Small Molecule Inhibitor Of Aurora Kinases Research Articles

Combined inhibition of aurora kinases and Bcl-xL induces apoptosis through select BH3-only proteins

Aurora Kinases (AURK) are mitotic kinases important for regulating cell cycle progression. Small molecule inhibitors of AURK have shown promising antitumor effects in multiple cancers; however, the utility of these inhibitors as inducers of cancer cell death has thus far been limited. Here, we examined the role of the Bcl-2 family proteins in AURK inhibition-induced apoptosis in colon cancer cells. We found that Alisertib and Danusertib, two small molecule inhibitors of AURK, are inefficient inducers of apoptosis in HCT116 and DLD-1 colon cancer cells, the survival of which requires at least one of the two anti-apoptotic Bcl-2 family proteins, Bcl-xL and Mcl-1. We further identified Bcl-xL as a major suppressor of Alisertib or Danusertib-induced apoptosis in HCT116 cells. We demonstrate that combination of a BH3-mimetic inhibitor (ABT-737), a selective inhibitor of Bcl-xL, Bcl-2, and Bcl-w, with Alisertib or Danusertib potently induces apoptosis through the Bcl-2 family effector protein Bax. In addition, we identified Bid, Puma, and Noxa, three of the BH3-only proteins of the Bcl-2 family, as mediators of Alisertib/ABT-737-induced apoptosis. We show while Noxa promotes apoptosis by constitutively sequestering Mcl-1, Puma becomes associated with Mcl-1 upon Alisertib treatment. On the other hand, we found Alisertib treatment causes activation of caspase-2, which promotes apoptosis by cleaving Bid into tBid, a suppressor of both Bcl-xL and Mcl-1. Together, these results define the Bcl-2 protein network critically involved in AURK inhibitor-induced apoptosis, and suggest that BH3-mimetics targeting Bcl-xL may help overcome resistance to AURK inhibitors in cancer cells.

Unweaving the mitotic spindle: A focus on Aurora kinase inhibitors in lung cancer.

Lung cancer is one of the most aggressive malignancies, classified into two major histological subtypes: non-small cell lung cancer (NSCLC), that accounts for about 85% of new diagnosis, and small cell lung cancer (SCLC), the other 15%. In the case of NSCLC, comprehensive genome sequencing has allowed the identification of an increasing number of actionable targets, which have become the cornerstone of treatment in the advanced setting. On the other hand, the concept of oncogene-addiction is lacking in SCLC, and the only innovation of the last 30 years has been the introduction of immune checkpoint inhibitors in extensive stage disease. Dysregulation of cell cycle is a fundamental step in carcinogenesis, and Aurora kinases (AURKs) are a family of serine/threonine kinases that play a crucial role in the correct advance through the steps of the cycle. Hyperexpression of Aurora kinases is a common protumorigenic pathway in many cancer types, including NSCLC and SCLC; in addition, different mechanisms of resistance to anticancer drugs rely on AURK expression. Hence, small molecule inhibitors of AURKs have been developed in recent years and tested in several malignancies, with different results. The aim of this review is to analyze the current evidences of AURK inhibition in lung cancer, starting from preclinical rationale to finish with clinical trials available up to now.

Screening of Drug Efficacy of Rosmarinic Acid Derivatives as Aurora Kinase Inhibitors by Computer-Aided Drug Design Method.

Aurora kinases (AKs) belong to the serine/threonine kinase family and play a crucial role in regulating the cell cycle. Therefore, AKs are the hopeful target for anticancer therapies and these findings have encouraged researchers to rigorously hunt small molecule aurora kinase inhibitors, not only for research articles but also for use as therapeutic agents. The present study helps us to identify and screen the best phytochemicals as potent inhibitors against AKs. These potent inhibitors come from the various substitution of rosmarinic acid (RA). In this paper, we choose different tested derivative compounds for designing anticancer drugs by substituting various functional groups of standard drug RA. In silico studies were carried out to appreciate better drug candidature of some of these derivative compounds. This study was performed on 56 derived compounds of the standard RA. DFT study was conducted using the UB3LYP/6-311++G(d,p) basis set to study HOMO-LUMO energies, dipole moments, using the Gaussian16 suite. Some of the derived parameters, like ionization potential, electron affinity, softness- hardness, chemical potential, and electrophilicity index were noted. A docking study was performed with AKs inhibiting receptor using AutoDock 4.2. ADME prediction was made with the preADMET web tool. Molecular descriptor properties were predicted with molinspiration and OSIRIS property explorer. Out of the 56 derivatives, 11 have passed all the rules of drug candidature, to serve as best AKs inhibitor, in a theoretical manner. This study should be supported by a new proposal to explore future studies with these 11 compounds against cancer.

