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Biochimica Et Biophysica Acta Apr 2014Observations that Glioma-associated transcription factors Gli1 and Gli2 (Gli1/2), executers of the Sonic Hedgehog (Shh) signaling pathway and targets of the Transforming...
Observations that Glioma-associated transcription factors Gli1 and Gli2 (Gli1/2), executers of the Sonic Hedgehog (Shh) signaling pathway and targets of the Transforming Growth Factor β (TGF-β) signaling axis, are involved in numerous developmental and pathological processes unveil them as attractive pharmaceutical targets. Unc-51-like serine/threonine kinase Ulk3 has been suggested to play kinase activity dependent and independent roles in the control of Gli proteins in the context of the Shh signaling pathway. This study aimed at investigating whether the mechanism of generation of Gli1/2 transcriptional activators has similarities regardless of the signaling cascade evoking their activation. We also elucidate further the role of Ulk3 kinase in regulation of Gli1/2 proteins and examine SU6668 as an inhibitor of Ulk3 catalytic activity and a compound targeting Gli1/2 proteins in different cell-based experimental models. Here we demonstrate that Ulk3 is required not only for maintenance of basal levels of Gli1/2 proteins but also for TGF-β or Shh dependent activation of endogenous Gli1/2 proteins in human adipose tissue derived multipotent stromal cells (ASCs) and mouse immortalized progenitor cells, respectively. We show that cultured ASCs possess the functional Shh signaling axis and differentiate towards osteoblasts in response to Shh. Also, we demonstrate that similarly to Ulk3 RNAi, SU6668 prevents de novo expression of Gli1/2 proteins and antagonizes the Gli-dependent activation of the gene expression programs induced by either Shh or TGF-β. Our data suggest SU6668 as an efficient inhibitor of Ulk3 kinase allowing manipulation of the Gli-dependent transcriptional outcome.
Topics: Animals; Cell Differentiation; Cells, Cultured; Gene Expression Regulation, Developmental; Hedgehog Proteins; Humans; Indoles; Kruppel-Like Transcription Factors; Leukocytes, Mononuclear; Mice; Multipotent Stem Cells; Neoplasms; Nuclear Proteins; Oxindoles; Propionates; Protein Serine-Threonine Kinases; Pyrroles; Signal Transduction; Transcription Factors; Transforming Growth Factor beta; Zinc Finger Protein GLI1; Zinc Finger Protein Gli2
PubMed: 24418624
DOI: 10.1016/j.bbamcr.2014.01.003 -
Chinese Clinical Oncology Dec 2013Approximately 32,000 patients die of primary liver cancer each year in Japan. The annual number of deaths from primary liver cancer in Japan ranks second only to that in...
Approximately 32,000 patients die of primary liver cancer each year in Japan. The annual number of deaths from primary liver cancer in Japan ranks second only to that in China in the world. In recent years, there has been a gradual trend towards decrease in the number of liver cancer patients from its peak in Japan, and this trend is expected to also continue in the future. The main reason for this decreasing trend was the establishment of screening of transfusion products for hepatitis B and hepatitis C viruses, which prevents transfusion-related transmission of the viral infection. Most patients with hepatocellular carcinoma (HCC) in Japan have underlying viral hepatitis, with hepatitis C accounting for about two-third of all the patients and hepatitis B accounting for about 15%. Regular screening of patients with viral hepatitis infection makes it possible to diagnose HCC early, and also enables effective loco-regional treatment, such as surgical resection, local ablative therapy and transcatheter arterial chemoembolization (TACE). However, HCC recurrence is encountered frequently even after these potentially effective treatments. After numerous loco-regional treatments for recurrent HCC, chemotherapy is administered for patients with highly advanced HCC. Among the modalities of chemotherapy, hepatic arterial infusion chemotherapy (HAIC) is employed more commonly than systemic chemotherapy, although no survival advantage has ever been demonstrated. Randomized controlled studies are currently under way to clarify the survival benefit of HAIC. Also, various novel systemic chemotherapeutic agents are currently under development in Japan, and further improvements in the treatment outcomes are expected.
PubMed: 25841919
DOI: 10.3978/j.issn.2304-3865.2013.09.01 -
Journal of Proteomics Jan 2014Recent advances in mass spectrometry-based chemical proteomics allow unbiased analysis of drug-target interactions under close to physiological conditions. In this...
