Recombinant DNA Advisory Committee - 09/14-15/92 
The LN vector has probably been safety tested more extensively than any of the other 
vectors reviewed to date. LN has been "ping-ponged" between various helper virus 
sensitive cell lines up to eight times. No detectable helper virus was demonstrated in 
these experiments. One of the older vectors, N2, produced helper virus within two 
passages. In addition, LN producing cell lines have been cultured for several weeks 
following supernatant harvest and no detectable helper virus was observed. Targeted 
Genetics, Inc., assays are only representative of helper virus. 
Dr. Murray inquired if the investigators were assaying for helper virus by the extended 
S + /L‘ assay. Dr. D. Miller said that although they are not performing the extended 
S + /L' assay, their safety testing is actually more stringent than other assay methods. The 
packaging cells are kept in continuous culture for up to four weeks following vector 
harvest. With this method, any replication competent particles originally present should 
be amplified to extremely high levels that would be readily detected by PCR. Dr. D. 
Miller noted that they have recently started to perform the Mus Dunni assay also on 
vector preparations. The viral stocks have been prepared, and assay are in storage. 
Helper virus has never been demonstrated with the PA317 cell line or in the co- 
cultivation and long-term marrow studies. 
Dr. Carmen submitted revised language for incorporation into the informed consent 
document. The word "bacterial" should be inserted prior to the word "gene" so that the 
patient will understand the source of that gene. 
Dr. Leventhal asked if the twin protocol requires autologous bone marrow (ABM) 
reinfusion or does it apply only to peripheral blood stem cells. Dr. Shuening answered 
that the twin study pertains only to the infusion of peripheral blood stem cells. 
Dr. Geiduschek inquired as to the number of transducing particles that are produced per 
producer cell. Dr. D. Miller responded that between one and ten particles of virus are 
produced per cell per day. This number of particles translates to approximately between 
10 6 and 10 7 colony forming units (CFUs) per milliliter. There are approximately 25 
milliliters per flask. 
Committee Motion 
A motion was made by Dr. Geiduschek to approve the protocol subject to a redefinition 
of the safety of the LN particles that verify that no helper virus is present in the 
equivalent of 300 milliliters of supernatant. Dr. Post suggested that the wording be 
changed to less than one particle per 300 milliliters. The investigators will be obligated 
to provide evidence of this data without actually testing all 300 milliliters of supernatant. 
This data should be obtained by extended culture of the producer cell. 
Recombinant DNA Research, Volume 16 
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