tumor lesions of the CNS in blood plasma of patients with glioblastoma has been tested. The obtained results' analysis shows the need to optimize the extraction of CFDNA and modify the testing system to improve the method's effectiveness.