UNLABELLED
Recent advances in mass spectrometry-based chemical proteomics allow unbiased analysis of drug-target interactions under close to physiological conditions. In this study, we designed and synthesized two small molecule probes targeting fibroblast growth factor receptors (FGFRs) and applied them to evaluate the selectivity profiles of the FGFR inhibitors Dovitinib and Orantinib. Probe F2 was capable of enriching all members of the FGF receptor family as well as other kinases involved in cancer such as KDR, FLT4 and RET from lysates of cancer cells or human placenta tissue. In combination with the established Kinobeads™ approach, probe F2 facilitated the identification of the target spectrum of the two inhibitors confirming many of the previously identified (off-) targets such as AURKA, FLT4-VEGFR3, IKBKE and PDGFRβ. The newly synthesized probe enlarges the arsenal of chemical proteomic tools for the expression profiling of kinases and selectivity profiling of their inhibitors. It will also be useful in applications aiming at a better understanding of a drug's cellular mechanisms of action as well as highlighting potential beneficial or adverse side effects.
BIOLOGICAL SIGNIFICANCE
The synthesis of a new chemical affinity probe targeting FGF-receptors and many other kinases improved the general scope of drug selectivity profiling by chemical proteomics. The application of the developed chemical tool identified most of the known targets for the advanced clinical kinase inhibitors Dovitinib and Orantinib thus exemplify the practical utility of the developed probe and the results obtained shed further light on how these drugs exert their anti-cancer activity in cells. More generally speaking, the significance of the work is that the molecular tools presented here extend the application scope of kinobeads based kinase profiling to FGFR/VEGFR/PDGFR families, which thus may be generically employed for selectivity profiling of kinase inhibitors using chemical proteomics. The overall aim of such studies is to improve our understanding of how target as well as off-target profiles can be used to assess or predict the therapeutic efficacy of a drug.
Topics: Female; Gene Expression Profiling; Humans; K562 Cells; Neoplasm Proteins; Neoplasms; Placenta; Pregnancy; Pregnancy Proteins; Proteomics; Receptors, Fibroblast Growth Factor
PubMed: 24184958
DOI: 10.1016/j.jprot.2013.10.031 -
British Journal of Cancer Oct 2013This study aimed to determine whether combination S-1 plus cisplatin (CDDP) therapy, the most widely used therapy for Japanese patients with advanced gastric cancer, and... (Randomized Controlled Trial)
Randomized Controlled Trial
BACKGROUND
This study aimed to determine whether combination S-1 plus cisplatin (CDDP) therapy, the most widely used therapy for Japanese patients with advanced gastric cancer, and the novel oral antiangiogenic agent TSU-68 could contribute to gastric cancer treatment.
METHODS
Ninety-three patients with chemotherapy-naïve unresectable or recurrent advanced gastric cancers were randomised into two groups: TSU-68 plus S-1/CDDP (group A) and S-1/CDDP (group B) groups. Both patient groups received identical S-1 and CDDP dosages. TSU-68 was orally administered for 35 consecutive days. Group B patients received S-1 orally twice daily for three consecutive weeks, followed by intravenous CDDP on day 8. The primary endpoint was progression-free survival (PFS).
RESULTS
Median PFS periods were 208 and 213 days in groups A and B, respectively (P=0.427). Median survival periods for groups A and B were 497.0 and 463.5 days, respectively (P=0.219). No statistically significant differences were noted for PFS, survival or the adverse event (AE) incidence rate. All AEs were expected according to previous reports for TSU-68, TS-1, and CDDP.
CONCLUSION
Combination therapy involving TSU-68, S-1, and CDDP was safe and well tolerated in patients with chemotherapy-naïve unresectable or recurrent advanced gastric cancers. However, factors related to therapeutic efficacy should be investigated further.
Topics: Aged; Angiogenesis Inhibitors; Antineoplastic Combined Chemotherapy Protocols; Cisplatin; Disease-Free Survival; Drug Combinations; Female; Humans; Indoles; Male; Middle Aged; Oxindoles; Oxonic Acid; Propionates; Pyrroles; Stomach Neoplasms; Survival Rate; Tegafur
PubMed: 24045669
DOI: 10.1038/bjc.2013.555 -
European Journal of Cancer (Oxford,... Sep 2013TSU-68 is an antitumour drug that acts by inhibiting angiogenesis. We evaluated the efficacy and safety of TSU-68 in combination with transarterial chemoembolisation... (Randomized Controlled Trial)
Randomized Controlled Trial
BACKGROUND
TSU-68 is an antitumour drug that acts by inhibiting angiogenesis. We evaluated the efficacy and safety of TSU-68 in combination with transarterial chemoembolisation (TACE) in patients with intermediate-stage hepatocellular carcinoma (HCC).
