Skip to main content
  • AACR Journals
    • Blood Cancer Discovery
    • Cancer Discovery
    • Cancer Epidemiology, Biomarkers & Prevention
    • Cancer Immunology Research
    • Cancer Prevention Research
    • Cancer Research
    • Clinical Cancer Research
    • Molecular Cancer Research
    • Molecular Cancer Therapeutics

AACR logo

  • Register
  • Log in
  • My Cart
Advertisement

Main menu

  • Home
  • About
    • The Journal
    • AACR Journals
    • Subscriptions
    • Permissions and Reprints
    • Reviewing
  • Articles
    • OnlineFirst
    • Current Issue
    • Past Issues
    • Meeting Abstracts
    • Collections
      • COVID-19 & Cancer Resource Center
      • Focus on Radiation Oncology
      • Novel Combinations
      • Reviews
      • Editors' Picks
      • "Best of" Collection
  • For Authors
    • Information for Authors
    • Author Services
    • Best of: Author Profiles
    • Submit
  • Alerts
    • Table of Contents
    • Editors' Picks
    • OnlineFirst
    • Citation
    • Author/Keyword
    • RSS Feeds
    • My Alert Summary & Preferences
  • News
    • Cancer Discovery News
  • COVID-19
  • Webinars
  • Search More

    Advanced Search

  • AACR Journals
    • Blood Cancer Discovery
    • Cancer Discovery
    • Cancer Epidemiology, Biomarkers & Prevention
    • Cancer Immunology Research
    • Cancer Prevention Research
    • Cancer Research
    • Clinical Cancer Research
    • Molecular Cancer Research
    • Molecular Cancer Therapeutics

User menu

  • Register
  • Log in
  • My Cart

Search

  • Advanced search
Molecular Cancer Therapeutics
Molecular Cancer Therapeutics
  • Home
  • About
    • The Journal
    • AACR Journals
    • Subscriptions
    • Permissions and Reprints
    • Reviewing
  • Articles
    • OnlineFirst
    • Current Issue
    • Past Issues
    • Meeting Abstracts
    • Collections
      • COVID-19 & Cancer Resource Center
      • Focus on Radiation Oncology
      • Novel Combinations
      • Reviews
      • Editors' Picks
      • "Best of" Collection
  • For Authors
    • Information for Authors
    • Author Services
    • Best of: Author Profiles
    • Submit
  • Alerts
    • Table of Contents
    • Editors' Picks
    • OnlineFirst
    • Citation
    • Author/Keyword
    • RSS Feeds
    • My Alert Summary & Preferences
  • News
    • Cancer Discovery News
  • COVID-19
  • Webinars
  • Search More

    Advanced Search

Small Molecule Therapeutics

Aurora A–Selective Inhibitor LY3295668 Leads to Dominant Mitotic Arrest, Apoptosis in Cancer Cells, and Shows Potent Preclinical Antitumor Efficacy

Jian Du, Lei Yan, Raquel Torres, Xueqian Gong, Huimin Bian, Carlos Marugán, Karsten Boehnke, Carmen Baquero, Yu-Hua Hui, Sonya C. Chapman, Yanzhu Yang, Yi Zeng, Sarah M. Bogner, Robert T. Foreman, Andrew Capen, Gregory P. Donoho, Robert D. Van Horn, Darlene S. Barnard, Jack A. Dempsey, Richard P. Beckmann, Mark S. Marshall, Li-Chun Chio, Yuewei Qian, Yue W. Webster, Amit Aggarwal, Shaoyou Chu, Shobha Bhattachar, Louis F. Stancato, Michele S. Dowless, Phillip W. Iversen, Jason R. Manro, Jennie L. Walgren, Bartley W. Halstead, Matthew Z. Dieter, Ricardo Martinez, Shripad V. Bhagwat, Emiko L. Kreklau, Maria Jose Lallena, Xiang S. Ye, Bharvin K.R. Patel, Christoph Reinhard, Gregory D. Plowman, David A. Barda, James R. Henry, Sean G. Buchanan and Robert M. Campbell
Jian Du
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • For correspondence: du_jian@lilly.com
Lei Yan
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Raquel Torres
2Eli Lilly and Company, Alcobendas (Madrid), Spain.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Xueqian Gong
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Huimin Bian
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Carlos Marugán
2Eli Lilly and Company, Alcobendas (Madrid), Spain.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Karsten Boehnke
2Eli Lilly and Company, Alcobendas (Madrid), Spain.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Karsten Boehnke
Carmen Baquero
2Eli Lilly and Company, Alcobendas (Madrid), Spain.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yu-Hua Hui
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sonya C. Chapman
3Eli Lilly and Company, Windlesham, United Kingdom.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Sonya C. Chapman
Yanzhu Yang
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yi Zeng
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sarah M. Bogner
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Robert T. Foreman
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Andrew Capen
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Gregory P. Donoho
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Robert D. Van Horn
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Darlene S. Barnard
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jack A. Dempsey
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Richard P. Beckmann
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Mark S. Marshall
4Ped-Hematology/Oncology, Indiana University School of Medicine, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Li-Chun Chio
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yuewei Qian
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Yue W. Webster
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Amit Aggarwal
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Shaoyou Chu
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Shobha Bhattachar
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Louis F. Stancato
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Michele S. Dowless
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Phillip W. Iversen
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jason R. Manro
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jennie L. Walgren
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bartley W. Halstead
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Matthew Z. Dieter
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Ricardo Martinez
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Shripad V. Bhagwat
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Emiko L. Kreklau
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Maria Jose Lallena
2Eli Lilly and Company, Alcobendas (Madrid), Spain.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Xiang S. Ye
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bharvin K.R. Patel
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Christoph Reinhard
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Gregory D. Plowman
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
David A. Barda
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
James R. Henry
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sean G. Buchanan
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Sean G. Buchanan
Robert M. Campbell
1Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, Indiana.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1158/1535-7163.MCT-18-0529 Published December 2019
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Abstract

Although Aurora A, B, and C kinases share high sequence similarity, especially within the kinase domain, they function distinctly in cell-cycle progression. Aurora A depletion primarily leads to mitotic spindle formation defects and consequently prometaphase arrest, whereas Aurora B/C inactivation primarily induces polyploidy from cytokinesis failure. Aurora B/C inactivation phenotypes are also epistatic to those of Aurora A, such that the concomitant inactivation of Aurora A and B, or all Aurora isoforms by nonisoform–selective Aurora inhibitors, demonstrates the Aurora B/C-dominant cytokinesis failure and polyploidy phenotypes. Several Aurora inhibitors are in clinical trials for T/B-cell lymphoma, multiple myeloma, leukemia, lung, and breast cancers. Here, we describe an Aurora A–selective inhibitor, LY3295668, which potently inhibits Aurora autophosphorylation and its kinase activity in vitro and in vivo, persistently arrests cancer cells in mitosis, and induces more profound apoptosis than Aurora B or Aurora A/B dual inhibitors without Aurora B inhibition–associated cytokinesis failure and aneuploidy. LY3295668 inhibits the growth of a broad panel of cancer cell lines, including small-cell lung and breast cancer cells. It demonstrates significant efficacy in small-cell lung cancer xenograft and patient-derived tumor preclinical models as a single agent and in combination with standard-of-care agents. LY3295668, as a highly Aurora A–selective inhibitor, may represent a preferred approach to the current pan-Aurora inhibitors as a cancer therapeutic agent.

This article is featured in Highlights of This Issue, p. 2183

Introduction

The Aurora kinases are composed of three highly conserved serine/threonine kinases: Aurora A, B, and C that regulate mitotic and meiotic processes (1–3). The proteins contain a highly conserved central kinase domain with a variable N-terminal domain critical for unique protein–protein interactions and a C-terminal domain carrying a protein stability regulation motif (4).

Aurora A is critical to centrosome maturation/separation, mitotic spindle formation, and mitotic spindle checkpoint activation (5–7). Aurora A mRNA/protein expression and kinase activity are cell-cycle regulated, with basal level at interphase and expression/activity dramatically increased from late G2 to telophase in mitosis. The kinase activity requires Aurora A autophosphorylation at Thr288 in the activation loop. Aurora A associates with centrosomes at interphase cells as well as with mitotic spindle in mitosis (8). Aurora A regulates mitotic progression by interacting with and phosphorylating multiple mitotic kinases, including targeting protein for Xklp2 (TPX2), p53, large tumor suppressor kinase 2 (LATS2), Rho-associated, coiled-coil–containing protein kinase (p160ROCK), and others (8–11).

Aurora B controls chromosome alignment in mitosis and cytokinesis as a catalytic unit in the chromosome passenger complex, which also contains Survivin, Borealin, and INCENP (12–14). Similar to Aurora A, Aurora B expression and localization are cell-cycle regulated, with expression and kinase activity peaking at mitosis and localization at kinetochores and centromeres from prophase to metaphase and then relocating to the cleavage furrow and midbody during cytokinesis. Meanwhile Aurora B kinase activity requires phosphorylation at Thr232 in the activation loop so that it can phosphorylate histone H3 at Ser10, and other substrates for chromosome condensation and spindle checkpoint (13, 15). Aurora C expression is restricted to the testis and functionally mimics Aurora B (16).

