Osteopontin mediates acquired resistance to hypoxia-inducing antiangiogenics and promotes anti–PD-L1 refractoriness in breast cancer models
Jose Luis Ruiz-Sepulveda, Maria J. Bueno, Silvana Mouron, Veronica Jimenez-Renard, Manuel Muñoz, Manuel Moradiellos, Leonardo D. Garma, Luis García-Jimeno, Adam W. Watson, Ghassan Mouneimne, Solip Park, Rebeca Jimeno, Miguel Quintela-Fandino
Jose Luis Ruiz-Sepulveda, Maria J. Bueno, Silvana Mouron, Veronica Jimenez-Renard, Manuel Muñoz, Manuel Moradiellos, Leonardo D. Garma, Luis García-Jimeno, Adam W. Watson, Ghassan Mouneimne, Solip Park, Rebeca Jimeno, Miguel Quintela-Fandino
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Research Article Immunology Oncology

Osteopontin mediates acquired resistance to hypoxia-inducing antiangiogenics and promotes anti–PD-L1 refractoriness in breast cancer models

Abstract

Resistance to antiangiogenics is a major challenge in cancer therapy. These agents can either normalize or exacerbate tumor vascular abnormality and hypoxia. The mechanisms of resistance remain unclear in the latter setting. By integrating data from mouse models and clinical trials, we showed that hypoxia-inducing anti-VEGF therapy upregulated programmed cell death ligand 1 (PD-L1), yet failed to sensitize tumors to PD-L1 blockade. Mechanistically, early hypoxic stress triggered epithelial osteopontin (SPP1) production, which recruited monocytes and skewed macrophages toward M2 states, suppressing T cell cytotoxicity. Pharmacological SPP1 depletion impeded the development of hypoxia, reduced M2 infiltration, restored T cell activity, and enabled synergy between antiangiogenics and anti–PD-L1. Genetic dissection — tumor-epithelial Spp1-KO grafts and bone marrow chimeras generated by lethal irradiation and reconstitution with Spp1–/– or WT hematopoietic donors — showed that myeloid SPP1 contributed only marginally compared with epithelial SPP1. These findings identified SPP1 as a central mediator of resistance to hypoxia-inducing antiangiogenics, contributed to a comprehensive model of antiangiogenic resistance, and supported SPP1-targeted strategies to personalize immunotherapy and antiangiogenic therapy according to tumor hypoxia.

Authors

Jose Luis Ruiz-Sepulveda, Maria J. Bueno, Silvana Mouron, Veronica Jimenez-Renard, Manuel Muñoz, Manuel Moradiellos, Leonardo D. Garma, Luis García-Jimeno, Adam W. Watson, Ghassan Mouneimne, Solip Park, Rebeca Jimeno, Miguel Quintela-Fandino

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Figure 1

Anti-VEGF treatment generates hypoxic and nonhypoxic response patterns associated with differential PD-L1 induction.

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Anti-VEGF treatment generates hypoxic and nonhypoxic response patterns a...
(A) Tumor growth and OS in response to B20-4.1.1 (n = 31) or isotype control (n = 36). Two-way ANOVA followed by Šidák’s multiple-comparison test. (B) Representative anti-pimonidazole (aPimonidazole) staining and quantification of tumor hypoxia over time in control- (n = 32) and B20-4.1.1–treated tumors (n = 90). Scale bars: 200 μm. Two-way ANOVA followed by Šidák’s multiple-comparison test. (C) Distribution of hypoxia values in individual tumors at the indicated time points, showing heterogeneous hypoxic responses to anti-VEGF treatment. Control (n = 28) or B20-4.1.1 at 6 weeks of treatment (n = 90), or Tend (n = 24). (D) Tumor growth and OS according to whether B20-4.1.1–treated tumors developed HH (n = 20 mice) or LH (n = 11 mice). Two-way ANOVA followed by Tukey’s multiple-comparison test. (E) Representative consecutive sections stained for pimonidazole and PD-L1, showing increased PD-L1 in hypoxic rims surrounding necrotic areas in HH tumors. Scale bars: 200 μm. (F) PD-L1 positivity in tumor cells and myeloid cells from vehicle-treated tumors (n = 5) and from LH or HH tumors after B20-4.1.1 (n = 5 each group). One-way ANOVA with Tukey’s post hoc test for multiple comparisons. Data are presented as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. n, necrotic areas; r, rims.