DataSheet_1_rAAV immunogenicity, toxicity, and durability in 255 clinical trials: A meta-analysis.xlsx

Recombinant Adeno-associated virus (rAAV) is one of the main delivery vectors for gene therapy. To assess immunogenicity, toxicity, and features of AAV gene therapy in clinical settings, a meta-analysis of 255 clinical trials was performed. A total of 7,289 patients are planned to be dosed. AAV2 was the most dominantly used serotype (29.8%, n=72), and 8.3% (n=20) of trials used engineered capsids. 38.7% (n=91) of trials employed neutralizing antibody assays for patient enrollment, while 15.3% (n=36) used ELISA-based total antibody assays. However, there was high variability in the eligibility criteria with cut-off tiers ranging from 1:1 to 1:1,600. To address potential immunogenicity, 46.3% (n=118) of trials applied immunosuppressants (prophylactic or reactive), while 32.7% (n=18) of CNS and 37.5% (n=24) of ocular-directed trials employed immunosuppressants, possibly due to the immune-privileged status of CNS and retina. There were a total of 11 patient deaths across 8 trials, and 18 out of 30 clinical holds were due to toxicity findings in clinical studies. 30.6% (n=78) of trials had treatment-emergent serious adverse events (TESAEs), with hepatotoxicity and thrombotic microangiopathy (systemic delivery) and neurotoxicity (CNS delivery) being the most prominent. Additionally, the durability of gene therapy may be impacted by two distinct decline mechanisms: 1) rapid decline presumably due to immune responses; or 2) gradual decline due to vector dilution. The durability varied significantly depending on disease indication, dose, serotypes, and patient individuals. Most CNS (90.0%) and muscle trials (73.3%) achieved durable transgene expression, while only 43.6% of ocular trials had sustained clinical outcomes. The rAAV production system can affect rAAV quality and thus immunogenicity and toxicity. Out of 186 trials that have disclosed production system information, 63.0% (n=126) of trials used the transient transfection of the HEK293/HEK293T system, while 18.0% (n=36) applied the baculovirus/Sf9 (rBac/Sf9) system. There were no significant differences in TESAEs and durability between AAV generated by rBac/Sf9 and HEK293/HEK293T systems. In summary, rAAV immunogenicity and toxicity poses significant challenges for clinical development of rAAV gene therapies, and it warrants collaborative efforts to standardize monitoring/measurement methods, design novel strategies to overcome immune responses, and openly share relevant information.

重组腺相关病毒（recombinant Adeno-associated virus, rAAV）是基因治疗的主要递送载体之一。为评估AAV基因治疗在临床场景中的免疫原性、毒性及相关特征，本研究对255项临床试验开展了荟萃分析。计划纳入的给药患者总计7289例。AAV2是最常用的血清型（占比29.8%，n=72），另有8.3%（n=20）的试验采用了工程化衣壳。38.7%（n=91）的试验采用中和抗体检测进行患者入组，另有15.3%（n=36）的试验采用基于酶联免疫吸附试验（Enzyme-Linked Immunosorbent Assay, ELISA）的总抗体检测方案。但入组标准的截断值差异显著，范围跨度从1:1至1:1600。为应对潜在的免疫原性，46.3%（n=118）的试验使用了免疫抑制剂（预防性或反应性给药）；其中中枢神经系统（Central Nervous System, CNS）试验中有32.7%（n=18）、眼部靶向试验中有37.5%（n=24）采用了免疫抑制剂策略，这可能与中枢神经系统和视网膜的免疫豁免特性相关。共有8项试验出现了11例患者死亡病例，30项临床暂停中有18项是因临床研究中观察到毒性结果所致。30.6%（n=78）的试验出现了治疗突发严重不良事件（treatment-emergent serious adverse events, TESAEs），其中最为突出的不良事件包括肝毒性、血栓性微血管病（全身性递送载体）以及神经毒性（中枢神经系统递送载体）。此外，基因治疗的持久性可能受到两种截然不同的衰退机制影响：1）推测由免疫反应介导的快速衰退；或2）由载体稀释导致的缓慢衰退。持久性因疾病适应症、给药剂量、血清型以及患者个体差异而存在显著异质性。多数中枢神经系统试验（90.0%）和肌肉组织试验（73.3%）实现了持久的转基因表达，而仅有43.6%的眼部试验获得了持续的临床结局。重组腺相关病毒的生产系统会影响其产品质量，进而影响免疫原性与毒性。在186项披露了生产系统信息的试验中，63.0%（n=126）的试验采用了HEK293/HEK293T细胞瞬时转染系统，另有18.0%（n=36）采用了杆状病毒/Sf9（rBac/Sf9）表达系统。采用rBac/Sf9系统与HEK293/HEK293T系统生产的AAV，在治疗突发严重不良事件发生率与转基因表达持久性方面均无显著差异。综上，重组腺相关病毒的免疫原性与毒性为重组腺相关病毒基因治疗的临床开发带来了重大挑战，亟需通过协同合作统一监测与检测方法、设计新型策略以克服免疫反应，并公开共享相关研究数据与信息。