AAV载体基因药物临床生物样本分析要点及挑战

摘要/Abstract

摘要： 细胞和基因治疗已进入高速发展期，多种病毒和非病毒载体被应用于细胞和基因治疗领域。其中，腺相关病毒（adeno-associated virus，AAV）载体因其对治疗基因的高效转导率、持久的表达能力和良好的安全性等特点，在遗传病的体内基因治疗领域得到广泛应用。随着越来越多的AAV载体基因药物进入临床研究阶段，AAV载体药物的生物分析也越来越受关注，涉及药物代谢动力学（pharmacokinetics，PK）、药效学（pharmacodynamics，PD）、免疫原性、病毒脱落和基因整合监测等方面，目前均缺乏标准化的分析方法、标准品和对照品，在分析方法验证方案、接受标准等方面亦尚无统一规定。囿于AAV载体药物组成和作用机制的复杂性，无论是分析方法、结果解释，还是监管机构的建议和要求等方面均不同于传统小分子和蛋白药物。本研究结合AAV载体基因药物特点、监管机构建议和已发表的临床研究，概述AAV载体基因药物临床生物样本给药前、给药后分析工作要点及面临的挑战，以期为临床上AAV载体基因治疗药物的研究开发提供借鉴和参考。

关键词: font-size:medium, ">AAV载体；基因治疗；生物分析；临床试验

Abstract: Cell and gene therapy has entered a period of rapid development， and a variety of viral and non-viral vectors have been applied to the field of cell and gene therapy. Among them， adeno-associated virus(AAV) vectors have been widely used in the field of in vivo gene therapy for genetic diseases because of their potent transduction efficiency， durable expression ability and good safety. As more and more AAV vector gene therapy products enter the clinical trials， the bioanalysis related to AAV vector has also attracted more and more attention， which involves pharmacokinetics (PK)， pharmacodynamics (PD)， immunogenicity， viral shedding and gene integration monitoring， etc. At present， there are no standardized analytical methods， standards and reference materials， and there are no uniform regulations on the verification scheme and acceptance standard of analytical methods.Due to the complexity of the structure and mechanism of AAV vector products， the bioanalytic methods， interpretation of results or regulatory recommendations is different from the bioanalysis of traditional small molecule and protein drugs. This study overviewed the key points and challenges of clinical bioanalysis products before and after administration of AAV vectors gene therapy based on the characteristics of AAV vector， the current regulatory recommendations and the published data of clinical trails， to provide reference for the development of AAV vector gene therapy in clinic.

Key words: font-size:medium, ">AAV vector； Gene therapy； Bioanalysis； Clinical trial

中图分类号:

余双庆, 吴小兵. AAV载体基因药物临床生物样本分析要点及挑战[J]. 中国医药导刊, 2023, 25(7): 669-676.

YU Shuangqing, WU Xiaobing. The Challenges and Key-points of Clinical Bioanalysis of AAV Vector Gene Therapy[J]. CHINESE JOURNAL OF MEDICINAL GUIDE, 2023, 25(7): 669-676.