Toward In Vitro Epigenetic Drug Design for Thyroid Cancer: The Promise of PF-03814735, an Aurora Kinase Inhibitor.

Thyroid cancer (TC) is a very common malignancy worldwide. Chief among the innovative molecular drug targets for TC are epigenetic modifications. Increased telomerase activity in cancer cells makes telomerase a novel target for epigenetic anticancer drug innovation. Recently, telomerase reverse transcriptase (TERT) gene promoter (TERTp) mutations (C228T and C250T) were reported at high frequency in TC cell lines and tumor biopsies. In this study, three representative TC cell lines, mutant TERTp (TPC1), mutant BRAF/TERTp (KTC2), and wild-type TERTp (WRO), were screened with a drug library composed of 51 epigenetic drugs: 14 Aurora kinase inhibitors; 23 histone deacetylase inhibitors; 5 sirtuin modifiers; 3 hypoxia-inducible factor inhibitors; 2 DNA methyltransferase inhibitors; 2 histone methyltransferase inhibitors, a histone demethylase inhibitor, and a bromodomain inhibitor. Effects of the drugs on cell growth at 48 and 72 h were compared. PF-03814735, a small-molecule inhibitor of Aurora kinase A (IC50 = 0.8 nM) and B (IC50 = 5 nM), was the most potent on KTC2 cells, whereas CUDC-101, a multitarget inhibitor, was effective on both WRO and KTC2 cells. Notably, PF-03814735 was found to be the most effective epigenetic drug on cell lines harboring the C228T mutation. In conclusion, these new findings offer specific guidance on dose and time course selection to design novel therapeutic interventions against TC using PF-03814735, and specifically target cells carrying the TERTpC228T mutation. In a larger context of drug discovery science, these findings inform new strategies to forecast optimal treatment regimens for TC, particularly with Aurora kinase inhibitors and in ways guided by epigenetic drug design.

Abstract 3008: Preclinical characterization of AMG 900, a pan-aurora kinase inhibitor, alone and in combination with taxanes in ovarian cancer

Abstract BACKGROUND: Aurora kinases (AK) A and B play essential roles in multiple stages of mitosis and are frequently overexpressed in a subset of human cancers, including ovarian cancer (OC). AMG 900, a potent and highly selective small molecule inhibitor of AKs, showed promising single-agent activity in heavily pretreated patients with advanced OC in a Phase 1b clinical trial. In this study, we report the preclinical effects of AMG 900 in a panel of well-characterized human cancer cell lines representing clinically-relevant OC subtypes. METHODS: The anti-proliferative effects of AMG 900 were evaluated using a 5-day cell count assay. Cell lines were classified as sensitive to AMG 900 when lethality was > 15% at 10 nM. Molecular markers were profiled including TP53 mutation status, AURKA, CCNE1, MYC copy number, and p53, p21 and cyclin E1 protein by reverse phase protein array. Flow cytometry and imaging methods were used to evaluate the mechanism of action of AMG 900 alone and in combination with chemotherapy. The combination of AMG 900 plus docetaxel was evaluated in an IGROV-1 ovarian endometrioid carcinoma xenograft model. RESULTS: One third of the cell lines (11 of 35) were classified as sensitive to AMG 900 and showed enrichment for TP53 mutations and serous OC subtype. However, 10 of 24 resistant cell lines harbored TP53 mutations, indicating that TP53 mutational status alone was not sufficient for predicting AMG 900 sensitivity. Inhibition of AK activity by AMG 900 in OC cells resulted in aborted cell division leading to polyploidy and cell death (suggestive of aurora-B dominant phenotype). Re-plating of remnant cells after AMG 900 treatment showed an attenuation of cell regrowth, where TP53mut IGROV-1 cells showed minimal regrowth compared to TP53wt OVCAR-5 cells. AMG 900 inhibited proliferation at low nanomolar concentrations in the majority of OC cell lines and enhanced the effects of paclitaxel, carboplatin, and doxorubicin in IGROV-1 cells. In tumor-bearing mice, administration of AMG 900 at 7.5 mg/kg (PO) for two days per week or docetaxel at 10 mg/kg (IP) weekly for four cycles significantly inhibited the growth of IGROV-1 tumor xenografts (P < .0001 vs. vehicle alone). Notably, co-administration of AMG 900 with docetaxel enhanced efficacy and induced a delay in tumor regrowth compared to docetaxel alone. Single-agent-treated mice showed minimal body weight loss (BWL), whereas combination-treated mice showed moderate BWL (average < 10%) that was largely reversible (2 of 12 animals removed due to toxicity). CONCLUSIONS: AMG 900 alone or in combination with chemotherapy such as paclitaxel may be a promising clinical strategy to treat patients with ovarian cancer. Citation Format: Ondrej Kalous, Dylan Conklin, Kanthinh Manivong, William Wayne, Kelly Hanestad, Jude Canon, Robert Loberg, Gregory Friberg, Erick Gamelin, Florian D. Vogl, Gloria Juan, Angela Coxon, Dennis Slamon, Richard Finn, Marc Payton. Preclinical characterization of AMG 900, a pan-aurora kinase inhibitor, alone and in combination with taxanes in ovarian cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3008.

