827. Integration and Biodistribution Studies of Retroviral Vector in Mouse Gonad Tissues

Abstract

Listen

Translate article

Top of pageAbstract Retroviruses have been used as efficient gene delivery vehicles because they are able to stably integrate into genome of host cells. This is particularly important safety concern if vector distributes to gonads, raising the possibility of inadvertent germ line integration(transmission). However, targeting integration to specific location in the human genome remains an met challenge. The goal of this study is evaluate the reproductive toxicity of ex vivo and/or in vivo retrovirus mediated gene transfer with appropriate methods, to this end we studied the biodistribution, integration pattern into chromosome and potential germ-line transmission of eGFP-expressing retrovirus (MT-eGFP). Both male and female C57BL/6 mice were transplanted with hematopoietic stem cells infected MT-eGFP. The PCR analysis showed that there were detectable vector sequences in blood, spleen, thymus, while liver, testis, and ovary observed. weak signal. Real time-PCR results showed that integration into chromosome of retroviral vector occurred somewhat variable integration range from 0.1 to 0.00001 depend on tissues. By using specific ligation-mediated PCR(LM-PCR) followed by sequencing of vector integration site, we found that integrations were occurred most chromosome including sex chromosomes X and Y. Vector-administered mice into gonad(testis and ovary) directly were mated and their offspring were evaluated for germ-line transmission of retroviral vector. The PCR analysis showed no evidence of germ-line transmission, although vector integration were identified from gonad tissues with LM-PCR. Our data suggested that the risk of the inadvertent germline transmission of retroviral vector following ex vivo and in vivo administration is extremely low and MT-vecor have a some specific integration site. Defining the different integration sites for retrovirus will improve the safety of retroviral vectors for use in gene therapy and may aid in our understanding of the factors which will be correlate with reproductive problems.