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Scientific Reports Aug 2017In recent years genome-wide RNAi screens have revealed hundreds of cellular factors required for influenza virus infections in human cells. The long-term goal is to...
In recent years genome-wide RNAi screens have revealed hundreds of cellular factors required for influenza virus infections in human cells. The long-term goal is to establish some of them as drug targets for the development of the next generation of antivirals against influenza. We found that several members of the polo-like kinases (PLK), a family of serine/threonine kinases with well-known roles in cell cycle regulation, were identified as hits in four different RNAi screens and we therefore studied their potential as drug target for influenza. We show that knockdown of PLK1, PLK3, and PLK4, as well as inhibition of PLK kinase activity by four different compounds, leads to reduced influenza virus replication, and we map the requirement of PLK activity to early stages of the viral replication cycle. We also tested the impact of the PLK inhibitor BI2536 on influenza virus replication in a human lung tissue culture model and observed strong inhibition of virus replication with no measurable toxicity. This study establishes the PLKs as potential drug targets for influenza and contributes to a more detailed understanding of the intricate interactions between influenza viruses and their host cells.
Topics: A549 Cells; Animals; Antimitotic Agents; Cell Cycle Proteins; Dogs; Glycine; HEK293 Cells; Humans; Influenza A virus; Influenza, Human; Madin Darby Canine Kidney Cells; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Pteridines; RNA Interference; Sulfones; Tumor Suppressor Proteins; Virus Replication; Polo-Like Kinase 1
PubMed: 28819179
DOI: 10.1038/s41598-017-08942-7 -
Cell Apr 2016Oncogenic activation of RAS genes via point mutations occurs in 20%-30% of human cancers. The development of effective RAS inhibitors has been challenging, necessitating...
Oncogenic activation of RAS genes via point mutations occurs in 20%-30% of human cancers. The development of effective RAS inhibitors has been challenging, necessitating new approaches to inhibit this oncogenic protein. Functional studies have shown that the switch region of RAS interacts with a large number of effector proteins containing a common RAS-binding domain (RBD). Because RBD-mediated interactions are essential for RAS signaling, blocking RBD association with small molecules constitutes an attractive therapeutic approach. Here, we present evidence that rigosertib, a styryl-benzyl sulfone, acts as a RAS-mimetic and interacts with the RBDs of RAF kinases, resulting in their inability to bind to RAS, disruption of RAF activation, and inhibition of the RAS-RAF-MEK pathway. We also find that ribosertib binds to the RBDs of Ral-GDS and PI3Ks. These results suggest that targeting of RBDs across multiple signaling pathways by rigosertib may represent an effective strategy for inactivation of RAS signaling.
Topics: Amino Acid Sequence; Animals; Cell Cycle Proteins; Cell Transformation, Neoplastic; Crystallography, X-Ray; Dimerization; Glycine; Humans; MAP Kinase Signaling System; Mice; Mice, Nude; Models, Molecular; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Pancreatic Neoplasms; Phosphorylation; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins B-raf; RNA-Binding Proteins; Sequence Alignment; Signal Transduction; Sulfones; ras Proteins; Polo-Like Kinase 1
PubMed: 27104980
DOI: 10.1016/j.cell.2016.03.045 -
Leukemia Sep 2013
Topics: Aged; Antineoplastic Agents; Enzyme Inhibitors; Female; Glycine; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Lymphoma, B-Cell; Male; Middle Aged; Phosphoinositide-3 Kinase Inhibitors; Recurrence; Sulfones; Treatment Outcome
PubMed: 23486532
DOI: 10.1038/leu.2013.79 -
Journal of Clinical Oncology : Official... Dec 2008We conducted a first-in-man (to our knowledge) phase I study to determine the dose-limiting toxicities (DLTs), characterize the pharmacokinetic profile, and document any...
PURPOSE
We conducted a first-in-man (to our knowledge) phase I study to determine the dose-limiting toxicities (DLTs), characterize the pharmacokinetic profile, and document any antitumor activity of ON 01910.Na, a new chemical entity that arrests cancer cells in G(2)/M by modulating mitotic regulatory pathways including polo-like kinase 1 (Plk1).
