STUDY OF A MURINE RETROVIRUS FROM A PACKAGING CELL LINE USED IN GENE THERAPY

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Replication-competent murine retroviruses (RCRs) can arise, in some case, unexpectedly and spontaneiously from retrovirus-based packaging cell systems used in human gene therapy products. RCRs continuing an amphotropic murine leukemia virus (MuLV) envelope possess a broad host range and can infect a wide variety of primate cells including human. Since the risk associated with RCRs in man is unknown, rigorous testing for RCRs is recommended to exclude possible contamination of product. Although previous studies have shown that RCRs were cleared when inoculated into normal and moderately immunosuppressed monkeys (Cornetta et al, 1991), a recent study reported RCR to be associated with development of lymphomas in severely immunosuppressed rhesus monkeys (Donahue et al, 1992). To evaluate the potential of RCR infection in humans, 4 normal, juvenile rhesus monkeys were inocualted with a vector virus preparation which contained RCR (provided by Genetic Therapy Inc., Gaithersburg, MD). The monkeys were monitored regularly for virus infection by 1) isolation of infectious MuLV from the PBMCs; 2) Western blot analysis for antibody response; and 3) DNA PCR for persistence of viral sequences in the host. Clinical changes were monitored by serum chemistry, hematology and physical examination. The results of a 3-year analysis idicate that all 4 animals were infected with RCR. There was early, transient virus isolation; persistence of viral DNA sequences in the PBMCs and long-term antibody response. Each animal responded uniquely with respect to the kinetics of infection and antibody response (manuscript in prep.). Current efforts are directed towards analyzing the state of the long-term persisting RCR sequences in the infected animals. We are studying the activation of the latent sequences for infectious virus using various known retrovirus inducers. These results will be important in evaluating the potential risk of RCR infection in man.