Although the three Aurora kinases share high sequence similarity, especially at the kinase domain, they function distinctly in cell-cycle progression (17). Aurora A is a mitotic kinase whose depletion primarily leads to mitotic spindle formation defects and consequently prometaphase arrest in human cancer cell lines, evidenced by the increase of histone H3 Ser10 phosphorylation as a mitotic marker. Aurora B/C function as chromosome passenger proteins such that kinase inactivation primarily induces polyploid cells. This ploidy is caused by cytokinesis failure, as evidenced by the failure of daughter cell separation and the Aurora B–mediated inhibition of histone Ser10 phosphorylation (18–20). Aurora B/C inactivation phenotypes are also epistatic to these of Aurora A. Concomitant inactivation of Aurora A and B, or all Aurora isoforms, demonstrates the Aurora B/C-dominant cytokinesis failure and polyploidy phenotypes. Polyploidy and subsequent chromosome segregation error-induced genomic instability are potential mechanisms of tumorigenesis and drug resistance. Thus, Aurora B/C inhibitory activity may counter the beneficial effects of Aurora A inhibition and should be avoided with a selective Aurora A inhibitor.

Although more than a dozen inhibitors have been developed, for example MLN8237 (alisertib), AZD1152 (barasertib), and others (21–25), most are either pan-Aurora or Aurora B/C inhibitors (26). As we show here, the most advanced Aurora inhibitor alisertib (MLN8237) has narrow Aurora A versus Aurora B selectivity, and at reported clinically efficacious concentrations (27), the Aurora B phenotype would dominate (27). Here we report development of a highly Aurora A–selective kinase inhibitor, LY3295668, which potently inhibits proliferation in a broad panel of cancer cell lines, induces apoptosis, and is highly efficacious in vivo, for example, in SCLC and other xenograft and patient-derived tumor (PDX) preclinical tumor models.

Materials and Methods

Cell lines and antibodies

NCI-H446, NCI-H69 (both from small-cell lung carcinoma), HeLa (cervical adenocarcinoma), Calu6, NCI-H358 (both from lung carcinoma), and Detroit 562 (pharyngeal carcinoma) were purchased from ATCC first in 2012 and later as needed. The panel of cell lines tested in Fig. 4 is listed in the Supplementary Information. The cell lines were pathogen tested and authenticated by short tandem repeat (STR)-based DNA profiling. All cell lines were cultured in conditions recommended by manufacturers at 37°C with or without 5% CO2 in a humidified atmosphere and used within 2 months from thaw. Growth curves were determined for all cell lines. Cell densities were optimized to achieve robust cell growth during assays.

The following antibodies against corresponding proteins were purchased and used according to the manufacturer's instructions: α-tubulin, Centrin 1 were from Abcam; actin, β-tubulin, Aurora A P-Thr288, Aurora B, P-Aurora A/B/C (P-Thr288/Thr232/Thr198, respectively), E-cadherin, vimentin, Sox2, cyclin B1, cleaved PARP, and Histone H3 were from Cell Signaling Technology; histone H3 P-Ser10 was from EMD Millipore, Aurora A was from R&D Systems; β-tubulin, actin, and GAPDH were from Sigma and Aldrich; horseradish peroxidase (HRP)-conjugated anti-rabbit and anti-mouse immunoglobulin G (IgG) were from GE Healthcare; and goat anti-mouse IgM Alexa Fluor-488, anti-mouse Alexa Fluor-647, anti-rabbit Alexa Fluor-488, and Hoechst 33342 from Thermo Fisher Scientific. The immunofluorescence microscopy was done with Leica DMI6000B or Zeiss LSM880 microscopes. For quantification of P-Aurora A and P-H3 in mitotic cells, images were acquired with the 40× water objective of an Opera Phenix and analyzed using Columbus software (Perkin Elmer).

Test compounds

LY3285668 was synthesized at Lilly Research Laboratories chemistry labs (example 1 in ref. 28). MLN8237 (LI-R034) was purchased from Shanghai ChemPartner Co. Ltd. AZD1152 (HM5015062) from Hutchison MediPharma Ltd., flavopiridol from AmFineCom, Inc. TGFβ was from R&D Systems and SB431542 from Sigma-Aldrich. All compounds were dissolved in DMSO as 10 mmol/L stock solution for in vitro assays. For in vivo animal studies, LY3295668 was formulated as a solution in 20% 2-hydroxypropyl-β -cyclodextrin (HPBCD) in 25 mmol/L phosphate buffer pH 2, adjusted with hydrochloric acid.

Aurora A and Aurora B enzymatic and kinase profiling assays

For details, see Supplementary Data.

Cell and tumor lysate preparations

For details, see Supplementary Data.

Aurora A inhibition by Aurora A P-Thr288 MSD and Aurora B inhibition by histone H3 P-Ser10 inhibition (1 hour) high-throughput cell-based assays

For details, please see Supplementary Data.

Mitotic arrest phenotypic and cell proliferation inhibition by histone H3 P-Ser10 and DNA content multiplexing (24 hours) cell-based assay

See Supplementary Data and ref. 40 for details.

Aurora A P-Thr288 and Aurora B histone H3 P-Ser10 in vivo target inhibition assays

NCI-H446 xenograft tumor lysates were used for in vivo target inhibition (IVTI). For details, see Supplementary Data.

Cell proliferation and apoptosis assays

For details, see Supplementary Data.

Epithelial-to-mesenchymal transition and cancer cell stemness assays

For details, see Supplementary Data and ref. 28.

Antitumor growth efficacy with small-cell lung cancer xenograft, PDX, and other tumor models

All in vivo studies were performed according to the Institutional Animal Care and Use Protocols of the party or provider conducting the experiments. NCI-H446, NCI-H69, and Detroit 562 xenograft and the small-cell lung cancer (SCLC) PDX models (in Oncotest GmbH; Charles River Laboratories) were used. For details, see Supplementary Data.

Western blot and immunofluorescence analyses

For details, see Supplementary Data.

Flow cytometry

For details, see Supplementary Data.

Results

LY3295668 is a potent Aurora A–selective inhibitor

LY3295668 [(2R,4R)-1-[(3-chloro-2-fluoro-phenyl)methyl]-4-[[3-fluoro-6-[(5-methyl-IH-pyrazol-3-yl)amino]-2-pyridyl]methyl]- 2-methylpiperidine-4-carboxylic acid] was identified during a medicinal chemistry program seeking to discover highly selective inhibitors of Aurora A kinase (Fig. 1A; ref. 29). It is a reversible ATP-competitive inhibitor with high aqueous solubility at 1.89 mg/mL. It also shows excellent physiochemical properties with LogD (pH 7.4) of 2.38 and measured fraction unbound of 1.0%, 1.1%, 1.4%, and 4.5% in human, dog, rat, and mouse sera, respectively.

Figure 1.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 1.

LY3295668 is a potent and highly selective Aurora A (AurA) inhibitor. A, LY3295668 chemical structure. B, Kinase binding profile from DiscoverX. All kinases with IC50 less than 0.1 μmol/L are listed. C, LY3295668 dose–response curves on Aurora A, Aurora B enzymatic (left), cell (middle) inhibitions, and mitotic phenotypic (right) profile. Enzymatically, Aurora A inhibition IC50 = 0.00112 ± 0.000148 μmol/L (n = 4, Embedded Image); Aurora B (AurB) IC50 1.51 ± 0.668 μmol/L (n = 5, Embedded Image). In cells, Aurora A inhibition IC50 = 0.00059 ± 0.000784 μmol/L (n = 4, Embedded Image); IC90 of 0.00623 ± 0.00318 μmol/L (n = 4, Embedded Image); and Aurora B inhibition IC50 = 1.42 ± 0.416 μmol/L (n = 4, Embedded Image). Mitotic index (MI; P-H3 increase) IC50 0.108 ± 0.015 μmol/L (n = 4, Embedded Image on left y-axis); G2–M arrest (4N DNA content increase) IC50 = 0.108 ± 0.027 μmol/L (n = 4, Δ on left y-axis); and cell growth inhibition IC50 = 0.053 ± 0.012 μmol/L (n = 4, Embedded Image on right y-axis). D, Multiple regression analysis for Aurora A (left), B (middle), and mitotic arrest (right) cell phenotype potency (on 448, 435, and 627 SAR cpds with available data points, respectively) with Aurora A and B enzymatic activity correlation.

LY3295668 potently inhibited Aurora A and showed excellent kinase selectivity in binding of a panel of 489 protein kinases in vitro, and carries significant binding activity for only 3 other protein kinases with IC50s less than 0.1 μmol/L (Fig. 1B). Enzymatically, LY3295668 showed remarkable selectivity against Aurora B (1,339-fold; Fig. 1C, left). In NCI-H446 cells, LY3295668 was a potent (IC50 = 0.00059 μmol/L) inhibitor on Aurora A kinase autophosphorylation at Thr288 (Fig. 1C, middle). Aurora B inhibition potency, as determined by histone H3 Ser10 phosphorylation, was very poor (IC50 = 1.42 μmol/L; Fig. 1C, middle), indicating a >2,000-fold selectivity of Aurora A over Aurora B in this cell-based assay. Consistent with LY3295668 Aurora A inhibitory activity, 24-hour treatment led to an increase in histone H3 Ser10 phosphorylation (mitotic arrest) as well as in G2–M DNA content (Fig. 1C, right) in HeLa cells. Accordingly, NCI-H446 and HeLa cell proliferations were inhibited (Fig. 1C, right; Supplementary Table S1). The correlation of Aurora A biochemical IC50 with P-H3 elevation, 4N DNA content increase, and cell proliferation inhibition indicates that LY3295668 Aurora A inhibition leads to mitotic arrest as well as cell proliferation inhibition.

