Stable Integration of Transgenes Delivered by a Retrotransposon–Adenovirus Hybrid Vector

Helper-dependent adenoviruses show great promise as gene delivery vectors. However, because they do not integrate into the host chromosome, transgene expression cannot be maintained indefinitely. To overcome these limitations, we have inserted an L1 retrotransposon/transgene element into a helper-dependent adenovirus to create a novel chimeric gene delivery vector. Efficient adenovirus-mediated delivery of the L1 element into cultured human cells results in subsequent retrotransposition and stable integration of the transgene. L1 retrotransposition frequency was found to correlate with increasing multiplicity of infection by the chimeric vector, and further retrotransposition from newly integrated elements was not observed on prolonged culture. Therefore, this vector, which utilizes components of low immunogenic potential, represents a novel two-stage gene delivery system capable of achieving high titers via the initial helper-dependent adenovirus stage and permanent transgene integration via the retrotransposition stage.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63154/1/104303401750298571.pd

Characterization of Autosomal Dominant Hypercholesterolemia Caused by PCSK9 Gain of Function Mutations and its Specific Treatment with Alirocumab, a PCSK9 Monoclonal Antibody

BACKGROUND: Patients with PCSK9 gene gain of function (GOF) mutations have a rare form of autosomal dominant hypercholesterolemia. However, data examining their clinical characteristics and geographic distribution are lacking. Furthermore, no randomized treatment study in this population has been reported. METHODS AND RESULTS: -We compiled clinical characteristics of PCSK9 GOF mutation carriers in a multinational retrospective, cross-sectional, observational study. We then performed a randomized placebo-phase, double-blind study of alirocumab 150 mg administered subcutaneously every 2 weeks to 13 patients representing four different PCSK9 GOF mutations with low-density lipoprotein cholesterol (LDL-C) >70 mg/dL on their current lipid-lowering therapies at baseline. Observational study: Among 164 patients, 16 different PCSK9 GOF mutations distributed throughout the gene were associated with varying severity of untreated LDL-C levels. Coronary artery disease was common (33%; average age of onset 49.4 years) and untreated LDL-C concentrations were higher compared with matched carriers of mutations in the LDLR (n=2126) or apolipoprotein B (n=470) genes. Intervention study: In PCSK9 GOF mutation patients randomly assigned to receive alirocumab, mean percent reduction in LDL-C at 2 weeks was 62.5% (P<0.0001) from baseline, 53.7% compared to placebo-treated PCSK9 GOF mutation patients (P=0.0009; primary endpoint). After all subjects received 8 weeks of alirocumab treatment, LDL-C was reduced by 73% from baseline (P<0.0001). CONCLUSIONS: -PCSK9 GOF mutation carriers have elevated LDL-C levels and are at high risk for premature cardiovascular disease. Alirocumab, a PCSK9 antibody, markedly lowers LDL-C levels and appears to be well tolerated in these patients. Clinical Trial Registration-www.clinicaltrials.gov; Unique Identifier: NCT01604824These studies were sponsored by Regeneron Pharmaceuticals Inc. and Sanofi

Amplification dynamics of miniature inverted‐repeat transposable elements and their impact on rice trait variability

Transposable elements (TEs) are a rich source of genetic variability. Among TEs, miniature inverted-repeat TEs (MITEs) are of particular interest as they are present in high copy numbers in plant genomes and are closely associated with genes. MITEs are deletion derivatives of class II transposons, and can be mobilized by the transposases encoded by the latter through a typical cut-and-paste mechanism. However, MITEs are typically present at much higher copy numbers than class II transposons. We present here an analysis of 103 109 transposon insertion polymorphisms (TIPs) in 738 Oryza sativa genomes representing the main rice population groups. We show that an important fraction of MITE insertions has been fixed in rice concomitantly with its domestication. However, another fraction of MITE insertions is present at low frequencies. We performed MITE TIP-genome-wide association studies (TIP-GWAS) to study the impact of these elements on agronomically important traits and found that these elements uncover more trait associations than single nucleotide polymorphisms (SNPs) on important phenotypes such as grain width. Finally, using SNP-GWAS and TIP-GWAS we provide evidence of the replicative amplification of MITEs