Risk assessment of germline transmission of adeno-associated virul vector of chimeric serotype PHP.eB into mouse oocytes

Aim. To assess the risk of germline transmission of an adeno-associated virul vector of the chimeric serotype PHP. eB into the germ cells of female mice based on the development of an original method for obtaining mouse oocytes devoid of somatic cells and free vector particles. Materials and methods. The vector under study was administered intravenously to female outbred CD-1 mice at a dose of 5 × 10¹⁰ vector genomes/mouse using an original technique which included hormonal superovulation of exposed animals, oocyte isolation, their purification from contaminating somatic cells, and direct quantitative PCR on lysed oocytes. The vector DNA content in the brain tissue, ovaries, and oocytes was assessed on days 1, 3, 7, 14, 30, and 90 after vector administration. Results. Using the developed technique, we examined the ability of the adeno-associated virus vector of the chimeric serotype PHP.eB to undergo germline transmission into mouse oocytes. It was established that, despite persistence in brain and ovarian tissue for up to 3 months, vector DNA was not detected in oocytes at any time after vector administration. Conclusion. The obtained data demonstrate the absence of germline transmission of the studied genetic construct into mouse oocytes. The detection of the adeno-associated vector in the ovaries, despite its absence in oocytes, confirms the effectiveness of the developed technique for obtaining mouse oocytes uncontaminated with somatic cells.