Using a chemoproteomics platform (KiNativ; ref. 30) to detect the compound kinase-binding profile in NCI-H446 cancer cells, LY3295668 treatment at 1 μmol/L for 24 hours demonstrated an excellent Aurora A potency with inhibition efficiency at 98%, as well as kinase selectivity with only one other kinase (EphA2) showing significant inhibition (92%) among 203 kinases tested (Supplementary Table S2). Under this condition, PLK1, Aurora B, and EGFR kinase binding increased 256%, 265%, and 851% (n = 2), respectively. Both PLK1 and Aurora B expression are cell-cycle regulated, and it is expected that they would increase dramatically in mitosis (Supplementary Table S2), indicating LY3295668 treatment induces G2–M arrest.

We further used the KiNativ chemoproteomics platform to examine the LY3295668 kinase inhibition profile in vivo with the NCI-H446 xenograft model. When the animals were treated at 50 mg/kg every day for 3 hours, only Aurora A and Aurora B showed evidence of significant inhibition out of 195 kinases detectable (Supplementary Fig. S1A, left; Supplementary Table S3). When mice were treated with LY3295668 at 50 mg/kg twice a day (at 0 and 12 hours) and then tumors collected at 24 hours, the compound only inhibited Aurora A significantly, whereas PLK1, AURKB, and three other kinase expression increased significantly (Supplementary Fig. S1A, right), again indicating LY3295668 treatment in vivo led to Aurora A inhibition-induced mitotic arrest. In the mouse spleen, LY3295668 treatment at 50 mg/kg every day for 3 hours only significantly inhibited Aurora A (Supplementary Fig. S1B; Supplementary Table S4). Two treatments of LY3295668 (50 mg/kg twice a day, at 0 and 12 hours) again demonstrated that Aurora A is the only kinase significantly inhibited and a Serine/Threonine-Protein Kinase A-Raf is the only kinase with expression significantly increased (Supplementary Table S4). Taken together, LY3295668 consistently demonstrates selective Aurora A inhibition in vitro and in vivo.

Several different Aurora A and Aurora B kinase substrates in cells have been identified and used to characterize Aurora kinase inhibitors (26). Here, we are the first to use an Aurora A P-Thr288 inhibition assay to screen for an Aurora A–selective inhibitor. Pairwise analysis from 2,940 compounds from our structure–activity relationship (SAR) library indicates that Aurora A cell-based P-T288 inhibition is most highly correlated with Aurora A enzymatic assay not Aurora B (Spearman ρ = 0.7264; Supplementary Table S5). The more revealing correlation comes from multiple linear regression analysis. For Aurora A cell assay (Fig. 1D, left), improving Aurora A biochemical potency 10-fold (1 log unit) would have an expected improvement 10-fold (log Aurora A has estimate of 101.083) in Aurora A cell potency, while Aurora B biochemical 10-fold improvement can only contribute 1.6-fold (log Aurora B has estimate of 100.202). For Aurora B cell-based activity (Fig. 1D, middle), Aurora A enzymatic activity potency increase does not contribute at all (10−0.043), whereas 10-fold Aurora B enzymatic potency increase contributes to 5-fold increase in the Aurora B cell potency (100.639). So both pairwise and multiple regression correlation analyses support our flow scheme to identify Aurora A–selective inhibitors (Fig. 1D; Supplementary Table S5).

Aurora A and B inhibition are not equal in inducing mitotic arrest (Fig. 1D, right): improving Aurora A biochemical potency 10-fold contributes to mitotic arrest 2.9-fold (100.457) while Aurora B only 1.9-fold (100.268). These data indicate that Aurora A inhibition was much more effective in mitotic arrest induction, whereas Aurora B inhibition–induced endoreduplication and polyploidy phenotype may interfere with the mitotic arrest phenotype.

To determine the Aurora A inhibition potency required inducing significant cell proliferation inhibition and thus significant antitumor efficacy in vitro and in vivo, a statistical analysis was performed which indicates that 90% Aurora A inhibition (Aurora A P-T288 IC90) can effectively induce cell proliferation inhibition as well as mitotic arrest (Supplementary Fig. S2).

Aurora A–selective inhibition by LY3295668 produces mitotic arrest and apoptosis in cancer cells

The potency and selectivity of LY3295668 against Aurora A translated to mitotic arrest measured by histone H3 P-Ser10 marker and an increase in 4N DNA content in HeLa, NCI-H446, Calu6, and other tumor cell lines (Figs. 1B and 2A, left; Supplementary Fig. S3). Several Aurora inhibitors are being tested clinically, notably alisertib (MLN8237) and barasertib (AZD1152; refs. 21–23, 25). Each of these Aurora inhibitors was compared with LY3295668 in cell-based assays (Fig. 2). Alisertib is an Aurora A/B dual inhibitor with approximately 16-fold Aurora A selectivity versus Aurora B in cell-based assay (Fig. 2A, middle). Although mitotic arrest from LY3295668 treatment was still evident up to 20 μmol/L (the highest tested concentration), alisertib only showed Aurora A–selective inhibition between 0.05 and 0.25 μmol/L, but exhibited Aurora B–related P-H3 inhibition at concentrations above 0.25 μmol/L (Fig. 2A, middle). Barasertib also demonstrated an Aurora B–selective inhibitor dominated phenotype (Fig. 2A, right).

Figure 2.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 2.

LY3295668 selective Aurora A (AurA) inhibition leads to most significant apoptosis in HeLa cells. A, In comparison of alisertib and barasertib, LY3295668 showed the most persistent Aurora A inhibition–induced mitotic arrest. LY3295668 cell potency is listed in 1C (middle). Alisertib showed Aurora A cell IC50 0.004 ± 0.004 μmol/L (n = 5); Aurora B cell IC50 0.051 ± 0.010 μmol/L (n = 7); and mitotic index (MI; P-H3 increase) IC50 0.066 ± 0.039 μmol/L (n = 7). Barasertib exhibited Aurora A cell IC50 > 20 μmol/L; Aurora B cell IC50 0.011 ± 0.006 μmol/L (n = 11); and mitotic index (MI; P-H3 increase) IC50 = 0.022 ± 0.007 μmol/L (n = 4). B, LY3295668, alisertib and barasertib cell proliferation inhibition (left), and apoptosis induction (CASP3/7, right) in IncuCyte assay. Each square represents the cell confluency or the Caspase 3/7–positive green dot numbers (y-axis) in the cell culture well from 0 to 48 hours (x-axis). The percentage of cell growth or apoptosis-positive cell numbers is plotted below.

All three Aurora inhibitors potently inhibited HeLa cell proliferation (Fig. 2B, left). LY3295668 also demonstrated potent apoptosis induction with significant caspase 3/7 increase reaching maximal at 0.087 μmol/L and higher (Fig. 2B, right), correlating to its Aurora A inhibition, mitotic arrest, and cell proliferation inhibition potency (Fig. 2A, left). In contrast, alisertib showed most dramatic apoptosis induction only from 0.087 to 0.250 μmol/L, matching its Aurora A–selective cell activity range (Fig. 2A, middle). At concentrations of alisertib shown to be dominated by effects of Aurora B inhibition (>0.25 μmol/L), the apoptosis induction actually slightly decreased (Fig. 2B, right). Barasertib did not induce considerable apoptosis, reflected by less caspase 3/7 induction (Fig. 2B, right).

The cell growth inhibition and apoptosis induction difference among LY3295668 and other Aurora inhibitors (2.2 μmol/L) can also be observed from cell morphology images (Supplementary Fig. S4) taken from the same cell culture plates used for IncuCyte quantitation in Fig. 2B. The majority (96.4%) of HeLa cells were killed by LY3295668 showing caspase 3/7 positivity (green dots). However, there were still significant numbers of surviving cells showing enlarged/multinuclei and cytoplasm in alisertib or barasertib treatment, indicating they had gone through endoreduplication and cytokinesis failure mediated by Aurora B inhibition (Supplementary Fig. S4).

Immunofluorescent staining of HeLa cells treated with 1 μmol/L of LY3295668 clearly indicates mitotic arrest at prometaphase with abnormally shortened microtubule spindle network, disorganized arrays of microtubules (β-tubulin staining and Aurora A staining, which localizes to mitotic spindle and centrosomes), as well as misaligned condensed chromosomes (DAPI staining; Fig. 3A, top and middle). LY3295668 treatment also inhibited Aurora A P-Thr288 (P-AurA) staining at 94% (±0.468), with only a minor effect on histone H3 P-Ser10 (P-H3, 16% ± 0.2093), consistent with its Aurora A–selective inhibition (Fig. 3A, top and middle and B). Alisertib treatment also arrested cells in mitosis with disarrayed microtubule spindle and misaligned chromosomes, but abolished both Aurora A P-Thr288 (95% ± 0.433) and histone H3 P-Ser10 (93% ± 0.313) staining. This is indicative of an Aurora A/B dual inhibition phenotype (Fig. 3A, top and middle and B). Although the gross network of mitotic spindle was near normal, barasertib treatment led to misaligned chromosomes as well as corrupted mitotic spindle (Fig. 3A, top and middle). Barasertib inhibited histone H3 P-Ser10 (90% ± 4.054), but only marginally on Aurora A P-Thr288 (21% ± 2.976), representative of an Aurora B–dominant phenotype (Fig. 3A, top and middle, Fig. 3B).

