Human Recombinant Alkaline Phosphatase (Ilofotase Alfa) Protects Against Kidney Ischemia-Reperfusion Injury in Mice and Rats Through Adenosine Receptors

Diane L Rosin; J Perry Hall; Shuqiu Zheng; Liping Huang; Silvia Campos-Bilderback; Ruben Sandoval; Andrea Bree; Kevin Beaumont; Emily Miller; Jennifer Larsen; Ghazal Hariri; Neelu Kaila; Iain M Encarnacion; Jeremy D Gale; Andrea van Elsas; Bruce A Molitoris; Mark D Okusa

doi:10.3389/fmed.2022.931293

Human Recombinant Alkaline Phosphatase (Ilofotase Alfa) Protects Against Kidney Ischemia-Reperfusion Injury in Mice and Rats Through Adenosine Receptors

Front Med (Lausanne). 2022 Jul 28:9:931293. doi: 10.3389/fmed.2022.931293. eCollection 2022.

Authors

Diane L Rosin¹, J Perry Hall², Shuqiu Zheng³, Liping Huang³, Silvia Campos-Bilderback⁴, Ruben Sandoval⁴, Andrea Bree², Kevin Beaumont⁵, Emily Miller⁶, Jennifer Larsen⁷, Ghazal Hariri⁸, Neelu Kaila⁹, Iain M Encarnacion³, Jeremy D Gale², Andrea van Elsas¹⁰, Bruce A Molitoris⁴, Mark D Okusa³

Affiliations

¹ Department of Pharmacology, University of Virginia, Charlottesville, VA, United States.
² Inflammation and Immunology Research Unit, Pfizer Inc., Cambridge, MA, United States.
³ Division of Nephrology, Center for Immunity, Inflammation and Regeneration, University of Virginia, Charlottesville, VA, United States.
⁴ Division of Nephrology, Department of Medicine, Indiana University School of Medicine, Indiana Center for Biological Microscopy, Roudebush VA Medical Center, Indianapolis, IN, United States.
⁵ BioMedicine Design, Pfizer Inc., Cambridge, MA, United States.
⁶ BioMedicine Design, Pfizer Inc., Groton, CT, United States.
⁷ Early Clinical Development, Pfizer Inc., Groton, CT, United States.
⁸ Drug Product Development, Pfizer Inc., Cambridge, MA, United States.
⁹ Medicinal Chemistry, Pfizer Inc., Cambridge, MA, United States.
¹⁰ AM-Pharma B.V., Utrecht, Netherlands.

Abstract

Adenosine triphosphate (ATP) released from injured or dying cells is a potent pro-inflammatory "danger" signal. Alkaline phosphatase (AP), an endogenous enzyme that de-phosphorylates extracellular ATP, likely plays an anti-inflammatory role in immune responses. We hypothesized that ilofotase alfa, a human recombinant AP, protects kidneys from ischemia-reperfusion injury (IRI), a model of acute kidney injury (AKI), by metabolizing extracellular ATP to adenosine, which is known to activate adenosine receptors. Ilofotase alfa (iv) with or without ZM241,385 (sc), a selective adenosine A_2A receptor (A_2AR) antagonist, was administered 1 h before bilateral IRI in WT, A_2AR KO (Adora2a^-/- ) or CD73^-/- mice. In additional studies recombinant alkaline phosphatase was given after IRI. In an AKI-on-chronic kidney disease (CKD) ischemic rat model, ilofotase alfa was given after the three instances of IRI and rats were followed for 56 days. Ilofotase alfa in a dose dependent manner decreased IRI in WT mice, an effect prevented by ZM241,385 and partially prevented in Adora2a^-/- mice. Enzymatically inactive ilofotase alfa was not protective. Ilofotase alfa rescued CD73^-/- mice, which lack a 5'-ectonucleotidase that dephosphorylates AMP to adenosine; ZM241,385 inhibited that protection. In both rats and mice ilofotase alfa ameliorated IRI when administered after injury, thus providing relevance for therapeutic dosing of ilofotase alfa following established AKI. In an AKI-on-CKD ischemic rat model, ilofotase alfa given after the third instance of IRI reduced injury. These results suggest that ilofotase alfa promotes production of adenosine from liberated ATP in injured kidney tissue, thereby amplifying endogenous mechanisms that can reverse tissue injury, in part through A_2AR-and non-A_2AR-dependent signaling pathways.

Keywords: ATP; CD73; IRI; acute kidney injury; adenosine; alkaline phosphatase; recAP.

Abstract

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