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1.
Expert Opin Investig Drugs ; 29(1): 5-14, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31815551

RESUMO

Introduction: Kinesin spindle protein (KSP) is indispensable for the proper separation of spindle poles during mitosis. Importantly, this protein is expressed only in cells undergoing cell division and hence represents an appealing target for the treatment of cancer. Many KSP inhibitors have demonstrated a strong antitumoral effect in vitro, however, they have exhibited only limited activity in clinical trials. By contrast, the KSP inhibitor filanesib has demonstrated clinical efficacy in patients with multiple myeloma (MM).Areas covered: This article provides a comprehensive overview about the progress to date in the preclinical and clinical development of filanesib for the treatment of cancer, and particularly, MM.Expert opinion: Responses observed with filanesib alone or in combination with dexamethasone were encouraging in MM. However, the subsequent appearance of highly effective novel agents such as monoclonal antibodies, has hindered the development of agents such as filanesib that exhibit a more limited activity. Nevertheless, filanesib has shown interesting results for some patients when combined with carfilzomib and pomalidomide. Most importantly, the availability of a biomarker of response such as alpha 1-acid glycoprotein (AAG), could be key to the identification of patients that could benefit most from these combinations.


Assuntos
Antimitóticos/administração & dosagem , Mieloma Múltiplo/tratamento farmacológico , Tiadiazóis/administração & dosagem , Animais , Antimitóticos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica , Biomarcadores Tumorais/metabolismo , Desenvolvimento de Medicamentos , Humanos , Cinesina/antagonistas & inibidores , Mieloma Múltiplo/patologia , Tiadiazóis/farmacologia
2.
Comput Biol Chem ; 83: 107104, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31546212

RESUMO

In this paper, we have established methylenebis (4-hydroxy-2H-chromen-2-one) as a promising anticancer scaffold with kinesin spindle protein (KSP) inhibitory activity under malignant condition. A series of biscoumarin derivatives (MN1 to MN30) with different substituent were synthesized, and their anticancer activity was explored. Six biscoumarin derivatives that were found active were further selected to formulate organic nanoparticles (ONPs). Anticancer activity of both the forms (viz conventional and ONPs) was compared. MN30 was found most potent whereby MN10 showed good anticancer activity in both, i.e., conventional and ONP form; the structural activity relationship (SAR) study has been established. Computational investigation revealed biscoumarin scaffold as a suitable pharmacophore to bind against KSP protein. Molecular dynamics simulation studies revealed protein-ligand stability and dynamic behavior of biscoumarin-KSP complex. Finally, accruing signal transduction model was formulated to explain the observed MTT trend of conventional and ONP form. The model seems useful towards solving population specific varied results of chemotherapeutic agents. According to the model, MN10 and MN30 derivatives have good pharmacodynamics inertia and therefore, both the molecules were able to provide dose-dependent cytotoxic results.


Assuntos
Antineoplásicos/farmacocinética , Cromonas/farmacologia , Simulação por Computador , Transdução de Sinais/efeitos dos fármacos , Células A549 , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cromonas/síntese química , Cromonas/química , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Células HeLa , Humanos , Cinesina/antagonistas & inibidores , Cinesina/metabolismo , Modelos Moleculares , Relação Estrutura-Atividade , Células Tumorais Cultivadas
3.
Monoclon Antib Immunodiagn Immunother ; 38(4): 162-170, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31260385

RESUMO

Kinesin-like protein KIF20B, originally named M-phase phosphoprotein 1 (MPP1), is a plus-end-directed kinesin-related protein that exhibits in vitro microtubule-binding and -bundling properties as well as microtubule-stimulated ATPase activity. It has been characterized as a slow molecular motor that moves toward the plus-end of microtubules. Human autoantibodies directed against KIF20B have been described in up to 25% of patients with idiopathic ataxia and less commonly in other neuropathies and autoinflammatory conditions. One of the limitations of research into the structure and function of KIF20B has been a reliable monoclonal antibody that can be used in a variety of applications. To establish a reference standard for anti-KIF20B immunoassays and facilitate studies on the role of KIF20B in developmental cell biology, we developed an IgG1 monoclonal antibody, 10C7, which reacts with the cognate KIF20B protein in Western immunoblots and in addressable laser bead immunoassays. In HEp2 cells, leptomeningeal pericytes, and transfected HEK293T cells, indirect immunofluorescence studies showed that reactivity was mainly localized to a proportion of interphase nuclei, but during metaphase, it was redistributed throughout the cytoplasm and perichromatin mass. Later in telophase/anaphase, KIF20B was localized to the stem body and midzone of the midbody. 10C7 also showed remarkable staining of a subset of cells in the cerebellum, ovary, and testis tissues. KIF20B was shown to have extensive coiled-coil domains. The monoclonal antibody, 10C7, will be of value to diagnostic laboratory scientists interested in having a reliable reference standard for anti-KIF20B immunoassays as well as cell, molecular, and developmental biology researchers.


