Objectives:Adenoviruses (Ads) are among the most used vectors for gene therapy; human Ad serotype 5-derived (HuAd5) vectors are the most frequently used for gene transfer applications. However, Ad5 infection is endemic in humans, and 20% of the Western population has neutralizing antibodies (nAbs). Pre-existing immunity against HuAd5 represents a major issue for many gene therapy applications. In our study, we evaluated several Ad serotypes derived from chimpanzees (ChAds) in vitro and in vivo to assess their transduction efficiency in various cell types and tissues. We aimed at identifying Ad serotypes able either to transduce “challenging” cell types or to represent a possible alternative to Ad5-derived vectors with comparable infectivity and tropism.Methods:We evaluated the efficacy of transduction of twelve ChAds vectors expressing enhanced green fluorescent protein (EGFP) in human embryonic kidney cells, as well as human leukemic and human mesenchymal stem cells, using flow cytometry to determine the percentage of EGFP-expressing cells and their mean fluorescent intensity (MFI). We observed the highest transduction efficiency in the serotype CV1 ChAd; therefore, we proceeded to evaluate toxicity and biodistribution in vivo.Results:After in vitro evaluation of twelve ChAds serotypes, we observed that the CV1 serotype was the most efficient in transducing both leukemia cell lines (HL-60 and NB-4) and human mesenchymal stem cells. Furthermore, in vivo analysis of the CV1 serotype induced an inflammatory reaction similar to what was observed after HuAd5 administration.Conclusions:ChAds vectors represent an effective alternative for the transduction of cells resistant to HuAd5 infection, such as mesenchymal stem cells and leukemic cells. In addition, we observed that the CV1 ChAd serotype presented a transduction profile similar to HuAd5 in vitro and induced a similar inflammatory response in vivo; therefore, CV1 ChAd-derived vectors represent an interesting alternative for gene therapy applications.
目的：腺病毒是目前基因治疗中最常用的载体之一，其中人腺病毒5型载体在基因转移应用中使用最为广泛。然而，腺病毒5型感染在人群中较为普遍，约20%的西方人群体内存在中和抗体。针对人腺病毒5型的预存免疫力是许多基因治疗方案面临的主要挑战。本研究通过体外与体内实验，评估了多种源自黑猩猩的腺病毒血清型在不同细胞和组织中的转导效率，旨在发现能够有效转导"难治性"细胞类型、或可作为腺病毒5型替代载体且具有相似感染性与趋向性的腺病毒血清型。
方法：我们通过流式细胞术检测了12种表达增强型绿色荧光蛋白的黑猩猩腺病毒载体在人胚胎肾细胞、人白血病细胞及人间充质干细胞中的转导效果，统计了绿色荧光蛋白阳性细胞百分比及其平均荧光强度。结果显示CV1血清型转导效率最高，随后我们对其进行了体内毒性及生物分布评估。
结果：对12种黑猩猩腺病毒血清型的体外实验表明，CV1血清型在转导白血病细胞系（HL-60和NB-4）及人间充质干细胞时效率最高。体内实验进一步显示，CV1血清型引发的炎症反应与人腺病毒5型给药后观察到的反应相似。
结论：黑猩猩腺病毒载体可作为有效替代方案，用于转导对人腺病毒5型感染具有抵抗性的细胞（如间充质干细胞和白血病细胞）。CV1黑猩猩腺病毒血清型在体外表现出与人腺病毒5型相似的转导特征，在体内诱导类似的炎症反应，因此CV1来源的载体有望成为基因治疗领域具有潜力的替代载体。
肿瘤(癌症)患者之家