关于重组药物生产用细胞库单克隆性的审评思考
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摘要
用于重组蛋白药物生产的细胞库应来源于单个克隆,以保持重组蛋白药物在整个生命周期中质量的一致性。多种技术可用于转染后细胞群的单克隆化分离及确认。本文以CHO细胞为例,综述了工业界和监管机构对于生产用细胞库单克隆性的认知和了解,提出了基于目前技术水平对申报临床及申报上市阶段单克隆性的一般要求,并对非单克隆性来源细胞库的质量控制策略及审评要求提出建议,希望能够为研发机构成功建立重组蛋白药物生产用细胞库提供参考。
To ensure the consistency of quality in recombinant protein production, the cell bank for biologics should be derived from a single clone. A number of techniques have been used for cloning and assurance from the cellular pool after transfection with a target gene. Here, using CHO cell as an example, we summarize the knowledge and understanding of monoclonality of production cell bank from both industries and regulatory authorities, and propose general considerations on the requirements of monoclonality for clinical trial application and new drug application based on current techniques. Furthermore, we suggest quality control strategies and assessment methods for those cell banks from non-single clones.
To ensure the consistency of quality in recombinant protein production, the cell bank for biologics should be derived from a single clone. A number of techniques have been used for cloning and assurance from the cellular pool after transfection with a target gene. Here, using CHO cell as an example, we summarize the knowledge and understanding of monoclonality of production cell bank from both industries and regulatory authorities, and propose general considerations on the requirements of monoclonality for clinical trial application and new drug application based on current techniques. Furthermore, we suggest quality control strategies and assessment methods for those cell banks from non-single clones.
引文
[1]Walsh G.Biopharmaceutical benchmarks 2018[J].Nat Biotech‐nol,2018,36:1136-1145.
[2]Datta P,Linhardt RJ,Sharfstein ST.An'omics approach towards CHO cell engineering[J].Biotechnol Bioeng,2013,110:1255-1271.
[3]Evans K,Albanetti T,Venkat R,et al.Assurance of monoclonality in one round of cloning through cell sorting for single cell depo‐sition coupled with high resolution cell imaging[J].Biotechnol Prog,2015,31:1172-1178.
[4]Koziol JA,Ferrari C,Chisari FV.Evaluation of monoclonality of cell lines from sequential dilution assays[J].J Immunol Methods,1987,105:139-143.
[5]Frye C,Deshpande R,Estes S,et al.Industry view on the relative importance of"clonality"of biopharmaceutical-producing cell lines[J].Biologicals,2016,44:117-122.
[6]Wurm FM,Wurm MJ.Cloning of CHO cells,productivity and genetic stability-a discussion[J].Processes,2017,5:20.
[7]Tharmalingam T,Barkhordarian H,Tejeda N,et al.Characteriza‐tion of phenotypic and genotypic diversity in subclones derived from a clonal cell line[J].Biotechnol Prog,2018,34:613-623.
[8]Vcelar S,Melcher M,Auer N,et al.Changes in chromosome counts and patterns in CHO cell lines upon generation of recom‐binant cell lines and subcloning[J].Biotechnol J,2018,13:e1700495.
[9]Aebischer-Gumy C,Moretti P,Little T,et al.Analytical assess‐ment of clonal derivation of eukaryotic/CHO cell populations[J].J Biotechnol,2018,286:17-26.
[10]O'Callaghan PM,Berthelot ME,Young RJ,et al.Diversity in host clone performance within a Chinese hamster ovary cell line[J].Biotechnol Prog,2015,31:1187-1200.
[11]ICH Q5D.ICH harmonised tripartite guideline:derivation and characterisation of cell substrates used for production of biotech‐nological/biological products[EB/OL].1997[2018-12-31].https://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guide‐lines/Quality/Q5D/Step4/Q5D_Guideline.pdf.
[12]WHO.WHO Expert Committee on biological standardization[EB/OL].2013[2018-12-31].https://www.who.int/biologicals/expert_committee/TRS_978_61st_report.pdf.
[13]Kennett S.Establishing clonal cell lines-a regulatory perspective black cell,cell blue,cell old,new cell[EB/OL].WCBP2014[2018-12-31].https://c.ymcdn.com/sites/www.casss.org/resource/resmgr/WCBP_Speaker_Slides/2014_WCBP_Sarah_Kennett.pdf.
[14]Novak R.Regulatory perspective on the evaluation of clonality of mammalian cell banks[EB/OL].2017[2018-12-31].https://cdn.ymaws.com/www.casss.org/resource/resmgr/cmc_no_am_jan_spkr_slds/2017_CMCJ_Novak_Rachel.pdf.
[15]Welch J.Tilting at clones:a regulatory perspective on the impor‐tance of"clonality"of mammalian cell banks[EB/OL].2017[2018-12-31].https://www.topbiox.com/wp-content/uploads/2017/06/Joel-Welch.pdf-Amsterdam-April-2017.pdf.
[16]Zhou Y,Shaw D,Lam C,et al.Beating the odds:the poisson distribution of all input cells during limiting dilution grossly underestimates whether a cell line is clonally-derived or not[J].Biotechnol Prog,2018,34:559-569.
[17]Fieder J,Schulz P,Gorr I,et al.A single-step FACS sorting strat‐egy in conjunction with fluorescent vital dye imaging efficiently assures clonality of biopharmaceutical production cell lines[J].Biotechnol J,2017.DOI:10.1002/biot.201700002.