Assuntos
Anticorpos Monoclonais/imunologia , Mapeamento de Epitopos , Hibridomas/imunologia , Cinesina/antagonistas & inibidores , Cinesina/imunologia , Neoplasias Laríngeas/metabolismo , Proteínas Recombinantes/imunologia , Anticorpos Monoclonais/biossíntese , Anticorpos Monoclonais/farmacologia , Formação de Anticorpos , Humanos , Neoplasias Laríngeas/diagnóstico , Neoplasias Laríngeas/imunologia , Células Tumorais Cultivadas
4.
J Exp Clin Cancer Res ; 38(1): 188, 2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31072351

RESUMO

BACKGROUND: Forkhead box M1 (FOXM1) is a proliferation-associated transcription factor of the forkhead box proteins superfamily, which includes four isoforms FOXM1a, b, c, and d. FOXM1 has been implicated in hepatocellular carcinoma (HCC) progression, but the underlying molecular mechanism remains elusive. In this study, we aim to clarify the molecular basis for FOXM1-mediated HCC progression. METHODS: Bioinformatic analysis was used to explore the differentially expressed genes predicting HCC proliferation. The expression of FOXM1 and kinesin family member (KIF)4A was confirmed by western blotting and immunohistochemistry in HCC tissues. Kaplan-Meier survival analysis was conducted to analyze the clinical impact of FOXM1 and KIF4A on HCC. The effect of FOXM1 on the regulation of KIF4A expression was studied in cell biology experiments. The interaction between KIF4A and FOXM1 was analyzed by chromatin immunoprecipitation and luciferase experiments. A series of experiments was performed to explore the functions of FOXM1/KIF4A in HCC progression, such as cell proliferation, cell growth, cell viability, and cell cycle. A xenograft mouse model was used to explore the regulatory effect of FOXM1-KIF4A axis on HCC tumor growth. RESULTS: FOXM1 and KIF4A were overexpressed in human primary HCC tissues compared to that in matched adjacent normal liver tissue and are significant risk factors for HCC recurrence and shorter survival. We found that KIF4A was dominantly regulated by FOXM1c among the four isoforms, and further identified KIF4A as a direct downstream target of FOXM1c. Inhibiting FOXM1 decreased KIF4A expression in HCC cells, whereas its overexpression had the opposite effect. FOXM1-induced HCC cell proliferation was dependent on elevated KIF4A expression as KIF4A knockdown abolished FOXM1-induced proliferation of HCC cells both in vitro and in vivo. CONCLUSION: The FOXM1-KIF4A axis mediates human HCC progression and is a potential therapeutic target for HCC treatment.


Assuntos
Carcinoma Hepatocelular/genética , Proteína Forkhead Box M1/genética , Cinesina/genética , Neoplasias Hepáticas/genética , Adulto , Animais , Carcinoma Hepatocelular/patologia , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Estimativa de Kaplan-Meier , Cinesina/antagonistas & inibidores , Fígado/metabolismo , Fígado/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Transdução de Sinais/genética , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Chemistry ; 25(35): 8208-8213, 2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-30869180

RESUMO

Many antibody-drug conjugates (ADCs) have failed to achieve a sufficient therapeutic window in clinical studies either due to target-mediated or off-target toxicities. To achieve an additional safety level, a new class of antibody-prodrug conjugates (APDCs) directed against different targets in solid tumors is here described. The tumor-associated lysosomal endopeptidase legumain with a unique cleavage sequence was utilized for APDC metabolism. Legumain-activatable APDCs were as potent as their cathepsin B-activatable analogues. The peptide sequence susceptible to legumain cleavage was optimized for further discrimination of the formation of active metabolites within tumor cells versus healthy tissues, leveraging different tissue-specific legumain activities. Optimized APDCs with slow legumain-mediated conversion reduced preclinically the levels of active metabolite in healthy organs while retaining high activity against different TWEAKR- and B7H3-expressing tumors.


