几种常用病毒载体在猕猴脑内的表达效率的比较

Abstract

Viral vector transfection systems are among the simplest of biological agents with the ability to transfer genes into the central nervous system. In brain research, a series of powerful and novel gene editing technologies are based on these systems. Although many viral vectors are used in rodents, their full application has been limited in non-human primates. To identify viral vectors that can stably and effectively express exogenous genes within non-human primates, eleven commonly used recombinant adeno-associated viral and lentiviral vectors, each carrying a gene to express green or red fluorescence, were injected into the parietal cortex of four rhesus monkeys. The expression of fluorescent cells was used to quantify transfection efficiency. Histological results revealed that recombinant adeno-associated viral vectors, especially the serotype 2/9 coupled with the cytomegalovirus, human synapsin Ⅰ, or Ca2+/calmodulin-dependent protein kinase Ⅱ promoters, and lentiviral vector coupled with the human ubiquitin C promoter, induced higher expression of fluorescent cells, representing high transfection efficiency. This is the first comparison of transfection efficiencies of different viral vectors carrying different promoters and serotypes in non-human primates (NHPs). These results can be used as an aid to select optimal vectors to transfer exogenous genes into the central nervous system of non-human primates.

AAV Gene Transfer with Tandem Promoter Design Prevents Anti-transgene Immunity and Provides Persistent Efficacy in Neonate Pompe Mice

“…Previous reports have shown the activity of the selected single transcriptional regulatory elements in large animal models and human cells;48, 50, 53 this is consistent with the observed activity of the LiMP in human hepatocyte and myoblasts. However, one limitation of the current work is the lack of studies in large animal models aimed at defining transgene persistence in neonate animals and therapeutic vector doses.…”

“…To achieve transgene expression in muscle, we selected the short synthetic spC5.12 promoter (C5.12),55, 56 which is active in both cardiac and skeletal muscle (type I and II fibers) and has been successfully used in large animal models of muscular dystrophy (Figure 1A; Figure S1). 50 For transgene expression in the CNS, we selected the pan-neuron human synapsin promoter (hSYN; Figure 1A; Figure S1) because it has been reported to be neuron-specific in both the brain and spinal cord in small and large animal models 51, 52, 53. Based on this, we used the full sequence of each tissue-specific promoter to generate three tandem promoters: the enhanced C5.12 promoter (Enh.C5.12, Apolipoprotein E [ApoE]-C5.12), the full liver-muscle promoter (LiMP, ApoE-hAAT-C5.12), and the full liver-neuron promoter (LiNeuP, ApoE-hAAT-hSYN) (Figure 1A).…”

“…Here we developed a novel tandem promoter expression cassette design that resulted in combined hepatic and extrahepatic tissue-specific transgene expression and prevented anti-transgene immune responses when used in the context of systemic AAV gene therapy. Tandem promoters were developed by rational design through multiplexing of selected tissue-specific regulatory elements widely reported to be active in hepatocytes,48, 49 muscle, 50 and neurons 51, 52, 53. The efficacy of AAV gene therapy using the tandem promoters was validated in the mouse model of PD.…”

A novel missense variant in ACAA1 contributes to early-onset Alzheimer’s disease, impairs lysosomal function, and facilitates amyloid-β pathology and cognitive decline

“… 47 The AAV overexpression system is safe for gene delivery to the brains of rodents, monkeys, and humans. 48 – 50 The AAV php.eb vectors expressing either green fluorescent protein (GFP) (AAV-Vector) or codon-optimized GFP-tagged human ACAA1 WT (AAV-ACAA1 WT) and ACAA1 p.N299S (AAV-ACAA1 N299S) under the control of the cytomegalovirus (CMV) promoter (Supplementaty Fig. S6a ) were bilaterally injected into the hippocampus region of presymptomatic APP/PS1ΔE9 mice and WT littermates (3-month-old), respectively.…”

AAV9-Retro mediates efficient transduction with axon terminal absorption and blood–brain barrier transportation

“…Non-human primates are deal experimental animals for human disease modeling [ 74 , 75 ], some recombinant adeno-associated viruses (rAAVs) can undergo retrograde transport in non-human primates, but the efficiency of retrograde transport is low, limiting the analysis of functional circuits and implications for disease modeling [ 76 ]. AAV2-Retro can mediate extensive retrograde transport in the rhesus macaque brain [ 77 ], but whether AAV9-Retro can achieve high-efficiency retrograde labelling in non-human primates brain is unknown.…”