PATIENTS AND METHODS
In this multicenter, open-label phase II study, we randomised patients with HCC who had been treated with a single session of TACE to receive either 200mg TSU-68 twice daily or no medication. The primary end-point was progression-free survival (PFS).
RESULTS
A total of 103 patients were enrolled. Median PFS was 157.0days (95% confidence interval [CI], 124.0-230.0days) in the TSU-68 group and 122.0days (95% CI, 73.0-170.0days) in the control group. The hazard ratio was 0.699 (95% CI, 0.450-1.088). Fatigue, elevated aspartate aminotransferase (AST), elevated alkaline phosphatase, oedema and anorexia were more frequent in the TSU-68 group than in the control group. The most frequent grade 3/4 adverse events were AST elevation (46% of patients in the TSU-68 group and 12% of controls) and alanine aminotransferase elevation (26% of patients in the TSU-68 group and 8% of controls). Two deaths, grade 5 hepatic failure and melena were noted in the TSU-68 group.
CONCLUSION
This exploratory study shows a trend towards prolonged PFS with TSU-68 treatment after a single session of TACE, but this observation was not statistically significant. The two deaths were related to the study treatment. These results suggest that further examination of the study design is necessary to determine whether TSU-68 has any clinical benefits when combined with TACE.
Topics: Aged; Angiogenesis Inhibitors; Carcinoma, Hepatocellular; Chemoembolization, Therapeutic; Disease-Free Survival; Female; Humans; Indoles; Japan; Kaplan-Meier Estimate; Liver Neoplasms; Male; Middle Aged; Oxindoles; Propionates; Proportional Hazards Models; Pyrroles; Time Factors; Treatment Outcome
PubMed: 23764238
DOI: 10.1016/j.ejca.2013.05.011 -
Structure (London, England : 1993) Jul 2013Tank-binding kinase 1 (TBK1) is a serine/threonine protein-kinase mediating innate antimicrobial immunity. TBK1 is involved in the signaling of TLRs, RLRs, and...
Tank-binding kinase 1 (TBK1) is a serine/threonine protein-kinase mediating innate antimicrobial immunity. TBK1 is involved in the signaling of TLRs, RLRs, and STING-mediated sensing of cytosolic DNA. Stimulation of these receptors results in the activation of TBK1, which phosphorylates interferon regulatory factor (IRF)-3. Phosphorylated IRF-3 translocates into the nucleus to initiate the transcription of the interferon (IFN)-β gene. Here, we show that TBK1 is activated by autophosphorylation at residue Ser172. Structures of TBK1 bound to two inhibitors showed that TBK1 has the IκB kinase fold with three distinct domains: the kinase domain, the ubiquitin-like domain, and the scaffold and dimerization domain. However, the overall structures of the TBK1 monomer and its dimer are different from IKKβ in the arrangements of the three domains and in dimer formation. Phosphorylation of IRF-3 by TBK1 in vitro results in its oligomerization, and phosphorylation of residue Ser386 plays a key role in IRF-3 activation.
Topics: Amino Acid Sequence; Animals; Bacterial Infections; Catalytic Domain; Crystallography, X-Ray; Enzyme Activation; Humans; Hydrogen Bonding; Hydrophobic and Hydrophilic Interactions; Immunity, Innate; Indoles; Interferon Regulatory Factor-3; Mice; Models, Molecular; Molecular Sequence Data; Oxindoles; Phosphorylation; Propionates; Protein Binding; Protein Interaction Domains and Motifs; Protein Kinase Inhibitors; Protein Multimerization; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Protein Structure, Quaternary; Protein Structure, Secondary; Pyrroles; Structural Homology, Protein
PubMed: 23746807
DOI: 10.1016/j.str.2013.04.025 -
Molecular & Cellular Proteomics : MCP Sep 2012The most common mutation in cystic fibrosis (CF) is a deletion of Phe at position 508 (ΔF508-CFTR). ΔF508-CFTR is a trafficking mutant that is retained in the ER,...