Figure 3.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 3.

LY3295668 Aurora A (AurA) inhibition–induced mitotic arrest leads to apoptosis, not polyploidy. A, Immunofluorescent staining on mitotic spindle (β-tubulin), Aurora A, Aurora P-Thr288, histone H3 P-Ser10, and Centrin 1 localization on Hela cells treated with 1 μmol/L of LY3295668, alisertib, or barasertib for 24 hours. B, Quantification of P-Aurora A and P-H3 inhibition by the Aurora inhibitors from three repeated immunofluorescence experiments. Statistical significance is indicated by ***P < 0.001. C, Western blot analysis on mitotic and apoptosis markers with HeLa cells treated by indicated Aurora inhibitors for 24 (left) and 48 (right) hours. D, Western blot analysis on mesenchymal marker E-cadherin and stemness marker Sox2 in NCI-H358 cells. E, DNA histogram by flow cytometry on mitotic arrest and tetraploidy induction from indicated Aurora inhibitor treatment. HeLa cells were treated with 1 μmol/L of each compound for 24 (top) or 48 (bottom) hours.

PLK4 inhibition was also observed with LY3295668 (Fig. 1C) and can produce mitotic arrest. To distinguish between the Aurora A versus PLK4 inhibition–induced mitotic phenotype, cells were stained with Centrin 1 because centriole duplication is PLK4 dependent (31). Neither LY3295668, nor alisertib or barasertib perturbed centriole duplication and maturation with duplicated Centrin staining localized at the center of centrosomes in all the mitotic cells (Fig. 3A, bottom). These data indicate that although LY3295668 may inhibit PLK4 enzyme activity in vitro, this activity did not translate into significant inhibition of PLK4 in cells (Fig. 3A, bottom).

Western blot analysis further revealed the Aurora A versus Aurora B inhibition difference on mitotic arrest and apoptosis (Fig. 3C). At 0.25, 1, and 5 μmol/L, LY3295668-treated cells for 24 hours showed Aurora A, Aurora B, histone H3 P-Ser10, and cyclin B1 increases in a dose-dependent manner, indicating it effectively induced mitotic arrest (Fig. 3C, left). Although there was a transient increase in the expression of these endpoints by alisertib at 0.25 μmol/L (Aurora A inhibition), the increases were quickly lost due to transition to an Aurora B inhibition–dominant phenotype at concentrations >0.25 μmol/L. Cyclin B1 levels decreased from those at 0.25 μmol/L to those at 1 and 5 μmol/L correlated with its cytokinesis defects with Aurora B inhibition, because cyclin B1 degradation is cell-cycle regulated toward telophase and cytokinesis. Barasertib as an Aurora B–selective inhibitor did not show these target gene expression increase, and quite opposite, it induced histone H3 P-Ser10 decrease from its potent Aurora B kinase inhibition (Fig. 3C, left). LY3295668 exhibited dose-dependent PARP cleavage (apoptosis induction) across the concentration range tested (0.25–5 μmol/L), whereas alisertib induced apoptosis dose-dependently up to 0.25 μmol/L (most Aurora A–selective concentration), which then decreased at 1 and 5 μmol/L (Aurora B inhibition), and barasertib only induced moderately (Fig. 3C). Longer treatment for 48 hours showed similar trend (Fig. 3C, right).

Several recent studies show Aurora A functions in EMT and tumor stemness (32–34). We tested whether LY3295668 could also prevent EMT and reduce cancer stemness in a validated model cell line, NCI-H358 (28). As demonstrated previously, TGFβ inhibitor SB431542 reversed the TGFβ-induced epithelial marker E-cadherin decrease (Fig. 3D). LY3295668, but not alisertib or barasertib, was able to partially prevent the TGFβ-induced E-cadherin decrease at 0.1 μmol/L (Fig. 3D). Mesenchymal marker vimentin also showed a similar trend as seen with E-cadherin, albeit to a lesser degree (Supplementary Fig. S5). LY3295668 and alisertib inhibited Sox2 expression at the same concentration (Fig. 3D), indicating Aurora A inhibitors can reduce cancer cell stemness. However, Aurora B inhibitor barasertib did not cause a significant Sox2 decrease at the same concentration (Fig. 3D).

The Aurora A versus Aurora B selectivity of these compounds also impacted genomic stability as determined by flow cytometry analysis (Fig. 3E). LY3295668 induced strong G2–M arrest after 24 hours in HeLa cells (4N peak). The mitotic arrest persisted up to 48 hours without significant progression to 8N DNA content (tetraploid from the second round of genome replication). Although both alisertib and barasertib initially caused mitotic arrest by 24 hours treatment, they accumulated tetraploid (8N) cells after 48-hour treatment (2 doublings for HeLa cells) through endoreduplication and cytokinesis failure; a profile attributed to Aurora B inhibition. These polyploid cells can survive multiple rounds of endoreduplication without fully engaging apoptosis or other cell death pathway activation, which explains the surviving cells in Supplementary Fig. S4.

LY3295668 inhibits SCLC and breast cancer cell viability

LY3295668 showed significant inhibition of cancer cell viability across a panel of 80 tumor lines (Fig. 4), mostly from breast and lung cancers. The potency of LY3295668 in cell lines ranged from IC50 single-digit nmol/L to greater than 20 μmol/L (Fig. 4A). Fifty-five of 80 (68%) cell lines displayed significant sensitivity (IC50 < 1 μmol/L) to LY3295668 with an average IC50 of 0.048 μmol/L (Fig. 4A). Twenty-eight of 34 (82%) of breast, as well as 18/27 (67%) of lung cancer cell lines, are sensitive to LY3295668 treatment and have IC50s below 1 μmol/L. The average IC50s were 0.039 and 0.034 μmol/L, respectively, for the sensitive breast and lung panels (Fig. 4B and C). In breast cancer cells, triple-negative breast cancer was the most sensitive subtype with average IC50 0.164 μmol/L, and 13/15 (87%) lines had IC50 below 1 μmol/L, while luminal was the most insensitive with an average IC50 3.653 μmol/L and 13/19 (68%) had IC50 less than 1 μmol/L (Fig. 4B). In lung cancer cells, SCLC was the most sensitive tumor type with an average IC50 of 0.016 μmol/L and all 6 (100%) SCLC cell lines were sensitive to LY3295668 (Fig. 4C). Twelve of 21 (57%) of non–small cell lung cancer were sensitive to LY3295668 with an average IC50 of 4.694 μmol/L (Fig. 4C).

Figure 4.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 4.

LY3295668 shows potent cell viability inhibition in a large panel of cancer cells. A, Waterfall plot of LY3295668 cell viability inhibition potency log(IC50). The cell lines are indicated at the bottom of the plot. B, LY3295668 cell viability inhibition potency log(IC50) in breast cancer cell line subpanel. C, LY3295668 log(IC50) in lung cancer cell line subpanel. The cells at exponent growth phase were treated with LY3295668 for two doubling times and the cell viability inhibition was measured by CellTiter Glo assay.

A series of 10 compounds with Aurora A and/or Aurora B activity were tested in a bone marrow colony formation assay to better understand the heme toxicity relative Aurora kinase selectivity. LY3295668 showed an IC90 of 1.4 μmol/mL, while alisertib and barasertib gave IC90s at 0.2 and 0.1 μmol/mL, respectively. Multiple regression analysis indicates that human bone marrow cell inhibition was correlation with Aurora B cell potency, but not Aurora A (Supplementary Fig. S6). This would suggest that the heme toxicity observed in rodents and clinically (alisertib, barasertib) may be driven by the Aurora B activity. As such, an Aurora A–selective inhibitor might be better tolerated (also see ref. 35).

LY3295668 is a very potent Aurora A inhibitor in vivo

LY3295668 demonstrated potent Aurora A inhibition in vivo in the NCI-H446 xenograft model using nude mouse and rat (Fig. 5A) at 3 hours after oral dosing. At 30 mg/kg oral dosing in mice, LY3295668 also showed persistent Aurora A target inhibition (>90% P-Aurora A inhibition) for more than 8 hours and induced significant mitotic arrest as indicated by increases in mitotic maker histone H3 Ser10 phosphorylation (Fig. 5B, left). At 50 mg/kg, LY3295668 effectively inhibited Aurora A > 90% for 12 hours (96.0%, 95.1%, 94.5%, 90.7%, and 77.5% at 1, 4, 8, 12, and 24 hours, respectively). Correspondingly, mitotic arrest persisted up to 16 hours postdosing with histone H3 P-Ser10 increasing by 237%, 323%, 330%, 245%, and 328% at 1, 4, 8, 12, and 24 hours, respectively. LY3295668 has a calculated threshold effective dose (TED50: dose that produces an effect equal to 50% P-AurA inhibition) at 2.13 mg/kg and threshold effective concentration (TEC50: concentration that produces an effect equal to 50% P-AurA inhibition) of 0.07 μmol/L. We estimated that with twice-a-day dosing schedule, a dose of 30 mg/kg might be expected to achieve >90% Aurora A target inhibition for more than 16 hours daily, whereas a dose of 50 mg/kg would achieve this level of target engagement for the entire dosing period. At exposures comparable with those in the mouse, a 5 mg/kg oral dose in rats, LY3295668 showed persistent Aurora A target inhibition and a corresponding histone H3 P-Ser10 increase (Fig. 5B, right).

Figure 5.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 5.

