Evaluation of the effectiveness and the possibility of optimizing the methodology for analyzing the plasma microRNA-371-3 (HSA-miR-371-3p) level for the diagnosis and monitoring of the effect of therapy of germ cell tumors in men

Background. The choice of optimal tactics for treating patients with germ cell tumors (GCTs) is based on determining the totality of prognostic factors. Almost all patients with GCTs with a good prognosis can be cured by a rational combination of drug therapy and surgical methods. At the same time, the results of treatment of patients with a poor prognosis remain extremely unsatisfactory. One way to increase the effectiveness of therapy is to search for modern molecular biological factors that can optimize and personalize the treatment approach. Objective: Development of RT-PCR system (polymerase chain reaction with reverse transcription) for miR-351-3 analysis, preliminary assessment of the diagnostic effectiveness of analyzing the concentration of this molecule, determination of the possibility (s) of optimizing the technology and the prospects of introducing into practice a new method for monitoring GCTs based on RT-PCR analysis of miR-371-3. Methods. The design, synthesis and analysis of the RT-PCR system for miR-371-3 detection was carried out using domestic reagents, the diagnostic value of the method was evaluated using the material (plasma) of patients with a verified diagnosis of GCTs (n=13) and healthy donors (n=15). To develop a method for the specific isolation of miR-371-1 from plasma, super-paramagnetic particles (SPMP) and a DNA probe complementary to miR-371-3 were used. Results. An original system for the quantitative analysis of miR-371-3 by the RT-PCR method (ttTR-PCR) was created; the analytical efficiency of this system was shown in a wide range of concentrations of the detected molecule. The plasma miR-371-3 concentration in patients with GCTs and healthy donors was estimated. To increase the sensitivity of the technology, a technique for the specific isolation of miR-371-3 from plasma using SPMP was developed and tested. Conclusion. A qualitative assessment of plasma miR-371-3 level seems to be a promising method for monitoring the effectiveness of GCT therapy, but the technology for analyzing this molecule requires optimization and large-scale clinical validation.

testicular germ cell tumor, miRNA, miR-371-3, reverse transcription polymerase chain reaction