Assuntos
Anticorpos/química , Antineoplásicos/química , Cisteína Endopeptidases/metabolismo , Imunoconjugados/química , Cinesina/antagonistas & inibidores , Oligopeptídeos/química , Pró-Fármacos/química , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/metabolismo , Antígenos B7/genética , Antígenos B7/imunologia , Antígenos B7/metabolismo , Linhagem Celular Tumoral , Xenoenxertos , Humanos , Imunoconjugados/administração & dosagem , Imunoconjugados/metabolismo , Camundongos
6.
J Biochem ; 166(2): 129-137, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-30785183

RESUMO

Among 40 plant-derived biflavonoids with inhibitory potential against Eg5, morelloflavone from Garcinia dulcis leaves was selected for further testing based on in silico analysis of binding modes, molecular interactions, binding energies and functional groups that interact with Eg5. Computational models predicted that morelloflavone binds the putative allosteric pocket of Eg5, within the cavity surrounded by amino acid residues of Ile-136, Glu-116, Glu-118, Trp-127, Gly-117, Ala-133, Glu-215, Leu-214 and Tyr-211. Binding energy was -8.4 kcal/mol, with a single hydrogen bond formed between morelloflavone and Tyr-211. The binding configuration was comparable to that of a reference inhibitor, S-trityl-L-cysteine. Subsequent biochemical analysis in vitro confirmed that morelloflavone inhibited both the basal and microtubule-activated ATPase activity of Eg5 in a manner that does not compete with ATP binding. Morelloflavone also suppressed Eg5 gliding along microtubules. These results suggest that morelloflavone binds the allosteric binding site in Eg5 and thereby inhibits ATPase activity and motor function of Eg5.


Assuntos
Biflavonoides/farmacologia , Inibidores Enzimáticos/farmacologia , Cinesina/antagonistas & inibidores , Mitose , Biflavonoides/química , Inibidores Enzimáticos/química , Garcinia/química , Humanos , Cinesina/metabolismo , Modelos Moleculares , Estrutura Molecular , Folhas de Planta/química
7.
FASEB J ; 33(1): 388-399, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-29944446

RESUMO

Keratin intermediate filaments (IFs) are the major cytoskeletal component in epithelial cells. The dynamics of keratin IFs have been described to depend mostly on the actin cytoskeleton, but the rapid transport of fully polymerized keratin filaments has not been reported. In this work, we used a combination of photoconversion experiments and clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated protein 9 genome editing to study the role of microtubules and microtubule motors in keratin filament transport. We found that long keratin filaments, like other types of IFs, are transported along microtubules by kinesin-1. Our data revealed that keratin and vimentin are nonconventional kinesin-1 cargoes because their transport did not require kinesin light chains, which are a typical adapter for kinesin-dependent cargo transport. Furthermore, we found that the same domain of the kinesin heavy chain tail is involved in keratin and vimentin IF transport, strongly suggesting that multiple types of IFs move along microtubules using an identical mechanism.-Robert, A., Tian, P., Adam, S. A., Kittisopikul, M., Jaqaman, K., Goldman, R. D., Gelfand, V. I. Kinesin-dependent transport of keratin filaments: a unified mechanism for intermediate filament transport.


Assuntos
Filamentos Intermediários/metabolismo , Queratina-18/metabolismo , Queratina-8/metabolismo , Cinesina/fisiologia , Microtúbulos/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Vimentina/metabolismo , Citoesqueleto de Actina/metabolismo , Animais , Sistemas CRISPR-Cas , Células Cultivadas , Cinesina/antagonistas & inibidores , Camundongos , Camundongos Knockout , Microscopia de Fluorescência
8.
Oncogene ; 38(3): 406-420, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30115976

