As gatekeepers of the cell cycle, the cyclin-dependent kinases are often implicated in the progression of cancer and make prime targets for therapy.
As gatekeepers of the cell cycle, the cyclin-dependent kinases are often implicated in the progression of cancer and make prime targets for therapy. Mirroring the cellular process that they target, two decades of CDK inhibitor development has cycled through excitement to disappointment, back to potential success.
Dual inhibitors of CDK4/6 are showing particular promise for the treatment of advanced breast cancer, and several agents are vying for success in the late stages of clinical development. Meanwhile, a pan-CDK inhibitor, dinaciclib, has worked its way through to phase III in chronic lymphocytic leukemia (CLL).
The translation of phase II success stories into the final stages of clinical development is eagerly anticipated, and it seems the stage may finally be set for approval of a CDK inhibitor for anticancer therapy.
The cell cycle is a highly regulated series of steps that takes the cell from a noncycling, quiescent state (G0) through an initial growth phase (G1), which is a regulatory and preparatory period leading up to a period of DNA replication (S), followed by a second growth phase (G2), involving rapid growth and protein synthesis, and ending in mitosis (Figure 1), during which the genetic material is condensed into chromosomes and divided equally into two genetically identical daughter cells (Figure 2).
CDK4/6 indicates cyclin-dependent kinases 4 and 6; E2F, transcription factors of E2F gene; G, growth phases, M, mitosis; RB, retinoblastoma protein; S, DNA replication.
Progression through the cell cycle is controlled by a wealth of specific cell cycle-associated proteins that act as “gatekeepers,” ensuring that the transition between the distinct phases of the cell cycle occur only at the appropriate time.
One particularly important group of gatekeeper proteins are the cyclin-dependent kinases (CDKs). The human genome encodes 21 CDKs, although only 7 (CDK1, 2, 3, 4, 6, 10, and 11) have been shown to have a direct role in the cell cycle. Other CDKs play an indirect, though no less important, role via activation of fellow CDKs or regulation of transcription.
The activity of CDKs is meticulously regulated via a number of activating and inhibitory mechanisms, but primarily through binding to cyclins. Cyclins were originally named for the cycling of their activity over the course of the cell cycle in response to growth signals in the cellular environment, which occurs in a distinct pattern for each cyclin.
In the process of mitosis, chromosomes replicate and produce two identical nuclei in preparation for cell division. Aberrant CDK4/6 activity ultimately dysregulates mytosis. Courtesy of the National Human Genome Research Institute. “Talking Glossary of Genetic Terms.”
The best-known and most important cyclins in cell cycle transitions are cyclins A, B, D, and E. D-type cyclins, partnered with CDK4 and CDK6, function in mid-to-late G1 and, in concert with cyclin E/CDK2, primarily regulate the G1—S transition. Cyclin A activates CDK1 and CDK2 in S phase and regulates the S/G2 transition, and the B-type cyclins, particularly cyclin B1 partnered with CDK1, drive entry into mitosis.
The transition from G1—S phase is particularly important as it is at this point that the cell commits to entering the cell cycle—the so-called restriction point, beyond which cells proceed into S phase regardless of external stimuli. The cyclin D-dependent kinases CDK4 and CDK6, as well as cyclin E/CDK2, are the primary regulators of this transition through activation of downstream signaling components. It is now appreciated that an array of downstream targets are activated and that there is significant crosstalk between them, such that CDK4/6 forms a central node in a complex network of signaling pathways. However, the best-characterized downstream target is the retinoblastoma protein (pRB).
pRB forms multiprotein complexes with a variety of other signaling proteins, including the E2F transcription factors, which it maintains in an inactive state. Cyclin D activates CDK4/6, which phosphorylates pRB and removes the repression of E2F transcription factors. This activates transcription of E2F target genes, which include many that are important for G1—S transition, ultimately pushing the cell past the restriction point.
