Kinase display

Screening methodologies for kinase inhibitors have been revolutionized in their breadth with the advent of bacteriophage libraries of kinase display [6], enabling... 

ATP-mimicking ligand is in a fixed phase within a chromatographic column. Thus, the affinity of the test compound is measured relative to that of the fixed phase ligand through its ability to displace the latter and hence enable the kinase display bacteriophage to elute from the column. 

Fig. 8.5 High-throughput screening at 10mMforWBZ 4 (red) andimatinib (STI 571, blue, control) over a battery of human kinases displayed in a T7-bacteriophage library (Ambit Biosciences, San Diego, CA). Hit values are reported as percentage-bound kinase. Reprinted from the [14], copyright 2007 with permission from the American Society for Clinical Investigation...

Comparison with drug screening/ kinase-display panels... 

As shown in Chap. 8, WBZ 4 is a re-engineered version of imatinib with an enhanced specificity needed to curb imatinib s potential cardiotoxicity. The inhibitory impact of WBZ 4 is restricted to KIT, PDGFR, and JNK, all with nanomolar affinity, as corroborated by screening against a T7-bacteriophage library of kinase displays (Ambit Biosciences, CA) [13],... 

Morrogh, L. M., Hinshelwood, S., Costello, E, Cory, G. O., and Kinnon, C. (1999). The SH3 domain of Bruton s tyrosine kinase displays altered ligand binding properties when auto-phosphorylated in vitro. Eur. J. Immunol. 29, 2269-2279. 

Karlsson, R., Zheng, J., Zheng, N.-H., Taylor, S. S., Sowadski, J. M. Structure of the mamalian catalytic subunit of cAMP-dependent protein kinase and an inhibitor peptide displays an open conformation. Acta Cryst. D 49 (1993) 381-388. 

Apelin receptors activate several signalling pathways including coupling through inhibitory G-proteins (G ) and Ras-independent activation of extracellular-regulated kinases (ERKs) via protein kinase C (PKC). The apelin receptor is one of number of G-protein-coupled receptors that can act as an alternative coreceptor for entry into cells of HIV and simian immunodeficiency vims (SIV) strains in human U87 cells expressing CD4 in vitro. Apelin peptides blocks entry of HIV but display different potencies, with apelin-36 being more effective than shorter sequences [3]. 

In chronic myelogenous leukemia (CML) as well as in a subset of acute lymphoblastic leukemia (ALL) Bcr-Abl, a fusion protein of c-Abl and the breakpoint cluster region (bcr), is expressed in the cytosol of leukemic cells. This fusion protein forms homo-oligomeric complexes that display elevated kinase activity and is the causative molecular abnormality in CML and certain ALL. The transforming effect of Bcr-Abl is mediated by numerous downstream signaling pathways, including protein kinase C (PKC), Ras-Raf-ERK MAPK, JAK-STAT (see below), and PI3-kinase pathways. 

However, acyclic nucleotide analogs (acyclic nucleoside phosphonates) have been developed, which carry one phosphonate moiety and require only the two subsequent phosphorylation steps (De Clercq et al. 1978). Independent of virus-encoded kinases, they display a broader spectrum of efficacy. This class comprises important drugs against HIV (tenofovir) and HBV (adefovir, tenofovir), as well as cidofovir, which is approved for use against CMV retinitis, but also displays an exceptionally broad efficacy profile against many herpesviruses, adenovirus, poxviruses, and papillomaviruses (De Clercq and Holy 2005). 

Pyruvate kinase (PK) is one of the three postulated rate-controlling enzymes of glycolysis. The high-energy phosphate of phosphoenolpyruvate is transferred to ADP by this enzyme, which requires for its activity both monovalent and divalent cations. Enolpyruvate formed in this reaction is converted spontaneously to the keto form of pyruvate with the synthesis of one ATP molecule. PK has four isozymes in mammals M, M2, L, and R. The M2 type, which is considered to be the prototype, is the only form detected in early fetal tissues and is expressed in many adult tissues. This form is progressively replaced by the M( type in the skeletal muscle, heart, and brain by the L type in the liver and by the R type in red blood cells during development or differentiation (M26). The M, and M2 isozymes display Michaelis-Menten kinetics with respect to phosphoenolpyruvate. The Mj isozyme is not affected by fructose-1,6-diphosphate (F-1,6-DP) and the M2 is al-losterically activated by this compound. Type L and R exhibit cooperatively in... 

As described in several monographs [4], bryostatin 1 exhibits significant in vitro and in vivo antineoplastic activity against a range of tumor cell lines including murine leukemia, B-cell lymphoma, reticulum cell sarcoma, ovarian carcinoma, and melanoma. It is also effective in the modulation of apoptotic function [5], the reversal of multidrug resistance [6], and stimulation of the immune system [7]. These unique features displayed by bryostatin 1 are attributed to its high affinity for protein kinase C (PKC) isozymes and its ability to selectively modulate their functions [8]. PKCs are a type of intracellular serine and threonine kinase that... 

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