Gene therapy continues to be a promising contender for the treatment of cystic fibrosis (CF) airway disease. We have previously demonstrated that airway conditioning with lysophosphatidylcholine (LPC) followed by delivery of a HIV-1–based lentiviral (LV) vector functionally corrects the CF transmembrane conductance regulator (CFTR) defect in the nasal airways of CF mice. In our earlier pilot study we showed that our technique can transduce marmoset lungs acutely; this study extends that work to examine gene expression in this nonhuman primate (NHP) 1 month after gene vector treatment. A mixture of three separate HIV-1 vesicular stomatitis virus G (VSV-G)–pseudotyped LV vectors containing the luciferase (Luc), LacZ, and hCFTR transgenes was delivered into the trachea through a miniature bronchoscope. We examined whether a single-dose delivery of LV vector after LPC conditioning could increase levels of transgene expression in the trachea and lungs compared with control (phosphate-buffered saline [PBS]) conditioning. At 1 month, bioluminescence was detected in vivo in the trachea of three of the six animals within the PBS control group, compared with five of the six LPC-treated animals. When examined ex vivo there was weak evidence that LPC improves tracheal Luc expression levels. In the lungs, bioluminescence was detected in vivo in four of the six PBS-treated animals, compared with five of the six LPC-treated animals; however, bioluminescence was present in all lungs when imaged ex vivo. LacZ expression was predominantly observed in the alveolar regions of the lung. hCFTR was detected by qPCR in the lungs of five animals. Basal cells were successfully isolated and expanded from marmoset tracheas, but no LacZ-positive colonies were detected. There was no evidence of an inflammatory response toward the LV vector at 1 month postdelivery, with cytokines remaining at baseline levels. In conclusion, we found weak evidence that LPC conditioning improved gene transduction in the trachea, but not in the marmoset lungs. We also highlight some of the challenges associated with translational lung gene therapy studies in NHPs.

中文翻译：

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对囊性纤维化慢病毒气道基因递送的人类翻译：在狨猴中进行的为期一个月的 CFTR 和报告基因研究

基因疗法仍然是治疗囊性纤维化 (CF) 气道疾病的有希望的竞争者。我们之前已经证明，使用溶血磷脂酰胆碱 (LPC) 进行气道调节，然后递送基于 HIV-1 的慢病毒 (LV) 载体可以在功能上纠正 CF 小鼠鼻气道中的 CF 跨膜电导调节器 (CFTR) 缺陷。在我们早期的试点研究中，我们表明我们的技术可以敏锐地传导狨肺。这项研究扩展了这项工作，以检查基因载体处理后 1 个月在这种非人类灵长类动物 (NHP) 中的基因表达。三种单独的 HIV-1 水疱性口炎病毒 G (VSV-G) 的混合物——含有荧光素酶(Luc)、LacZ和hCFTR的假型 LV 载体转基因通过微型支气管镜进入气管。我们检查了与对照（磷酸盐缓冲盐水 [PBS]）调理相比，LPC 调理后单剂量递送 LV 载体是否可以增加气管和肺中的转基因表达水平。在 1 个月时，在 PBS 对照组的六只动物中的三只的气管中检测到体内生物发光，而六只 LPC 治疗的动物中有五只。当离体检查时，有微弱的证据表明 LPC 改善了气管 Luc 表达水平。在肺中，在体内检测到生物发光在六只 PBS 处理的动物中的四只中，与六只 LPC 处理的动物中的五只相比；然而，在离体成像时，所有肺部都存在生物发光。LacZ 表达主要在肺的肺泡区域观察到。hCFTR通过 qPCR 在五只动物的肺中检测到。从狨猴气管中成功分离和扩增基底细胞，但未检测到 LacZ 阳性菌落。在分娩后 1 个月，没有证据表明对 LV 载体有炎症反应，细胞因子保持在基线水平。总之，我们发现弱证据表明 LPC 调节改善了气管中的基因转导，但在狨猴肺中没有。我们还强调了与 NHP 中的肺转化基因治疗研究相关的一些挑战。