A Novel Recombinant Modified Vaccinia Ankara Virus expressing Interleukin-13 Receptor α2 Antigen for Potential Cancer Immunotherapy

Yuki Sato; Ramjay Vatsan; Bharat H Joshi; Syed R Husain; Raj K Puri

doi:10.2174/1566524023666230331085007

A Novel Recombinant Modified Vaccinia Ankara Virus expressing Interleukin-13 Receptor α2 Antigen for Potential Cancer Immunotherapy

Curr Mol Med. 2024;24(6):758-770. doi: 10.2174/1566524023666230331085007.

Authors

Yuki Sato^{1

2}, Ramjay Vatsan³, Bharat H Joshi¹, Syed R Husain^{1

4}, Raj K Puri^{1

4}

Affiliations

¹ Tumor Vaccines and Biotechnology Branch, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD, 20993, USA.
² Department of Research Promotion, Division of Cancer Research, Japan Agency for Medical Research and Development, 1-7-1, Otemachi, Chiyoda, Tokyo 100- 0004, Japan.
³ Gene Therapy Branch, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD, 20993, USA.
⁴ Iovance Biotherapeutics, 825 Industrial Road, Suite 400, San Carlos, CA, California, 94070, USA.

PMID: 36999709
DOI: 10.2174/1566524023666230331085007

Abstract

Background: Genetically altered recombinant poxviruses hold great therapeutic promise in animal models of cancer. Poxviruses can induce effective cellmediated immune responses against tumor-associated antigens. Preventive and therapeutic vaccination with a DNA vaccine expressing IL-13Rα2 can mediate partial regression of established tumors in vivo, indicating that host immune responses against IL-13Rα2 need further augmentation.

Objective: The aim of the study is developing a recombinant modified vaccinia Ankara (MVA) expressing IL-13Rα2 (rMVA-IL13Rα2) virus and study in vitro infectivity and efficacy against IL-13Rα2 positive cell lines.

Methods: We constructed a recombinant MVA expressing IL-13Rα2 and a green fluorescent protein (GFP) reporter gene. Purified virus titration by infection of target cells and immunostaining using anti-vaccinia and anti-IL-13Rα2 antibodies was used to confirm the identity and purity of the rMVA-IL13Rα2.

Results: Western Blot analysis confirmed the presence of IL-13Rα2 protein (~52 kDa). Flow cytometric analysis of IL-13Rα2 negative T98G glioma cells when infected with rMVA-IL13Rα2 virus demonstrated cell-surface expression of IL-13Rα2, indicating the infectivity of the recombinant virus. Incubation of T98G-IL13Rα2 cells with varying concentrations (0.1-100 ng/ml) of interleukin-13 fused to truncated Pseudomonas exotoxin (IL13-PE) resulted in depletion of GFP⁺ fluorescence in T98G-IL13Rα2 cells. IL13-PE (10-1000 ng/ml) at higher concentrations also inhibited the protein synthesis in T98G-IL13Rα2 cells compared to cells infected with the control pLW44-MVA virus. IL13- PE treatment of rMVA-IL13Rα2 infected chicken embryonic fibroblast and DF-1 cell line reduced virus titer compared to untreated cells.

Conclusion: rMVA-IL13Rα2 virus can successfully infect mammalian cells to express IL-13Rα2 in a biologically active form on the surface of infected cells. To evaluate the efficacy of rMVA-IL13Rα2, immunization studies are planned in murine tumor models.

Keywords: GFP; IL-13Rα2; IL13-PE; MVA; green fluorescent protein; interleukin-13 fused to mutated form of Pseudomonas exotoxin.; interleukin-13 receptor α2; modified vaccinia ankara.

MeSH terms

Animals
Cancer Vaccines / genetics
Cancer Vaccines / immunology
Cell Line, Tumor
Exotoxins / genetics
Exotoxins / immunology
Genetic Vectors / genetics
Green Fluorescent Proteins / genetics
Green Fluorescent Proteins / metabolism
Humans
Immunotherapy* / methods
Interleukin-13 Receptor alpha2 Subunit* / genetics
Interleukin-13 Receptor alpha2 Subunit* / immunology
Interleukin-13 Receptor alpha2 Subunit* / metabolism
Neoplasms / immunology
Neoplasms / therapy
Pseudomonas aeruginosa Exotoxin A
Vaccinia virus* / genetics
Vaccinia virus* / immunology
Virulence Factors / genetics
Virulence Factors / immunology

Substances

Interleukin-13 Receptor alpha2 Subunit
Cancer Vaccines
Exotoxins
Pseudomonas aeruginosa Exotoxin A
Green Fluorescent Proteins
Virulence Factors

Supplementary concepts

Modified Vaccinia Ankara virus