Oxygen-carrying nanoemulsions and respiratory hyperoxia eliminate tumor hypoxia–induced immunosuppression
Katarina Halpin-Veszeleiova, Michael P. Mallouh, Lucy M. Williamson, Ashley C. Apro, Nuria R. Botticello-Romero, Camille Bahr, Maureen Shin, Kelly M. Ward, Laura Rosenberg, Vladimir B. Ritov, Michail V. Sitkovsky, Edwin K. Jackson, Bruce D. Spiess, Stephen M. Hatfield
Katarina Halpin-Veszeleiova, Michael P. Mallouh, Lucy M. Williamson, Ashley C. Apro, Nuria R. Botticello-Romero, Camille Bahr, Maureen Shin, Kelly M. Ward, Laura Rosenberg, Vladimir B. Ritov, Michail V. Sitkovsky, Edwin K. Jackson, Bruce D. Spiess, Stephen M. Hatfield
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Research Article Immunology Oncology

Oxygen-carrying nanoemulsions and respiratory hyperoxia eliminate tumor hypoxia–induced immunosuppression

Abstract

Hypoxia/hypoxia-inducible factor 1α–driven immunosuppressive transcription and cAMP-elevating signaling through A2A adenosine receptors (A2ARs) represent a major tumor-protecting pathway that enables immune evasion. Recent promising clinical outcomes due to the blockade of the adenosine-generating enzyme CD73 and A2AR in patients refractory to all other therapies have confirmed the importance of targeting hypoxia-adenosinergic signaling. We report a feasible approach to target the upstream stage of hypoxia-adenosinergic immunosuppression using an oxygen-carrying nanoemulsion (perfluorocarbon blood substitute). We show that oxygenation agent therapy (a) eliminates tumor hypoxia, (b) improves efficacy of endogenously developed and adoptively transferred T cells, and thereby (c) promotes regression of tumors in different anatomical locations. We show that both T cells and NK cells avoid hypoxic tumor areas and that reversal of hypoxia by oxygenation agent therapy increases intratumoral infiltration of activated T cells and NK cells due to reprogramming of the tumor microenvironment (TME). Thus, repurposing oxygenation agents in combination with supplemental oxygen may improve current cancer immunotherapies by preventing hypoxia-adenosinergic suppression, promoting immune cell infiltration and enhancing effector responses. These data also suggest that pretreating patients with oxygenation agent therapy may reprogram the TME from immunosuppressive to immune-permissive prior to adoptive cell therapy, or other forms of immunotherapy.

Authors

Katarina Halpin-Veszeleiova, Michael P. Mallouh, Lucy M. Williamson, Ashley C. Apro, Nuria R. Botticello-Romero, Camille Bahr, Maureen Shin, Kelly M. Ward, Laura Rosenberg, Vladimir B. Ritov, Michail V. Sitkovsky, Edwin K. Jackson, Bruce D. Spiess, Stephen M. Hatfield

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Figure 4

Oxygenation agent therapy induces tumor regression and improves survival.

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Oxygenation agent therapy induces tumor regression and improves survival...
C57BL/6 and BALB/c mice were injected i.d. with MCA205 fibrosarcoma or CT26 colon carcinoma and treated with or without Perflubron (PFC) and respiratory hyperoxia (60% O2). (A) Schematic illustration of the experimental design and treatment regimen in the MCA205 intradermal model. C57BL/6 mice with 9-day-established MCA205 fibrosarcoma intradermal tumors were housed in 60% O2 chambers or maintained at 21% O2 (normoxia) and treated with or without oxygenated Perflubron (15 mL/kg) 3 times per week until study completion. Created with BioRender. (B) Oxygenation agent therapy (PFC + 60% O2) induces the strongest tumor regression compared with control or either treatment alone. Tumors were measured 3 times per week using Vernier calipers and tumor volume was calculated by the following formula: π/6 × H × W × L. *P < 0.05, **P < 0.005. Data presented as mean ± SEM, n ≥ 7. (C) Survival curves for MCA205 tumor–bearing mice treated with oxygenation agent therapy (PFC + 60% O2) versus control (P < 0.001, n ≥ 9). (D) Schematic illustration of the experimental design and treatment regimen in the CT26 intradermal tumor model. BALB/c mice with 11-day-established CT26 intradermal tumors were housed in 60% O2 chambers or maintained at 21% O2 with or without oxygenated Perflubron (10 mL/kg) 3 times per week until study completion. Created with BioRender. (E) Oxygenation agent therapy (PFC + 60% O2) induces the strongest tumor regression compared with control or either treatment alone. Evaluation of tumor growth kinetics and analysis is identical to B. *P < 0.05, **P < 0.005 for control versus PFC + 60% O2; #P < 0.05 for control versus 60% O2. Data are presented as mean ± SEM, n ≥ 6. Inset: Representative image of ulceration, necrosis, and large holes observed in tumors from mice treated with oxygenation agent therapy. (F) Survival curves for CT26 tumor–bearing mice treated with oxygenation agent therapy (PFC + 60% O2) versus control. ***P < 0.001, n ≥ 6. Statistical significance was assessed using 2-way ANOVA with Tukey’s multiple-comparison test (B, C, E, and F).