Stability and Compatibility of Recombinant Adeno-Associated Virus Under Conditions Commonly Encountered in Human Gene Therapy Trials
Stability and Compatibility of Recombinant Adeno-Associated Virus Under Conditions Commonly Encountered in Human Gene Therapy Trials
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DOI：
10.1089/hgtb.2015.040
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发表时间：
2015-04-01
影响因子：
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通讯作者：
Flotte, Terence R.
中科院分区：
文献类型：
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作者：
Gruntman, Alisha M.;Su, Lin;Flotte, Terence R.
重组腺相关病毒（rAAV）载体正迅速成为人类基因治疗研究的首选，因为临床疗效已在几项人体试验中得到证实，概念验证数据已被证明用于纠正许多其他人。在FDA研究性新药（IND）申请的支持下进入人体使用时，有必要证明载体材料在人体使用时在各种储存、稀释和给药条件下的稳定性。目前文献中关于载体相容性和稳定性的数据有限，导致大多数IND申办者重复所有必要的研究。目前的研究解决了这个问题与rAAV载体（rAAV 1-CB-chAATmyc）含有AAV 2反向末端重复序列包装到AAV 1衣壳。将载体的等分试样暴露于各种温度、稀释剂、容器成分和其它环境条件，并且通过在小鼠中肌内注射后测量转基因表达来评估其功能性生物活性（在这些各种处理后）。发现rAAV在4 ℃至55 ℃的温度范围内（在70 ℃下20分钟后仅部分丧失效力），在5.5至8.5的pH范围内，在与小鼠或人血清（有或没有补体耗尽）或与钆接触后，以及在与玻璃、聚苯乙烯、聚乙烯、聚丙烯和不锈钢接触后，非常稳定。导致载体活性几乎完全丧失（10，000倍损失）的唯一暴露是UV暴露10 min。rAAV 1制剂的稳定性预示着这种治疗方式的未来传播。
Recombinant adeno-associated virus (rAAV) vectors are rapidly becoming the first choice for human gene therapy studies, as clinical efficacy has been demonstrated in several human trials and proof-of-concept data have been demonstrated for correction of many others. When moving into human use under the auspices of an FDA Investigational New Drug (IND) application, it is necessary to demonstrate the stability of vector material under various conditions of storage, dilution, and administration when used in humans. Limited data are currently available in the literature regarding vector compatibility and stability, leading most IND sponsors to repeat all necessary studies. The current study addresses this issue with an rAAV vector (rAAV1-CB-chAATmyc) containing AAV2-inverted terminal repeat sequences packaged into an AAV1 capsid. Aliquots of vector were exposed to a variety of temperatures, diluents, container constituents, and other environmental conditions, and its functional biological activity (after these various treatments) was assessed by measuring transgene expression after intramuscular injection in mice. rAAV was found to be remarkably stable at temperatures ranging from 4 degrees C to 55 degrees C (with only partial loss of potency after 20 min at 70 degrees C), at pH ranging from 5.5 to 8.5, after contact with mouse or human serum (with or without complement depletion) or with gadolinium and after contact with glass, polystyrene, polyethylene, polypropylene, and stainless steel. The only exposure resulting in near-total loss of vector activity (10,000-fold loss) was UV exposure for 10 min. The stability of rAAV1 preparations bodes well for future dissemination of this therapeutic modality.