LY3295668 inhibits Aurora A in vivo. A, In mouse and rat NCI-H446 xenograft models, animals were treated with LY3295668 at indicated doses for 3 hours. Aurora A P-Thr288 inhibition in tumors (Embedded Image y-axis at left) and compound exposure in animal plasma (Embedded Image y-axis on right) were measured. B, Animals were treated with LY3295668 once. At indicated time points, tumor and plasma samples were collected for Aurora A P-Thr288, P-H3 (Embedded Image and Embedded Image, y-axis on left), and compound exposure (Embedded Image y-axis on right). TED50, threshold effective dose (dose that produces an effect that equals to 50% P-Aurora A inhibition). TEC50, threshold effective concentration (plasma concentration that produces an effect that equals to 50% P-Aurora A inhibition).

LY3295668 at these doses did not inhibit histone H3 P-Ser10 at the 1-hour time point (Fig. 5B), indicating it did not show Aurora B inhibition in xenograft tumors. Taken together with the previous kinase profiling data (Fig. 1; Supplementary Fig. S1), LY3295668 consistently demonstrated that it is a potent and selective Aurora A inhibitor in vivo.

LY3295668 demonstrates potent and durable responses in preclinical tumor models

Inhibition of cell proliferation inhibition by LY3295668 was time-dependent, that is potency increased with longer incubation time (4–48 hours) and maximal inhibition reached after 24 to 48 hours (Supplementary Table S1; Supplementary Fig. S7). After 48 hours of LY3295668 treatment, proliferation IC50s were 0.090 ± 0.021 (n = 2), 0.108 ± 0.034 (n = 2), and 0.076 ± 0.033 (n = 2) μmol/L, respectively, in NCI-H446, HeLa, and Calu6 cells (Supplementary Table S1). The time-dependent cell proliferation inhibition in combination with the notion that 90% phospho-Aurora A inhibition was needed (Supplementary Fig. S2) enabled us to design an optimized in vivo efficacy schedule. We subsequently conducted in vivo efficacy studies by achieving >90% phospho-Aurora A inhibition for at least 16 hours/day (30 mg/kg) or 24 hours/day (50 mg/kg) in mice (Fig. 6A and B).

Figure 6.
  • Download figure
  • Open in new tab
  • Download powerpoint
Figure 6.

LY3295668 is efficacious in preclinical tumor models. A, LY3295668 was dosed at indicated doses and durations with NCI-H446 (left) and NCI-H69 (middle and right) models. The dosing durations for each treatment is indicated by the red line (left) and lines color coded to match the dosing schedules (middle and right). At the indicated time (red arrow), the average tumor growth inhibitions for each treatment group are plotted in the insert. B, LY3295668 efficacy with SCLC PDX models. LY3295668 was dosed at 50 mg/kg by orally twice a day for 28 days, whereas the standard-of-care was dosed with etoposide 30 mg/kg subcutaneously Q7D×4 (every 7 days for four times) plus cisplatin 3.2 mg/kg s.c. Q7D×4. The dosing duration is indicated by the red line. C, LY3295668 in combination with SoC showed better efficacy in SCLC PDX and head and neck xenograft models. For LXFS 2156 and 573 SCLC PDX models, LY3295668 was dosed at 40 mg/kg orally for 28 days. Standard of care was etoposide at 30 mg/kg s.c. Q7D×3 (every 7 days for three times) plus cisplatin 3.2 mg/kg s.c. Q7D×3. For Detroit 562 xenograft model, LY3295668 was dose at 30 mg/kg orally twice a day for 28 days. Cisplatin was used at 4 mg/kg Q7D×4 and cetuximab was at 20 mg/kg twice a week ×4. The dosing duration is indicated by the red line. The insert plot of tumor response rates at the measured time point are indicated by the red arrow, and the statistical significance is indicated by the P value.

LY3295668 demonstrated significant tumor growth inhibition in SCLC NCI-H446 xenograft mode in a dose-dependent manner (Fig. 6A, left). Tumors treated with 30 mg/kg LY3295668 regrew once dosing had stopped, whereas the tumors receiving 50 mg/kg LY3295668 did not show significant regrowth until 14 days postdosing (day-55 total duration, Fig. 6A, left). LY3295668 was well tolerated, with no significant body weight loss in the study.

We varied dosing schedule to understand the duration of LY3295668 dosing needed to reach durable efficacy in SCLC NCI-H69 xenograft model. LY3295668 at 50 mg/kg, dosed either as (BID×7, rest 14) × 2, (BID×14, rest 7) × 2, or (BID×21) × 2 schedules, all produced significant tumor growth inhibition (Fig. 6A, middle). Although the effect of (BID×7, rest 14) schedule did not persist after dosing stopped, both (BID14, rest 7) and BID×21 dosing schedules similarly produced sustained tumor growth inhibition; lasting more than 40 days (109-day total duration) postdosing (Fig. 6A, middle). We also tested comparable daily dose scenarios by dosing LY3295668 at 90 mg/kg BID×7, 50 mg/kg BID×14, or 30 mg/kg BID×21 (Fig. 6A, right). The results confirmed that constant, complete target inhibition (50 mg/kg BID×14) produced the most robust antitumor efficacy (Fig. 6A right) that persisted for at least 40 days postdosing (day-80 total duration). Should a dosing holiday be required, LY3295668 administered at concentrations achieving >90% Aurora A inhibition on a 14-day-on/7-day-off schedule would still be expected to reach maximal efficacy in SCLC (Fig. 6A).

We further demonstrated LY3295668 antitumor efficacy using SCLC patient-derived xenografts (PDX) models. In LXFS538, LY3295668 treatment induced 61.1% of tumor regression at day 56 of treatment whereas the standard-of care (etoposide + cisplatin) was only moderate at 43.1% tumor growth inhibition (Fig. 6B, left). When there was a significant tumor regrowth from 1 of the 5 tumors at day 70, LY3295668 retreatment at 50 mg/kg showed tumor regression again (Fig. 6B, left). In LXFS2156, LY3295668 treatment at 50 mg/kg BID×28 showed 97.2% of tumor growth inhibition whereas the standard-of-care did not show any significant tumor inhibition (Fig. 6B, right). However, the tumor inhibition in LY3295668-treated group did not persist beyond dosing period (Fig. 6B, right).

Efficacy of LY3295668 was also explored in combination with various standards of care. In LXFS2156, LY3295668 at 40 mg/kg showed significant efficacy, whereas standard of care (Etoposide + Cisplatin) did not significantly affect tumor growth (Fig. 6C, left). LY3295668 in combination with standard of care produced better-than-additive effect than either monotherapy (Fig. 6C, left). In LXFS573, LY3295668 also showed significant combination benefit with standard-of-care agents (Fig. 6C, middle). In these studies, 10% to 15% of body weight losses were observed in combination groups.

Multiple lines of data indicate potential synergism between Aurora A and EGFR inhibitors (36). Consequently, LY3295668 was investigated for potential synergy in combination with cetuximab and cisplatin in a head and neck squamous cell carcinoma (HNSCC), the Detroit 562 xenograft model (Fig. 6C, right). At the end of dosing period (day 44 total duration; 4-day postdosing regimen completed), LY3295668 at 30 mg/kg alone showed significant tumor growth inhibition, so did cetuximab or cetuximab plus cisplatin treatment. More importantly, LY3295668 in combination with cetuximab or with both cetuximab plus cisplatin exhibited a synergistic effect in comparison with either monotherapy, produced much prolonged tumor growth inhibition beyond the dosing period, especially LY3295668 with cetuximab + cisplatin group (Fig. 6C, right).

Discussion

Although Aurora kinases A, B, and C are highly homologous, they are not functional orthologs. Aurora A–selective inhibition leads to mitotic arrest in many cancer cells. The mitotic arrest often further progresses to cancer cell apoptosis/death in sensitive cancer cell lines. We contend that an Aurora A–selective inhibitor such as LY3295668 may be more effective than Aurora B or Aurora A/B dual inhibitors (alisertib, barasertib), as it would avoid DNA endoreduplication and polyploidy, which could persist for multiple cell divisions without completely killing the cancer cells (37). A couple of Aurora A inhibitors have been identified over the years; however, they suffer either the lack of Aurora A inhibition potency or lack of enough selectivity against Aurora B (Fig. 2; refs. 24, 26). LY3295668 is a potential best-in-class Aurora A inhibitor with >1,000-fold Aurora A versus Aurora B selectivity. Furthermore, we contend that at the pharmacologically relevant concentrations, LY3295668 only inhibits Aurora A in vitro and in vivo, not Aurora B.

Using LY3295668, we discovered that Aurora A versus Aurora B/C inhibitors show profound differences in mitotic arrest phenotypes and especially apoptosis induction. Although LY3295668, alisertib, and barasertib each can potently inhibit HeLa cell growth, only LY3295668 consistently promoted apoptosis across the range of tested concentrations, as well as inhibits EMT and cancer cell stemness markers (Figs. 2 and 3). Interestingly, alisertib inhibits Aurora A selectively in a range of 0.080–0.250 μmol/L. This corresponds to the same range of concentrations needed to induce maximal apoptosis in HeLa cells. At concentrations >0.25 μmol/L, alisertib reduced its apoptosis induction capacity as it switched to an Aurora B–dominant phenotype (Figs. 2 and 3). So at a clinically relevant human plasma exposure of 1 to 2 μmol/L (38–41), alisertib behaves phenotypically as an Aurora B inhibitor in vivo (42). This phenotype has been observed previously in sarcoma cells, although alisertib showed an Aurora A inhibition phenotype (increased phospho-H3) in their model (42). Although alisertib is a dual Aurora A/B inhibitor in vitro, the appearance of the Aurora B phenotype is exposure-dependent, that is, at lower plasma concentrations, only Aurora A may be inhibited.

