Kinase-Akt

Sato, S., Fujita, N., and Tsuruo, T. 2000. Modulation of Akt kinase activity by binding to Hsp. Proc Natl Acad Sci 97 10832-10837. 

Scheme 6.261 Synthesis of quinoxalines as allosteric Akt kinase inhibitors.

Sah JF, Balasubramanian S, Eckert RL and Rorke EA. 2004. Epigallocatechin-3-gallate inhibits epidermal growth factor receptor signaling pathway. Evidence for direct inhibition of ERK1/2 and AKT kinases. J Biol Chem 279(13)42755-12762. 

Protein kinase Cd, Akt kinase, calcium/calmodulin-dependent protein kinase IV, mitogen-activated protein kinase kinase (MEKK-1), focal adhesion kinase (FAK), protein phosphatase (PP)2A, calcineurin... 

PI3K/AKT Kinase Pathway Inhibitors in Pre-Clinical Development 366... 

AKT kinases participate in pathways that regulate several cellular processes, including survival, proliferation, tissue invasion, and metabolism [5-7]. Hyperactivation of the three isoforms of AKT kinases is a common finding in human malignancies. Increased AKT-1 activity has been observed in about 40% of breast and ovarian cancers and >50% of prostate carcinomas. Activation of... 

PI3K/AKT KINASE PATHWAY INHIBITORS IN PRE-CLINICAL DEVELOPMENT... 

Turek-Etienne, T.C., Lei, M., Terracciano, J.S., Langsdorf, E.F., Bryant, R.W., Hart, R.F., and Horan, A.C., Use of red-shifted dyes in a fluorescence polarization AKT kinase assay for detection of biological activity in natural product extracts, /. Biomol. Screen., 9, 52,2004. 

Matsuzaki H, TamataniM, MitsudaN, et al. Activation of Akt kinase inhibits apoptosis and changes in B cl- 2 and B ax expres sion induced by nitric oxide in primary hippocampal neurons. J Neu rochem... 

Fig. 1.57. Model of the regulation of translation by insulin. Insulin ( and other growth factors) activates the Akt kinase pathway (see ch. 10), whose final result is the phosphorylation of 4E-BPl, a regulatory protein of translation initiation. The 4E-BP1 protein inactivates the initation factor eIF-4E by complex formation. eIE-4E is required, together with the proteins eIE-4A and p220, for the binding of the 40S subunit of the ribosome to the cap structure of the mRNA. If the 4E-BP1 protein becomes phosphorylated as a result of insulin-mediated activation of the PI3 kinase/Akt kinase cascade, then eIE-4E is liberated from the inactive eIP-4E 4E-BPl complex and protein biosynthesis can begin.

The regulation of translation is accomplished in this system via a specific inhibitory protein and an initiation factor of translation. The binding activity of the inhibitor protein is regulated by protein phosphorylation, and thus, by protein kinases. The activity of protein kinases can be regulated in a multitude of ways. A signal initiating from insulin, for example, can activate the PI3-kinase and the Akt kinase pathway (see 6.6.2), resulting in phosphorylation of 4E-BP1. 

A further susceptible point for insulin-regulated signaling pathways is the ribosomal protein S6. Under the influence of insulin, S6 is phosphorylated by a specific protein kinase, the p70 kinase, resulting in increased levels of translation of certain mRNAs. Several pathways including the MAPK/ERK pathway (see chapter 10) and the Akt kinase pathway can contribute to the activation of the p70 kinase. 

Other members of class I of the PI3-kinases, such as PI3-kinase of the y subtype, are stimulated by interaction with Pycomplexes (see Chapter 5.5.7) and have their own regulatory subunit. It is interesting that both a lipid kinase activity and a protein kinase activity have been identified in the catalytic domain of the P13-kinase y subtype in brain (Bondeva et al., 1998). Activation of the MAPK pathway (see Chapter 10) may take place via the protein kinase activity, so that this enzyme can produce a bifurcated signal the lipid kinase activity stimulates the Akt kinase (see below), the protein kinase the MAPK pathway. Proliferation promoting signals are transmitted via both pathways. 

An important target protein of Ptdlns(3,4,5)P3 is Akt kinase, also known as protein kinase B (PKB). The signahng pathway for Akt kinase shown in Fig. 6.9b illustrates the role of P13-kinase and Ptdlns(3,4,5)P3 in growth factor controlled signal pathways that lead from the cell membrane into the cytosol and the nucleus. 

In the Akt signaling pathway (review Downward, 1998), first an extracellular growth factor activates the corresponding transmembrane receptor (e.g., PGDF receptor, see 8.1). Consequently, tyrosine phosphorylation takes place on the cytoplasmic domain of the receptor. The tyrosine residues serve as docking sites for the SH2 domain of the p85 subimit of the PI3-kinase. The associated translocation of PI3-kinase is synonymous with its activation. The PtdIns(3,4,5)P3 formed binds to the PH domain of the signal protein next in sequence, the Akt kinase, which recruits the latter to the membrane. 

