A tumor-intrinsic PD-L1/NLRP3 inflammasome signaling pathway drives resistance to anti–PD-1 immunotherapy
Balamayoora Theivanthiran, … , Alisha Holtzhausen, Brent A. Hanks
Balamayoora Theivanthiran, … , Alisha Holtzhausen, Brent A. Hanks
Published February 4, 2020
Citation Information: J Clin Invest. 2020;130(5):2570-2586. https://doi.org/10.1172/JCI133055.
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Research Article Immunology Oncology

A tumor-intrinsic PD-L1/NLRP3 inflammasome signaling pathway drives resistance to anti–PD-1 immunotherapy

Abstract

An in-depth understanding of immune escape mechanisms in cancer is likely to lead to innovative advances in immunotherapeutic strategies. However, much remains unknown regarding these mechanisms and how they impact immunotherapy resistance. Using several preclinical tumor models as well as clinical specimens, we identified a mechanism whereby CD8+ T cell activation in response to programmed cell death 1 (PD-1) blockade induced a programmed death ligand 1/NOD-, LRR-, and pyrin domain–containing protein 3 (PD-L1/NLRP3) inflammasome signaling cascade that ultimately led to the recruitment of granulocytic myeloid-derived suppressor cells (PMN-MDSCs) into tumor tissues, thereby dampening the resulting antitumor immune response. The genetic and pharmacologic inhibition of NLRP3 suppressed PMN-MDSC tumor infiltration and significantly augmented the efficacy of anti–PD-1 antibody immunotherapy. This pathway therefore represents a tumor-intrinsic mechanism of adaptive resistance to anti–PD-1 checkpoint inhibitor immunotherapy and is a promising target for future translational research.

Authors

Balamayoora Theivanthiran, Kathy S. Evans, Nicholas C. DeVito, Michael Plebanek, Michael Sturdivant, Luke P. Wachsmuth, April K.S. Salama, Yubin Kang, David Hsu, Justin M. Balko, Douglas B. Johnson, Mark Starr, Andrew B. Nixon, Alisha Holtzhausen, Brent A. Hanks

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Figure 4

CD8+ T cells induce tumor HSP70 release in a NLRP3-dependent manner in response to anti–PD-1 Ab immunotherapy.

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CD8+ T cells induce tumor HSP70 release in a NLRP3-dependent manner in r...
(A) Schema illustrating coculture of OT-I CD8+ T cells with OVA-expressing BRAFV600E PTEN–/– melanoma cells followed by HSP70 Western blot analysis of isolated supernatant. Harvested supernatant was coincubated at increasing concentrations with WT BRAFV600E PTEN–/– melanoma cells followed by Wnt5a Western blot analysis. Blots are representative of 2 independent experiments. (B) Flow cytometric analysis of PMN-MDSCs and CD8+ T cells from resected autochthonous BRAFV600E PTEN–/– melanoma tissues following anti–PD-1 Ab or IgG isotype control therapy. Results are expressed per gram of tumor tissue (n = 6). (C) Flow cytometric analysis of tumor-infiltrating PMN-MDSCs from autochthonous BRAFV600E PTEN–/– melanomas following anti–PD-1 Ab versus IgG isotype control therapy with or without anti-CD8 Ab. Data were normalized to IgG control–treated tumors (n = 3). (D) HSP70 and β-actin Western blot analysis following treatment of BRAFV600E PTEN–/– melanoma cells with increasing concentrations of dacarbazine. Blots are representative of 3 independent experiments. (E) Tumor growth curve of syngeneic BRAFV600E PTEN–/– melanomas following vehicle control or low-dose (lo) (50 mg/kg i.p. q.o.d.) or high-dose (hi) (75 mg/kg i.p. q.o.d.) dacarbazine therapy (n = 5). (F) Flow cytometric analysis of PMN-MDSCs from BRAFV600E PTEN–/– melanomas following vehicle control or dacarbazine therapy (n = 5). Flow cytometric analysis of CD8+ T cells from BRAFV600E PTEN–/– melanomas following vehicle control or dacarbazine therapy (n = 5). (G) HSP70 Western blot analysis of supernatant and tumor cell lysates following ATP stimulation of BRAFV600E PTEN–/– melanoma cells at different time points, with or without treatment with the NLRP3 inhibitor (NLRP3i) MCC950. Blots are representative of 3 independent experiments. (H) HSP70 Western blot following coincubation of OT-1 CD8+ T cells and OVA-expressing BRAFV600E PTEN–/– melanoma cells with or without increasing concentrations of NLRP3 inhibitor. Blots are representative of 3 independent experiments. Spearman’s correlation calculation was performed in B. *P < 0.05 and ***P < 0.0005, by Student’s t test ( C), 1-way ANOVA with Sidak’s post hoc multiple comparisons test (E and F). See also Supplemental Figure 5.