PATIENTS AND METHODS
Patients had solid tumors refractory to standard therapy. ON 01910.Na was administered as a 2-hour infusion on days 1, 4, 8, 11, 15, and 18 in 28-day cycles. The starting dose was 80 mg, and an accelerated titration schedule (single-patient cohorts) was used for escalation. Pharmacokinetics were studied on days 1 and 15 of cycle 1.
RESULTS
Twenty patients (11 women and nine men; age 46 to 73 years) were enrolled onto the study. Dose levels of 80, 160, 320, 480, 800, 1,280, 2,080, and 3,120 mg were evaluated in single-patient cohorts. A DLT and additional grade 2 toxicities made the 4,370-mg dose (n = 6) not tolerable, and the next lower dose cohort (3,120 mg) was expanded to six assessable patients. Toxicities were skeletal, abdominal, and tumor pain; nausea; urge to defecate; and fatigue. Hematologic toxicity was infrequent and mild. ON 01910.Na pharmacokinetics were characterized by a rapid distribution phase (distribution half-life, 1 hour) and a relatively slow elimination phase (elimination half-life, 27 hours). A refractory ovarian cancer patient had an objective response after four cycles and remained progression free for 24 months.
CONCLUSION
ON 01910.Na showed a distinct but moderate toxicity pattern. The recommended phase II dose of ON 01910.Na with this schedule of administration is 3,120 mg. Single-agent activity was documented in an ovarian cancer patient.
Topics: Aged; Antineoplastic Agents; Cell Cycle Proteins; Cell Division; Dose-Response Relationship, Drug; Female; G2 Phase; Gene Expression Regulation, Neoplastic; Glycine; Humans; Male; Middle Aged; Neoplasms; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Sulfones; Time Factors; Treatment Outcome; Polo-Like Kinase 1
PubMed: 18955447
DOI: 10.1200/JCO.2008.17.9788 -
Biomolecules Jan 2023Tubulin is a protein that plays a critical role in maintaining cellular structure and facilitating cell division. Inhibiting tubulin polymerization has been shown to be...
Tubulin is a protein that plays a critical role in maintaining cellular structure and facilitating cell division. Inhibiting tubulin polymerization has been shown to be an effective strategy for inhibiting the proliferation of cancer cells. In the past, identifying compounds that could inhibit tubulin polymerization has required the use of in vitro assays utilizing purified tubulin or immunofluorescence of fixed cells. This study presents a novel approach for identifying tubulin polymerization inhibitors using a CRISPR-edited cell line that expresses fluorescently tagged β-tubulin and a nuclear protein, enabling the visualization of tubulin polymerization dynamics via high-content imaging analysis (HCI). The cells were treated with known tubulin polymerization inhibitors, colchicine, and vincristine, and the resulting phenotypic changes indicative of tubulin polymerization inhibition were confirmed using HCI. Furthermore, a library of 429 kinase inhibitors was screened, resulting in the identification of three compounds (ON-01910, HMN-214, and KX2-391) that inhibit tubulin polymerization. Live cell tracking analysis confirmed that compound treatment leads to rapid tubulin depolymerization. These findings suggest that CRISPR-edited cells with fluorescently tagged endogenous β-tubulin can be utilized to screen large compound libraries containing diverse chemical families for the identification of novel tubulin polymerization inhibitors.
Topics: Humans; Tubulin; Tubulin Modulators; Histones; Polymerization; Clustered Regularly Interspaced Short Palindromic Repeats; Cell Line; Antineoplastic Agents; Cell Proliferation; Cell Line, Tumor; Molecular Structure
PubMed: 36830618
DOI: 10.3390/biom13020249 -
PloS One 2013Lymphoma-specific biomarkers contribute to therapeutic strategies and the study of tumorigenesis. Diffuse large B-cell lymphoma (DLBCL) is the most common type of...