The membrane-associated Akt kinase is now a substrate for protein kinase PDKl that phosphorylates a specific Thr and Ser residue of Akt kinase. The double phosphorylation converts Akt kinase to the active form. It is assumed that the Akt kinase now dissociates from the membrane and phosphorylates cytosolic substrates such as glycogen synthase kinase, 6-phosphofructo-2-kinase and ribosomal protein S6 kinase, p70 . According to this mechanism, Akt kinase regulates central metabolic pathways of the cell. Furthermore, it has a promoting influence on cell division and an inhibitory influence on programmed cell death, apoptosis. A role in apoptosis is suggested by the observation that a component of the apoptotic program. Bad protein (see Chapter 15) has been identified as a substrate of Akt kinase. 

The great importance of PtdIns(3,4,5)P3 metabolism for growth regulation is illustrated by the observation that an enzyme of PtdIns(3,4,5)P3 metabolism has been identified as a tumor suppressor protein (Wu et al., 1998). PTEN tumor suppressor protein has lipid phosphatase activity that is specific for hydrolysis of PtdIns(3,4,5)P3. It is assumed that PTEN lipid phosphatase is a negative regulator of the Akt pathway by lowering the concentration of PtdIns(3,4,5)P3 and counteracting stimulation of Akt kinase. 

Ras-mediated activation of P13-kinase also links the Ras protein to Akt kinase (see 6.6.1), which mediates antiapoptotic signals. 

Fig. 15.9. Antiapoptotic signalling by the PI3-kinase/Akt kinase pathway The PI3 kinase/Akt kinase pathway influences apoptosis via phosphorylation of the Bad protein, which is a member of the family of Bcl-2 proteins. Activation of the PI3-kinase pathway leads to Akt-kinase-catalyzed phosphorylation of Bad protein. Bad protein in its unphosphorylated form participates in activation of initiator caspases and thus has a proapoptotic effect. Phosphorylation of Bad protein by Akt kinase (or related kinases) has an antiapoptotic effect since phosphoryla-ted Bad protein is a binding substrate of 14-3-3 proteins. Bad is thus sequestered in an inactive state and is not available for triggering of apoptosis.

PI3-kinase (see 6.6) can mediate antiapoptotic signals, in addition to growth-promoting signals (Fig. 14.9). The antiapoptotic signal conduction starts at PI3-kinase to Akt kinase, which is activated by the messenger substance PtdInsPs formed by PI3-kinase. The Bad protein has been identified as a substrate of Akt kinase. The Bad protein is a proapoptotic member of the Bcl-2 family. It is phosphorylated by Akt kinase at several Ser residues and its proapoptotic effect is thus inhibited (Datta et al., 1997). Experimental evidence exists that the 14-3-3 proteins are involved in this inhibition these bind to phosphoserine residues of Bad protein and thus inactivate its proapoptotic fim-ction. 

Glutamate-mediated Ca2+ entry through NMDA at the plasma membrane level and mobilization of Ca2+from intracellular stores through PLC-mediated generation of PtdIns-3/J is indispensable for the basal NF-kB activity. Three cytosolic Ca2+ sensors, calmodulin, protein kinases C (PKC), and the p2 l(ras)/phosphatidylinositol 3-kinase (Ptdlns-3K)/Akt pathways, are simultaneously involved in the steps linking the Ca2+ to NF-kB activity (Lilienbaum and Israel, 2003 Marchetti et al., 2004 Lubin et al., 2005). Calmodulin modulates calcineurin, a Ca2+-dependent protein phosphatase, which plays a role in the basal NF-kB activity, whilst stimulation of both the calmodulin kinase II and Akt kinase pathways results in the up-regulation of the transcriptional potential of the p65 subunit of NF-/cB (Lilienbaum and Israel,... 

In the synthesis of a compound library of allosteric Akt kinase inhibitors 39, Lindsley and coworkers employed different HTS techniques (Scheme 24) [54]. A polymer-supported base and a fluorous thiol scavenger were used in the alkylation reaction of 40. F-SPE purified intermediate was then used for microwave-assisted cycloaddition of 41. Similar intermediates have been used for generation of an unnatural canthine alkaloid library 42 by performing cycloaddition reactions with an indo-tethered acyl hy-drazide [55]. 

K. Yamashita, J. Kajstura, D.J. Dischcr, B.J. Wasserlauf, N.H. Bishopric, P. Anversa and K.A. Webster, Reperfusion-activated Akt kinase prevents apoptosis in transgenic mouse hearts overexpressing insulinlike growth factor-1, Circ. Res. 88, 609-614 (2001). 

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