Aurora B inhibition–induced endoreduplication and subsequent polyploidy are retinoblastoma protein-dependent and can lead to chromosome segregation errors and thus genomic instability through aberrant mitosis (Figs. 2 and 3; refs. 17, 18, 23, 26, 35, 43). As such, Aurora B inhibitor–induced tetraploid cells, which show better survival after compound treatment (Figs. 2 and 3), could significantly increase the refractory/resistant cell population, especially when the inhibitor in vivo exposure is at subthreshold levels during dosing intervals. Aneuploidy/genomic instability is one of the most prevalent signatures of cancer (44, 45). Polyploidy, including tetraploidy, is thought to be a major pathway to induce aneuploidy and genomic instability. For example, RAS oncogene induces tetraploidy as an intermediate step in its tumorigenesis (46). As for cancer therapy, Aurora A–selective inhibitor may be the best strategy and Aurora B activity should be avoided. LY3295668 has now advanced to phase I clinical testing to test the translation of this hypothesis in patients (NCT03092934).

Disclosure of Potential Conflicts of Interest

J. Du is a research advisor at Eli Lilly & Company. L. Yan is a research scientist at Eli Lilly & Company. R. Torres is a senior research scientist Eli Lilly & Company. X. Gong is a principal research scientist at Eli Lilly & Company. C. Marugán is a research associate at Eli Lilly & Company. C. Baquero is a research associate at Eli Lilly & Company. S.C. Chapman is a senior research scientist and has ownership interest (including patents) in Eli Lilly & Company. Y. Zeng is a consultant at Eli Lilly & Company. G.P. Donoho is a director, oncology research, In Vivo Pharmacology and has ownership interest (including patents) in Eli Lilly & Company. R.D. Van Horn is a consultant biologist and has ownership interest (including patents) in Eli Lilly & Company. D.S. Barnard is a consultant biologist and has ownership interest (including patents) in Eli Lilly & Company. R.P. Beckmann is a research scientist and has ownership interest (including patents) in Eli Lilly & Company. L.-C. Chio is a biology consultant at Eli Lilly & Company. A. Aggarwal is a Head, Oncology Genomics and Bioinformatics and has ownership interest (including patents) in Eli Lilly & Company. L.F. Stancato is a research fellow at Eli Lilly & Company. P.W. Iversen is a senior research advisor at Eli Lilly & Company. J.R. Manro is a senior research scientist and has ownership interest (including patents) in Eli Lilly & Company. J.L. Walgren is an advisor of regulatory affairs at Eli Lilly & Company. B.W. Halstead is a consultant oncology BDS&E at Eli Lilly & Company. M.Z. Dieter is a principal research scientist at Eli Lilly & Company. S.V. Bhagwat is a senior research advisor at Eli Lilly & Company. E.L. Kreklau is a senior director at Eli Lilly & Company. B.K.R. Patel is a research fellow and has ownership interest (including patents) in Eli Lilly & Company. C. Reinhard is a senior director at Eli Lilly & Company. G.D. Plowman is a vice president, oncology research at Eli Lilly & Company. D.A. Barda is a research scientist and has ownership interest (including patents) in Eli Lilly & Company. J.R. Henry is a senior research fellow and has ownership interest (including patents) in Eli Lilly & Company. S.G. Buchanan has ownership interest (including patents) in Eli Lilly & Company. R.M. Campbell is a research fellow and has ownership interest (including patents) in Eli Lilly & Company. No potential conflicts of interest were disclosed by the other authors.

Authors' Contributions

Conception and design: J. Du, L. Yan, X. Gong, S.C. Chapman, R.P. Beckmann, M.S. Marshall, P.W. Iversen, E.L. Kreklau, M.J. Lallena, X.S. Ye, C. Reinhard, G.D. Plowman, D.A. Barda, J.R. Henry, S.G. Buchanan, R.M. Campbell

Development of methodology: J. Du, L. Yan, R. Torres, X. Gong, H. Bian, K. Boehnke, R.T. Foreman, R.P. Beckmann, M.S. Marshall, L.-C. Chio, S. Chu, S. Bhattachar, L.F. Stancato, J.L. Walgren, S.V. Bhagwat, E.L. Kreklau, M.J. Lallena, X.S. Ye

Acquisition of data (provided animals, acquired and managed patients, provided facilities, etc.): J. Du, R. Torres, X. Gong, H. Bian, C. Marugán, K. Boehnke, C. Baquero, Y.-H. Hui, Y. Yang, Y. Zeng, S.M. Bogner, A. Capen, G.P. Donoho, R.D. Van Horn, D.S. Barnard, M.S. Marshall, Y. Qian, A. Aggarwal, M.S. Dowless, J.L. Walgren, M.Z. Dieter, M.J. Lallena

Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis): J. Du, L. Yan, R. Torres, X. Gong, C. Marugán, K. Boehnke, C. Baquero, Y.-H. Hui, S.C. Chapman, R.D. Van Horn, Y.W. Webster, A. Aggarwal, L.F. Stancato, P.W. Iversen, J.R. Manro, J.L. Walgren, B.W. Halstead, R. Martinez, M.J. Lallena, B.K.R. Patel, D.A. Barda, J.R. Henry, S.G. Buchanan, R.M. Campbell

Writing, review, and/or revision of the manuscript: J. Du, R. Torres, Y.-H. Hui, S.C. Chapman, G.P. Donoho, R.P. Beckmann, A. Aggarwal, L.F. Stancato, P.W. Iversen, J.R. Manro, B.W. Halstead, M.Z. Dieter, R. Martinez, E.L. Kreklau, M.J. Lallena, X.S. Ye, B.K.R. Patel, G.D. Plowman, D.A. Barda, J.R. Henry, S.G. Buchanan, R.M. Campbell

Administrative, technical, or material support (i.e., reporting or organizing data, constructing databases): J. Du, R. Torres, Y.-H. Hui, J.A. Dempsey, J.R. Manro, D.A. Barda

Study supervision: J. Du, A. Capen, G.P. Donoho, X.S. Ye, C. Reinhard, G.D. Plowman, S.G. Buchanan, R.M. Campbell

Acknowledgments

We would like to thank Drs. James Starling, Alfonso de Dios, Michal Vieth, Daphne L. Farrington, Harold B. Brooks, John Toth, Farhana Merzoug, Karen L. Cox, Susan A. Konicek, and Teresa Burke for their guidance and support. This work was supported by Eli Lilly & Company.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Footnotes

  • Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

  • Mol Cancer Ther 2019;18:2207–19

  • Received May 15, 2018.
  • Revision received April 29, 2019.
  • Accepted September 6, 2019.
  • Published first September 17, 2019.
  • ©2019 American Association for Cancer Research.

