Clinical-Grade Oncolytic Adenovirus Purification Using Polysorbate 20 as an Alternative for Cell Lysis.

Abstract

Introduction: Oncolytic virus therapy is currently considered as a promising therapeutic ap-proach for cancer treatment. Adenovirus is well-known and extensively characterized as an oncolytic agent. The increasing number of clinical trials using this virus generates the demand for the development of a well-established purification approach. Triton X-100 is commonly used in cell lysis buffer prepara-tions. The addition of this surfactant in the list of substances with the very high concern of the Registra-tion, Evaluation, Authorization and Restriction of Chemicals (REACH) regulation promoted the research for effective alternatives.Methods: In this work, a purification strategy for oncolytic adenovirus compatible with phase I clinical trials, using an approved surfactant – Polysorbate 20 was developed. The proposed downstream train, composed by clarification, concentration using tangential flow filtration, intermediate purification with anion exchange chromatography, followed by a second concentration and a final polishing step was evaluated for both Triton X-100 and Polysorbate 20 processes. The impact of cell lysis with Polysorb-ate20 and Triton X-100 for each downstream step was evaluated in terms of product recovery and impu-rities removal. Overall, 61 ± 4% of infectious viral particles were recovered. Depletion of host cell pro-teins and ds-DNA was 99.9% and 97.1%, respectively.Results & Conclusion: The results indicated that Polysorbate 20 can be used as a replacement for Triton X-100 during cell lysis with no impact on product recovery, potency, and purity. Moreover, the devel-oped process is scalable and able to provide a highly purified product to be used in phase I and II clinical trials.