The most common mutation in cystic fibrosis (CF) is a deletion of Phe at position 508 (ΔF508-CFTR). ΔF508-CFTR is a trafficking mutant that is retained in the ER, unable to reach the plasma membrane. To identify compounds and drugs that rescue this trafficking defect, we screened a kinase inhibitor library enriched for small molecules already in the clinic or in clinical trials for the treatment of cancer and inflammation, using our recently developed high-content screen technology (Trzcinska-Daneluti et al. Mol. Cell. Proteomics 8:780, 2009). The top hits of the screen were further validated by (1) biochemical analysis to demonstrate the presence of mature (Band C) ΔF508-CFTR, (2) flow cytometry to reveal the presence of ΔF508-CFTR at the cell surface, (3) short-circuit current (Isc) analysis in Ussing chambers to show restoration of function of the rescued ΔF508-CFTR in epithelial MDCK cells stably expressing this mutant (including EC(50) determinations), and importantly (4) Isc analysis of Human Bronchial Epithelial (HBE) cells harvested from homozygote ΔF508-CFTR transplant patients. Interestingly, several inhibitors of receptor Tyr kinases (RTKs), such as SU5402 and SU6668 (which target FGFRs, VEGFR, and PDGFR) exhibited strong rescue of ΔF508-CFTR, as did several inhibitors of the Ras/Raf/MEK/ERK or p38 pathways (e.g. (5Z)-7-oxozeaenol). Prominent rescue was also observed by inhibitors of GSK-3β (e.g. GSK-3β Inhibitor II and Kenpaullone). These results identify several kinase inhibitors that can rescue ΔF508-CFTR to various degrees, and suggest that use of compounds or drugs already in the clinic or in clinical trials for other diseases can expedite delivery of treatment for CF patients.
Topics: Animals; Benzazepines; Cell Line; Cricetinae; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Dogs; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; HEK293 Cells; Humans; Indoles; Ion Transport; Membrane Proteins; Oxindoles; Propionates; Protein Kinase Inhibitors; Protein Transport; Protein-Tyrosine Kinases; Pyrroles; RNA Interference; RNA, Small Interfering; Receptor, Fibroblast Growth Factor, Type 1; Receptors, Platelet-Derived Growth Factor; Receptors, Vascular Endothelial Growth Factor; Sequence Deletion; Signal Transduction; Zearalenone
PubMed: 22700489
DOI: 10.1074/mcp.M111.016626 -
International Journal of Clinical... Aug 2013TSU-68 is a novel multiple tyrosine kinase inhibitor that inhibits vascular endothelial growth factor receptor-2, platelet-derived growth factor receptor and fibroblast...
A multicenter phase II study of TSU-68, a novel oral multiple tyrosine kinase inhibitor, in patients with metastatic breast cancer progressing despite prior treatment with an anthracycline-containing regimen and taxane.
PURPOSE
TSU-68 is a novel multiple tyrosine kinase inhibitor that inhibits vascular endothelial growth factor receptor-2, platelet-derived growth factor receptor and fibroblast growth factor receptor. TSU-68 demonstrated a strong anti-tumor effect against established human breast cancer xenografts in nude mice without apparent toxicity. We conducted a phase II study to evaluate the efficacy and safety of TSU-68 monotherapy in patients with metastatic breast cancer progressing despite prior treatment with an anthracycline-containing regimen and taxane.
METHODS
TSU-68 was administered daily at a dose of 400 mg twice a day after meals in 20 patients. The primary endpoint was objective overall response rate according to the Response Evaluation Criteria in Solid Tumors guideline version 1.0. Secondary endpoints included clinical benefit rate (complete response, partial response and stable disease lasting for at least 24 weeks), exploratory assessments of change in mRNA levels of biological markers associated with angiogenesis in tumor tissue at the end of Cycle 1, and safety of TSU-68.
RESULTS
Twenty patients were enrolled into the study from October 2002 through April 2003. TSU-68 monotherapy produced objective overall response in none of the patients; however, clinical benefit was seen in 5 % of the patients. The mRNA levels of CD31, Flt-1 and Flk-1/KDR showed a decreasing trend in all 4 patients who provided additional written informed consent for collection of tumor tissue. However, no significant difference was observed in the change in mRNA level due to the small sample size. The most common adverse drug reaction (ADR) was tumor pain (60 %); hematological ADRs rarely occurred, and they were mild in severity. Only one patient experienced grade 2 rash and no patient experienced hypertension. No patients experienced a grade 4 ADR and no episode of death related to the study treatment occurred in the 20 patients.
CONCLUSIONS
TSU-68 monotherapy produced clinical benefit in only 5 % of the patients and did not produce objective overall response; however, the treatment was well tolerated. Further evaluation of the efficacy of TSU-68 will be worthwhile because the mRNA levels of CD31, Flt-1 and Flk-1/KDR decreased in 4 patients.
