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CF33-hNIS-antiPDL1 virus primes pancreatic ductal adenocarcinoma for enhanced anti-PD-L1 therapy

Abstract

Immunotherapeutic strategies that combine oncolytic virus (OV) and immune checkpoint inhibitors have the potential to overcome treatment resistance in pancreatic ductal adenocarcinoma (PDAC), one of the least immunogenic solid tumors. Oncolytic viral chimera, CF33-hNIS-antiPDL1 genetically modified to express anti-human PD-L1 antibody and CF33-hNIS-Δ without the anti-PD-L1 gene, were used to investigate the immunogenic effects of OVs and virus-delivered anti-PD-L1 in PDAC in vitro. Western blot, flow cytometry, and immunofluorescence microscopy were used to evaluate the effects of CF33-hNIS-Δ and IFNγ on PD-L1 upregulation in AsPC-1 and BxPC-3 cells, and CF33-hNIS-antiPDL1 production of anti-PD-L1 and surface PD-L1 blockade of AsPC-1 and BxPC-3 with or without cocultured activated T cells. The cytosolic and cell surface levels of PD-L1 in PDAC cell lines varied; only BxPC-3 showed high cell surface expression. Treatment of these cells with CF33-hNIS-Δ and IFNγ significantly upregulated PD-L1 expression and translocation of PD-L1 from the cytosol onto the cell surface. Following coculture of activated T cells and BxPC-3 with CF33-hNIS-antiPDL1, the cell surface PD-L1 blockade on BxPC-3 cells by virus-delivered anti-PD-L1 antibody increased granzyme B release and prevented virus-induced decrease of perforin release from activated CD8+ T cells. Our results suggest that CF33-IOVs can prime immune checkpoint inhibition of PDAC and enhance antitumor immune killing.

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Fig. 1: PD-L1 is found mostly in the cytosol of human pancreatic ductal adenocarcinoma cells (PDACs).
Fig. 2: CF33-hNIS-Δ increases the translocation/expression of PD-L1 in human PDAC cell lines.
Fig. 3: IFNγ induces PD-L1 translocation/upregulation from the cytosol onto the surface of human PDAC cell lines.
Fig. 4: Expression of anti-PD-L1 antibody encoded by CF33-hNIS-antiPDL1 in PDACs over time.
Fig. 5: Anti-PD-L1 antibody encoded by the virus blocks CF33-hNIS-Δ-induced surface PD-L1/CD274 binding of PDACs.
Fig. 6: Virus-encoded anti-PD-L1 antibody blocks IFNγ-induced surface PD-L1 binding of PDACs.
Fig. 7: CF33-hNIS-antiPDL1 induces BxPC-3 cell killing and increases T cell granzyme B and perforin release in coculture with activated T cells in vitro.

Data availability

All data relevant to the study are provided in the article or uploaded as supplementary information.

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Acknowledgements

The authors are grateful to Imugene Limited for providing approval to use their licensed oncolytic virus CF33 in the studies reported in this paper. Research reported in this publication included work performed in the Pathology Core, Flow Cytometry Core, Light Microscopy Core, and Integrative Genomics and Bioinformatics core facilities supported by the National Cancer Institute of the National Institutes of Health under grant number P30CA033572. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The authors would also like to thank Dr Jinhui Wang in Integrative Genomics Core, Lucy Brown in Flow Cytometry core, and Dr Brian Armstrong in the Light Microscopy Core of City of Hope for supporting the work.

The authors would like to thank Byungwook Kim, Martha Magallanes, Seonah Kang, and Dr Maria Hahn in our laboratory and Dr Chunyan Zhang in Department of Immuno-Oncology for technical support, and Supriya Deshpande, PhD, for assistance with manuscript editing.

SGW and SC are supported through the generosity of Natalie and David Roberts. These authors wish to thank them for their philanthropy. The authors would also like to thank Samuel Kuo and Grace Liu of Samson Holding, Ltd. for their generosity.

Funding

Department of Defense E01 Award W81XWH-19-1-0225 (CA180425).

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ZZ, YW, and YF conceived the experiments. ZZ, AY, SC, AKP, JL, and S-IK performed the experiments, researched, and interpreted data. ZZ, AY, Y-CY, ZL, and YW analyzed data. YW, SGW, HH, DVH, and YF secured funding and interpreted the data. ZZ, YW, AY, and YF wrote the manuscript. All authors edited and approved the final manuscript.

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Correspondence to Yanghee Woo.

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Zhang, Z., Yang, A., Chaurasiya, S. et al. CF33-hNIS-antiPDL1 virus primes pancreatic ductal adenocarcinoma for enhanced anti-PD-L1 therapy. Cancer Gene Ther (2021). https://doi.org/10.1038/s41417-021-00350-4

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