alvocidib and Bone-Neoplasms

Flavopiridol is the first cyclin-dependent kinase (cdk) inhibitor to enter clinical trials. Serum levels of flavopiridol obtained during phase I studies were sufficient to inhibit in vitro cancer cell growth. Because responses were observed in kidney cancer patients in the phase I trials, we performed a phase II trial of flavopiridol in this patient population.. Thirty-five minimally pretreated patients were accrued using a standard two-step mechanism. Flavopiridol (50 mg/m(2)/d) was administered by continuous infusion for 72 hours every 2 weeks, and response was evaluated every 8 weeks. Peripheral blood mononuclear cells (PBMCs) were collected at baseline, at completion of drug infusion, and on day 7 of the first therapy cycle, and cell cycle parameters after phytohemagglutinin and interleukin-2 stimulation were assessed.. There were two objective responses (response rate = 6%, 95% confidence interval, 1% to 20%). The most common toxicities were asthenia, occurring in 83% of patients (grade 3 or 4 in 9%), and diarrhea, occurring in 77% of patients (grade 3 or 4 in 20%). Also, nine patients (26%) experienced grade 3 or 4 vascular thrombotic events, including one myocardial infarction, two transient neurologic ischemic attacks, four deep venous thrombosis, and two pulmonary emboli. Cell cycle studies did not reveal any effect of flavopiridol on stimulated PBMCs.. Flavopiridol, at the dose and schedule administered in this trial, is ineffective in metastatic renal cancer. In addition to the diarrhea observed in phase I studies, we also observed a higher incidence of asthenia and serious vascular thrombotic events than expected.

Topics: Adult; Aged; Antineoplastic Agents; Asthenia; Bone Neoplasms; Carcinoma, Renal Cell; Diarrhea; Drug Administration Schedule; Female; Flavonoids; Humans; Kidney Neoplasms; Liver Neoplasms; Lung Neoplasms; Lymphatic Metastasis; Male; Middle Aged; Piperidines; Thrombosis; Treatment Outcome

Selective cyclin-dependent kinase (cdk) 2 inhibition is readily compensated. However, reduced cdk2 activity may have antiproliferative effects in concert with other family members. Here, inducible RNA interference was used to codeplete cdk2 and cdk1 from NCI-H1299 non-small cell lung cancer and U2OS osteosarcoma cells, and effects were compared with those mediated by depletion of either cdk alone. Depletion of cdk2 slowed G1 progression of NCI-H1299 cells and depletion of cdk1 slowed G2-M progression in both cell lines, with associated endoreduplication in U2OS cells. However, compared with the incomplete cell cycle blocks produced by individual depletion, combined depletion had substantial consequences, with G2-M arrest predominating in NCI-H1299 cells and apoptosis the primary outcome in U2OS cells. In U2OS cells, combined depletion affected RNA polymerase II expression and phosphorylation, causing decreased expression of the antiapoptotic proteins Mcl-1 and X-linked inhibitor of apoptosis (XIAP), effects usually mediated by inhibition of the transcriptional cdk9. These events do not occur after individual depletion of cdk2 and cdk1, suggesting that reduction of cdk2, cdk1, and RNA polymerase II activities all contribute to apoptosis in U2OS cells. The limited cell death induced by combined depletion in NCI-H1299 cells was significantly increased by codepletion of cdk9 or XIAP or by simultaneous treatment with the cdk9 inhibitor flavopiridol. These results show the potency of concomitant compromise of cell cycle and transcriptional cdk activities and may guide the selection of clinical drug candidates.

Topics: Apoptosis; Bone Neoplasms; Carcinoma, Non-Small-Cell Lung; CDC2 Protein Kinase; Cell Division; Cell Line, Tumor; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 9; Flavonoids; G1 Phase; G2 Phase; Humans; Lung Neoplasms; Neoplasms; Osteosarcoma; Piperidines; RNA Polymerase II; RNA, Small Interfering

Breast cancer micrometastases in the bone marrow are resistant to chemotherapy. They can remain dormant for years before some begin to proliferate. We seek to understand survival mechanisms and develop targeted approaches to eliminating these cells.. In an in vitro model of dormancy, basic fibroblast growth factor 2 (FGF-2), abundant in the bone marrow, inhibits the growth of well-differentiated cells in the 2- to 10-cell stage and up-regulates integrin alpha(5)beta(1). Through this integrin, cells bind fibronectin, spread out, and acquire a survival advantage, partly through activation of the phosphatidylinositol 3-kinase/Akt pathway. We investigated the effects of Taxotere, flavopiridol, and mitogen-activated protein/extracellular signal-regulated kinase (ERK) kinase and p38 inhibitors on survival of dormant clones and that of flavopiridol on expression of integrins, adhesion strength, and phosphorylation of Akt, ERK 1/2, and p38.. Dormant MCF-7 and T-47D cell clones were resistant to Taxotere concentrations 10-fold higher than needed to eliminate growing clones but were almost completely eradicated by 200 nmol/L flavopiridol. Flavopiridol caused a decrease in FGF-2-induced expression of integrins, including alpha(5) and beta(1), and decreased FGF-2-induced specific adhesion to fibronectin. It diminished Akt phosphorylation, but reexpression of active Akt was not sufficient to reverse dormant clone inhibition. Flavopiridol did not affect phosphorylation of ERK 1/2 and p38 but diminished total protein levels. Chemical inhibition of these pathways partially abrogated dormant clone survival.. Flavopiridol has pleiotropic effects on key targets involved with survival of dormant breast cancer cells and may represent a useful approach to eliminating cells dependent on multiple signal pathways for survival.