A Cell Biologist's Field Guide to Aurora Kinase Inhibitors.

Aurora kinases are essential for cell division and are frequently misregulated in human cancers. Based on their potential as cancer therapeutics, a plethora of small molecule Aurora kinase inhibitors have been developed, with a subset having been adopted as tools in cell biology. Here, we fill a gap in the characterization of Aurora kinase inhibitors by using biochemical and cell-based assays to systematically profile a panel of 10 commercially available compounds with reported selectivity for Aurora A (MLN8054, MLN8237, MK-5108, MK-8745, Genentech Aurora Inhibitor 1), Aurora B (Hesperadin, ZM447439, AZD1152-HQPA, GSK1070916), or Aurora A/B (VX-680). We quantify the in vitro effect of each inhibitor on the activity of Aurora A alone, as well as Aurora A and Aurora B bound to fragments of their activators, TPX2 and INCENP, respectively. We also report kinome profiling results for a subset of these compounds to highlight potential off-target effects. In a cellular context, we demonstrate that immunofluorescence-based detection of LATS2 and histone H3 phospho-epitopes provides a facile and reliable means to assess potency and specificity of Aurora A versus Aurora B inhibition, and that G2 duration measured in a live imaging assay is a specific readout of Aurora A activity. Our analysis also highlights variation between HeLa, U2OS, and hTERT-RPE1 cells that impacts selective Aurora A inhibition. For Aurora B, all four tested compounds exhibit excellent selectivity and do not significantly inhibit Aurora A at effective doses. For Aurora A, MK-5108 and MK-8745 are significantly more selective than the commonly used inhibitors MLN8054 and MLN8237. A crystal structure of an Aurora A/MK-5108 complex that we determined suggests the chemical basis for this higher specificity. Taken together, our quantitative biochemical and cell-based analyses indicate that AZD1152-HQPA and MK-8745 are the best current tools for selectively inhibiting Aurora B and Aurora A, respectively. However, MK-8745 is not nearly as ideal as AZD1152-HQPA in that it requires high concentrations to achieve full inhibition in a cellular context, indicating a need for more potent Aurora A-selective inhibitors. We conclude with a set of “good practice” guidelines for the use of Aurora inhibitors in cell biology experiments.

Aurora Kinase Inhibitors: Current Status and Outlook.

The Aurora kinase family comprises of cell cycle-regulated serine/threonine kinases important for mitosis. Their activity and protein expression are cell cycle regulated, peaking during mitosis to orchestrate important mitotic processes including centrosome maturation, chromosome alignment, chromosome segregation, and cytokinesis. In humans, the Aurora kinase family consists of three members; Aurora-A, Aurora-B, and Aurora-C, which each share a conserved C-terminal catalytic domain but differ in their sub-cellular localization, substrate specificity, and function during mitosis. In addition, Aurora-A and Aurora-B have been found to be overexpressed in a wide variety of human tumors. These observations led to a number of programs among academic and pharmaceutical organizations to discovering small molecule Aurora kinase inhibitors as anti-cancer drugs. This review will summarize the known Aurora kinase inhibitors currently in the clinic, and discuss the current and future directions.

Preclinical testing of selective Aurora kinase inhibitors on a medullary thyroid carcinoma-derived cell line.

