Journal Article•
Specific inhibition of cyclin-dependent kinase 4/6 by PD 0332991 and associated antitumor activity in human tumor xenografts
TL;DR: Results indicate that inhibition of Cdk4/6 alone is sufficient to cause tumor regression and a net reduction in tumor burden in some tumors.
Abstract: PD 0332991 is a highly specific inhibitor of cyclin-dependent kinase 4 (Cdk4) (IC50, 0.011 micromol/L) and Cdk6 (IC50, 0.016 micromol/L), having no activity against a panel of 36 additional protein kinases. It is a potent antiproliferative agent against retinoblastoma (Rb)-positive tumor cells in vitro, inducing an exclusive G1 arrest, with a concomitant reduction of phospho-Ser780/Ser795 on the Rb protein. Oral administration of PD 0332991 to mice bearing the Colo-205 human colon carcinoma produces marked tumor regression. Therapeutic doses of PD 0332991 cause elimination of phospho-Rb and the proliferative marker Ki-67 in tumor tissue and down-regulation of genes under the transcriptional control of E2F. The results indicate that inhibition of Cdk4/6 alone is sufficient to cause tumor regression and a net reduction in tumor burden in some tumors.
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TL;DR: Among patients with previously untreated ER-positive, HER2-negative advanced breast cancer, palbociclib combined with letrozole resulted in significantly longer progression-free survival than that with let rozole alone, although the rates of myelotoxic effects were higher with palbokiclib-letrozoles.
Abstract: BackgroundA phase 2 study showed that progression-free survival was longer with palbociclib plus letrozole than with letrozole alone in the initial treatment of postmenopausal women with estrogen-receptor (ER)–positive, human epidermal growth factor receptor 2 (HER2)–negative advanced breast cancer. We performed a phase 3 study that was designed to confirm and expand the efficacy and safety data for palbociclib plus letrozole for this indication. MethodsIn this double-blind study, we randomly assigned, in a 2:1 ratio, 666 postmenopausal women with ER-positive, HER2-negative breast cancer, who had not had prior treatment for advanced disease, to receive palbociclib plus letrozole or placebo plus letrozole. The primary end point was progression-free survival, as assessed by the investigators; secondary end points were overall survival, objective response, clinical benefit response, patient-reported outcomes, pharmacokinetic effects, and safety. ResultsThe median progression-free survival was 24.8 months (95...
1,737 citations
TL;DR: Palbociclib (PD-0332991) is an oral, small-molecule inhibitor of cyclin-dependent kinases (CDKs) 4 and 6 with preclinical evidence of growth-inhibitory activity in oestrogen receptor-positive breast cancer cells and synergy with anti-oestrogens as mentioned in this paper.
Abstract: Summary Background Palbociclib (PD-0332991) is an oral, small-molecule inhibitor of cyclin-dependent kinases (CDKs) 4 and 6 with preclinical evidence of growth-inhibitory activity in oestrogen receptor-positive breast cancer cells and synergy with anti-oestrogens. We aimed to assess the safety and efficacy of palbociclib in combination with letrozole as first-line treatment of patients with advanced, oestrogen receptor-positive, HER2-negative breast cancer. Methods In this open-label, randomised phase 2 study, postmenopausal women with advanced oestrogen receptor-positive and HER2-negative breast cancer who had not received any systemic treatment for their advanced disease were eligible to participate. Patients were enrolled in two separate cohorts that accrued sequentially: in cohort 1, patients were enrolled on the basis of their oestrogen receptor-positive and HER2-negative biomarker status alone, whereas in cohort 2 they were also required to have cancers with amplification of cyclin D1 ( CCND1 ), loss of p16 (INK4A or CDKN2A), or both. In both cohorts, patients were randomly assigned 1:1 via an interactive web-based randomisation system, stratified by disease site and disease-free interval, to receive continuous oral letrozole 2·5 mg daily or continuous oral letrozole 2·5 mg daily plus oral palbociclib 125 mg, given once daily for 3 weeks followed by 1 week off over 28-day cycles. The primary endpoint was investigator-assessed progression-free survival in the intention-to-treat population. Accrual to cohort 2 was stopped after an unplanned interim analysis of cohort 1 and the statistical analysis plan for the primary endpoint was amended to a combined analysis of cohorts 1 and 2 (instead of cohort 2 alone). The study is ongoing but closed to accrual; these are the results of the final analysis of progression-free survival. The study is registered with the ClinicalTrials.gov, number NCT00721409. Findings Between Dec 22, 2009, and May 12, 2012, we randomly assigned 165 patients, 84 to palbociclib plus letrozole and 81 to letrozole alone. At the time of the final analysis for progression-free survival (median follow-up 29·6 months [95% CI 27·9–36·0] for the palbociclib plus letrozole group and 27·9 months [25·5–31·1] for the letrozole group), 41 progression-free survival events had occurred in the palbociclib plus letrozole group and 59 in the letrozole group. Median progression-free survival was 10·2 months (95% CI 5·7–12·6) for the letrozole group and 20·2 months (13·8–27·5) for the palbociclib plus letrozole group (HR 0·488, 95% CI 0·319–0·748; one-sided p=0·0004). In cohort 1 (n=66), median progression-free survival was 5·7 months (2·6–10·5) for the letrozole group and 26·1 months (11·2–not estimable) for the palbociclib plus letrozole group (HR 0·299, 0·156–0·572; one-sided p Interpretation The addition of palbociclib to letrozole in this phase 2 study significantly improved progression-free survival in women with advanced oestrogen receptor-positive and HER2-negative breast cancer. A phase 3 trial is currently underway. Funding Pfizer.
