Packaging of endogenous retroviral sequences in retroviral vectors produced by murine and human packaging cells.
Interaction of retrovirus vectors and endogenous retroviruses present in packaging cell lines and target cells may result in unwanted events, such as the formation of recombinant viruses and the mobilization of therapeutic vectors. Using sensitive reverse transcriptase PCR assays, we investigated human and murine gene therapy packaging cell lines for incorporation of endogenous retrovirus transcripts into murine leukemia virus (MLV) vector particles and, conversely, whether vector genomes are incorporated into human endogenous retrovirus (HERV) particles. VL30 endogenous retrovirus sequences were efficiently packaged in particles produced by the murine AM12 packaging system. For every seven MLV-derived beta-galactosidase (beta-Gal) vector genomes present in the particles, one copy of VL30 was also packaged. Although human FLY packaging cells expressed several classes of HERV transcripts (HERV-K, HuRT, type C, and RTVL-H), none was detectable in the MLV vector particles released from the cells. Nonspecific packaging of the MLV Gag-Pol expression vector transcripts was detected in the FLY virions at a low level (1 in 17,000 sequences). These findings indicate that human packaging cells produce retrovirus particles far less contaminated by endogenous viral sequences than murine packaging cells. Human teratocarcinoma cells (GH cells), which produce HERV-K particles, were transduced with an MLV-derived beta-Gal vector. Although both HERV-K and RTVL-H sequences were found in association with the particles, beta-Gal transcripts were not detected, indicating that HERV Gag proteins do not efficiently package MLV-based vectors.
Patience
C
C
Chester Beatty Laboratories, Institute of Cancer Research, London, United Kingdom. clive@icr.ac.uk
Takeuchi
Y
Y
Cosset
F L
FL
Weiss
R A
RA
D000818:Animals; D002460:Cell Line; D015870:Gene Expression; D015750:Genes, gag; D005822:Genetic Vectors / Q000235:genetics / Q000502:physiology*; D006801:Humans; D009052:Leukemia Virus, Murine / Q000235:genetics / Q000502:physiology*; D051379:Mice; D016133:Polymerase Chain Reaction / Q000379:methods; D012190:Retroviridae / Q000235:genetics*; D012680:Sensitivity and Specificity; D014158:Transcription, Genetic; D019065:Virus Assembly