[18]Yim M,Shaw D.Achieving greater efficiency and higher confi‐dence in single-cell cloning by combining cell printing and plate imaging technologies[J].Biotechnol Prog,2018,34:1454-1459.
[19]Roy G,Miro-Quesada G,Zhuang L,et al.Sequential screening by ClonePix FL and intracellular staining facilitate isolation of high producer cell lines for monoclonal antibody manufacturing[J].J Immunol Methods,2017,451:100-110.
[20]Wright C,Groot J,Swahn S,et al.Genetic mutation analysis at early stages of cell line development using next generation sequencing[J].Biotechnol Prog,2016,32:813-817.
[21]Shaw D,Yim M,Tsukuda J,et al.Development and characteriza‐tion of an automated imaging workflow to generate clonallyderived cell lines for therapeutic proteins[J].Biotechnol Prog,2018,34:584-592.
[2]Datta P,Linhardt RJ,Sharfstein ST.An'omics approach towards CHO cell engineering[J].Biotechnol Bioeng,2013,110:1255-1271.
[3]Evans K,Albanetti T,Venkat R,et al.Assurance of monoclonality in one round of cloning through cell sorting for single cell depo‐sition coupled with high resolution cell imaging[J].Biotechnol Prog,2015,31:1172-1178.
[4]Koziol JA,Ferrari C,Chisari FV.Evaluation of monoclonality of cell lines from sequential dilution assays[J].J Immunol Methods,1987,105:139-143.
[5]Frye C,Deshpande R,Estes S,et al.Industry view on the relative importance of"clonality"of biopharmaceutical-producing cell lines[J].Biologicals,2016,44:117-122.
[6]Wurm FM,Wurm MJ.Cloning of CHO cells,productivity and genetic stability-a discussion[J].Processes,2017,5:20.
[7]Tharmalingam T,Barkhordarian H,Tejeda N,et al.Characteriza‐tion of phenotypic and genotypic diversity in subclones derived from a clonal cell line[J].Biotechnol Prog,2018,34:613-623.
[8]Vcelar S,Melcher M,Auer N,et al.Changes in chromosome counts and patterns in CHO cell lines upon generation of recom‐binant cell lines and subcloning[J].Biotechnol J,2018,13:e1700495.
[9]Aebischer-Gumy C,Moretti P,Little T,et al.Analytical assess‐ment of clonal derivation of eukaryotic/CHO cell populations[J].J Biotechnol,2018,286:17-26.
[10]O'Callaghan PM,Berthelot ME,Young RJ,et al.Diversity in host clone performance within a Chinese hamster ovary cell line[J].Biotechnol Prog,2015,31:1187-1200.
[11]ICH Q5D.ICH harmonised tripartite guideline:derivation and characterisation of cell substrates used for production of biotech‐nological/biological products[EB/OL].1997[2018-12-31].https://www.ich.org/fileadmin/Public_Web_Site/ICH_Products/Guide‐lines/Quality/Q5D/Step4/Q5D_Guideline.pdf.
[12]WHO.WHO Expert Committee on biological standardization[EB/OL].2013[2018-12-31].https://www.who.int/biologicals/expert_committee/TRS_978_61st_report.pdf.
[13]Kennett S.Establishing clonal cell lines-a regulatory perspective black cell,cell blue,cell old,new cell[EB/OL].WCBP2014[2018-12-31].https://c.ymcdn.com/sites/www.casss.org/resource/resmgr/WCBP_Speaker_Slides/2014_WCBP_Sarah_Kennett.pdf.
[14]Novak R.Regulatory perspective on the evaluation of clonality of mammalian cell banks[EB/OL].2017[2018-12-31].https://cdn.ymaws.com/www.casss.org/resource/resmgr/cmc_no_am_jan_spkr_slds/2017_CMCJ_Novak_Rachel.pdf.
[15]Welch J.Tilting at clones:a regulatory perspective on the impor‐tance of"clonality"of mammalian cell banks[EB/OL].2017[2018-12-31].https://www.topbiox.com/wp-content/uploads/2017/06/Joel-Welch.pdf-Amsterdam-April-2017.pdf.
[16]Zhou Y,Shaw D,Lam C,et al.Beating the odds:the poisson distribution of all input cells during limiting dilution grossly underestimates whether a cell line is clonally-derived or not[J].Biotechnol Prog,2018,34:559-569.
[17]Fieder J,Schulz P,Gorr I,et al.A single-step FACS sorting strat‐egy in conjunction with fluorescent vital dye imaging efficiently assures clonality of biopharmaceutical production cell lines[J].Biotechnol J,2017.DOI:10.1002/biot.201700002.
[18]Yim M,Shaw D.Achieving greater efficiency and higher confi‐dence in single-cell cloning by combining cell printing and plate imaging technologies[J].Biotechnol Prog,2018,34:1454-1459.
[19]Roy G,Miro-Quesada G,Zhuang L,et al.Sequential screening by ClonePix FL and intracellular staining facilitate isolation of high producer cell lines for monoclonal antibody manufacturing[J].J Immunol Methods,2017,451:100-110.
[20]Wright C,Groot J,Swahn S,et al.Genetic mutation analysis at early stages of cell line development using next generation sequencing[J].Biotechnol Prog,2016,32:813-817.
[21]Shaw D,Yim M,Tsukuda J,et al.Development and characteriza‐tion of an automated imaging workflow to generate clonallyderived cell lines for therapeutic proteins[J].Biotechnol Prog,2018,34:584-592.