References

  1. 1.↵
    1. Glover DM,
    2. Leibowitz MH,
    3. McLean DA,
    4. Parry H
    . Mutations in aurora prevent centrosome separation leading to the formation of monopolar spindles. Cell 1995;81:95–105.
    OpenUrlCrossRefPubMed
  2. 2.↵
    1. Bischoff JR,
    2. Plowman GD
    . The Aurora/IPL1 kinase family: regulators of chromosome segregation and cytokinesis. Trends Cell Biol 1999;9:454–9.
    OpenUrlCrossRefPubMed
  3. 3.↵
    1. Goldenson B,
    2. Crispino JD
    . The aurora kinases in cell cycle and leukemia. Oncogene 2014;34:537–45.
    OpenUrlPubMed
  4. 4.↵
    1. Karthigeyan D,
    2. Prasad SB,
    3. Shandilya J,
    4. Agrawal S,
    5. Kundu TK
    . Biology of Aurora A kinase: implications in cancer manifestation and therapy. Med Res Rev 2011;31:757–93.
    OpenUrlPubMed
  5. 5.↵
    1. Bischoff JR,
    2. Anderson L,
    3. Zhu Y,
    4. Mossie K,
    5. Ng L,
    6. Souza B,
    7. et al.
    A homologue of Drosophila aurora kinase is oncogenic and amplified in human colorectal cancers. EMBO J 1998;17:3052–65.
    OpenUrlAbstract/FREE Full Text
  6. 6.↵
    1. Zhou H,
    2. Kuang J,
    3. Zhong L,
    4. Kuo WL,
    5. Grey JW,
    6. Sahin A,
    7. et al.
    Tumor amplified kinase STK15/BTAK induces centrosome amplification, aneuploidy and transformation. Nat Genet 1998;20:189–93.
    OpenUrlCrossRefPubMed
  7. 7.↵
    1. Katayama H,
    2. Zhou H,
    3. Li Q,
    4. Tatsuka M
    . Interaction and feedback regulation between STK15/BTAK/Aurora-A kinase and protein phosphatase 1 through mitotic cell division cycle. J Biol Chem 2001;276:46219–24.
    OpenUrlAbstract/FREE Full Text
  8. 8.↵
    1. Kufer TA,
    2. Silljé HHW,
    3. Körner R,
    4. Gruss OJ,
    5. Meraldi P,
    6. Nigg EA
    . Human TPX2 is required for targeting Aurora-A kinase to the spindle. J Cell Biol 2002;158:617–23.
    OpenUrlAbstract/FREE Full Text
  9. 9.↵
    1. Liu Q,
    2. Kaneko S,
    3. Yang L,
    4. Feldman FI,
    5. Nicosia SV,
    6. Chen C,
    7. et al.
    Aurora-A abrogation of p53 DNA binding and transactivation activity by phosphorylation of serine-215. JBC 2004;279:52175–82.
    OpenUrlAbstract/FREE Full Text
  10. 10.↵
    1. Toji S,
    2. Yabuta N,
    3. Hosomi T,
    4. Nishihara S,
    5. Kobayashi T,
    6. Suzuki S,
    7. et al.
    The centrosomal protein Lats2 is a phosphorylation target of Aurora-A kinase. Genes Cells 2004;9:383–97.
    OpenUrlCrossRefPubMed
  11. 11.↵
    1. Du J,
    2. Hannon GJ
    . Suppression of p160ROCK bypasses cell cycle arrest after Aurora-A/STK15 depletion. Proc Natl Acad Sci U S A 2004;101:8975–80.
    OpenUrlAbstract/FREE Full Text
  12. 12.↵
    1. Hsu JY,
    2. Sun ZW,
    3. Li X,
    4. Beuben M,
    5. Tatchell K,
    6. Bishop DK,
    7. et al.
    Mitotic phosphorylation of histone H3 is governed by Ipl1/aurora kinase and Glc7/PP1 phosphatase in budding yeast and nematodes. Cell 2000;102:279–91.
    OpenUrlCrossRefPubMed
  13. 13.↵
    1. Giet R,
    2. Glove DM
    . Drosophila aurora B kinase is required for histone H3 phosphorylation and condensin recruitment during chromosome condensation and to organize the central spindle during cytokinesis. J Cell Biol 2001;152:669–82.
    OpenUrlAbstract/FREE Full Text
  14. 14.↵
    1. Adams RR,
    2. Maiato H,
    3. Earnshaw WC,
    4. Carmena M
    . Essential roles of Drosophila inner centromere protein (INCENP) and aurora B in histone H3 phosphorylation, metaphase chromosome alignment, kinetochore disjunction, and chromosome segregation. J Cell Biol 2001;153:865–80.
    OpenUrlAbstract/FREE Full Text
  15. 15.↵
    1. Yasui Y,
    2. Urano T,
    3. Kawajiri A,
    4. Nagata K,
    5. Tatsuka M,
    6. Saya H,
    7. et al.
    Autophosphorylation of a newly identified site of Aurora-B is indispensable for cytokinesis. J Biol Chem 2004;279:12997–3003.
    OpenUrlAbstract/FREE Full Text
  16. 16.↵
    1. Sasai K,
    2. Katayama H,
    3. Stenoien DL,
    4. Fujii S,
    5. Honda R,
    6. Kimura M,
    7. et al.
    Aurora-C kinase is a novel chromosomal passenger protein that can complement Aurora-B kinase function in mitotic cells. Cell Motil Cytoskeleton 2004;59:249–63.
    OpenUrlCrossRefPubMed
  17. 17.↵
    1. Hilton JF,
    2. Shapiro GI
    . Aurora kinase inhibition as an anticancer strategy. J Clin Oncol 2013;31:1–3.
    OpenUrlFREE Full Text
  18. 18.↵
    1. Yang H,
    2. Burke T,
    3. Dempsey J,
    4. Diaz B,
    5. Collins E,
    6. Toth J,
    7. et al.
    Mitotic requirement for aurora A kinase is bypassed in the absence of aurora B kinase. FEBS Lett 2005;579:3385–91.
    OpenUrlCrossRefPubMed
  19. 19.↵
    1. Warner SL,
    2. Munoz RM,
    3. Stafford P,
    4. Koller E,
    5. Hurley LH,
    6. Von Hoff DD,
    7. et al.
    Comparing Aurora A and Aurora B as molecular targets for growth inhibition of pancreatic cancer cells. Mol Cancer Ther 2006;5:2450–8.
    OpenUrlAbstract/FREE Full Text
  20. 20.↵
    1. Li Y,
    2. Xu FL,
    3. Lu J,
    4. Saunders WS,
    5. Prochownik EV
    . Widespread genomic instability mediated by a pathway involving glycoprotein Ib alpha and Aurora B kinase. J Biol Chem 2010;285:13183–92.
    OpenUrlAbstract/FREE Full Text
  21. 21.↵
    1. Gorgun G,
    2. Calabrese E,
    3. Hideshima T,
    4. Ecsedy J,
    5. Perrone G,
    6. Mani M,
    7. et al.
    A novel Aurora-A kinase inhibitor MLN8237 induces cytotoxicity and cell-cycle arrest in multiple myeloma. Blood 2010;115:5202–13.
    OpenUrlAbstract/FREE Full Text
  22. 22.↵
    1. Wilkinson RW,
    2. Odedra R,
    3. Heaton SP,
    4. Wedge SR,
    5. Keen NJ,
    6. Carfter C,
    7. et al.
    AZD1152, a selective inhibitor of Aurora B kinase, inhibits human tumor xenograft growth by inducing apoptosis. Clin Cancer Res 2007;13:3682–8.
    OpenUrlAbstract/FREE Full Text
  23. 23.↵
    1. Nair JS,
    2. Ho AL,
    3. Tse AN,
    4. Coward J,
    5. Cheema H,
    6. Ambrosini G,
    7. et al.
    Aurora B kinase regulates the postmitotic endoreduplication checkpoint via phosphorylation of the retinoblastoma protein at serine 780. Mol Biol Cell 2009;20:2218–28.
    OpenUrlAbstract/FREE Full Text
  24. 24.↵
    1. Nair JS,
    2. Ho AL,
    3. Schwartz GK
    . The induction of polyploidy or apoptosis by the Aurora A kinase inhibitor MK8745 is p53-dependent. Cell Cycle 2012;11:807–17.
    OpenUrlCrossRefPubMed
  25. 25.↵
    1. Nair JS,
    2. Schwartz GK
    . MLN-8237: a dual inhibitor of aurora A and B in soft tissue sarcomas. Oncotarget 2016;7:12893–903.
    OpenUrlPubMed
  26. 26.↵
    1. de Groot CO,
    2. Hsia JE,
    3. Anzola JV,
    4. Motamedi A,
    5. Yoon M,
    6. Wong YL,
    7. et al.
    A cell biologist's field guide to aurora kinase inhibitors. Front Oncol 2015;5:1–25.
    OpenUrlPubMed
  27. 27.↵
    1. Dickson MA,
    2. Mahoney MR,
    3. Tap WD,
    4. D'Angelo SP,
    5. Keohan ML,
    6. Van Tine BA,
    7. et al.
    Phase II study of MLN8237 (Alisertib) in advanced/metastatic sarcoma. Ann Oncol 2016;10:1855–60.
    OpenUrl
  28. 28.↵
    1. Thomson S,
    2. Petti F,
    3. Sujka-Kwok I,
    4. Epstein D,
    5. Haley JD
    . Kinase switching in mesenchymal-like non-small cell lung cancer lines contributes to EGFR inhibitor resistance through pathway redundancy. Clin Exp Metastasis 2008;25:843–54.
    OpenUrlCrossRefPubMed
  29. 29.↵
    1. Henry JR
    , inventor; Eli Lilly and Company, assignee. Preparation of aminopyridine compounds as Aurora A kinase inhibitors; May, 2017. United Statespatent 9637474 (example 1).
  30. 30.↵
    1. Patricelli MP,
    2. Szardenings AK,
    3. Liyanage M,
    4. Nomanbhoy TK,
    5. Wu M,
    6. Weissig H,
    7. et al.
    Functional interrogation of the kinome using nucleotide acyl phosphates. Biochemistry 2007;46:350–8.
    OpenUrlCrossRefPubMed
  31. 31.↵
    1. Habedanck R,
    2. Stierhof YD,
    3. Wilkinson CJ,
    4. Nigg EA
    . The Polo kinase Plk4 functions in centriole duplication. Nat Cell Biol 2005;7:1140–6.
    OpenUrlCrossRefPubMed
  32. 32.↵
    1. Ren BJ,
    2. Zhou ZW,
    3. Zhu DJ,
    4. Ju YL,
    5. Wu JH,
    6. Ouyang MZ,
    7. et al.
    Alisertib induces cell cycle arrest, apoptosis, autophagy and suppresses EMT in HT29 and Caco-2 Cells. Int J Mol Sci 2015;17:E41.
    OpenUrl
  33. 33.↵
    1. Chen C,
    2. Song G,
    3. Xiang J,
    4. Zhang H,
    5. Zhao S,
    6. Zhan Y
    . AURKA promotes cancer metastasis by regulating epithelial-mesenchymal transition and cancer stem cell properties in hepatocellular carcinoma. Biochem Biophys Res Commun 2017;486:514–20.
    OpenUrl
  34. 34.↵
    1. Opyrchal M,
    2. Gil M,
    3. Salisbury JL,
    4. Goetz MP,
    5. Suman V,
    6. Degnim A,
    7. et al.
    Molecular targeting of the Aurora-A/SMAD5 oncogenic axis restores chemosensitivity in human breast cancer cells. Oncotarget 2017;8:91803–16.
    OpenUrl
  35. 35.↵
    1. Gong X,
    2. Du J,
    3. Parsons SH,
    4. Merzoug FF,
    5. Webster Y,
    6. Iversen PW,
    7. et al.
    Aurora-A kinase inhibition is synthetic lethal with loss of the RB1 tumor suppressor gene. Cancer Discov 2019;9:248–263.
    OpenUrlAbstract/FREE Full Text
  36. 36.↵
    1. Astsaturov I,
    2. Ratushny V,
    3. Sukhanova A,
    4. Einarson MB,
    5. Bagnyukova T,
    6. Zhou Y,
    7. et al.
    Synthetic lethal screen of an EGFR-centered network to improve targeted therapies. Sci Signal 2010;3:1–18.
    OpenUrlCrossRefPubMed
  37. 37.↵
    1. Kaestner P,
    2. Stolz A,
    3. Bastians H
    . Determinants for the efficiency of anticancer drugs targeting either Aurora-A or Aurora-B kinases in human colon carcinoma cells. Mol Cancer Ther 2009;8:2046–56.
    OpenUrlAbstract/FREE Full Text
  38. 38.
    1. Venkatakrishnan K,
    2. Zhou X,
    3. Ecsedy J,
    4. Mould DR,
    5. Liu H,
    6. Danaee H,
    7. et al.
    Dose selection for the investigational anticancer agent alisertib (MLN8237): Pharmacokinetics, pharmacodynamics, and exposure-safety relationships. J Clin Pharmacol 2015;55:336–47.
    OpenUrl
  39. 39.↵
    1. Dees EC,
    2. Cohen RB,
    3. von Mehren M,
    4. Stinchcombe TE,
    5. Liu H,
    6. Venkatakrishnan K,
    7. et al.
    Phase I study of aurora A kinase inhibitor MLN8237 in advanced solid tumors: safety, pharmacokinetics, pharmacodynamics, and bioavailability of two oral formulations. Clin Cancer Res 2012;18:4775–84.
    OpenUrlAbstract/FREE Full Text
  40. 40.↵
    1. Cervantes A,
    2. Elez E,
    3. Roda D,
    4. Ecsedy J,
    5. Macarulla T,
    6. Venkatakrishnan K,
    7. et al.
    Phase I pharmacokinetic/pharmacodynamic study of MLN8237, an investigational, oral, selective aurora a kinase inhibitor, in patients with advanced solid tumors. Clin Cancer Res 2012;18:4764–74.
    OpenUrlAbstract/FREE Full Text
  41. 41.↵
    1. Matulonis UA,
    2. Sharma S,
    3. Ghamande S,
    4. Gordon MS,
    5. Del Prete SA,
    6. Ray-Coquard I,
    7. et al.
    Phase II study of MLN8237 (alisertib), an investigational Aurora A kinase inhibitor, in patients with platinum-resistant or -refractory epithelial ovarian, fallopian tube, or primary peritoneal carcinoma. Gynecol Oncol 2012;127:63–9.
    OpenUrlCrossRefPubMed
  42. 42.↵
    1. Dickson MA,
    2. Mahoney MR,
    3. Tap WD,
    4. D'Angelo SP,
    5. Keohan ML,
    6. Van Tine BA,
    7. et al.
    Phase II study of MLN8237 (alisertib) in advanced/metastatic sarcoma. Ann Oncol 2016;27:1855–60.
    OpenUrlCrossRefPubMed
  43. 43.↵
    1. Oser MG,
    2. Fonseca R,
    3. Chakraborty AA,
    4. Brough R,
    5. Spektor A,
    6. Jennings RB,
    7. et al.
    Cells lacking the RB1 tumor suppressor gene are hyperdependent on aurora B kinase for survival. Cancer Discov 2019;9:230–247.
    OpenUrlAbstract/FREE Full Text
  44. 44.↵
    1. Hanahan D,
    2. Weinberg RA
    . Hallmarks of cancer: the next generation. Cell 2011;144:646–74.
    OpenUrlCrossRefPubMed
  45. 45.↵
    1. Storchova Z,
    2. Kuffer C
    . The consequences of tetraploidy and aneuploidy. J Cell Sci 2008;121:3859–66.
    OpenUrlAbstract/FREE Full Text
  46. 46.↵
    1. Zeng X,
    2. Shaikh FY,
    3. Harrison MK,
    4. Adon AM,
    5. Trimboli AJ,
    6. Carroll KA,
    7. et al.
    The Ras oncogene signals centrosome amplification in mammary epithelial cells through cyclin D1/Cdk4 and Nek2. Oncogene 2010;29:5103–12.
    OpenUrlCrossRefPubMed
PreviousNext
Back to top
Molecular Cancer Therapeutics: 18 (12)
December 2019
Volume 18, Issue 12
  • Table of Contents
  • Table of Contents (PDF)
  • About the Cover
  • Editorial Board (PDF)