参考文献

[1]Wang D， Tai PWL， Gao G. Adeno-associated virus vector as a platform for gene therapy delivery[J]. Nat Rev Drug Discov， 2019， 18(5)：358-378.
     [2]Klamroth R， Hayes G， Andreeva T， et al. Global seroprevalence of pre-existing immunity against AAV5 and other AAV serotypes in people with hemophilia A[J]. Hum Gene Ther， 2022， 33(7-8)：432-441.
     [3]Leborgne C， Latournerie V， Boutin S， et al. Prevalence and long-term monitoring of humoral immunity against adeno-associated virus in duchenne muscular dystrophy patients[J]. Cell Immunol， 2019， 342： 103780.
     [4]Fitzpatrick Z， Leborgne C， Barbon E， et al. Influence of pre-existing anti-capsid neutralizing and binding antibodies on AAV vector transduction[J]. Mol Ther Methods Clin Dev， 2018， 9：119-129.
     [5]Louis Jeune V， Joergensen JA， Hajjar RJ， et al. Pre-existing anti-adeno-associated virus antibodies as a challenge in AAV gene therapy[J]. Hum Gene Ther Methods， 2013， 24(2)：59-67.
     [6]Wang L， Calcedo R， Bell P， et al. Impact of pre-existing immunity on gene transfer to nonhuman primate liver with adeno-associated virus 8 vectors[J]. Hum Gene Ther， 2011， 22(11)：1389-1401.
     [7]FDA. Human Gene Therapy for Neurodegenerative Diseases. Draft Guidance for Industry[EB/OL]. (2021) [2023-05-05]. https：//www. fda. gov/ media/ 144886/download.
     [8]EMA. Reflection paper on quality， non-clinical and clinical issues related to the development of recombinant adenoassociated viral vectors[EB/OL]. (2010) [2023-05-05]. https：//www. ema. europa. eu/en/documents/scientific-guideline/reflection-paper-quality-non-clinical-clinical-issues-related-development-recombinant-adeno_en. pdf.
     [9]FDA. Considerations for the design of early-phase clinical trials of cellular and gene therapy products[EB/OL]. (2015) [2023-05-05]. https：//www. fda. gov/regulatory-information/search-fda-guidance-documents/considerations-design-early-phase-clinical-trials-cellular-and-gene-therapy-products.
     [10]国家药品监督管理局药品审评中心. 基因治疗血友病临床试验设计技术指导原则[S]. 2023.
     [11]Falese L， Sandza K， Yates B， et al. Strategy to detect pre-existing immunity to AAV gene therapy[J]. Gene Ther， 2017， 24(12)：768-778.
     [12]Gorovits B， Azadeh M， Buchlis G， et al. Evaluation of the humoral response to adeno-associated virus-based gene therapy modalities using total antibody assays[J]. AAPS J， 2021， 23(6)：108.
     [13]Kuranda K， Alphonse PJ， Leborgne C， et al. Exposure to wild-type AAV drives distinct capsid immunity profiles in humans[J]. J Clin Invest， 2018， 128(12)：5267-5279.
     [14]Manno CS， Arruda VR， Pierce GF， et al. Successful transduction of liver in hemophilia by AAV-Factor IX and limitations imposed by the host immune response[J]. Nat Med， 2006， 12(3)：342-347.
     [15]Mingozzi F， High KA. Overcoming the host immune response to adeno-associated virus gene delivery vectors：the race between clearance， tolerance， neutralization， and escape[J]. Annu Rev Virol， 2017， 4(1)：511-534.
     [16]Mingozzi F， Maus MV， Hui DJ， et al. CD8(+) T-cell responses to adeno-associated virus capsid in humans[J]. Nat Med， 2007， 13(4)：419-422.
     [17]Berrier KL， Kazi ZB， Prater SN， et al. CRIM-negative infantile Pompe disease： characterization of immune responses in patients treated with ERT monotherapy[J]. Genet Med， 2015， 17(11)：912-918.
     [18]Chen N， Sun KF， Chemuturi NV， et al. The perspective of DMPK on recombinant adeno-associated virus-based gene therapy：past learning， current support， and future contribution[J]. AAPS J， 2022， 24(1)：31.
     [19]FDA. Preclinical assessment of investigational cellular and gene therapy products[EB/OL]. (2013) [2023-05-05]. https：//www. fda. gov/regulatory-information/search-fda-guidance-documents/preclinical-assessment-investigational-cellular-and-gene-therapy-products.
     [20]国家药品监督管理局药品审评中心. 基因治疗产品非临床研究与评价技术指导原则（试行）[S]. 2021.
     [21]Chowdhury EA， Tetanget GM， Chang HY， et al. Current progress and limitations of AAV mediated delivery of protein therapeutic genes and the importance of developing quantitative pharmacokinetic/pharmacodynamic (PK/PD) models[J]. Adv Drug Deliv Rev， 2021， 170： 214-237.
     [22]FDA. Rare diseases：common issues in drug development Guidance for Industry[EB/OL]. (2019) [2023-05-05]. https：//www. fda. gov/regulatory-information/search-fda-guidance-documents/rare-diseases-common-issues-drug-development-guidance-industry.
     [23]Gorovits B， Marshall JC， Smith J， et al. Bioanalysis of adeno-associated virus gene therapy therapeutics： regulatory expectations[J]. Bioanalysis， 2019， 11(21)：2011-2024.
     [24]Considerations I. General principles to address virus and vector shedding[EB/OL]. (2009) [2023-05-05]. https：//www. ema. europa. eu/en/documents/scientific-guideline/international-conference-harmonisation-technical-requirements-registration-pharmaceuticals-human-use_en-10. pdf.
