Immunostimulatory bacterial antigen–armed oncolytic measles virotherapy significantly increases the potency of anti-PD1 checkpoint therapy
Eleni Panagioti, … , Ianko D. Iankov, Evanthia Galanis
Eleni Panagioti, … , Ianko D. Iankov, Evanthia Galanis
Published July 1, 2021
Citation Information: J Clin Invest. 2021;131(13):e141614. https://doi.org/10.1172/JCI141614.
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Research Article Oncology

Immunostimulatory bacterial antigen–armed oncolytic measles virotherapy significantly increases the potency of anti-PD1 checkpoint therapy

Abstract

Clinical immunotherapy approaches are lacking efficacy in the treatment of glioblastoma (GBM). In this study, we sought to reverse local and systemic GBM-induced immunosuppression using the Helicobacter pylori neutrophil-activating protein (NAP), a potent TLR2 agonist, as an immunostimulatory transgene expressed in an oncolytic measles virus (MV) platform, retargeted to allow viral entry through the urokinase-type plasminogen activator receptor (uPAR). While single-agent murine anti-PD1 treatment or repeat in situ immunization with MV-s-NAP-uPA provided modest survival benefit in MV-resistant syngeneic GBM models, the combination treatment led to synergy with a cure rate of 80% in mice bearing intracranial GL261 tumors and 72% in mice with CT-2A tumors. Combination NAP-immunovirotherapy induced massive influx of lymphoid cells in mouse brain, with CD8+ T cell predominance; therapeutic efficacy was CD8+ T cell dependent. Inhibition of the IFN response pathway using the JAK1/JAK2 inhibitor ruxolitinib decreased PD-L1 expression on myeloid-derived suppressor cells in the brain and further potentiated the therapeutic effect of MV-s-NAP-uPA and anti-PD1. Our findings support the notion that MV strains armed with bacterial immunostimulatory antigens represent an effective strategy to overcome the limited efficacy of immune checkpoint inhibitor–based therapies in GBM, creating a promising translational strategy for this lethal brain tumor.

Authors

Eleni Panagioti, Cheyne Kurokawa, Kimberly Viker, Arun Ammayappan, S. Keith Anderson, Sotiris Sotiriou, Kyriakos Chatzopoulos, Katayoun Ayasoufi, Aaron J. Johnson, Ianko D. Iankov, Evanthia Galanis

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Figure 2

Infection with uPAR-retargeted MV-Edm strains results in cytopathic effects against syngeneic murine and patient-derived glioblastoma lines.

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Infection with uPAR-retargeted MV-Edm strains results in cytopathic effe...
(A) GBM6, GBM12, CT-2A, or GL261 cells were infected in vitro with MV-GFP-uPA or MV-GFP at MOI = 2. Expression of GFP-positive cells was measured by flow cytometry 48 hours after infection. (B) Copies of MV nucleoprotein mRNA per microgram of total RNA measured in CT-2A and GL261 cells infected at MOI = 2 with inactivated MV-s-NAP-uPA or MV-s-NAP-uPA. Where indicated, cells were pretreated with 5 μM ruxolitinib and remained in the presence of the drug until completion of the experiment. Results were obtained 48 hours after infection. (C) Murine and PDX glioblastoma cells were infected with the indicated viruses at MOI = 2. Where mentioned, cells were cotreated with 5 μM ruxolitinib. Following a 72-hour incubation, virus oncolytic activity was captured by crystal violet staining. Representative light microscopy pictures of virus infection. Vehicle-treated (Opti-MEM), DMSO-treated, and ruxolitinib-treated cells were included at the assay to serve as controls. Original magnification, ×100. (D) uPAR-retargeted MV infection cell viability time kinetics against CT-2A and GL261 glioblastomas measured by CellTiter-Blue assay at MOI = 0.1, 1, and 2 with and without cotreatment with ruxolitinib. (E) Expression of NAP transgene (ng/mL) examined by ELISA in the cell lysates of glioma cells following 72 hours of infection with uPAR-retargeted MV-s-NAP or preclinical grade MV-s-NAP at MOI = 2 in the presence or not of 5 μM ruxolitinib. MV-producing Vero cells infected with MV-s-NAP-uPA served as a positive control for NAP detection. All samples were run in triplicate and are representative of at least 2 independent experiments. Values represent mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 by 1-way ANOVA with Tukey’s multiple comparison test. NS, not significant.