1,466 citations
TL;DR: This cellular thermal shift assay (CETSA) is based on the biophysical principle of ligand-induced thermal stabilization of target proteins and validated drug binding for a set of important clinical targets and monitored processes of drug transport and activation, off-target effects and drug resistance in cancer cell lines, as well as drug distribution in tissues.
Abstract: The efficacy of therapeutics is dependent on a drug binding to its cognate target. Optimization of target engagement by drugs in cells is often challenging, because drug binding cannot be monitored inside cells. We have developed a method for evaluating drug binding to target proteins in cells and tissue samples. This cellular thermal shift assay (CETSA) is based on the biophysical principle of ligand-induced thermal stabilization of target proteins. Using this assay, we validated drug binding for a set of important clinical targets and monitored processes of drug transport and activation, off-target effects and drug resistance in cancer cell lines, as well as drug distribution in tissues. CETSA is likely to become a valuable tool for the validation and optimization of drug target engagement.
1,305 citations
TL;DR: How CDK inhibitors with high selectivity (particularly for both CDK4 and CDK6), in combination with patient stratification, have resulted in more substantial clinical activity is discussed.
Abstract: Cancer represents a pathological manifestation of uncontrolled cell division; therefore, it has long been anticipated that our understanding of the basic principles of cell cycle control would result in effective cancer therapies. In particular, cyclin-dependent kinases (CDKs) that promote transition through the cell cycle were expected to be key therapeutic targets because many tumorigenic events ultimately drive proliferation by impinging on CDK4 or CDK6 complexes in the G1 phase of the cell cycle. Moreover, perturbations in chromosomal stability and aspects of S phase and G2/M control mediated by CDK2 and CDK1 are pivotal tumorigenic events. Translating this knowledge into successful clinical development of CDK inhibitors has historically been challenging, and numerous CDK inhibitors have demonstrated disappointing results in clinical trials. Here, we review the biology of CDKs, the rationale for therapeutically targeting discrete kinase complexes and historical clinical results of CDK inhibitors. We also discuss how CDK inhibitors with high selectivity (particularly for both CDK4 and CDK6), in combination with patient stratification, have resulted in more substantial clinical activity.
1,276 citations
TL;DR: The incidence of melanoma is rising steadily in western populations — the number of cases worldwide has doubled in the past 20 years — and these advances are being exploited to provide targeted drugs and new therapeutic approaches.
Abstract: Melanoma is a cancer that arises from melanocytes, specialized pigmented cells that are found predominantly in the skin. The incidence of melanoma is rising steadily in western populations--the number of cases worldwide has doubled in the past 20 years. In its early stages malignant melanoma can be cured by surgical resection, but once it has progressed to the metastatic stage it is extremely difficult to treat and does not respond to current therapies. Recent discoveries in cell signalling have provided greater understanding of the biology that underlies melanoma, and these advances are being exploited to provide targeted drugs and new therapeutic approaches.
1,262 citations
References
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TL;DR: Unlike other quantitative PCR methods, real-time PCR does not require post-PCR sample handling, preventing potential PCR product carry-over contamination and resulting in much faster and higher throughput assays.
Abstract: We have developed a novel "real time" quantitative PCR method. The method measures PCR product accumulation through a dual-labeled fluorogenic probe (i.e., TaqMan Probe). This method provides very accurate and reproducible quantitation of gene copies. Unlike other quantitative PCR methods, real-time PCR does not require post-PCR sample handling, preventing potential PCR product carry-over contamination and resulting in much faster and higher throughput assays. The real-time PCR method has a very large dynamic range of starting target molecule determination (at least five orders of magnitude). Real-time quantitative PCR is extremely accurate and less labor-intensive than current quantitative PCR methods.
6,367 citations
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TL;DR: Genetic alterations affecting p16INK4a and cyclin D1, proteins that govern phosphorylation of the retinoblastoma protein and control exit from the G1 phase of the cell cycle, are so frequent in human cancers that inactivation of this pathway may well be necessary for tumor development.
