中国药物警戒 ›› 2023, Vol. 20 ›› Issue (1): 27-33.
DOI: 10.19803/j.1672-8629.20220341

• 基因治疗产品安全性评价专栏 • 上一篇    下一篇

基因治疗药物AAV5-脂蛋白脂酶变异体在食蟹猴体内的毒性研究

侯田田1, 夏艳2∆, 潘东升1, 霍桂桃1, 马雪梅2, 刘子洋2, 孙立1, 刘艺2, 闫建奥2, 吴小兵2, 周晓冰1#, 刘国庆2,*, 耿兴超1   

  1. 1中国食品药品检定研究院国家药物安全评价监测中心,药物非临床安全评价研究北京市重点实验室,北京100176;
    2北京锦篮基因科技有限公司,北京100176
  • 收稿日期:2022-06-22 发布日期:2023-01-19
  • 通讯作者: *刘国庆,男,博士,教授·博导,脂代谢紊乱和动脉粥样硬化研究。E-mail: liuguoqing@bj-genecradle.com
    #为共同通信作者。
  • 作者简介:侯田田,女,硕士,助理研究员,药物临床前安全性评价。
    △为并列第一作者。
  • 基金资助:
    国家重点研发计划（2021YFA1101602）

Toxicity of gene therapy drug AAV5-LPLS447X (GC304) in cynomolgus monkeys

HOU Tiantian1, XIA Yan2∆, PAN Dongsheng1, HUO Guitao1, MA Xuemei2, LIU Ziyang2, SUN Li1, LIU Yi2, YAN Jian'ao2, WU Xiaobing2, ZHOU Xiaobing1#, LIU Guoqing2,*, GENG Xingchao1   

  1. 1National Center for Safety Evaluation of Drugs, National Institutes for Food and Drug Control, Beijing Key Laboratory for Safety Evaluation of Drugs, Beijing 100176, China;
    2Genecradle Therapeutics Inc, Beijing 100176, China
  • Received:2022-06-22 Published:2023-01-19

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摘要： 目的 评估基因治疗药物AAV5-脂蛋白脂酶变异体（GC304）在食蟹猴体内的毒性。方法 30只食蟹猴,分为溶媒对照组（静脉注射溶媒）、GC304低剂量组（3×1013 vg·kg-1）和高剂量组（1×1014 vg·kg-1）。试验期间,所有动物进行临床症状和注射部位观察,每周进行摄食量和体重测定。在给药前后不同时间点进行免疫原性检测（包含抗AAV5结合抗体、中和抗体及抗LPL结合抗体）、血液学、血清生化、血凝和尿生化检查,并同时进行T淋巴细胞分型、细胞因子、心电、血压、体温和眼科检查。给药后4周和6个月解剖,动物进行大体观察、脏器称重和组织病理学检查,同时进行GC304的生物分布研究及表达产物测定,并在不同时间点对血液中目的基因DNA进行检测。结果 受试物对食蟹猴临床症状、注射部位、体重、摄食量、眼睛各项指标、体温、血压、心电图、血凝、血清生化、T淋巴细胞分型、尿液、细胞因子、脏器重量等指标未见影响。高剂量组动物给药后4周血小板下降。与给予受试物相关的组织病理学改变为脾脏和腹股沟淋巴结生发中心活跃伴易染体巨噬细胞增多。低剂量和高剂量均可引起食蟹猴产生抗AAV5结合与中和抗体,给药后6个月2种抗体仍维持在较高的水平,但未见明显的剂量-效应关系。给予GC304后,未显著诱导食蟹猴产生抗LPL抗体。qPCR检测结果显示,受试物在食蟹猴外周血及脏器广泛分布和表达,但主要集中在肝脏。结论 食蟹猴单次静脉给予GC304,动物耐受性良好,未见明显毒性反应,且主要在肝脏中分布和表达,为该药物进入临床试验提供参考。

关键词: 基因治疗, AAV5-脂蛋白脂酶变异体（GC304）, 食蟹猴, 毒性研究

Abstract: Objective To determine the toxicity of gene therapy drug AAV5-LPLS447X (GC304) in cynomolgus monkeys. Methods Thirty cynomolgus monkeys were divided into the vehicle control group, low-dose group and high-dose group before being injected with vehicle, 3×1013 vg·kg-1 and 1×1014 vg·kg-1 GC304 respectively. During the study, All animals had their clinical symptoms and injection sites observed while food consumption and body weight were measured weekly. Immunogenicity tests (involving anti-AAV5-binding antibodies, anti-AAV5-neutralizing antibodies, and anti-LPL-binding antibodies), hematology, clinical chemistry, coagulation parameters and urinalysis were performed at different time points before and after administration, while T lymphocyte typing and cytokines, ECG, blood pressure, body temperature and ophthalmology were observed. At 4 weeks and 6 months after administration, cynomolgus monkeys were dissected for gross observation, organ weight and histopathological examination. At the same time, biodistribution of GC304 was studied and expression products were determination, and the target gene DNA in blood was detected at different time points. Results The clinical symptoms, injection sites, body weight, food consumption, eye indexes, body temperature, blood pressure, electrocardiogram, blood coagulation, clinical chemistry, T lymphocyte typing, urine, cytokines and organ weight were normal. Platelets in the high-dose group decreased 4 weeks after administration. Histopathological changes associated with the test article were active germinal centers in the spleen and inguinal lymph nodes with increased chromosomal macrophages. Both a low dose and a high dose caused cynomolgus monkeys to produce anti-AAV5 binding and anti-AAV5 neutralizing antibodies, and the two antibodies remained at high levels 6 months after administration, but there was no obvious “dose-effect” relationship. Anti-LPL antibody production in cynomolgus monkeys was not significantly induced after administration of GC304. The results of qPCR detection showed that the test article was widely distributed and expressed in the peripheral blood and organs of cynomolgus monkeys, but concentrated in the liver. Conclusion Cynomolgus monkeys can be singly dosed with GC304 via intravenous injection, which is well tolerated without obvious toxic reactions, and GC304 is mainly distributed and expressed in the liver, which can provide data for subsequent clinical trials.

Key words: gene therapy, GC304, cynomolgus monkeys, toxicity study

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