Deregulated expression of the Aurora kinases (Aurora-A, B, and C) is thought to be involved in cell malignant transformation and genomic instability in several cancer types. Over the last decade, a number of small-molecule inhibitors of Aurora kinases have been developed, which have proved to efficiently restrain malignant cell growth and tumorigenicity. Regarding medullary thyroid carcinoma (MTC), we previously showed the efficacy of a pan-Aurora kinase inhibitor (MK-0457) in impairing growth and survival of the MTC-derived cell line TT. In the present study, we sought to establish if one of the Aurora kinases might represent a preferential target for MTC therapy. The effects of selective inhibitors of Aurora-A (MLN8237) and Aurora-B (AZD1152) were analyzed on TT cell proliferation, apoptosis, cell cycle, and ploidy. The two inhibitors reduced TT cell proliferation in a time- and dose-dependent manner, with IC50 of 19.0 ± 2.4 nM for MLN8237 and 401.6 ± 44.1 nM for AZD1152. Immunofluorescence experiments confirmed that AZD1152 inhibited phosphorylation of histone H3 (Ser10) by Aurora-B, while it did not affect Aurora-A autophosphorylation. MLN8237 inhibited Aurora-A autophosphorylation as expected, but at concentrations required to achieve the maximum antiproliferative effects it also abolished H3 (Ser10) phosphorylation. Cytofluorimetry experiments showed that both inhibitors induced accumulation of cells in G2/M phase and increased the subG0/G1 fraction and polyploidy. Finally, both inhibitors triggered apoptosis. We demonstrated that inhibition of either Aurora-A or Aurora-B has antiproliferative effects on TT cells, and thus it would be worthwhile to further investigate the therapeutical potential of Aurora kinase inhibitors in MTC treatment.

Bioactive compounds of Eriocaulon sieboldianum blocking proliferation and inducing apoptosis of HepG2 cells might be involved in Aurora kinase inhibition.

Eriocaulon sieboldianum (Sieb. & Zucc. ex Steud.) is an edible and medicinal plant used in traditional Chinese medicine. Often in combination with other herbs, it is processed into healthcare beverages for expelling wind-heat, protecting eyes, and reducing blood lipids. Besides, its water decoction together with other herbs has been utilized to treat cancer in China. However, the active ingredients and the precise cellular mechanisms of E. sieboldianum remain to be elucidated. The Aurora kinase family plays critical roles in the regulation of cell division and has attracted great attention to the identification of small-molecule Aurora kinase inhibitors for potential treatment of cancer. A molecular docking study was employed for docking of the most bioactive compounds. Hispidulin (HPDL) and quercetin-3-O-(6''-O-galloyl)-β-D-galactopyranoside (QGGP) were singled out as potent inhibitors of Aurora kinase. Their inhibitory activity towards Aurora kinase was further confirmed by the obvious decrease in autophosphorylation of Aurora-A (Thr288) and Aurora-B (Thr232). Moreover, the induction of cell cycle arrest in HepG2 cells and the suppressed phosphorylation of histone H3 were also consistent with the inhibition of Aurora kinase. The data indicate that the E. sieboldianum extract and its two active compounds, HPDL and QGGP, could effectively induce apoptosis via p53, MAPKs and the mitochondrial apoptotic pathways. These findings could improve the understanding and enhance the development of drugs based on E. sieboldianum and raise its application value in anticancer therapy or prevention. In addition, our results indicated that Aurora kinase might be a novel target of HPDL and QGGP.

A Phase I Study of the Aurora a Kinase Inhibitor Alisertib in Combination with 7+3 Induction Chemotherapy in Patients with Acute Myeloid Leukemia