RESUMO

Hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide. The poor survival may be due to a high proportions of tumor recurrence and metastasis. Kinesin family member C1 (KIFC1) is highly expressed in a variety of neoplasms and is a potential marker for non-small cell lung cancer or ovarian adenocarcinoma metastasis. Nevertheless, the role of KIFC1 in HCC metastasis remains obscure. We investigated this in the present study using HCC cell lines and clinical specimens. Our results indicated that increased levels of KIFC1 were associated with poor prognosis and metastasis in HCC. In addition, KIFC1 induced epithelial-to-mesenchymal transition (EMT) and HCC metastasis both in vitro and in vivo. This tumorigenic effect depended on gankyrin; inhibiting gankyrin activity reversed EMT via activation of protein kinase B (AKT)/Twist family BHLH transcription factor 1 (AKT/TWIST1). We also found that KIFC1 was directly regulated by the microRNA miR-532-3p, whose downregulation was associated with metastatic progression in HCC. These results denote that a decrease in miR-532-3p levels results in increased KIFC1 expression in HCC, leading to metastasis via activation of the gankyrin/AKT/TWIST1 signaling pathway.


Assuntos
Carcinoma Hepatocelular/secundário , Transição Epitelial-Mesenquimal/fisiologia , Cinesina/fisiologia , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/secundário , MicroRNAs/fisiologia , Proteínas de Neoplasias/fisiologia , RNA Neoplásico/fisiologia , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/mortalidade , Linhagem Celular Tumoral , Regulação para Baixo , Xenoenxertos , Humanos , Estimativa de Kaplan-Meier , Cinesina/antagonistas & inibidores , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/mortalidade , Neoplasias Pulmonares/fisiopatologia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/biossíntese , MicroRNAs/genética , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas Nucleares/fisiologia , Prognóstico , Complexo de Endopeptidases do Proteassoma/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Interferência de RNA , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Transdução de Sinais , Proteína 1 Relacionada a Twist/fisiologia
9.
Bioorg Med Chem Lett ; 29(2): 148-154, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30528696

RESUMO

The mitotic spindle is a microtubule-based machine that segregates a replicated set of chromosomes during cell division. Many cancer drugs alter or disrupt the microtubules that form the mitotic spindle. Microtubule-dependent molecular motors that function during mitosis are logical alternative mitotic targets for drug development. Eg5 (Kinesin-5) and Kif15 (Kinesin-12), in particular, are an attractive pair of motor proteins, as they work in concert to drive centrosome separation and promote spindle bipolarity. Furthermore, we hypothesize that the clinical failure of Eg5 inhibitors may be (in part) due to compensation by Kif15. In order to test this idea, we screened a small library of kinase inhibitors and identified GW108X, an oxindole that inhibits Kif15 in vitro. We show that GW108X has a distinct mechanism of action compared with a commercially available Kif15 inhibitor, Kif15-IN-1 and may serve as a lead with which to further develop Kif15 inhibitors as clinically relevant agents.


Assuntos
Inibidores Enzimáticos/farmacologia , Cinesina/antagonistas & inibidores , Sondas Moleculares/farmacologia , Oxindois/farmacologia , Quinazolinonas/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Cinesina/metabolismo , Sondas Moleculares/síntese química , Sondas Moleculares/química , Estrutura Molecular , Oxindois/síntese química , Oxindois/química , Quinazolinonas/síntese química , Quinazolinonas/química , Relação Estrutura-Atividade
11.
Bioorg Chem ; 84: 493-504, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30594885

RESUMO

Human kinesin Eg5 is a potential inhibiting site for cancer chemotherapy. Blocking metaphase by binding foreign inhibitors with Eg5 eventually leads to apoptotic cell death. Here, we report the pyrazolopyrimidine derivates as potent inhibitors of Eg5 that prevents mitotic kinesin progression. IC50 values were evaluated against the motor domain of Eg5 using steady-state ATPase assay. To better understanding, we have performed molecular docking simulation. It reveals that the interactions of the proposed inhibitors with both the allosteric sites (helices α2, α3 and loopL5, and helices α4 & α6). Out of fifteen pyrazolopyrimidine derivates, three compounds (12, 25, and 27) have shown significant inhibition of Eg5. The synthesized compounds (12, 25, and 27) were tested for their in-vitro anticancer activity against cervical cancer cell line (HeLa).