Abstract: Uncontrolled cell proliferation is the hallmark of cancer, and tumor cells have typically acquired damage to genes that directly regulate their cell cycles. Genetic alterations affecting p16(INK4a) and cyclin D1, proteins that govern phosphorylation of the retinoblastoma protein (RB) and control exit from the G1 phase of the cell cycle, are so frequent in human cancers that inactivation of this pathway may well be necessary for tumor development. Like the tumor suppressor protein p53, components of this "RB pathway," although not essential for the cell cycle per se, may participate in checkpoint functions that regulate homeostatic tissue renewal throughout life.
5,509 citations
TL;DR: Although the Ki‐67 protein is well characterized on the molecular level and extensively used as a proliferation marker, the functional significance still remains unclear; there are indications, however, that Ki‐ 67 protein expression is an absolute requirement for progression through the cell‐division cycle.
Abstract: The expression of the human Ki-67 protein is strictly associated with cell proliferation. During interphase, the antigen can be exclusively detected within the nucleus, whereas in mitosis most of the protein is relocated to the surface of the chromosomes. The fact that the Ki-67 protein is present during all active phases of the cell cycle (G(1), S, G(2), and mitosis), but is absent from resting cells (G(0)), makes it an excellent marker for determining the so-called growth fraction of a given cell population. In the first part of this study, the term proliferation marker is discussed and examples of the applications of anti-Ki-67 protein antibodies in diagnostics of human tumors are given. The fraction of Ki-67-positive tumor cells (the Ki-67 labeling index) is often correlated with the clinical course of the disease. The best-studied examples in this context are carcinomas of the prostate and the breast. For these types of tumors, the prognostic value for survival and tumor recurrence has repeatedly been proven in uni- and multivariate analysis. The preparation of new monoclonal antibodies that react with the Ki-67 equivalent protein from rodents now extends the use of the Ki-67 protein as a proliferation marker to laboratory animals that are routinely used in basic research. The second part of this review focuses on the biology of the Ki-67 protein. Our current knowledge of the Ki-67 gene and protein structure, mRNA splicing, expression, and cellular localization during the cell-division cycle is summarized and discussed. Although the Ki-67 protein is well characterized on the molecular level and extensively used as a proliferation marker, the functional significance still remains unclear. There are indications, however, that Ki-67 protein expression is an absolute requirement for progression through the cell-division cycle.
4,359 citations
"Specific inhibition of cyclin-depen..." refers background in this paper
...Moreover, there was a concomitant elimination of staining for Ki-67, a commonly measured marker of proliferation (83)....
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...This effect also was associated with a substantial reduction in staining for Ki-67, a well-known marker of proliferation (83)....
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TL;DR: The 5'----3' exonuclease activity of the thermostable enzyme Thermus aquaticus DNA polymerase may be employed in a polymerase chain reaction product detection system to generate a specific detectable signal concomitantly with amplification.
Abstract: The 5'----3' exonuclease activity of the thermostable enzyme Thermus aquaticus DNA polymerase may be employed in a polymerase chain reaction product detection system to generate a specific detectable signal concomitantly with amplification. An oligonucleotide probe, nonextendable at the 3' end, labeled at the 5' end, and designed to hybridize within the target sequence, is introduced into the polymerase chain reaction assay. Annealing of probe to one of the polymerase chain reaction product strands during the course of amplification generates a substrate suitable for exonuclease activity. During amplification, the 5'----3' exonuclease activity of T. aquaticus DNA polymerase degrades the probe into smaller fragments that can be differentiated from undegraded probe. The assay is sensitive and specific and is a significant improvement over more cumbersome detection methods.
3,050 citations
TL;DR: The eukaryotic protein kinases make up a large superfamily of homologous proteins, and a classification scheme can be founded on a kinase domain phylogeny, which reveals families of enzymes that have related substrate specificities and modes of regulation.
Abstract: The eukaryotic protein kinases make up a large superfamily of homologous proteins. They are related by virtue of their kinase domains (also known as catalytic domains), which consist of approximately 250-300 amino acid residues. The kinase domains that define this group of enzymes contain 12 conserved subdomains that fold into a common catalytic core structure, as revealed by the 3-dimensional structures of several protein-serine kinases. There are two main subdivisions within the superfamily: the protein-serine/threonine kinases and the protein-tyrosine kinases. A classification scheme can be founded on a kinase domain phylogeny, which reveals families of enzymes that have related substrate specificities and modes of regulation.
2,730 citations
"Specific inhibition of cyclin-depen..." refers background in this paper
...PD 0332991 was tested against 39 individual serine, threonine, and tyrosine kinases, representing most of the primary protein kinase families (84)....
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