10. Initial Critique and Responses B . This criteria will make this trial not comparable to other trials in RA, which require six active joints. The primary purpose of this study is to evaluate the safety of intra-articular injections of tgAAC94. It is not necessary to change the entry criteria to match those used in other clinical trials, because there are no plans to compare the results to other trials .
11. Initial Critique and Responses C . This criteria will decrease the ability to detect responses in other joints (“contralateral effect”). [Other TNF-inhibitor] trials were conducted in patients with active RA involving multiple joints, hence the requirement for involvement of at least six joints. In contrast, this trial is aimed at patients who have reasonably well-controlled disease in most joints, but one or two persistently inflamed and swollen joints which are at risk for cumulative damage.
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13. Initial Critique and Responses In the non-human primate model, after a single intra-articular injection, biodistribution of vector DNA was detected in the spleen , left (non-injected) femoral tibial joint tissues and right iliac lymph node by the limit of detection PCR assay (1 in 104 cells to 1 in 103 cells). At the intra-articular injection site (bone, cartilage, synovium, tendon and ligament) gene transfer was documented in a quantitative DNA PCR assay and the amount of vector DNA was dose dependent, with the highest amounts being … 6000 copies/cell. In contrast, while vector DNA in the contralateral joint was quantitated in this assay, the amount of vector was not dose dependent and the highest amount of vector DNA (1.2 copies/cell) was three orders of magnitude lower than at the injection site. Human TNFR:Fc transgene mRNA was present at the injection site using a qualitative RNA-specific PCR assay. In samples from non-injection sites there was no consistent detectable expression, particularly in the spleen and lymph node. II A . Studies did show DNA in other organs, suggesting systemic spread of the virus.
14. Initial Critique and Responses In summary, the exact mechanism of the observed systemic effect after local intraarticular delivery (in the experimentally induced rat model) is not completely understood . We do not know if we will observe a similar systemic effect after intra-articular delivery in the proposed clinical trial. As mentioned above, recent clinical trials of intra-articular administration of etanercept at doses of up to 8 mg per injection were considered safe and did not result in any systemic effect (Bliddal et al., 2002). However, we plan to measure the circulating levels of TNFR:Fc protein. II B . The trials in rodents demonstrated an effect of inoculation into one joint on disease in the contralateral joint.
15. Initial Critique and Responses Our current pre-clinical data supports equivalent efficacy with both approaches in the rat SCW model, despite very different levels of circulating, systemic TNFR:Fc (approximately 2 log difference in serum TNFR:Fc). The benefit of intra-articular route of delivery is the potentially reduced exposure to systemic TNFR:Fc protein compared to either etanercept or intramuscular delivery of a rAAV-TNFR:Fc vector. II C . Ultimately, if this therapy appears successful, one question will be whether there is any benefit to intra-articular therapy over IM.
16. III. [The] protein that will be expressed, is immunogenic, despite having a human TNF-R fragment and the Fc portion of human IgG1. The hinge region, the junction between the two components, is thought to be the target. Perhaps the inflammation at the site of expression of the fusion construct will have an adjuvant-like effect. There is a risk that treatment with the active agent could induce antibodies that would reduce the efficacy of etanercept. Initial Critique and Responses
17. Initial Critique and Responses If we can develop and validate a reliable assay to detect antibodies to the human TNFR:Fc protein we will test the serum or joint fluid collected during the trial. III . [The] protein that will be expressed, is immunogenic…
18. IV. For some chimeric proteins, efficacy has been linked to particular allotypes of the FcR, which alter affinity. Initial Critique and Responses
19. Initial Critique and Responses This is a small study focused on safety. We will take this suggestion into consideration for future trials. IV . For some chimeric proteins, efficacy has been linked to particular allotypes of the FcR, which alter affinity.
20. V. There is theoretical possibility that adenovirus could reach the joint and rescue the AAV. There is no evidence that this is a problem from previous studies with AAV. Should there be restrictions on participation in this trial in subjects with RA and upper respiratory symptoms? Initial Critique and Responses
21. Initial Critique and Responses Subjects who are acutely ill should be excluded from the protocol under the last exclusion criterion, which includes “other concurrent medical condition that, in the opinion of the Investigator, would make the subject unsuitable for the study”. We do not believe that it is necessary to exclude subjects with upper respiratory symptoms , per se, because the tgAAC94 particles are non-replicating as they lack the wild-type AAV viral genes ( rep and cap ) required for viral replication and encapsidation of DNA. Thus, even in the presence of a helper virus infection (adenovirus 5) the particles are not capable of replication. Simultaneous joint infection with both wild-type AAV (containing rep and cap ) and adenovirus 5 is required to rescue the vector. V . … theoretical possibility that adenovirus could reach the joint and rescue the AAV.
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23. Initial Critique and Responses Although most patients will likely be receiving methotrexate, it is not necessary to standardize baseline treatment, because this is primarily a safety study . … However, baseline medications and changes in medications will be recorded , so it will be possible to perform exploratory retrospective analyses about at the conclusion of the trial to see if the “strength” of the DMARD at baseline, or addition of other medications has an impact on the outcome of the study. VI . DMARD selection criteria.
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