A Case Study of a Bispecific Antibody Manufacturability Assessment and Optimization During Discovery Stage and Its Implications

Overview

Journal Antib Ther

Publisher Oxford University Press

Date 2024 Jul 22

PMID 39036070

Authors

Shuang Wang

Weijie Zhang

Baotian Yang

Xudong Zhang

Jing Fang

Haopeng Rui

Zhijian Chen

Jijie Gu

Zhiqiang Chen

Jianqing Xu

Affiliations

Soon will be listed here.

Abstract

The manufacturability assessment and optimization of bispecific antibodies (bsAbs) during the discovery stage are crucial for the success of the drug development process, impacting the speed and cost of advancing such therapeutics to the Investigational New Drug (IND) stage and ultimately to the market. The complexity of bsAbs creates challenges in employing effective evaluation methods to detect developability risks in early discovery stage, and poses difficulties in identifying the root causes and implementing subsequent engineering solutions. This study presents a case of engineering a bsAb that displayed a normal solution appearance during the discovery phase but underwent significant precipitation when subjected to agitation stress during 15 L Chemistry, Manufacturing, and Control (CMC) production Leveraging analytical tools, structural analysis, prediction, and wet-lab validations, the key molecular origins responsible for the observed precipitation were identified and addressed. Sequence engineering to reduce protein surface hydrophobicity and enhance conformational stability proved effective in resolving agitation-induced aggregation. The refined bsAb sequences enabled successful mass production in CMC department. The findings of this case study contribute to the understanding of the fundamental mechanism of agitation-induced aggregation and offer a potential protein engineering procedure for addressing similar issues in bsAb. Furthermore, this case study emphasizes the significance of a close partnership between Discovery and CMC teams. Integrating CMC's rigorous evaluation methods with Discovery's engineering capability can facilitate a streamlined development process for bsAb molecules.

References

Ma H, OFagain C, OKennedy R . Antibody stability: A key to performance - Analysis, influences and improvement. Biochimie. 2020; 177:213-225. DOI: 10.1016/j.biochi.2020.08.019. View

Strickley R, Lambert W . A review of Formulations of Commercially Available Antibodies. J Pharm Sci. 2021; 110(7):2590-2608.e56. DOI: 10.1016/j.xphs.2021.03.017. View

Ito T, Tsumoto K . Effects of subclass change on the structural stability of chimeric, humanized, and human antibodies under thermal stress. Protein Sci. 2013; 22(11):1542-51. PMC: 3831669. DOI: 10.1002/pro.2340. View

Barthelemy P, Raab H, Appleton B, Bond C, Wu P, Wiesmann C . Comprehensive analysis of the factors contributing to the stability and solubility of autonomous human VH domains. J Biol Chem. 2007; 283(6):3639-3654. DOI: 10.1074/jbc.M708536200. View

Xu Y, Wang D, Mason B, Rossomando T, Li N, Liu D . Structure, heterogeneity and developability assessment of therapeutic antibodies. MAbs. 2018; 11(2):239-264. PMC: 6380400. DOI: 10.1080/19420862.2018.1553476. View

Zhang Y, Wang Y, Li Y . A method for improving protein A chromatography's aggregate removal capability. Protein Expr Purif. 2019; 158:65-73. DOI: 10.1016/j.pep.2019.02.017. View

Dyson M, Masters E, Pazeraitis D, Perera R, Syrjanen J, Surade S . Beyond affinity: selection of antibody variants with optimal biophysical properties and reduced immunogenicity from mammalian display libraries. MAbs. 2020; 12(1):1829335. PMC: 7592150. DOI: 10.1080/19420862.2020.1829335. View

Hughes J, Rees S, Kalindjian S, Philpott K . Principles of early drug discovery. Br J Pharmacol. 2010; 162(6):1239-49. PMC: 3058157. DOI: 10.1111/j.1476-5381.2010.01127.x. View

Razinkov V, Treuheit M, Becker G . Accelerated formulation development of monoclonal antibodies (mAbs) and mAb-based modalities: review of methods and tools. J Biomol Screen. 2015; 20(4):468-83. DOI: 10.1177/1087057114565593. View

10.

Kayser V, Chennamsetty N, Voynov V, Helk B, Trout B . Conformational stability and aggregation of therapeutic monoclonal antibodies studied with ANS and Thioflavin T binding. MAbs. 2011; 3(4):408-11. PMC: 3218538. DOI: 10.4161/mabs.3.4.15677. View

11.

Kemmish H, Fasnacht M, Yan L . Fully automated antibody structure prediction using BIOVIA tools: Validation study. PLoS One. 2017; 12(5):e0177923. PMC: 5436848. DOI: 10.1371/journal.pone.0177923. View

12.

Mahler H, Muller R, Friess W, Delille A, Matheus S . Induction and analysis of aggregates in a liquid IgG1-antibody formulation. Eur J Pharm Biopharm. 2005; 59(3):407-17. DOI: 10.1016/j.ejpb.2004.12.004. View

13.

Wolf Perez A, Lorenzen N, Vendruscolo M, Sormanni P . Assessment of Therapeutic Antibody Developability by Combinations of In Vitro and In Silico Methods. Methods Mol Biol. 2021; 2313:57-113. DOI: 10.1007/978-1-0716-1450-1_4. View

14.

Gamage C, Weis D, Walters B . Identification of Agitation-Induced Unfolding Events Causing Aggregation of Monoclonal Antibodies Using Hydrogen Exchange-Mass Spectrometry. J Pharm Sci. 2022; 111(8):2210-2216. DOI: 10.1016/j.xphs.2022.05.002. View

15.

Black S, Mould D . Development of hydrophobicity parameters to analyze proteins which bear post- or cotranslational modifications. Anal Biochem. 1991; 193(1):72-82. DOI: 10.1016/0003-2697(91)90045-u. View

16.

Sarciaux J, Mansour S, Hageman M, Nail S . Effects of buffer composition and processing conditions on aggregation of bovine IgG during freeze-drying. J Pharm Sci. 1999; 88(12):1354-61. DOI: 10.1021/js980383n. View

17.

Lowe D, Dudgeon K, Rouet R, Schofield P, Jermutus L, Christ D . Aggregation, stability, and formulation of human antibody therapeutics. Adv Protein Chem Struct Biol. 2011; 84:41-61. DOI: 10.1016/B978-0-12-386483-3.00004-5. View

18.

Ghazvini S, Kalonia C, Volkin D, Dhar P . Evaluating the Role of the Air-Solution Interface on the Mechanism of Subvisible Particle Formation Caused by Mechanical Agitation for an IgG1 mAb. J Pharm Sci. 2016; 105(5):1643-1656. DOI: 10.1016/j.xphs.2016.02.027. View

19.

Saito S, Hasegawa J, Kobayashi N, Tomitsuka T, Uchiyama S, Fukui K . Effects of ionic strength and sugars on the aggregation propensity of monoclonal antibodies: influence of colloidal and conformational stabilities. Pharm Res. 2013; 30(5):1263-80. DOI: 10.1007/s11095-012-0965-4. View

20.

Saito S, Namisaki H, Hiraishi K, Takahashi N, Iida S . A stable engineered human IgG3 antibody with decreased aggregation during antibody expression and low pH stress. Protein Sci. 2019; 28(5):900-909. PMC: 6459999. DOI: 10.1002/pro.3598. View