Cyclic adenosine 5'-diphosphate ribose analogs without a "southern" ribose inhibit ADP-ribosyl cyclase-hydrolase CD38.

Joanna M Swarbrick,Barry V L Potter,Hongmin Zhang,Richard Graeff,Quan Hao,Mark P Thomas

doi:10.1021/jm501037u

Joanna M Swarbrick, Barry V L Potter + Show 4 more

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https://doi.org/10.1021/jm501037u

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Abstract

Cyclic adenosine 5′-diphosphate ribose (cADPR) analogs based on the cyclic inosine 5′-diphosphate ribose (cIDPR) template were synthesized by recently developed stereo- and regioselective N1-ribosylation. Replacing the base N9-ribose with a butyl chain generates inhibitors of cADPR hydrolysis by the human ADP-ribosyl cyclase CD38 catalytic domain (shCD38), illustrating the nonessential nature of the “southern” ribose for binding. Butyl substitution generally improves potency relative to the parent cIDPRs, and 8-amino-N9-butyl-cIDPR is comparable to the best noncovalent CD38 inhibitors to date (IC50 = 3.3 μM). Crystallographic analysis of the shCD38:8-amino-N9-butyl-cIDPR complex to a 2.05 Å resolution unexpectedly reveals an N1-hydrolyzed ligand in the active site, suggesting that it is the N6-imino form of cADPR that is hydrolyzed by CD38. While HPLC studies confirm ligand cleavage at very high protein concentrations, they indicate that hydrolysis does not occur under physiological concentrations. Taken together, these analogs confirm that the “northern” ribose is critical for CD38 activity and inhibition, provide new insight into the mechanism of cADPR hydrolysis by CD38, and may aid future inhibitor design.

Highlights

Cyclic adenosine 5′-diphosphate ribose[1,2] is synthesized in biological systems from nicotinamide adenine dinucleotide (NAD+) by ADP-ribosyl cyclases (ADPRCs)
Formation of Cyclic adenosine 5′-diphosphate ribose (cADPR) is catalyzed by the multifunctional transmembrane glycoprotein ADPRC CD38.7 CD38 acts as an NAD+ glycohydrolase (NADase) and as a cADPR hydrolase to generate adenosine 5′-diphosphate ribose (ADPR), another Ca2+-releasing second messenger.[8−10] Under acidic conditions, CD38 generates the most potent Ca2+releasing second messenger known to date, nicotinic acid adenine dinucleotide phosphate (NAADP) from nicotinamide adenine dinucleotide phosphate (NADP).[11]
Crystallography of shCD38 has revealed the mechanism by. As both cADPR and ADPR are derived from a common intermediate,[24] we chose to design product-like inhibitors based which NAD+ is either cyclized to cADPR or hydrolyzed to ADPR.[39−41] Crystal structures obtained with shCD38 and on the cADPR structure to exploit inhibition of CD38 cADPR unnatural ligands 2 (PDB code 2PGJ),[42] cyclic adenosine 5′

Summary

■ INTRODUCTION

Cyclic adenosine 5′-diphosphate ribose (cADPR, 1, Figure 1)[1,2] is synthesized in biological systems from nicotinamide adenine. It acts as a second messenger, mobilizing intracellular calcium.[3−6]. CD38 is a marker in AIDS progression[12] and a negative prognostic marker of chronic lymphocytic leukemia.[13] It acts to regulate intracellular levels of NAD+, being implicated in energy homeostasis, signal transduction, and aging,[14−16] and recently has been shown to be critical for social behavior in mice.[17] The emerging role of CD38 in disease states is stimulating the search for modulators of activity for chemical biological studies and to provide structural clues for drug design and potential therapeutic candidates.[18] To date, CD38 inhibitors fall broadly into two categories: mechanism-based covalent inhibitors that bind to the catalytic residue, and reversible, competitive, noncovalent inhibitors. N9-butyl cIDPR 7 (carbons in pink) with critical residues in the binding pocket and additional predicted H-bond to Asp[155]

■ RESULTS AND DISCUSSION

■ CONCLUSION

■ ACKNOWLEDGMENTS

■ REFERENCES