Lymphoma-specific biomarkers contribute to therapeutic strategies and the study of tumorigenesis. Diffuse large B-cell lymphoma (DLBCL) is the most common type of malignant lymphoma. However, only 50% of patients experience long-term survival after current treatment; therefore, developing novel therapeutic strategies is warranted. Comparative proteomic analysis of two DLBCL lines with a B-lymphoblastoid cell line (LCL) showed differential expression of Ran GTPase-activating protein 1 (RanGAP1) between them, which was confirmed using immunoblotting. Immunostaining showed that the majority of DLBCLs (92%, 46/50) were RanGAP1(+), while reactive lymphoid hyperplasia (n = 12) was RanGAP1(+) predominantly in germinal centers. RanGAP1 was also highly expressed in other B-cell lymphomas (BCL, n = 180) with brisk mitotic activity (B-lymphoblastic lymphoma/leukemia: 93%, and Burkitt lymphoma: 95%) or cell-cycle dysregulation (mantle cell lymphoma: 83%, and Hodgkin's lymphoma 91%). Interestingly, serum RanGAP1 level was higher in patients with high-grade BCL (1.71 ± 2.28 ng/mL, n = 62) than in low-grade BCL (0.75 ± 2.12 ng/mL, n = 52) and healthy controls (0.55 ± 1.58 ng/mL, n = 75) (high-grade BCL vs. low-grade BCL, p = 0.002; high-grade BCL vs. control, p < 0.001, Mann-Whitney U test). In vitro, RNA interference of RanGAP1 showed no effect on LCL but enhanced DLBCL cell death (41% vs. 60%; p = 0.035) and cell-cycle arrest (G0/G1: 39% vs. 49%, G2/M: 19.0% vs. 7.5%; p = 0.030) along with decreased expression of TPX2 and Aurora kinases, the central regulators of mitotic cell division. Furthermore, ON 01910.Na (Estybon), a multikinase inhibitor induced cell death, mitotic cell arrest, and hyperphosphorylation of RanGAP1 in DLBCL cell lines but no effects in normal B and T cells. Therefore, RanGAP1 is a promising marker and therapeutic target for aggressive B-cell lymphoma, especially DLBCL.
Topics: Cell Cycle Checkpoints; Cell Line, Tumor; Enzyme Inhibitors; GTPase-Activating Proteins; Humans; Lymphoma, B-Cell; Phosphorylation; RNA Interference
PubMed: 24223200
DOI: 10.1371/journal.pone.0079863 -
Laboratory Investigation; a Journal of... Oct 2012Polo-like kinase 1 (Plk1) is a mitotic serine/threonine kinase and its kinase activity is closely interrelated to cell cycle progression, various types of cancer...
Polo-like kinase 1 (Plk1) is a mitotic serine/threonine kinase and its kinase activity is closely interrelated to cell cycle progression, various types of cancer development and often correlates with poor prognosis. Thus, it is of prime importance to characterize the phenotypes of Plk1 inhibition in cells for drug development and clinical application. Here, we report a novel kinase inhibitory chemical, CBB2001, which specifically inhibited Plk1 kinase activity in vitro with an IC(50) of 0.39 μM. In cervical carcinoma HeLa cells, we found that treatment of CBB2001 caused mitotic cell cycle arrest (EC(50)=0.72 μM) and induction of 'polo' cells (EC(50)=0.32 μM). Interestingly, the cell cycle arrest induced by CBB2001 was associated with accumulation of Plk1 (EC(50)=0.61 μM) and Geminin (EC(50)=0.43 μM) proteins, but distinct from the phenotypes induced by Aurora kinase inhibitors. The inhibitory effects of CBB2001 were phenocopied by RNA interferences of Plk1. We also confirmed the cell cycle inhibitory effects of CBB2001 in other cancer cells. Moreover, CBB2001 inhibited the growth of HeLa cells with an IC(50) of 0.85 μM in MTT assays, which is better than that of reported Plk1 inhibitory chemicals ON01910 (IC(50)=6.46 μM) and LFM-A13 (IC(50)=37.36 μM). CBB2001 also inhibited mouse xenograft tumor growth. Furthermore, CBB2001 inhibited mitotic exit and delayed degradation of APC/C substrates, Geminin, Cyclin B1 and Aurora A. These specific phenotypes may serve as specific features for Plk1 inhibition and for Plk1-based clinic trials.
Topics: Amides; Animals; Aurora Kinase A; Aurora Kinases; Benzimidazoles; Cell Cycle Proteins; Cell Division; Cell Line, Tumor; Cell Proliferation; Cyclin B1; Female; Geminin; Glycine; Humans; Inhibitory Concentration 50; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasms; Nitriles; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Sulfones; Thiazoles; Polo-Like Kinase 1
PubMed: 22890557
DOI: 10.1038/labinvest.2012.114 -
Leukemia Research Aug 2012We previously demonstrated upregulation of c-myc, survivin, and cyclin D1 in CD34+ bone marrow mononuclear cells (BMMNCs) of patients with trisomy 8 and monosomy 7...