Topics: Adult; Aged; Anthracyclines; Antineoplastic Combined Chemotherapy Protocols; Biomarkers, Tumor; Breast Neoplasms; Bridged-Ring Compounds; Drug Administration Schedule; Female; Gene Expression Regulation, Neoplastic; Humans; Indoles; Middle Aged; Neovascularization, Pathologic; Oxindoles; Propionates; Protein Kinase Inhibitors; Pyrroles; Taxoids; Treatment Outcome; Vascular Endothelial Growth Factor A
PubMed: 22585426
DOI: 10.1007/s10147-012-0421-9 -
Respirology (Carlton, Vic.) Aug 2012Malignant pleural mesothelioma (MPM) is an aggressive neoplasm of the mesothelium with high chemotherapeutic resistance. In this study, the preclinical therapeutic...
BACKGROUND AND OBJECTIVE
Malignant pleural mesothelioma (MPM) is an aggressive neoplasm of the mesothelium with high chemotherapeutic resistance. In this study, the preclinical therapeutic activity of the multiple tyrosine kinase inhibitor, SU6668, against MPM was examined.
METHODS
Two human MPM cell lines with different pro-angiogenic cytokine expression, Y-MESO-14 cells that express high levels of vascular endothelial growth factor (VEGF) and MSTO-211H cells that express high levels of basic fibroblast growth factor (bFGF), were orthotopically inoculated into the thoracic cavities of mice with severe combined immunodeficiency. The mice with MPM were treated or not treated with SU6668 (200 mg/kg/day).
RESULTS
SU6668 abrogated the proliferation of endothelial cells stimulated by VEGF or bFGF, but did not directly affect the growth of human MPM cells in vitro. In this orthotopic implantation model, treatment with SU6668 effectively reduced tumour weight and pleural effusion volumes, in association with inhibition of the growth of tumour vasculature. More importantly, treatment with SU6668 significantly prolonged survival time in mice with MPM.
CONCLUSIONS
These findings suggest that SU6668 has a promising therapeutic effect on the progression of MPM in vivo through its anti-angiogenic effects.
Topics: Animals; Antineoplastic Agents; Cell Line, Tumor; Cell Proliferation; Fibroblast Growth Factor 2; Humans; Indoles; Male; Mesothelioma; Mice; Mice, SCID; Neovascularization, Pathologic; Oxindoles; Pleural Effusion, Malignant; Pleural Neoplasms; Propionates; Protein Kinase Inhibitors; Pyrroles; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays
PubMed: 22574723
DOI: 10.1111/j.1440-1843.2012.02193.x -
Breast Cancer (Tokyo, Japan) Jan 2014TSU-68 is a novel multiple tyrosine kinase inhibitor that inhibits vascular endothelial growth factor receptor-2, platelet-derived growth factor receptor, and fibroblast...
A multicenter phase II study of TSU-68, an oral multiple tyrosine kinase inhibitor, in combination with docetaxel in metastatic breast cancer patients with anthracycline resistance.
BACKGROUND
TSU-68 is a novel multiple tyrosine kinase inhibitor that inhibits vascular endothelial growth factor receptor-2, platelet-derived growth factor receptor, and fibroblast growth factor receptor. This open-label, non-comparative, multicenter phase II study evaluated TSU-68 in combination with docetaxel in patients with metastatic breast cancer that had relapsed within 1 year despite prior treatment with an anthracycline-containing regimen.
METHODS
TSU-68 was orally administered on days 1-21, and docetaxel was intravenously delivered on day 1. The regimen was repeated every 21 days. Primary endpoint was objective response rate according to the RECIST guidelines version 1.0.
RESULTS
TSU-68 in combination with docetaxel produced objective responses in 21.1% and clinical benefits in 42.1% of the patients, respectively (1 complete response, 3 partial response, and 4 stable disease for at least 24 weeks, n = 19). Median time to progression was 148 days, and median overall survival was 579 days. The common adverse drug reactions were leukopenia, neutropenia, nail disorder, malaise, dysgeusia, alopecia, and edema.
CONCLUSIONS
TSU-68 in combination with docetaxel showed a promising antitumor response with manageable toxicity in patients with anthracycline-resistant metastatic breast cancer. Further studies are warranted in a different population of breast cancer or other solid cancers.
Topics: Administration, Intravenous; Administration, Oral; Adult; Aged; Anthracyclines; Antineoplastic Combined Chemotherapy Protocols; Breast Neoplasms; Docetaxel; Drug Interactions; Drug Resistance, Neoplasm; Female; Humans; Indoles; Middle Aged; Oxindoles; Propionates; Protein Kinase Inhibitors; Pyrroles; Taxoids; Treatment Outcome; Vascular Endothelial Growth Factor A
PubMed: 22382811
DOI: 10.1007/s12282-012-0344-3