Chapter title |
Use of Inosine Monophosphate Dehydrogenase Activity Assay to Determine the Specificity of PARP-1 Inhibitors
|
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Chapter number | 22 |
Book title |
Poly(ADP-Ribose) Polymerase
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Published in |
Methods in molecular biology, July 2017
|
DOI | 10.1007/978-1-4939-6993-7_22 |
Pubmed ID | |
Book ISBNs |
978-1-4939-6992-0, 978-1-4939-6993-7
|
Authors |
Anthony, Sajitha, Peterson, Jeffrey R., Ji, Yingbiao, Sajitha Anthony, Jeffrey R. Peterson, Yingbiao Ji |
Abstract |
Inosine monophosphate dehydrogenase (IMPDH) is a rate-limiting enzyme involved in purine nucleotide biosynthesis. It is responsible for catalyzing the oxidation of inosine monophosphate (IMP) into xanthosine monophosphate (XMP). Concurrently, the cofactor NAD(+) is reduced to NADH. Poly(ADP-ribose) polymerase 1 (PARP-1) also utilizes NAD(+) as a substrate to synthesize poly(ADP-ribose). It has been demonstrated that inhibition of PARP-1 activity can be an effective cancer therapeutic. However, most PARP-1 inhibitors, including olaparib, were developed as NAD(+) analogs. Therefore, these inhibitors likely interfere with other NAD(+)-dependent pathways such as the one involved in de novo purine metabolism. In this chapter, we describe a method to quantitatively measure IMPDH activity by taking advantage of the autofluorescence of the product NADH. We use this method to analyze the effects of olaparib and non-NAD(+)-like PARP-1 inhibitor (5F02) on IMPDH activity. We found that olaparib, unlike 5F02, significantly inhibits IMPDH activity in a dose-dependent manner. Our results suggest that IMPDH inhibition is an off-target effect of olaparib treatment. The consequences of this effect should be addressed by future clinical studies. |
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