A Rev-Independent gag/pol Eliminates Detectable psi-gag Recombination in Lentiviral Vectors

Lentiviral vectors (LVs) are being developed for clinical use in humans for applications including gene therapy and immunotherapy. A safety concern for use of LVs in humans is the generation of replication-competent lentivirus (RCL), which may arise due to recombination between the split genomes of third-generation LVs. Although no RCL has been detected to date, design optimizations that minimize recombination events between split genome vectors would provide an added safety benefit that may further reduce the risk of RCL formation. Here we describe design elements introduced to the gag/pol plasmid with the intention of eliminating psi-gag recombination between the vector genome and gag/pol. These design changes, consisting of codon optimization of the gag/pol sequence and the deletion of the Rev-responsive element, abrogate the requirement for Rev in expression of Gag protein, thus the resulting gag/pol construct being Rev independent (RI gag/pol). We show that generating vector using the RI gag/pol construct has no effect on particle production or transduction titers. The RI and wild-type gag/pol vectors function equivalently as antigen-specific immunotherapy, potently inducing antigen-specific CD8 T cells that protect against challenge with vaccinia virus. Most importantly, the designed RI gag/pol eliminated detectable psi-gag recombination. Interestingly, we detected recombination between the vector genome and gag/pol from regions without sequence homology. Our findings imply that although unpredictable recombination events may still occur, the RI gag/pol design is sufficient to prevent psi-gag recombination

Emanuel Miller Lecture: Attachment insecurity, disinhibited attachment, and attachment disorders: where do research findings leave the concepts?

Background: Despite the evidence on anomalous attachment patterns, there has been a tendency to interpret most of these as reflecting differences in security/insecurity.Methods: Empirical research findings are reviewed in relation to attachment/insecurity as evident in both infancy and later childhood, disorganised attachment, inhibited attachment disorder, and disinhibited attachment disorder.Findings: Substantial differences are found in the correlates and meaning of these different features, as well as in the patterns associated with conditions such as autism, psychopathy, and Williams syndrome.Conclusions: It is seriously misleading to view all of these patterns through the lens of security/insecurity. This heterogeneity in social relationship features necessarily has implications for the assessment measures for social relationships that need to be used