Sign up for alerts

View this article with LENS

Open full page PDF
Article Alerts
Sign In to Email Alerts with your Email Address
Email Article

Thank you for sharing this Molecular Cancer Therapeutics article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Aurora A–Selective Inhibitor LY3295668 Leads to Dominant Mitotic Arrest, Apoptosis in Cancer Cells, and Shows Potent Preclinical Antitumor Efficacy
(Your Name) has forwarded a page to you from Molecular Cancer Therapeutics
(Your Name) thought you would be interested in this article in Molecular Cancer Therapeutics.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Citation Tools
Aurora A–Selective Inhibitor LY3295668 Leads to Dominant Mitotic Arrest, Apoptosis in Cancer Cells, and Shows Potent Preclinical Antitumor Efficacy
Jian Du, Lei Yan, Raquel Torres, Xueqian Gong, Huimin Bian, Carlos Marugán, Karsten Boehnke, Carmen Baquero, Yu-Hua Hui, Sonya C. Chapman, Yanzhu Yang, Yi Zeng, Sarah M. Bogner, Robert T. Foreman, Andrew Capen, Gregory P. Donoho, Robert D. Van Horn, Darlene S. Barnard, Jack A. Dempsey, Richard P. Beckmann, Mark S. Marshall, Li-Chun Chio, Yuewei Qian, Yue W. Webster, Amit Aggarwal, Shaoyou Chu, Shobha Bhattachar, Louis F. Stancato, Michele S. Dowless, Phillip W. Iversen, Jason R. Manro, Jennie L. Walgren, Bartley W. Halstead, Matthew Z. Dieter, Ricardo Martinez, Shripad V. Bhagwat, Emiko L. Kreklau, Maria Jose Lallena, Xiang S. Ye, Bharvin K.R. Patel, Christoph Reinhard, Gregory D. Plowman, David A. Barda, James R. Henry, Sean G. Buchanan and Robert M. Campbell
Mol Cancer Ther December 1 2019 (18) (12) 2207-2219; DOI: 10.1158/1535-7163.MCT-18-0529

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Share
Aurora A–Selective Inhibitor LY3295668 Leads to Dominant Mitotic Arrest, Apoptosis in Cancer Cells, and Shows Potent Preclinical Antitumor Efficacy
Jian Du, Lei Yan, Raquel Torres, Xueqian Gong, Huimin Bian, Carlos Marugán, Karsten Boehnke, Carmen Baquero, Yu-Hua Hui, Sonya C. Chapman, Yanzhu Yang, Yi Zeng, Sarah M. Bogner, Robert T. Foreman, Andrew Capen, Gregory P. Donoho, Robert D. Van Horn, Darlene S. Barnard, Jack A. Dempsey, Richard P. Beckmann, Mark S. Marshall, Li-Chun Chio, Yuewei Qian, Yue W. Webster, Amit Aggarwal, Shaoyou Chu, Shobha Bhattachar, Louis F. Stancato, Michele S. Dowless, Phillip W. Iversen, Jason R. Manro, Jennie L. Walgren, Bartley W. Halstead, Matthew Z. Dieter, Ricardo Martinez, Shripad V. Bhagwat, Emiko L. Kreklau, Maria Jose Lallena, Xiang S. Ye, Bharvin K.R. Patel, Christoph Reinhard, Gregory D. Plowman, David A. Barda, James R. Henry, Sean G. Buchanan and Robert M. Campbell
Mol Cancer Ther December 1 2019 (18) (12) 2207-2219; DOI: 10.1158/1535-7163.MCT-18-0529
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Jump to section

  • Article
    • Abstract
    • Introduction
    • Materials and Methods
    • Results
    • Discussion
    • Disclosure of Potential Conflicts of Interest
    • Authors' Contributions
    • Acknowledgments
    • Footnotes
    • References
  • Figures & Data
  • Info & Metrics
  • PDF
Advertisement

Related Articles

Cited By...

More in this TOC Section

  • Antineoplastic Effects of a Novel CDK2/9 Inhibitor CYC065
  • MTX-23, a Novel PROTAC That Degrades AR-V7 and AR-FL
  • Glutaminase Inhibition Improves Efficacy of Immunotherapies
Show more Small Molecule Therapeutics
  • Home
  • Alerts
  • Feedback
  • Privacy Policy
Facebook  Twitter  LinkedIn  YouTube  RSS

Articles

  • Online First
  • Current Issue
  • Past Issues
  • Meeting Abstracts

Info for

  • Authors
  • Subscribers
  • Advertisers
  • Librarians

About MCT

  • About the Journal
  • Editorial Board
  • Permissions
  • Submit a Manuscript
AACR logo

Copyright © 2021 by the American Association for Cancer Research.

Molecular Cancer Therapeutics
eISSN: 1538-8514
ISSN: 1535-7163

Advertisement