The aurora kinases play an essential role in regulating mitosis and cell division. Over-expression of aurora kinases A and B has been associated with poor-risk features in acute myeloid leukemia (AML). Preclinical studies suggest that small molecule aurora kinase inhibitors have activity in AML cell lines. Alisertib (MLN8237), a novel Aurora-A kinase inhibitor, has been generally well tolerated in early clinical trials of hematological malignancies, including AML. This study sought to evaluate tolerability of alisertib combined with standard “7+3” induction chemotherapy for AML.In this phase I, 3+3 cohort dose-escalation study, alisertib was administered with standard 7+3 induction for newly diagnosed AML. Those with acute promyelocytic leukemia or with AML and core-binding factor alterations were excluded. All patients received 7+3 induction (continuous infusion cytarabine 200mg/m2 x 7 days and intravenous idarubicin 12mg/m2 x 3 days), after which on day 8, alisertib was administered twice daily (BID) for 7 days. Dose escalation occurred via three cohorts (10mg BID, 20mg BID, 30mg BID). All underwent a mid-induction bone marrow biopsy, following the course of alisertib, to assess for residual disease, which if present, was to be treated with 5+2 re-induction (cytarabine 200mg/m2 x 5 days, idarubicin 12mg/m2 x 2 days) without alisertib. Following induction, up to four cycles of consolidation were allowed, using cytarabine (3g/m2 BID days 1,3,5 for those aged <60 and 2g/m2 daily days 1-5 for those aged ≥60) followed by alisertib, according to dose cohort, on days 6-12. After consolidation, alisertib maintenance was initiated, at cohort dose level, for days 1-7 of 21 day cycles, to be continued for 12 months or until disease progression. Those eligible for stem cell transplant (SCT), following induction and/or consolidation courses, were removed from study for this purpose.Currently, 14 patients are enrolled on study (n=3, 10mg BID; n=7, 20mg BID; n=4, 30mg BID). The median age was 62 (range 43-75); 9 (64%) were male, and all but 3 were Caucasian. One patient (7%) had therapy-related AML and six (43%) had underlying myelodysplasia. FLT3 mutations were detected in 3 (21%), NPM1 mutations in 2 (14%), CEBPA mutations in 2 (14%), and IDH1/IDH2 mutations in 4 patients (28.4%). One patient in cohort 2 died of sepsis, deemed unrelated to study drug, prior to completion of the toxicity assessment period and was replaced. Five patients have gone on to SCT. All patients received induction, 7 patients have undergone first consolidation, 3 a second consolidation, 2 a third consolidation, 1 a fourth consolidation, and 1 patient has started maintenance therapy. Alisertib has been well tolerated. All patients have experienced expected grade 4 toxicities of anemia, thrombocytopenia, and febrile neutropenia seen during the induction phase of treatment. At initial dose of 10mg BID, no dose-limited toxicities (DLTs) were encountered. In the 20mg BID cohort, one DLT was encountered, of delayed thrombocytopenia (G4 at day 40). The final cohort, at 30mg BID, is currently accruing (4 of 6 enrolled). No other DLTs have been detected, and other toxicities on study will be presented. The maximum tolerated dose (MTD) will be established upon completion of this cohort. To date, 11 of 12 (92%) evaluable patients have achieved remission following induction (8 cases of CR and 3 of CRp). During the course of the study, one patient has experienced relapse, and none have died. Thirteen of 14 evaluable patients had an ablated mid-induction marrow biopsy, with the remaining patient showing 6% blasts in a 20% cellular marrow at mid-induction. None of the patients have undergone re-induction with 5+2 at mid-induction. We anticipate that the study will be fully enrolled and closed to further accrual by the time of presentation.Alisertib, a novel aurora A kinase inhibitor, appears to be well tolerated in conjunction with standard induction chemotherapy in newly diagnosed AML. Currently, 13 of 14 treated patients have demonstrated marrow ablation at the mid-induction point of therapy, and none have required re-induction with 5+2. Eleven of 12 evaluable patients have achieved remission following induction. These results suggest that alisertib is well-tolerated and may hold promise as a therapeutic agent in AML, when combined with induction chemotherapy. Ongoing efforts seek to define the MTD and establish appropriate dosing for phase II studies. DisclosuresFathi:Millennium: Research Funding; Seattle Genetics: Advisory Board, Advisory Board Other; Agios: Advisory Board, Advisory Board Other. Attar:Agios: Employment.

The aurora kinases in cell cycle and leukemia

The Aurora kinases, which include Aurora A (AURKA), Aurora B (AURKB) and Aurora C (AURKC), are serine/threonine kinases required for the control of mitosis (AURKA and AURKB) and meiosis (AURKC). Since their discovery nearly twenty years ago, Aurora kinases have been studied extensively in cell and cancer biology 1. Several early studies found that Aurora kinases are amplified and overexpressed at the transcript and protein level in various malignancies, including several types of leukemia. These discoveries and others provided a rationale for the development of small molecule inhibitors of Aurora kinases as leukemia therapies. The first generation of Aurora kinase inhibitors did not fare well in clinical trials, owing to poor efficacy and high toxicity. However, the creation of second generation, highly selective Aurora kinase inhibitors has increased the enthusiasm for targeting these proteins in leukemia. This review will describe the functions of each Aurora kinase, summarize their involvement in leukemia and discuss inhibitor development and efficacy in leukemia clinical trials.

P53 deficiency enhances mitotic arrest and slippage induced by pharmacological inhibition of Aurora kinases

A number of small-molecule inhibitors of Aurora kinases have been developed and are undergoing clinical trials for anti-cancer therapies. Different Aurora kinases, however, behave as very different targets: while inhibition of Aurora A (AURKA) induces a delay in mitotic exit, inhibition of Aurora B (AURKB) triggers mitotic slippage. Furthermore, while it is evident that p53 is regulated by Aurora kinase-dependent phosphorylation, how p53 may in turn regulate Aurora kinases remains mysterious. To address these issues, isogenic p53-containing and -negative cells were exposed to classic inhibitors that target both AURKA and AURKB (Alisertib and ZM447439), as well as to new generation of inhibitors that target AURKA (MK-5108), AURKB (Barasertib) individually. The fate of individual cells was then tracked with time-lapse microscopy. Remarkably, loss of p53, either by gene disruption or small interfering RNA-mediated depletion, sensitized cells to inhibition of both AURKA and AURKB, promoting mitotic arrest and slippage respectively. As the p53-dependent post-mitotic checkpoint is also important for preventing genome reduplication after mitotic slippage, these studies indicate that the loss of p53 in cancer cells represents a major opportunity for anti-cancer drugs targeting the Aurora kinases.

