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Design and in vivo activity of A3 adenosine receptor agonist prodrugs

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Abstract

Prodrugs (MRS7422, MRS7476) of highly selective A3 adenosine receptor (AR) agonists Cl-IB-MECA and MRS5698, respectively, were synthesized by succinylation of the 2′ and 3′ hydroxyl groups, and the parent, active drug was shown to be readily liberated upon incubation with liver esterases. The prodrug MRS7476 had greatly increased aqueous solubility compared with parent MRS5698 and was fully efficacious and with a longer duration than MRS7422 in reversing mouse neuropathic pain (chronic constriction injury model, 3 μmol/kg, p.o.), a known A3AR effect. MRS7476 (5 mg/kg, p.o., twice daily) was found to protect against non-alcoholic steatohepatitis (NASH) in the STAM mouse model, indicated by the NAFLD activity score. Hepatocyte ballooning, IL-10 production, and liver histology were significantly normalized in the MRS7476-treated mice, but not liver fibrosis (no change in ACTA2 levels) or inflammation. Hepatic expression of ADORA3 in human NAFLD patients was 1.9-fold lower compared to normal controls. Adora3 expression determined by qPCR in primary mouse liver was associated with the stellate cells, and its mouse full body A3AR knockout worsened liver markers of inflammation and steatosis. Thus, we have introduced a reversible prodrug strategy that enables water solubility and in vivo activity of masked A3AR agonists in models of two disease conditions.

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Abbreviations

ACTA2:

Smooth muscle α actin

ALT:

Alanine transaminase

AR:

Adenosine receptor

AUC:

Area under the curve

CCI:

Chronic constriction injury

Cl-IB-MECA:

2-Chloro-N6-(3-iodobenzyl)-adenosine-5′-N-methyluronamide

GPCR:

G protein-coupled receptor

HCC:

Hepatocellular carcinoma

IB-MECA:

N6-(3-iodobenzyl)-adenosine-5′-N-methyluronamide

IL-10:

Interleukin 10

MRS5698:

(1S,2R,3S,4R,5S)-4-(6-((3-chlorobenzyl)amino)-2-((3,4-difluorophenyl)ethynyl)-9H-purin-9-yl)-2,3-dihydroxy-N-methylbicyclo[3.1.0]hexane-1-carboxamide

NAFLD:

Non-alcoholic fatty liver disease

NAS:

NAFLD activity score

NASH:

Non-alcoholic steatohepatitis

NLRP3:

Nucleotide-binding oligomerization domain-like receptor protein 3 (inflammasome)

OGTT:

Oral glucose tolerance test

SAR:

Structure activity relationship

TGFβ:

Transforming growth factor β

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Funding

This work has been supported by funds from SLU (Salvemini Start-Up Funds) and NIDDK Intramural Research Program (Jacobson, ZIADK31117; Rotman, ZIADK75013).

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Authors and Affiliations

  1. Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bldg. 8A, Rm. B1A-19, 9000 Rockville Pike, Bethesda, MD, 20892-0810, USA

    R. Rama Suresh, Shanu Jain, Dilip K. Tosh & Kenneth A. Jacobson

  2. Department of Pharmacology and Physiology, Saint Louis University, St. Louis, MO, USA

    Zhoumou Chen & Daniela Salvemini

  3. Henry and Amelia Nasrallah Center for Neuroscience, Saint Louis University, St. Louis, MO, USA

    Zhoumou Chen & Daniela Salvemini

  4. Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, NIH, Bethesda, MD, USA

    Yanling Ma, Maren C. Podszun & Yaron Rotman

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  1. R. Rama Suresh
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  2. Shanu Jain
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Correspondence to Kenneth A. Jacobson.

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All procedures were conducted in accordance with the International Association for the Study of Pain, the National Institutes of Health guidelines on laboratory animal welfare guidelines, and the approval of Saint Louis University and NIDDK Institutional Animal Care and Use Committees.

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Suresh, R.R., Jain, S., Chen, Z. et al. Design and in vivo activity of A3 adenosine receptor agonist prodrugs. Purinergic Signalling 16, 367–377 (2020). https://doi.org/10.1007/s11302-020-09715-0

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