Development of a Robust Cell-based Potency Assay for a Coxsackievirus A21 Oncolytic Virotherapy

Overview

Journal Heliyon

Specialty Social Sciences

Date 2024 Apr 1

PMID 38560158

Authors

Venkateswarlu Chamcha

Li He

Jenny Xu

Andrew R Swartz

Erin Green-Trexler

Kevin Gurney

Tessie McNeely

Affiliations

Soon will be listed here.

Abstract

Oncolytic viruses (OV) are part of a burgeoning field of investigational oncolytic therapy (OT), in which lytic viruses dissolve advanced tumors productively and specifically. One such OT is a Coxsackievirus A21 (CVA21) based OV that is currently under clinical evaluation. A tissue culture infectious dose (TCID50) assay was used for CVA21 potency release and stability testing in early clinical development. The titer measured in this method was an extrapolated value from cytopathic effect (CPE) observed during the serial dilution but doesn't represent direct viral killing of cells. Moreover, the assay was not deemed to be optimal to carry into late phase clinical development due to limitations in assay precision, turn-around time, and sample throughput. To address these points, we developed a plaque assay to measure viral plaque forming units to measure the potency value for drug substance (DS), drug product (DP) and virus seed (master and working) stocks. In this manuscript, we describe the steps taken to develop this plaque assay for the late-stage clinical development, which include the assay qualification, validation, and robustness protocols, and describe statistical methods for data analysis. Moreover, the method was validated for linearity, accuracy, precision, and specificity. Furthermore, the plaque assay quantifies OV infectivity with better precision (32% vs 58%), with higher sample throughput (22 samples/week vs 3 samples/week) and shorter assay turnaround time (4 days vs 7 days) than the TCID50 method. This assay development strategy can provide guidance for the development of robust cell-based potency methods for OVs and other infectious viral products.

References

Smither S, Lear-Rooney C, Biggins J, Pettitt J, Lever M, Olinger Jr G . Comparison of the plaque assay and 50% tissue culture infectious dose assay as methods for measuring filovirus infectivity. J Virol Methods. 2013; 193(2):565-71. DOI: 10.1016/j.jviromet.2013.05.015. View

Peraman R, Bhadraya K, Reddy Y . Analytical quality by design: a tool for regulatory flexibility and robust analytics. Int J Anal Chem. 2015; 2015:868727. PMC: 4332986. DOI: 10.1155/2015/868727. View

Verch T, Trausch J, Shank-Retzlaff M . Principles of vaccine potency assays. Bioanalysis. 2018; 10(3):163-180. DOI: 10.4155/bio-2017-0176. View

Supian N, Ng K, Chook J, Takebe Y, Chan K, Tee K . Genetic diversity of Coxsackievirus A21 associated with sporadic cases of acute respiratory infections in Malaysia. BMC Infect Dis. 2021; 21(1):446. PMC: 8130276. DOI: 10.1186/s12879-021-06148-x. View

Holly J, Fogelova M, Jakubcova L, Tomcikova K, Vozarova M, Vareckova E . Comparison of infectious influenza A virus quantification methods employing immuno-staining. J Virol Methods. 2017; 247:107-113. DOI: 10.1016/j.jviromet.2017.06.004. View

Annels N, Mansfield D, Arif M, Ballesteros-Merino C, Simpson G, Denyer M . Phase I Trial of an ICAM-1-Targeted Immunotherapeutic-Coxsackievirus A21 (CVA21) as an Oncolytic Agent Against Non Muscle-Invasive Bladder Cancer. Clin Cancer Res. 2019; 25(19):5818-5831. DOI: 10.1158/1078-0432.CCR-18-4022. View

Lawler S, Speranza M, Cho C, Chiocca E . Oncolytic Viruses in Cancer Treatment: A Review. JAMA Oncol. 2016; 3(6):841-849. DOI: 10.1001/jamaoncol.2016.2064. View

Macedo N, Miller D, Haq R, Kaufman H . Clinical landscape of oncolytic virus research in 2020. J Immunother Cancer. 2020; 8(2). PMC: 7552841. DOI: 10.1136/jitc-2020-001486. View

Working P, Lin A, Borellini F . Meeting product development challenges in manufacturing clinical grade oncolytic adenoviruses. Oncogene. 2005; 24(52):7792-801. DOI: 10.1038/sj.onc.1209045. View

10.

Bradley S, Jakes A, Harrington K, Pandha H, Melcher A, Errington-Mais F . Applications of coxsackievirus A21 in oncology. Oncolytic Virother. 2016; 3:47-55. PMC: 4918364. DOI: 10.2147/OV.S56322. View

11.

Ramirez-Fort M, Downing C, Doan H, Benoist F, Oberste M, Khan F . Coxsackievirus A6 associated hand, foot and mouth disease in adults: clinical presentation and review of the literature. J Clin Virol. 2014; 60(4):381-6. DOI: 10.1016/j.jcv.2014.04.023. View

12.

Fujimoto T, Iizuka S, Enomoto M, Abe K, Yamashita K, Hanaoka N . Hand, foot, and mouth disease caused by coxsackievirus A6, Japan, 2011. Emerg Infect Dis. 2012; 18(2):337-9. PMC: 3310456. DOI: 10.3201/eid1802.111147. View

13.

Zou L, Yi L, Song Y, Zhang X, Liang L, Ni H . A cluster of coxsackievirus A21 associated acute respiratory illness: the evidence of efficient transmission of CVA21. Arch Virol. 2016; 162(4):1057-1059. PMC: 7087265. DOI: 10.1007/s00705-016-3201-4. View

14.

Muller L, Holmes M, Michael J, Scott G, West E, Scott K . Plasmacytoid dendritic cells orchestrate innate and adaptive anti-tumor immunity induced by oncolytic coxsackievirus A21. J Immunother Cancer. 2019; 7(1):164. PMC: 6604201. DOI: 10.1186/s40425-019-0632-y. View

15.

Au G, Lindberg A, Barry R, Shafren D . Oncolysis of vascular malignant human melanoma tumors by Coxsackievirus A21. Int J Oncol. 2005; 26(6):1471-6. DOI: 10.3892/ijo.26.6.1471. View