Uncontrolled cellular proliferation was recognized by Douglas Hanahan and Robert Weinberg as one of the hallmarks of cancer. Therefore, it is unsurprising that components of the cell cycle, particularly CDK4 and CDK6, which regulate entry into the cell cycle, are highly dysregulated in cancer.
In fact, amplifications, mutations, chromosomal translocations, and deletions of the CDKs and their regulatory subunits are among some of the most common genetic alterations observed in cancer. The CDK inhibitor p16 (also known as multiple tumor suppressor 1), which acts upstream of CDK4/6 to inhibit their activation, is the most frequently deleted locus across all human cancers.
Amplifications in the cyclin D1 gene (CCND1) are common in breast cancer, melanoma, and head and neck squamous cell carcinoma (HNSCC); a chromosomal translocation that affects CCND1 is found in all cases of mantle cell lymphoma (MCL). CDK4 gene amplification is commonly observed in glioblastomas and breast cancer, in particular the luminal B and HER2- expressing subtypes.
Dysregulation of CDKs in cancer is not limited to genetic alterations and can also occur as a result of alterations in upstream signaling pathways, leading to hyperactivation of CDKs.
Sustained activation of CDKs like CDK4/6 allows the cancer cell to continuously trigger the G1—S switch to enter the cell cycle, leading to uncontrolled tumor cell proliferation.
As a result of their key role in driving oncogenesis and the fact that they are readily druggable kinases, CDKs are a prime target for therapeutic intervention. Inhibitors targeting the protein kinase enzymatic activity of CDKs have been in development for more than two decades, and hundreds of potential molecules have been tested.
Flavopiridol (alvocidib), a flavonoid alkaloid intravenously administered inhibitor of CDK2, 7, and 9, was the first to enter clinical trials. However, flavopiridol and other first-generation CDK inhibitors that emerged shortly afterward, such as roscovitine (seliciclib) , SNS-032, and AT7519 showed disappointing activity as single agents. They often displayed a low therapeutic index or significant toxicity, which hindered their development, although several continue to be evaluated in phase I and II clinical trials with modified dosing regimens or in combination with other anticancer agents (Table).
The field has undergone a renaissance of late and, following success in phase II trials, several newer agents have progressed to phase III. These agents predominantly target CDK4 and CDK6, and are much more specific and potent inhibitors than their predecessors.
The three front-runners are Pfizer’s palbociclib (PD-0332991), Novartis’ LEE011, and Eli Lilly’s abemaciclib (LY2835219). All are currently being evaluated in phase III trials in patients with breast cancer and phase I and II trials in a range of other malignancies (Table).
Results for both palbociclib and LY2835219 recently were reported at the American Association for Cancer Research (AACR) Annual Meeting.
The PALOMA-1 phase II study of palbociclib in combination with letrozole in postmenopausal women with estrogen receptor-positive (ER+)/HER2-negative, locally advanced metastatic breast cancer was presented by Richard Finn, MD, David Geffen School of Medicine at UCLA. The reported near-doubling of progression-free survival (PFS) at 20.2 months for palbociclib/letrozole versus 10.2 months for letrozole alone was somewhat disappointing compared with earlier reports that suggested a tripling of PFS; a lack of statistically significant improvement in overall survival (OS) was also shown. However, given the aggressive nature of the disease, the results are impressive nonetheless.
Furthermore, the secondary endpoints of duration of treatment and clinical benefit rate also appear superior in the combination arm, and the treatment was well tolerated. The most common serious adverse events (AEs) included leukopenia, anemia, thrombocytopenia, and metabolic evidence of tumor lysis in 15% of patients, researchers reported.
In mid-May, Pfizer said that PALOMA-1 results would be submitted to the FDA later this year as the basis of a new drug application for palbociclib plus letrozole as a first-line treatment for patients similar to those in the study cohort.