Topics: Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Bone Marrow; Bone Neoplasms; Breast Neoplasms; Cell Adhesion; Cell Survival; Docetaxel; Extracellular Signal-Regulated MAP Kinases; Female; Fibroblast Growth Factor 2; Flavonoids; Gene Expression Regulation, Neoplastic; Humans; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Piperidines; Taxoids; Tumor Cells, Cultured

Flavopiridol is the potent inhibitor of cdks sharing its function with endogenous cdk inhibitors, and causes arrest at both the G1 and G2 phases of the cell cycle resulting in apoptosis in various tumor cell lines. Cyclin-dependent kinase inhibitor p16INK4a induces cell cycle arrest in G1 or G2 or both, and is inactivated in many malignant tumors. In this study, we focused on the effects of flavopiridol on chemically-induced rat lung adenocarcinoma, osteosarcoma and malignant fibrous histiocytoma (MFH) cell lines showing different pattern of p16INK4a status. The data demonstrated that flavopiridol inhibited cellular growth in a dose- and time-dependent manner, inducing apoptosis within 24 h in all cell lines at a concentration of 300 nM. The growth inhibition rate was the greatest for lung adenocarcinoma cells, lacking p16INK4a expression associated with methylation-mediated gene silencing; 83% at a concentration of 300 nM for 72-h treatment; while the growth of osteosarcoma and MFH cells, both expressing p16INK4a, were inhibited at similar levels; 54-61% for osteosarcoma and 61-64% for MFH cell lines. Then, we further investigated the influence of p16INK4a induction upon the effect of flavopiridol in p16INK4a-deficient lung adenocarcinoma cells. 5-aza 2'-deoxycytidine (5-Aza-CdR) induced p16INK4a expression and inhibited cellular growth in lung adenocarcinoma at a similar level to that with flavopiridol treatment. After the induction of p16INK4a expression by 5-Aza-CdR, the growth inhibition rates of flavopiridol in the p16INK4a-induced lung adenocarcinoma cells could not achieve comparable inhibition to that in the p16INK4a-deficient cells; the efficacy was reduced compared to original p16INK4a-deficient cells at each concentration of 50, 100 and 500 nM for 72-h treatment. These data indicate that flavopiridol shows cell type specific inhibition and possibly acts in a more compensatory manner for endogenous p16INK4a function in tumor cells having the aberrations of p16INK4a gene.

Topics: Adenocarcinoma; Animals; Apoptosis; Bone Neoplasms; Cell Division; Cell Line, Tumor; Cyclin D1; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinase Inhibitor p16; Cyclin-Dependent Kinases; DNA Methylation; Flavonoids; Gene Expression Regulation, Neoplastic; Histiocytoma, Benign Fibrous; Lung Neoplasms; Osteosarcoma; Piperidines; Promoter Regions, Genetic; Proto-Oncogene Proteins; Rats; RNA, Messenger

Transformed cells are selectively sensitized to apoptosis induced by the cyclin-dependent kinase inhibitor flavopiridol after their recruitment to S phase. During S phase, cyclin A-dependent kinase activity neutralizes E2F-1 allowing orderly S phase progression. Inhibition of cyclin A-dependent kinase by flavopiridol could cause inappropriately persistent E2F-1 activity during S phase traversal and exit. Transformed cells, with high baseline levels of E2F-1 activity, may be particularly sensitive to cyclin A-dependent kinase inhibition, as the residual level of E2F-1 activity that persists may be sufficient to induce apoptosis. Here, we demonstrate that flavopiridol treatment during S phase traversal results in persistent expression of E2F-1. The phosphorylation of E2F-1 is markedly diminished, whereas that of the retinoblastoma protein is minimally affected, so that E2F-1/DP-1 heterodimers remain bound to DNA. In addition, manipulation of E2F-1 levels leads to predictable outcomes when cells are exposed to flavopiridol during S phase. Tumor cells expressing high levels of ectopic E2F-1 are more sensitive to flavopiridol-induced apoptosis during S phase compared with parental counterparts, and high levels of ectopic E2F-1 expression are sufficient to sensitize nontransformed cells to flavopiridol. Furthermore, E2F-1 activity is required for flavopiridol-induced apoptosis during S phase, which is severely compromised in cells homozygous for a nonfunctional E2F-1 allele. Finally, the response to flavopiridol during S phase is blunted in cells expressing a nonphosphorylatable E2F-1 mutant incapable of binding cyclin A, suggesting that the modulation of E2F-1 activity produced by flavopiridol-mediated cyclin-dependent kinase inhibition is critical for the apoptotic response of S phase cells.

Topics: Antineoplastic Agents; Apoptosis; Bone Neoplasms; Carcinoma, Non-Small-Cell Lung; Cell Cycle; Cell Cycle Proteins; Cell Line, Transformed; Cell Line, Tumor; DNA-Binding Proteins; DNA, Neoplasm; Drug Synergism; E2F Transcription Factors; E2F1 Transcription Factor; Enzyme Inhibitors; Flavonoids; Humans; Lung Neoplasms; Osteosarcoma; Phosphorylation; Piperidines; Protein Kinase Inhibitors; Protein Kinases; S Phase; Transcription Factor DP1; Transcription Factors