     [25]FDA. Design and analysis of shedding studies for virus or bacteria-based gene therapy and oncolytic products. Guidance for industry[EB/OL]. (2015) [2023-05-05]. https：//www. fda. gov/regulatory-information/search-fda-guidance-documents/design-and-analysis-shedding-studies-virus-or-bacteria-based-gene-therapy-and-oncolytic-products.
     [26]国家药品监督管理局药品审评中心. 基因治疗产品长期随访临床研究技术指导原则（试行）[S]. 2021.
     [27]Gorovits B， Azadeh M， Buchlis G， et al. Evaluation of cellular immune response to adeno-associated virus-based gene therapy[J]. AAPS J， 2023， 25(3)：47.
     [28]Zhu J， Huang X， Yang Y. The TLR9-MyD88 pathway is critical for adaptive immune responses to adeno-associated virus gene therapy vectors in mice[J]. J Clin Invest， 2009， 119(8)：2388-2398.
     [29]Rogers GL， Martino AT， Aslanidi GV， et al. Innate immune responses to AAV vectors[J]. Front Microbiol， 2011， 2： 194.
     [30]Muhuri M， Maeda Y， Ma H， et al. Overcoming innate immune barriers that impede AAV gene therapy vectors[J]. J Clin Invest， 2021， 131(1)：e143780.
     [31]Srivastava A. AAV vectors： are they safe?[J]. Hum Gene Ther， 2020， 31(13-14)：697-699.
     [32]Wilson JM， Flotte TR. Moving forward after two deaths in a gene therapy trial of myotubular myopathy[J]. Hum Gene Ther， 2020， 31(13-14)：695-696.
     [33]Stone D， Liu Y， Li ZY， et al. Biodistribution and safety profile of recombinant adeno-associated virus serotype 6 vectors following intravenous delivery[J]. J Virol， 2008， 82(15)：7711-7715.
     [34]Russell S， Bennett J， Wellman JA， et al. Efficacy and safety of voretigene neparvovec (AAV2-hRPE65v2) in patients with RPE65-mediated inherited retinal dystrophy： a randomised， controlled， open-label， phase 3 trial[J]. Lancet， 2017， 390(10097)：849-860.
     [35]FDA. Human Gene Therapy for Retinal Disorders Guidance for Industry[EB/OL]. (2009) [2023-05-05]. https：// www. fda. gov/ media/ 124641/ download.
     [36]EMA. Guideline on follow-up of patients administered with gene therapy medicinal products[EB/OL]. (2009) [2023-05-05]. https：//www. ema. europa. eu/en/documents/scientific-guideline/guideline-follow-patients-administered-gene-therapy-medicinal-products_en.pdf.
     [37]FDA. Human Gene Therapy for Hemophilia Guidance for Industry[EB/OL]. (2020) [2023-05-05]. https：// www. fda. gov/ media/ 113799/ download.
     [38]FDA. Human Gene Therapy for Rare Diseases Guidance for Industry[EB/OL]. (2020) [2023-05-05]. https：// www. fda. gov/ media/ 113807/ download.
     [39]Long BR， Veron P， Kuranda K， et al. Early phase clinical immunogenicity of valoctocogene roxaparvovec， an AAV5-mediated gene therapy for hemophilia A[J]. Mol Ther， 2021， 29(2)：597-610.
     [40]Mendell JR， Campbell K， Rodino-Klapac L， et al. Dystrophin immunity in Duchenne′s muscular dystrophy[J]. N Engl J Med， 2010， 363(15)：1429-1437.
     [41]Vandamme C， Adjali O， Mingozzi F. Unraveling the complex story of immune responses to AAV vectors trial after trial[J]. Hum Gene Ther， 2017， 28(11)：1061-1074.
     [42]Ferreira V， Twisk J， Kwikkers K， et al. Immune responses to intramuscular administration of alipogene tiparvovec (AAV1-LPL(S447X)) in a phase II clinical trial of lipoprotein lipase deficiency gene therapy[J]. Hum Gene Ther， 2014， 25(3)：180-188.
     [43]Gil-Farina I， Fronza R， Kaeppel C， et al. Recombinant AAV integration is not associated with hepatic genotoxicity in nonhuman primates and patients[J]. Mol Ther， 2016， 24(6)：1100-1105.
     [44]Chandler RJ， Sands MS， Venditti CP. Recombinant adeno-associated viral integration and genotoxicity： insights from animal models[J]. Hum Gene Ther， 2017， 28(4)：314-322.
     [45]Bell P， Wang L， Lebherz C， et al. No evidence for tumorigenesis of AAV vectors in a large-scale study in mice[J]. Mol Ther， 2005， 12(2)：299-306.
     [46]Li H， Malani N， Hamilton SR， et al. Assessing the potential for AAV vector genotoxicity in a murine model[J]. Blood， 2011， 117(12)：3311-3319.
     [47]Arruda VR， Schuettrumpf J， Herzog RW， et al. Safety and efficacy of factor IX gene transfer to skeletal muscle in murine and canine hemophilia B models by adeno-associated viral vector serotype 1[J]. Blood， 2004， 103(1)：85-92.
     [48]Nichols TC， Whitford MH， Arruda VR， et al. Translational data from adeno-associated virus-mediated gene therapy of hemophilia B in dogs[J]. Hum Gene Ther Clin Dev， 2015， 26(1)：5-14.
     [49]Niemeyer GP， Herzog RW， Mount J， et al. Long-term correction of inhibitor-prone hemophilia B dogs treated with liver-directed AAV2-mediated factor IX gene therapy[J]. Blood， 2009， 113(4)：797-806.
     [50]Nathwani AC， Rosales C， McIntosh J， et al. Long-term safety and efficacy following systemic administration of a self-complementary AAV vector encoding human FIX pseudotyped with serotype 5 and 8 capsid proteins[J]. Mol Ther， 2011， 19(5)：876-885.

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