butyrolactone-i and olomoucine

The eukaryotic cell division cycle is regulated by a family of protein kinases, the cyclin-dependent kinases (cdk's), constituted of at least two subunits, a catalytic subunit (cdk1-7) associated with a regulatory subunit (cyclin A-H). Transient activation of cdk's is responsible for transition through the different phases of the cell cycle. Major abnormalities of cdk's expression and regulation have been described in human tumours. Enzymatic screening is starting to uncover chemical inhibitors of cdk's with anti-mitotic activities. This review summarizes our knowledge of these first inhibitors, their mechanism of action, their effects on the cell cycle, and discusses the potential of such type of inhibitors as anti-tumour agents.

Topics: 4-Butyrolactone; Animals; Cell Cycle; Cyclin-Dependent Kinases; Enzyme Inhibitors; Flavonoids; Humans; Kinetin; Piperidines; Purines; Staurosporine; Suramin

Cyclin-dependent kinases (CDKs) mediate proliferation and neuronal development, while aberrant CDK activity is associated with cancer and neurodegeneration. Consequently, pharmacologic inhibitors, such as 2,6,9-trisubstituted purines, which potently inhibit CDKs 1, 2, and 5, were developed to combat these pathologies. One agent, R-roscovitine (CYC202), has advanced to clinical trials as a potential cancer therapy. In primary neuronal cultures, these agents have been used to delineate the physiologic and pathologic functions of CDKs, and associated signaling pathways. Herein we demonstrate that three 2,6,9-trisubstituted purines: olomoucine, roscovitine, and purvalanol, used at concentrations ascribed by others to potently inhibit CDKs 1, 2, and 5, are powerful triggers of death in maturing cerebellar granule neurons, assessed by loss of mitochondrial reductive capacity and differential staining with fluorescent indicators of living/dead neurons. Based on several criteria, including delayed time course and establishment of an irreversible commitment point of death, pyknotic cell and nuclear morphology, and caspase-3 cleavage, the death process is apoptotic. However, pharmacological and biochemical data indicate that apoptosis is independent of CDK 1, 2, or 5 inhibition. This is based on the pattern of changes in c-jun mRNA, c-Jun protein, and Ca(2+)/cAMP response element binding protein (CREB) phosphorylation, and also, the ineffectiveness of structurally distinct CDK 1, 2, and 5 inhibitors butyrolactone-1 and PNU112445A to induce apoptosis. Collectively, our results, and those of others, indicate that the CDK regulation of transcription (CDKs 7 and 9) should be examined as a target of these agents, and as an indirect mediator of neuronal fate.

Topics: 4-Butyrolactone; Animals; Apoptosis; CDC2 Protein Kinase; CDC2-CDC28 Kinases; Cerebellum; Cyclic AMP Response Element-Binding Protein; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase 5; Cyclin-Dependent Kinase 9; Cyclin-Dependent Kinases; Enzyme Inhibitors; JNK Mitogen-Activated Protein Kinases; Kinetin; Mitogen-Activated Protein Kinases; Neurons; Phosphorylation; Proto-Oncogene Proteins; Purines; Pyrimidines; Rats; Rats, Sprague-Dawley; Roscovitine; Signal Transduction; Sulfonamides

The present study was aimed to investigate whether or not cyclin-dependent kinases (CDKs) participate in different cascades leading to apoptosis. We examined the effects of two CDK inhibitors, olomoucine (OLM) and butyrolactone-I (BL-I), on apoptosis induced in two kinds of Drosophila cell lines. Increases of caspase activity induced by actinomycin D, cycloheximide, H-7 or A23187 in a Drosophila neuronal cell line, ML-DmBG2-c2, and induced by excessive expression of a Drosophila cell death gene, reaper, in Drosophila S2 cells were suppressed by 24-h pretreatment of each CDK inhibitor. Concomitant with the suppression of the caspase activity, fragmentations of cells and DNA, representatives of apoptosis, were also inhibited. These results suggest that CDK(s) participates in progression of apoptosis. However, these effects of the CDK inhibitors were also observed even at lower doses which did not affect cell proliferation. Therefore, it was shown that apoptosis is not always related to cell cycle in Drosophila cells. It was also suggested that the target(s) of the CDK inhibitors locates upstream of caspase in the cascade(s) of apoptosis.

Topics: 4-Butyrolactone; Animals; Apoptosis; Cells, Cultured; Cyclin-Dependent Kinases; Dose-Response Relationship, Drug; Drosophila melanogaster; Drosophila Proteins; Enzyme Inhibitors; Gene Expression Regulation; Kinetin; Peptides; Purines; Time Factors

Cyclin-dependent kinases (CDKs) mediate cell-cycle phase transitions. Recently, CDKs have been associated with non-cell-cycle roles such as DNA repair, transcription, and phosphate metabolism in yeast. The cyclical processes, circadian rhythms and the eukaryotic cell cycle, are similar in many respects. It is possible that a kinase like CDK is involved in the control of circadian rhythms. In this study, the effects of CDK inhibitors (olomoucine, roscovitine, and butyrolactone I) on the Aplysia ocular circadian rhythm were investigated. Continuous treatments with olomoucine (10 microM) lengthened the free-running period of the rhythm, and pulse treatments of olomoucine (6 h, 100 microM) delayed the rhythm. The effects of olomoucine on the rhythm were qualitatively similar to those of a reversible inhibitor of transcription, 5,6-dichloro-beta-1-ribobenzimidazole. Subsequently, olomoucine was found to inhibit RNA synthesis in the eye of Aplysia and Bulla. All of the other CDK inhibitors used in this study also inhibited transcription in the eye of Aplysia, and their effects on transcription correlated with their effects on the circadian rhythm. This study adds substantial evidence to that previously obtained by using 5,6-dichloro-beta-1-ribobenzimidazole for a role of transcription in the mechanism responsible for circadian rhythmicity in the eye of Aplysia. Also, these results indicate that caution is warranted in interpreting results obtained by using CDK inhibitors, because these drugs appear to inhibit transcription as well as CDKs.

Topics: 4-Butyrolactone; Action Potentials; Animals; Aplysia; Circadian Rhythm; Cyclin-Dependent Kinases; Electrophysiology; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Kinetin; Macromolecular Substances; Neural Conduction; Ocular Physiological Phenomena; Purines; RNA, Messenger; Roscovitine; Transcription, Genetic