Pediatric Phase I Trial and Pharmacokinetic Study of MLN8237, an Investigational Oral Selective Small-Molecule Inhibitor of Aurora Kinase A: A Children's Oncology Group Phase I Consortium Study

MLN8237, a selective small-molecule inhibitor of Aurora kinase A, has activity in a broad range of preclinical pediatric cancer models. We conducted a phase I trial in children with refractory/recurrent solid tumors to define the maximum-tolerated dose, toxicities, and pharmacokinetic properties of MLN8237. MLN8237 was administered orally either once daily or divided twice daily for seven days, every 21 days. Using a rolling-six design, four dose levels (45, 60, 80, and 100 mg/m(2)/day) were evaluated on the once-daily schedule, and two dose levels (60 and 80 mg/m(2)/d) on the twice-daily schedule. Pharmacokinetic studies were conducted with the initial dose and trough drug concentrations also measured at the steady state. Thirty-seven patients were enrolled. On the once-daily dosing schedule, myelosuppression was dose limiting in three of four patients at 100 mg/m(2), and one of six patients had dose-limiting mood alteration at 80 mg/m(2). At 45 mg/m(2), one of six patients experienced dose-limiting mucositis. Mucositis and myelosuppression were dose limiting at 80 mg/m(2) on the twice-daily schedule, and one of five patients at 60 mg/m(2) on the twice-daily schedule experienced a dose-limiting alkaline phosphatase. Five of 11 patients experienced hand-foot-skin syndrome with twice-daily dosing versus one of 21 after once-daily dosing. There was one partial response and six with prolonged stable disease among 33 evaluable subjects. The twice-daily dose regimen is well tolerated in adults; however, children experienced a greater frequency of myelosuppression and hand-foot-skin syndrome on this schedule. Children tolerated a higher dose and the recommended pediatric phase II dose is 80 mg/m(2)/d once daily for seven days.

The induction of polyploidy or apoptosis by the Aurora A kinase inhibitor MK8745 is p53-dependent

Aurora kinases are mitotic serine/threonine protein kinases and are attractive novel targets for anticancer therapy. Many small-molecule inhibitors of Aurora kinases are currently undergoing clinical trials. Aurora A kinase is essential for successful mitotic transition. MK8745 is a novel and selective small-molecule inhibitor of Aurora A kinase. MK8745 induced apoptotic cell death in a p53-dependent manner when tested in vitro in cell lines of multiple lineages. Cells expressing wild-type p53 showed a short delay in mitosis followed by cytokinesis, resulting in 2N cells along with apoptosis. However, cells lacking or with mutant p53 resulted in a prolonged arrest in mitosis followed by endoreduplication and polyploidy. Cytokinesis was completely inhibited in p53-deficient cells, as observed by the absence of 2N cell population. The induction of apoptosis in p53-proficient cells was associated with activation of caspase 3 and release of cytochrome c but was independent of p21. Exposure of p53 wild-type cells to MK8745 resulted in the induction of p53 phosphorylation (ser15) and an increase in p53 protein expression. p53-dependent apoptosis by MK8745 was further confirmed in HCT 116 p53-/- cells transfected with wild-type p53. Transient knockdown of Aurora A by specific siRNA recapitulated these p53- dependent effects, with greater percent induction of apoptosis in p53 wild-type cells. In conclusion, our studies show p53 as a determining factor for induction of apoptosis vs. polyploidy upon inhibition of Aurora A.

Danusertib, an aurora kinase inhibitor

Introduction: Drugs that interfere with the normal progression of mitosis belong to the most successful cytotoxic agents currently used for anticancer treatment. Aurora kinases are serine/threonine kinases that function as key regulators of mitosis and are frequently overexpressed in human cancers. The use of several small molecule aurora kinase inhibitors as potential anticancer therapeutic is being investigated. Danusertib (formerly PHA-739358) is a small ATP competitive molecule that inhibits aurora A, B and C kinases. Interestingly, danusertib also inhibits several receptor tyrosine kinases such as Abl, Ret, FGFR-1 and TrkA. These tyrosine kinases are involved in the pathogenesis of a variety of malignancies and the observed multi-target inhibition may increase the antitumor activity resulting in extending the indication. Danusertib was one of the first aurora kinase inhibitors to enter the clinic and has been studied in Phase I and II trials.Areas covered: This review provides an updated summary of preclinical and clinical experience with danusertib up to July 2011.Expert opinion: Future studies with danusertib should focus on the possibility of combining this agent with other targeted anticancer agents, chemotherapy or radiotherapy. As a single agent, danusertib may show more promise in the treatment of leukemias than in solid tumors.

