Manufacturing Considerations

Although any consideration of how pharmaceutical companies would be able to manufacture gene therapy vectors was an irrelevant concern in the 1990s, this has now become a real issue. Retroviral vectors are biological agents that can only be made by living cells, and such systems are not easy for carrying out good manufacturing practice and quality assurance/quality control (QA/QC) assays. Furthermore, the large-scale production of retroviral vectors requires the establishment of producer cell lines that maintain a stable arrangement of vector sequences and sufficiently high vector production levels during the procedure. In addition, the subsequent purification of vectors from the supernatant of producer cells is a relatively cumbersome procedure and some loss of titer is inevitable. For vectors pseudotyped with the amphotropic MuLV Env, much of the loss of activity is due to the relative lability of the Env protein (the 2 subunits are noncovalently attached and can dissociate under conditions of sheer stress). In part, this problem could be countered by the use of a single polypeptide fusion protein such as VSV-G, although the inherent cytotoxicity of this protein makes the use of stable producer cell lines difficult (see Section II.C).

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