Selectivity Determination of a Small Molecule Chemical Probe Using Protein Microarray and Affinity Capture Techniques.

Small molecule selectivity is an essential component of candidate drug selection and target validation. New technologies are required to better understand off-target effects, with particular emphasis needed on broad protein profiling. Here, we describe the use of a tritiated chemical probe and a 9000 human protein microarray to discern the binding selectivity of an inhibitor of the mRNA decapping scavenger enzyme DcpS. An immobilized m7GTP resin was also used to assess the selectivity of a DcpS inhibitor against mRNA cap-associated proteins in whole cell extracts. These studies confirm the exquisite selectivity of diaminoquinazoline DcpS inhibitors, and highlight the utility of relatively simple protein microarray and affinity enrichment technologies in drug discovery and chemical biology.

[1]  Heather L. Plasterer,et al.  The DcpS inhibitor RG3039 improves motor function in SMA mice. , 2013, Human molecular genetics.

[2]  V. Cowling,et al.  Cap-binding complex (CBC) , 2013, The Biochemical journal.

[3]  Nigel Greene,et al.  Physicochemical drug properties associated with in vivo toxicological outcomes: a review , 2009, Expert opinion on drug metabolism & toxicology.

[4]  Betsy S. Pierce,et al.  Cellular thermal shift and clickable chemical probe assays for the determination of drug-target engagement in live cells. , 2016, Organic & biomolecular chemistry.

[5]  M. Gurney,et al.  Synthesis and biological evaluation of novel 2,4-diaminoquinazoline derivatives as SMN2 promoter activators for the potential treatment of spinal muscular atrophy. , 2008, Journal of medicinal chemistry.

[6]  Paul J. Chirik,et al.  Iron-catalysed tritiation of pharmaceuticals , 2016, Nature.

[7]  S. Haggarty,et al.  Finding new components of the target of rapamycin (TOR) signaling network through chemical genetics and proteome chips. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[8]  L. Jones.  An industry perspective on drug target validation , 2016, Expert opinion on drug discovery.

[9]  Xinfu Jiao,et al.  The scavenger mRNA decapping enzyme DcpS is a member of the HIT family of pyrophosphatases , 2002, The EMBO journal.

[10]  G. M. Di Guglielmo,et al.  Synthetic Triterpenoids Target the Arp2/3 Complex and Inhibit Branched Actin Polymerization* , 2010, The Journal of Biological Chemistry.

[11]  A. Narayanan,et al.  Rational targeting of active-site tyrosine residues using sulfonyl fluoride probes. , 2015, ACS chemical biology.

[12]  Heather L. Plasterer,et al.  The DcpS inhibitor RG3039 improves survival, function and motor unit pathologies in two SMA mouse models. , 2013, Human molecular genetics.

[13]  A. Rai,et al.  Immune response biomarker profiling application on ProtoArray protein microarrays. , 2010, Methods in molecular biology.

[14]  Martin J. Simard,et al.  The human decapping scavenger enzyme DcpS modulates microRNA turnover , 2015, Scientific Reports.

[15]  M. Gurney,et al.  Effects of 2,4-diaminoquinazoline derivatives on SMN expression and phenotype in a mouse model for spinal muscular atrophy. , 2010, Human molecular genetics.

[16]  R. Rhoads,et al.  Purification of the messenger RNA cap-binding protein using a new affinity medium. , 1984, Biochemistry.

[17]  M. Gurney,et al.  DcpS as a therapeutic target for spinal muscular atrophy. , 2008, ACS chemical biology.

[18]  W. Merrick,et al.  eIF4F: A Retrospective* , 2015, The Journal of Biological Chemistry.

[19]  Mark E Bunnage,et al.  Target validation using chemical probes. , 2013, Nature chemical biology.