Abstract C202: Discovery and initial characterization of the clinical compound ABT-348, a potent inhibitor of the VEGF, PDGF, and Aurora kinase families.

Abstract The Aurora kinases are a family of serine/threonine kinases that mediate multiple events in cell division. Humans have three Aurora kinases, A, B and C, that are differentially localized and mediate distinct functions during mitosis. Because the Aurora kinases play a key role in mitosis and are overexpressed in multiple human tumor types, there has been considerable interest in developing Aurora kinase inhibitors as antitumor agents. A number of small-molecule Aurora kinase inhibitors have been reported and there are several compounds currently in Phase I/II clinical trials for cancer. ABT-348 is a novel, potent and orally bioavailable inhibitor of the Aurora kinases as well as the VEGF and PDGF families of receptor tyrosine kinases and is currently in Phase I clinical trials. Herein we describe the discovery and preparation of this molecule including the SAR work that provided improved Aurora kinase potency, aqueous solubility and oral bioavailability with reduced hERG and CYP liabilities relative to earlier analogs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C202.

Discovery of a Potent, Selective, and Orally Bioavailable Pyridinyl-Pyrimidine Phthalazine Aurora Kinase Inhibitor

The discovery of aurora kinases as essential regulators of cell division has led to intense interest in identifying small molecule aurora kinase inhibitors for the potential treatment of cancer. A high-throughput screening effort identified pyridinyl-pyrimidine 6a as a moderately potent dual inhibitor of aurora kinases -A and -B. Optimization of this hit resulted in an anthranilamide lead (6j) that possessed improved enzyme and cellular activity and exhibited a high level of kinase selectivity. However, this anthranilamide and subsequent analogues suffered from a lack of oral bioavailability. Converting the internally hydrogen-bonded six-membered pseudo-ring of the anthranilamide to a phthalazine (8a-b) led to a dramatic improvement in oral bioavailability (38-61%F) while maintaining the potency and selectivity characteristics of the anthranilamide series. In a COLO 205 tumor pharmacodynamic assay measuring phosphorylation of the aurora-B substrate histone H3 at serine 10 (p-histone H3), oral administration of 8b at 50 mg/kg demonstrated significant reduction in tumor p-histone H3 for at least 6 h.

An integrated pharmacokinetic–pharmacodynamic model for an Aurora kinase inhibitor

The spindle assembly checkpoint is a cell cycle surveillance mechanism that ensures the proper separation of chromosomes prior to cell division at mitosis. Aurora kinases play critical roles in mitotic progression and hence small-molecule inhibitors of Aurora kinases have been developed as a new class of potential anti-cancer drugs. In this paper we present for the first time an integrated pharmacokinetic-pharmacodynamic model of the functional effects of CYC116 (a known inhibitor of Aurora kinases A and B) on the spindle assembly checkpoint. We use the model to simulate two common experimental systems: cell culture and p.o. dosing of mice and present predictions of the effects of CYC116 for a range of doses and drug scheduling regimes. The model reveals that a critical peak drug concentration is required to cause aberrant kinetochore-microtubule attachments. The model also predicts that provided this threshold concentration is exceeded, a high total oral dose causes a high number of aberrant attachments within any given damaged cell. However, the proportion of cells which enter anaphase with aberrant attachments is associated with the total length of time for which the plasma concentration is maintained above the threshold. Moreover, our model reveals that the length of prometaphase/metaphase is a nonlinear function of drug dose and this time period can be extended or shortened. Finally, a strong saturation effect on CYC116 efficacy is predicted by the model. We discuss how these predictions may have implications for further drug trials using CYC116 and other similar AK inhibitors.