Assuntos
Antineoplásicos/química , Cinesina/antagonistas & inibidores , Pirazóis/química , Pirimidinas/química , Sítio Alostérico , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Sítios de Ligação , Células HeLa , Humanos , Cinesina/metabolismo , Simulação de Acoplamento Molecular , Pirimidinas/metabolismo , Pirimidinas/farmacologia , Relação Estrutura-Atividade
12.
Chemotherapy ; 64(4): 187-196, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-32074608

RESUMO

Kinesin family (KIF) members have vital roles in mitosis, meiosis, and transport of macromolecules in eukaryotic cells. In this study, we aimed to investigate the role of KIF15 in osteosarcoma. Immunohistochemical staining was performed to determine expression levels of KIF15 in osteosarcoma tissues and adjacent normal tissues. Tissue microarray analysis showed a correlation between the expression of KIF15 and pathological features of patients. Subsequently, lentivirus was used to inhibit the expression of KIF15 in osteosarcoma cells. An MTT assay was performed to examine cell proliferation. Transwell and wound healing assays were used to estimate the invasion and migration of osteosarcoma cells, respectively. Flow cytometric analysis was employed to define the apoptosis of osteosarcoma cells. Our results showed that KIF15 expression was significantly upregulated in osteosarcoma tissues compared with adjacent normal tissues. The Mann-Whitney U test and Spearman correlation analysis showed that the expression levels of KIF15 in osteosarcoma tissues were positively correlated with tumor infiltrate, a pathological characteristic of patients. The expression of KIF15 was successfully suppressed by shKIF15, and the knockdown efficiency reached 80 and 69% in MNNG/HOS and U2OS cells, respectively. Subsequently, knockdown of KIF15 significantly inhibited the capacity of cell proliferation, colony formation, invasion, and migration, but enhanced G2 phase arrest and partially enhanced cell apoptosis. This study preliminarily showed KIF15 to be a critical regulatory molecule involved in osteosarcoma cell proliferation. Consequently, KIF15 can be a potential diagnostic and therapeutic target for osteosarcoma.


Assuntos
Apoptose , Neoplasias Ósseas/patologia , Proliferação de Células , Cinesina/metabolismo , Osteossarcoma/patologia , Adulto , Neoplasias Ósseas/metabolismo , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular , Feminino , Humanos , Cinesina/antagonistas & inibidores , Cinesina/genética , Masculino , Osteossarcoma/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
13.
PLoS One ; 13(12): e0209296, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30557316

RESUMO

Cultured cells easily develop resistance to kinesin-5 inhibitors (K5Is) often by overexpressing a related motor protein, kinesin-12/KIF15, or by acquiring mutations in the N-terminal motor domain of kinesin-5/KIF11 itself. We aimed to identify novel mechanisms responsible for resistance to S-trityl L-cysteine (STLC), one of the K5Is, using human osteosarcoma cell lines. Among six lines examined, U-2OS and HOS survived chronic STLC treatment and gave rise to resistant cells with IC50s at least 10-fold higher than those of the respective parental lines. Depletion of KIF15 largely eliminated the acquired K5I resistance in both cases, consistent with the proposed notion that KIF15 is indispensable for it. In contrast to the KIF11-independent property of the cells derived from HOS, those derived from U-2OS still required KIF11 for their growth and, intriguingly, expressed a C-terminal truncated variant of KIF11 resulting from a frame shift mutation (S1017fs). All of the isolated clones harbored the same mutation, suggesting its clonal expansion in the cell population due to the growth advantage during chronic STLC treatment. Transgenic expression of KIF11S1017fs in the parental U-2OS cells, as well as in HeLa cells, conferred a moderate but reproducible STLC resistance, probably owing to STLC-resistant localization of the mutant KIF11 on mitotic spindle. Our observations indicate that both KIF15 and the C-terminal-truncated KIF11 contributes to the STLC resistance of the U-2OS derived cells.


Assuntos
Antimitóticos/farmacologia , Cisteína/análogos & derivados , Resistência a Medicamentos/fisiologia , Cinesina/antagonistas & inibidores , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisteína/farmacologia , Resistência a Medicamentos/genética , Dineínas/metabolismo , Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Cinesina/genética , Cinesina/metabolismo , Mutação , Membrana Nuclear/efeitos dos fármacos , Membrana Nuclear/metabolismo , Fuso Acromático/efeitos dos fármacos , Fuso Acromático/metabolismo
14.
Cell Physiol Biochem ; 51(1): 1-10, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30439711

