Clinical trials with retroviral gene vectors are performed with extensively tested vector lots. The FDA recommends that sensitive tests that will detect replication competent forms of retrovirus be employed in the production of gene vectors. After patients are treated with gene vector preparations, the FDA recommends that the patients be monitored for replication competent retrovirus, so-called RCR, in patients. Several methods have potential utility. To determine the most informative type of assay to recommend to sponsors using retroviral vectors, the sensitivity of the current methods for patient monitoring will be analyzed and compared. Current recommendations include analysis of the presence of antibodies to replication competent retrovirus (RCR) using Western blot and PCR amplification to detect nucleic acids of RCR. The first objective of this set of experiments is to determine the sensitivity of these assays. Second, using selected primers for PCR the goal is to determine whether the primer sequences affect either sensitivity or specificity of the assay. Finally, the plan is to develop and validate primer sets that could be provided to sponsors to standardize assay performance.
