基因治疗产品的生物分析：方法开发的策略

摘要/Abstract

摘要： 基因治疗产品采用独特的递送和治疗机制，评估其安全性和有效性需要设计并开发非传统和全面的生物分析方法。在临床前和临床生物分析研究涉及药代动力学、药效学、免疫原性、生物标志物分析等方面。基因治疗产品的药代动力学和药效学研究需要确定载体、目的基因、以及目的基因表达产物蛋白的存在、作用部位和生物分布。而对于免疫原性的研究，则需要监测机体针对基因治疗载体所产生的抗体，以及可能需要在接受治疗前评估受试者的预存抗体。由于基因治疗产品表达的外源产物蛋白也可能引发机体的免疫反应，因此针对表达产物的抗体也是免疫原性的研究内容。本文从药代动力学、药效学与免疫原性分析研究方向，综述基因治疗产品的生物分析方法开发的关键点、策略和实践。

关键词: font-size:medium, ">生物分析；基因治疗；药代动力学；药效学；免疫原性

Abstract: As gene therapy products employs unique delivery and therapeutic mechanisms， novel analytical methods are required for assessing their safety and efficacy. Preclinical and clinical bioanalysis includes pharmacokinetic， pharmacodynamic， immunogenicity， and biomarker analyses. For pharmacokinetic and pharmacodynamic studies of gene therapy products， it is necessary to determine the presence， site of action， and biodistribution of the vectors， transgene， and transgene product(s). For immunogenicity analyses， de novo anti-vector antibodies and most likely， pre-existing antibodies will need to be monitored in patients. In addition， as exposure to a transgene product may induce immune responses， the antibodies against a transgene product will also need to be monitored. This review will provide an overview of the critical considerations， strategies， and practical aspects of analytical method development for pharmacokinetic， pharmacodynamic， and immunogenicity analyses of gene therapy products.

Key words: font-size:medium, ">Bioanalysis； Gene therapy； Pharmacokinetics； Pharmacodynamics； Immunogenicity

中图分类号:

张贺峰, 梁宇, 刘颖, 邹灵龙, . 基因治疗产品的生物分析：方法开发的策略[J]. 中国药物评价, 2023, 40(5): 361-368.

ZHANG He-feng, LIANG Yu, LIU Ying, ZOU Ling-long, . Bioanalysis of Gene Therapy Products： Method Development Strategy[J]. CHINESE JOURNAL OF DRUG EVALUATION, 2023, 40(5): 361-368.