RESUMO

BACKGROUND/AIMS: Lung cancer is the leading cause of cancer-related deaths worldwide. The outcome of patients with non-small cell lung cancer remains poor; the 5-year survival rate for stage IV non-small cell lung cancer is only 1.0%. KIF15 is a tetrameric kinesin spindle motor that has been investigated for its regulation of mitosis. While the roles of kinesin motor proteins in the regulation of mitosis and their potentials as therapeutic targets in pancreatic cancer have been described previously, the role of KIF15 in lung cancer development remains unknown. METHODS: Paired lung carcinoma specimens and matched adjacent normal tissues were used for protein analysis. Clinical data were obtained from medical records. We first examined KIF15 messenger RNA expression in The Cancer Genome Atlas database, and then determined KIF15 protein levels using immunohistochemistry and western blotting. Differences between the groups were analyzed using repeated measures analysis of variance. Overall survival was analyzed using the Kaplan-Meier method. Cell-cycle and proliferation assays were conducted using A549, NCI-H1299, and NCI-H226 cells. RESULTS: KIF15 was significantly upregulated at both the messenger RNA and protein levels in human lung tumor tissues. In patients with lung adenocarcinoma, KIF15 expression was positively associated with disease stages; high KIF15 expression predicted a poor prognosis. KIF15 knockdown using short hairpin RNA in two human lung adenocarcinoma cell lines induced G1/S phase cell cycle arrest and inhibited cell growth, but there was no effect in human lung squamous cell carcinoma. CONCLUSION: Our findings show that KIF15 is involved in lung cancer carcinogenesis. KIF15 could therefore serve as a specific prognostic marker for patients with lung adenocarcinoma.


Assuntos
Adenocarcinoma/patologia , Cinesina/metabolismo , Neoplasias Pulmonares/patologia , Células A549 , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidade , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Pontos de Checagem da Fase G1 do Ciclo Celular , Humanos , Estimativa de Kaplan-Meier , Cinesina/antagonistas & inibidores , Cinesina/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/mortalidade , Masculino , Estadiamento de Neoplasias , Prognóstico , Interferência de RNA , RNA Interferente Pequeno/metabolismo
15.
Angew Chem Int Ed Engl ; 57(46): 15243-15247, 2018 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-30180286

RESUMO

The number of cytotoxic payload classes successfully employed in antibody-drug conjugates (ADCs) is still rather limited. The identification of ADC payloads with a novel mode of action will increase therapeutic options and potentially increase the therapeutic window. Herein, we describe the utilization of kinesin spindle protein inhibitors (KSPi) as a novel payload class providing highly potent ADCs against different targets, for instance HER-2 or TWEAKR/Fn14. Aspects of technical optimization include the development of different linker attachment sites, the stabilization of ADC linkage to avoid payload deconjugation and finally, the tailor-made design of active metabolites with a long lasting intracellular exposure in the tumor matching the mode of action of KSP inhibition. These KSPi-ADCs are highly potent and selective in vitro and demonstrate in vivo efficacy in a broad panel of tumor models including complete regressions in a patient-derived urothelial cancer model.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Imunoconjugados/química , Imunoconjugados/farmacologia , Cinesina/antagonistas & inibidores , Pirróis/química , Pirróis/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Descoberta de Drogas , Humanos , Imunoconjugados/uso terapêutico , Neoplasias/tratamento farmacológico , Pirróis/uso terapêutico
16.
Cancer Sci ; 109(11): 3450-3460, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30191636

RESUMO

Kinesin family member 20B (KIF20B, also known as MPHOSPH1) is a kinesin protein that plays a critical role in cytokinesis. Previously, we and others have demonstrated the oncogenic role of KIF20B in several cancers; however, the exact mechanisms underlying its tumorigenic effects remain unclear. Herein, we showed overexpression of KIF20B in human hepatocellular carcinoma (HCC) and reported a negative correlation between KIF20B level and prognosis of patients. Mechanistically, reducing KIF20B blockades mitotic exit of HCC cells at telophase in a spindle assembly checkpoint independent way. Importantly, reducing KIF20B acts synergistically with three microtubule-associated agents (MTA) to p53- or p14ARF-dependently suppress p53-wt or p53-null HCC cells. In addition to taxol, reducing KIF20B also enhanced the toxicity of two chemotherapeutic drugs, hydroxycamptothecin and mitomycin C. In conclusion, we found a novel mechanism in that blocking cytokinesis by KIF20B inhibition increases the efficacy of MTA; our results thus suggested a dual-mitotic suppression approach against HCC by combining MTA with KIF20B inhibition, which simultaneously blocks mitosis at both metaphase and telophase.