Amita Patnaik, MD, South Texas Accelerated Research Therapeutics (START), reported results from a phase I study of LY2835219 at the AACR meeting. The study of LY2835219 monotherapy is evaluating five tumor types (glioblastoma, melanoma, lung cancer, colorectal cancer, and metastatic breast cancer), although only results from patients with metastatic breast cancer were presented.
LY2835219 demonstrated durable single-agent activity, particularly in patients with hormone receptor—positive (HR+) disease. Among 47 patients, administered 150 mg to 200 mg orally every 12 hours, there was a partial response (PR) rate of 19% and stable disease (SD) in 51% of patients, while in patients with HR+ disease, a PR of 25% and SD of 55% were observed, corresponding to an overall response rate (ORR) of 25% for this population. The disease control rate was 81% and median PFS was 9.1 months in HR+ patients.
Across the entire study population, the most common AEs of grade 3/4 severity that may be related to treatment include neutropenia (11%), diarrhea (5%), and nausea (3%).
LY2835219 is currently being investigated in a phase III trial in combination with fulvestrant in patients with metastatic breast cancer (Table). Novartis’ LEE011 caused a recent stir when the company announced that the agent has already entered phase III clinical trials, despite the absence of reported data thus far from phase II trials.
Alvocidib was the first CDK inhibitor to be examined for the treatment of CLL. Though it demonstrated limited activity, it set the scene for other CDK inhibitors to be evaluated in this disease setting.
Dinaciclib (SCH727965; Merck), a broader CDK inhibitor that targets CDK 1, 2, 5, and 9, is currently the only other CDK inhibitor to be in phase III trials; it is being compared with the CD20 monoclonal antibody ofatumumab in refractory CLL. The results of a phase I dose-escalation trial of dinaciclib in patients with relapsed/ refractory CLL were reported by Joseph Flynn, DO, MPH, The Ohio State University Comprehensive Cancer Center, at the American Society of Hematology (ASH) Annual Meeting last year.
Dinaciclib appeared safe and effective, with an ORR of 58% among 52 patients and common AEs, including leukopenia, thrombocytopenia, and anemia, that were easily managed. In patients with high-risk disease, the PR rate was 57% for those with the 17p13.1 chromosomal deletion and 63% for those with 11q22 deletion.
Exciting preclinical studies suggest that dinaciclib may also have potential in the treatment of KRAS-mutant pancreatic cancers. The RAS family are among the most commonly mutated genes in human cancer; KRAS is mutated in up to 90% of pancreatic adenocarcinomas. Despite decades of research efforts, Ras has proved to be extremely difficult to target for the treatment of cancer. Recently, new strategies for both direct and indirect targeting of Ras and its related signaling pathways have begun to emerge. Among them is the discovery that CDK5 is an important downstream target of the RalGDS pathway, a Ras-specific downstream signaling cascade. In preclinical studies, dual inhibition of CDK5 with dinaciclib and Akt with MK-2206 resulted in a 90% reduction of tumor growth. As a result of these promising findings, the combination is now being studied in a phase I clinical trial in pancreatic cancer (NCT01783171).
As we edge closer to the potential FDA approval of a CDK inhibitor for the treatment of breast cancer and possibly CLL, researchers are now highlighting the need for biomarkers to aid in the selection of patients who will most benefit from these agents.
A variety of biomarkers are being examined with the hope that they will further enhance the efficacy of CDK inhibitors. These include truncated cyclin E, a shortened form of the cyclin E protein that is able to bind and activate CDK2, but is resistant to proteolytic degradation, making it much more stable and active.
Myc amplification has also been identified as a potential biomarker in patients with breast cancer and neuroblastoma, and CDK inhibition has been shown to be synthetically lethal with Myc overexpression. Finally, the Mcl1 survival factor, a Bcl2 protein family member, could prove to be an important biomarker in patients with CLL, multiple myeloma, neuroblastoma, and nasopharyngeal carcinoma.
Jane de Lartigue, PhD, is a freelance medical writer and editor based in the Davis, California.
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