BACKGROUND
We previously demonstrated upregulation of c-myc, survivin, and cyclin D1 in CD34+ bone marrow mononuclear cells (BMMNCs) of patients with trisomy 8 and monosomy 7 myelodysplastic syndromes (MDS). "Knockdown" of cyclin D1 by RNA interference decreased trisomy 8 cell growth, suggesting that this might be a therapeutic target in MDS.
EXPERIMENTAL DESIGN
We performed preclinical studies using BMMNCs from patients with MDS and AML to examine the effects of the styryl sulfone ON 01910.Na on cyclin D1 accumulation, aneuploidy, and CD34+ blast percentage. We next treated twelve patients with higher risk MDS and two trisomy 8 AML patients with ON 01910.Na on a phase I clinical protocol (NCT00533416).
RESULTS
ON 01910.Na inhibited cyclin D1 expression, and was selectively toxic to trisomy 8 cells in vitro. Flow cytometry studies demonstrated increased mature CD15+ myeloid cells and decreased CD34+ blasts. Three patients treated with ON 01910.Na on a clinical had decreased bone marrow blasts by ≥ 50%, and three patients had hematologic improvements, one of which was sustained for 33 months. Patients with hematologic responses to ON 01910.Na had decreased cyclin D1 expression in their CD34+ cells.
CONCLUSIONS
The preclinical results and responses of patients on a clinical trial warrant further investigation of ON 01910.Na as a potential novel targeted therapy for higher risk MDS patients.
Topics: Aged; Aged, 80 and over; Algorithms; Antineoplastic Agents; Bone Marrow Cells; Chromosomes, Human, Pair 8; Cyclin D1; Dose-Response Relationship, Drug; Female; Glycine; Humans; Male; Middle Aged; Molecular Targeted Therapy; Myelodysplastic Syndromes; Sulfones; Trisomy; Tumor Cells, Cultured
PubMed: 22524974
DOI: 10.1016/j.leukres.2012.04.002 -
Blood Cancer Journal Dec 2017
Randomized Controlled Trial
Validation of a post-hypomethylating agent failure prognostic model in myelodysplastic syndromes patients treated in a randomized controlled phase III trial of rigosertib vs. best supportive care.
Topics: Antineoplastic Agents; Azacitidine; Decitabine; Drug Resistance, Neoplasm; Glycine; Humans; Kaplan-Meier Estimate; Myelodysplastic Syndromes; Prognosis; Sulfones; Treatment Outcome
PubMed: 29238044
DOI: 10.1038/s41408-017-0018-7 -
Cell Cycle (Georgetown, Tex.) Jun 2017For almost a decade, there has been much interest in the development of chemical inhibitors of Polo-like kinase 1 (Plk1) protein interactions. Plk1 is a master regulator...
For almost a decade, there has been much interest in the development of chemical inhibitors of Polo-like kinase 1 (Plk1) protein interactions. Plk1 is a master regulator of the cell division cycle that controls numerous substrates. It is a promising target for cancer drug development. Inhibitors of the kinase domain of Plk1 had some success in clinical trials. However, they are not perfectly selective. In principle, Plk1 can also be inhibited by interfering with its protein interaction domain, the Polo-Box Domain (PBD). Selective chemical inhibitors of the PBD would constitute tools to probe for PBD-dependent functions of Plk1 and could be advantageous in cancer therapy. The discovery of Poloxin and thymoquinone as PBD inhibitors indicated that small, cell-permeable chemical inhibitors could be identified. Other efforts followed, including ours, reporting additional molecules capable of blocking the PBD. It is now clear that, unfortunately, most of these compounds are non-specific protein alkylators (defined here as groups covalently added via a carbon) that have little or no potential for the development of real Plk1 PBD-specific drugs. This situation should be minded by biologists potentially interested in using these compounds to study Plk1. Further efforts are needed to develop selective, cell-permeable PBD inhibitors.
Topics: Alkylation; Antineoplastic Agents; Benzoates; Benzoquinones; Cell Cycle Proteins; Glycine; Humans; Mitosis; Neoplasms; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Quinones; Sulfones; Polo-Like Kinase 1
PubMed: 28521657
DOI: 10.1080/15384101.2017.1325043