Drug-Resistant Aurora A Mutants for Cellular Target Validation of the Small Molecule Kinase Inhibitors MLN8054 and MLN8237

The Aurora kinases regulate multiple aspects of mitotic progression, and their overexpression in diverse tumor types makes them appealing oncology targets. An intensive research effort over the past decade has led to the discovery of chemically distinct families of small molecule Aurora kinase inhibitors, many of which have demonstrated therapeutic potential in model systems. These agents are also important tools to help dissect signaling pathways that are orchestrated by Aurora kinases, and the antiproliferative target of pan-Aurora inhibitors such as VX-680 has been validated using chemical genetic techniques. In many cases the nonspecific nature of Aurora inhibitors toward unrelated kinases is well established, potentially broadening the spectrum of cancers to which these compounds might be applied. However, unambiguously demonstrating the molecular target(s) for clinical kinase inhibitors is an important challenge, one that is absolutely critical for deciphering the molecular basis of compound specificity, resistance, and efficacy. In this paper, we have investigated amino acid requirements for Aurora A sensitivity to the benzazepine-based Aurora inhibitor MLN8054 and the close analogue MLN8237, a second-generation compound that is in phase II clinical trials. A crystallographic analysis facilitated the design and biochemical investigation of a panel of resistant Aurora A mutants, a subset of which were then selected as candidate drug-resistance targets for further evaluation. Using inducible human cell lines, we show that cells expressing near-physiological levels of a functional but partially drug-resistant Aurora A T217D mutant survive in the presence of MLN8054 or MLN8237, authenticating Aurora A as a critical antiproliferative target of these compounds.

Abstract 2754: Phase I and pharmacokinetic (PK) study of two regimens combining the aurora kinase inhibitor AS703569 and gemcitabine in patients with advanced solid tumors

Abstract Background: AS703569 is an orally available, ATP-competitive, small-molecule inhibitor of aurora kinases A, B, and C and other cancer-relevant kinases. AS703569 has shown in vitro and in vivo antitumor activity as a single agent and in combinations, including with gemcitabine (Gem). Phase I monotherapy studies in patients (pts) with solid tumors and hematologic malignancies are ongoing. Objective: To define the maximum tolerated dose (MTD) of 2 regimens (Rs) of sequential-combination AS703569 and Gem (fixed dose 1000 mg/m2/day [d]). Methods: This multicenter, dose-escalation study with a 3+3 design was performed in pts with advanced solid tumors. R1: AS703569 d2 and 9, Gem d1 and 8; R2: AS703569 d1 and 8, Gem d2 and 9 of 21-d cycles. MTD definition: dose level (DL) below that at which &amp;gt;1/3 or &amp;gt;1/6 pts experienced a dose-limiting toxicity (DLT) in cycle 1. Secondary objectives included PK evaluations and antitumor activity. AS703569 starting dose (DL1) was 10 mg/m2/d; other DLs assessed were 15, 21, 28, and 37 mg/m2/d (ongoing). Results: To date, 45 pts have been treated. R1 (19 pts): median (range) age 62 (31-75) years (y); 13 (68%) men; most common tumors: pancreatic (n=5), and lung, colorectal, and stomach (n=2 each); 1-16 cycles completed per pt. R2 (26 pts): median age 58 (20-80) y; 19 (73%) men; most common tumors: colorectal (n=5), liver (n=3), and lung, bladder, and pancreas (n=2 each); 1-10 cycles completed per pt. DLTs reported over the DLs studied to date: in R1, CTCAE Grade (Gr) 4 neutropenia ≥7 d and Gr 4 thrombocytopenia (37 mg/m2/d); in R2, Gr 4 neutropenia and septic shock (10 mg/m2/d), general status deterioration and asthenia (15 mg/m2/d), and Gr 4 neutropenia ≥7 d (28 and 37 mg/m2/d). Related treatment-emergent adverse events (TEAEs) ≥Gr 3 occurred in 13 pts (68%) in R1 and 19 pts (73%) in R2, most commonly neutropenia (R1 n=8; R2 n=13), lymphopenia (R1 n=11; R2 n=9), anemia (R1 n=4; R2 n=5), and thrombocytopenia (R1 n=2; R2 n=3). Other TEAEs ≥Gr 3 included nausea (R1 n=1), asthenia (R1 n=4; R2 n=2), and fatigue (R1 and R2 n=1). Two TEAEs led to deaths in R2 (10 mg/m2/d): 1 pt with Gr 4 neutropenia and septic shock; 1 pt with digestive hemorrhage. Preliminary PK data (n=34): peak AS703569 plasma concentrations (DL 10-37 mg/m2/d) were mostly reached between 2-4 (range 1-8) h. Cmax and AUClast increased with dose. PK data are comparable to those from monotherapy Phase I studies. Stable disease (SD; &amp;gt;4 cycles) was seen in 11/33 pts; 1 pt with non-small-cell lung cancer had a partial response lasting 6 months. Conclusion: AS703569 can be safely combined with standard-dose Gem. Most toxicities were hematologic and reversible. The MTD is being defined for both Rs and AS703569 exposure seems unaltered by Gem coadministration. Some pretreated pts experienced tumor responses and long-lasting SD. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2754.