参考文献

[1]　Katherine AH， Maria GR. Gene therapy[J]. N Engl J Med， 2019， 381(5)：455-464.
     [2]　Gupta A， Andresen JL， Manan RS， et al. Nucleic acid delivery for therapeutic applications[J]. Advanced Drug Delivery Reviews， 2021， 178：113834.
     [3]　Mendell JR， Al-Zaidy SA， Rodino-Klapac LR， et al. Current Clinical Applications of In Vivo Gene Therapy with AAVs[J]. Molecular Therapy， 2021， 29(2)：464-488.
     [4]　国家药品监督管理局. 体内基因治疗产品药学研究与评价技术指导原则[EB/OL]. (2022-05-31)[2023-08-01]. https：//www.cde.org.cn/main/news/viewInfoCommon/c0ec5e347ba84df67bf75e15f6ad3f3f.
     [5]　张贺峰，梁宇，刘颖，等.基因修饰细胞治疗产品的生物分析策略与方法学介绍[J]. 中国药物评价， 2023， 40(5)：369-376.
     [6]　Shi J， Chen X， Diao J， et al. Bioanalysis in the Age of New Drug Modalities[J]. AAPS Journal， 2021， 23(3)：64.
     [7]　FDA. Human Gene Therapy Products Incorporating Human Genome Editing[EB/OL]. (2022-03-21) [2023-08-01]. https：//www.fda.gov/media/156894/download.
     [8]　FDA.S12 Nonclinical Biodistribution Considerations for Gene Therapy Products[EB/OL]. (2023-05-02) [2023-08-01]. https：//www.fda.gov/media/167605/download.
     [9]　国家药品监督管理局.基因治疗产品非临床研究与评价技术指导原则[EB/OL]. (2021-12-03) [2023-08-01]. https：//www.cde.org.cn/main/news/viewInfoCommon/41bc557bec23a6ebfb0e148cc989f041.
     [10]　Stevenson L， Richards S， Pillutla R， et al. 2018 White Paper on Recent Issues in Bioanalysis： Focus on flow cytometry， gene therapy， cut points and key clarifications on BAV (Part 3-LBA/cell-based assays： Immunogenicity， biomarkers and PK assays)[J]. Bioanalysis， 2018， 10(24)：1973-2001.
     [11]　Guiner CLE， Moullier P， Arruda VR. Biodistribution and Shedding of AAV Vectors[J]. Methods in Molecular Biology， 2011， 807(4)：339-359.
     [12]　Bulcha JT， Wang Y， Ma H， et al. Viral vector platforms within the gene therapy landscape[J]. Signal Transduction and Targeted Therapy， 2021， 6(1)：53.
     [13]　国家药典委员会. 人用基因治疗制品总论（公示稿）[EB/OL]. (2019-06-01) [2023-08-01]. https：//www.chp.org.cn/upload/userfiles/20190610/67171560147543847.pdf.
     [14]　Biasco L. Integration Site Analysis in Gene Therapy Patients： Expectations and Reality[J]. Human Gene Therapy， 2017， 28(12)：1122-1129.
     [15]　FDA. Design and Analysis of Shedding Studies for Virus or Bacteria-Based Gene Therapy and Oncolytic Products： Guidance for Industry[EB/OL]. (2015-08-27)[2023-08-01]. https：//www.fda.gov/files/vaccines%2C%20blood%20%26%20biologics/published.
     [16]　Asaad W， Volos P， Maksimov D， et al. AAV genome modification for efficient AAV production[J]. Heliyon， 2023， 9(4)：e15071.
     [17]　FDA.Considerations for the Design of Early-Phase Clinical Trials of Cellular and Gene Therapy Products[EB/OL]. (2015-06-27)[2023-08-01]. https：//www.fda.gov/media/106369/download.
     [18]　国家药品监督管理局.基因治疗产品长期随访临床研究技术指导原则[EB/OL]. (2021-12-03) [2023-08-01]. https：//www.cde.org.cn/main/news/viewInfoCommon/c9de887410ddcc291ce5a1c039a241c6.
     [19]　Piccoli S， Mehta D， Vitaliti A， et al. 2019 White Paper on Recent Issues in Bioanalysis： FDA Immunogenicity Guidance， Gene Therapy， Critical Reagents， Biomarkers and Flow Cytometry Validation (Part 3-Recommendations on 2019 FDA Immunogenicity Guidance， Gene Therapy Bioanalytical Challenges， St[J]. Bioanalysis， 2019， 11(24)：2207-2244.
     [20]　Neubert H， Olah T， Lee A， et al. 2018 White Paper on Recent Issues in Bioanalysis： Focus on immunogenicity assays by hybrid LBA/LCMS and regulatory feedback Part 2-PK， PD & ADA assays by hybrid LBA/LCMS & regulatory agencies’inputs on bioanalysis， biomarkers and immunogenicity[J]. Bioanalysis， 2018， 10(23)：1897-1917.
     [21]　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.
     [22]　Ertl HCJ.Immunogenicity and toxicity of AAV gene therapy[J]. Frontiers in Immunology， 2022， 13(August)：1-9.
     [23]　Weber T.Anti-AAV Antibodies in AAV Gene Therapy： Current Challenges and Possible Solutions[J]. Frontiers in Immunology， 2021， 12(March)：1-5.
     [24]　Verdera HC， Kuranda K， Mingozzi F.AAV Vector Immunogenicity in Humans： A Long Journey to Successful Gene Transfer[J]. Molecular Therapy， 2020， 28(3)：723-746.
     [25]　Schulz M， Levy DI， Petropoulos CJ， et al. Binding and neutralizing anti-AAV antibodies： Detection and implications for rAAV-mediated gene therapy[J]. Molecular Therapy， 2023， 31(3)：616-630.
     [26]　Corsaro B， Yang TY， Murphy R， et al. 2020 White Paper on Recent Issues in Bioanalysis： Vaccine Assay Validation， qPCR Assay Validation， QC for CAR-T Flow Cytometry， NAb Assay Harmonization and ELISpot Validation (Part 3-Recommendations on Immunogenicity Assay Strategies， NAb Assays， Biosim[J]. Bioanalysis， 2021， 13(6)：415-463.
     [27]　Yang TY， Braun M， Lembke W， et al. Immunogenicity assessment of AAV-based gene therapies： An IQ consortium industry white paper[J]. Molecular Therapy-Methods and Clinical Development， 2022， 26(September)：471-494.
     [28]　Patton KS， Harrison MT， Long BR， et al. Monitoring cell-mediated immune responses in AAV gene therapy clinical trials using a validated IFN-γ ELISpot method[J]. Molecular Therapy-Methods and Clinical Development， 2021， 22：183-195.
     [29]　Sun K， Xu Y， Zhang L， et al. A Phase 2 Trial of Enhancing Immune Checkpoint Blockade by Stereotactic Radiation and In Situ Virus Gene Therapy in Metastatic Triple-Negative Breast Cancer[J]. Clinical Cancer Research， 2022， 28(20)：4392-4401.
     [30]　Buscail L， Bournet B， Vernejoul F， et al. First-in-man phase 1 clinical trial of gene therapy for advanced pancreatic cancer： Safety， biodistribution， and preliminary clinical findings[J]. Molecular Therapy， 2015， 23(4)：779-789.
     [31]　Loo L， Harris S， Milton M， et al. 2021 White Paper on Recent Issues in Bioanalysis： TAb/NAb， Viral Vector CDx， Shedding Assays； CRISPR/Cas9 & CAR-T Immunogenicity； PCR & Vaccine Assay Performance；ADA Assay Comparability & Cut Point Appropriateness (Part 3-Recommendations on Gene Therap[J]. Bioanalysis， 2022， 14(11)：737-793.