been tested extensively over several years for replicating retrovirus, which 
could theoretically develop by recorabination between vector and viral 
sequences in the packaging cell lines. To date, the multiple modifications in 
the vector and the packaging cell lines aimed at preventing recombination 
appear successful since replication virus has been detected only rarely after 
long periods of incubation. As a further check, each batch of supemate 
intended for clinical use is extensively screened for replication-competent virus. 
Even if this screening were to fail, data from primates have shown that intact 
MLV replicates poorly and transiently in primates in vivo even when these 
animals are injected with massive doses of virus. To date no clinical illness or 
sequelae has developed in these animals. 
Additional information exists in four animals exposed to high titer replication- 
competent retrovirus along with retroviral vector at the time of severe 
immunosuppression, and autologous bone marrow transplant. The 
transplanted monkeys were exposed to virus and vector at approximately equal 
titer and three of the four showed transient vector expression post transplant. 
Tests for replicating virus in serum, peripheral blood mononuclear cells, and 
bone marrow cells, performed two and three years post transplant have been 
negative. PCR performed on peripheral blood and bone marrow cells for the 
viral envelope region (sensitivity of detection 1 in 100,000 cells) were negative 
in all four animals. To date no animal has shown any clinical illness as a 
result of virus exposure. 
4.4 Insertional Mutagenesis 
The intact Moloney murine leukemia virus (MoMLV), the parent retrovirus 
of LNL-6, can cause neoplasia in mice. There is no evidence that this will be 
a problem in non-human primates or in humans. In 21 monkeys undergoing 
a retroviral-mediated gene transfer/autologous bone marrow transplantation 
protocol there has been no evidence of neoplasia (mean follow-up of 2.5 to 
4.5 years). There are also no known examples of transformation of primate 
cells by a murine retrovirus either in vitro or in vivo. Although clinical 
experience is limited, there has been no toxicity or pathology after 
approximately 72 patient-months in the TIL cell study. 
In fact, the mechanism by which the Moloney virus causes leukemia in mice 
is unlikely to be operative in man. The murine T-cell lymphoma associated 
with MoMLV is not due directly to MoMLV but is caused by a recombinant 
virus called a mink focus forming virus which results from recombination 
between MoMLV and endogenous murine leukemia virus sequences. These 
murine endogenous virus sequences are not present in humans. Moreover, 
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Recombinant DNA Research, Volume 14 
