Efficacy and Safety of Clinical-Grade Human Vascular Endothelial Growth Factor-DΔNΔC Gene Therapy Containing Residual Replication-Competent Adenoviruses

Biological bypass through induced angiogenesis by vascular endothelial growth factor D (VEGF-D) gene therapy (GT) is a new concept for the treatment of cardiac ischemia. Serotype 5 adenoviruses are used in the clinical trials for transferring the VEGF-D cDNA into the ischemic myocardium. However, the presence of replication-competent vectors in the adenovirus products is a widely recognized problem that may pose a potential safety risk to the treated patients. We compared three different VEGF-D GT production lots containing different levels of replication-competent adenoviruses (RCA) tested in 3 x 10(10) viral particles (vp): <10 RCA (VEGF-D L-RCA1), 10-100 RCA (VEGF-D H-RCA2), and 100-200 RCA (VEGF-D H-RCA3), as measured by a novel droplet digital polymerase chain reaction (PCR) RCA assay in a preclinical rabbit model (n = 21). beta-galactosidase encoding nonclinical-grade preparation was used as a nonangiogenic control. Each preparation was injected into the right semimembranosus muscle using dose of 1 x 10(11) vp. Efficacy of the products was tested by the combination of contrast pulse sequencing ultrasound and modified Miles assay as well as quantifying the total cross-sectional area of capillaries. Safety, immunogenicity, toxicity, biodistribution, and shedding were assessed by general histology, serial measurements of C-reactive protein, white blood cell count and body temperature as well as using quantitative real-time PCR with primers targeted to the VEGF-D and replication-permitting E1 sequences. We found no significant differences in the efficacy or safety between the study groups. Most importantly, no detectable presence of RCA-specific E1 sequence was found in any samples tested, indicating that no detectable vector replication took place in vivo. We conclude that relatively low levels of RCA in adenoviral GT products may not be as important major safety issue as previously anticipated.

Automated summary

This study compared three different levels of replication-competent adenoviruses (RCA) in VEGF-D gene therapy production lots in a preclinical rabbit model and found no significant differences in efficacy or safety between the groups. Importantly, no detectable presence of RCA-specific E1 sequence was found in any samples tested, indicating that no detectable vector replication occurred in vivo. This suggests that relatively low levels of RCA in adenoviral gene therapy products may not pose a major safety concern as previously thought.