Assuntos
Carcinoma Hepatocelular/metabolismo , Cinesina/antagonistas & inibidores , Neoplasias Hepáticas/metabolismo , RNA Interferente Pequeno/farmacologia , Moduladores de Tubulina/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citocinese/efeitos dos fármacos , Sinergismo Farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Mitose/efeitos dos fármacos , Prognóstico , Regulação para Cima/efeitos dos fármacos
17.
Lab Chip ; 18(20): 3196-3206, 2018 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-30204813

RESUMO

Molecular motors, essential to force-generation and cargo transport within cells, are invaluable tools for powering nanobiotechnological lab-on-a-chip devices. These devices are based on in vitro motility assays that reconstitute molecular transport with purified motor proteins, requiring a deep understanding of the biophysical properties of motor proteins and thorough optimization to enable motility under varying environmental conditions. Until now, these assays have been prepared manually, severely limiting throughput. To overcome this limitation, we developed an in vitro motility assay where sample preparation, imaging and data evaluation are fully automated, enabling the processing of a 384-well plate within less than three hours. We demonstrate the automated assay for the analysis of peptide inhibitors for kinesin-1 at a wide range of concentrations, revealing that the IAK domain responsible for kinesin-1 auto-inhibition is both necessary and sufficient to decrease the affinity of the motor protein for microtubules, an aspect that was hidden in previous experiments due to scarcity of data.


Assuntos
Movimento Celular , Ensaios de Triagem em Larga Escala/instrumentação , Cinesina/metabolismo , Sequência de Aminoácidos , Animais , Automação , Movimento Celular/efeitos dos fármacos , Proteínas de Drosophila/antagonistas & inibidores , Proteínas de Drosophila/metabolismo , Cinesina/antagonistas & inibidores , Microscopia de Fluorescência , Microtúbulos/efeitos dos fármacos , Microtúbulos/metabolismo , Peptídeos/química , Peptídeos/farmacologia
18.
Eur J Med Chem ; 156: 641-651, 2018 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-30031975

RESUMO

The thiadiazole scaffold is an important core moiety in a variety of clinical drug candidates targeting a range of diseases. For example, the 2,4,5-substituted 1,3,4-thiadiazole scaffold is present in a lead compound and at least two clinical candidates targeting the human motor protein Eg5, against neoplastic diseases. An inhibitor named K858 has in vivo activity in various mouse xenografts whereas the clinical candidates (S)-ARRY-520 and (R)-Litronesib have entered clinical trials with the former one in phase III clinical trials either alone or in combination with a proteasome inhibitor against relapsed/refractory multiple myeloma. Astonishingly, structural data are lacking for all thiadiazole-containing Eg5 inhibitors. Here we report the structure determination of two crystal forms of the ternary Eg5-ADP-K858 complex, locking the motor in the so-called final inhibitor bound state, thus blocking ADP release, a crucial stage for Eg5 activity. K858 acts at the established allosteric inhibitor-binding pocket formed of helix α2, loop L5 and helix α3. The structure of the complex has far reaching consequences for thiadiazole containing Eg5 inhibitors. For example, we could rationalise the structure-activity relationship in the crucial 5-position of the thiadiazole scaffold and the complex will serve in the future as a basis for strucutre-based drug design.


Assuntos
Desenho de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Cinesina/antagonistas & inibidores , Tiadiazóis/química , Tiadiazóis/farmacologia , Sítios de Ligação , Cristalografia por Raios X , Humanos , Cinesina/química , Cinesina/metabolismo , Simulação de Acoplamento Molecular , Ligação Proteica , Relação Estrutura-Atividade
19.
Bioorg Med Chem Lett ; 28(17): 2930-2938, 2018 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-30055887

RESUMO

A novel series of 1,3,4-thiadiazole-thiazolone hybrids 5a-v were designed, synthesized, characterized, and evaluated against the basal and the microtubule (MT)-stimulated ATPase activity of Eg5. From the evaluated derivatives, 5h displayed the highest inhibition with an IC50 value of 13.2 µM against the MT-stimulated Eg5 ATPase activity. Similarly, compounds 5f and 5i also presented encouraging inhibition with IC50 of 17.2 µM and 20.2 µM, respectively. A brief structure-activity relationship (SAR) analysis indicated that 2-chloro and 4-nitro substituents on the phenyl ring of the thiazolone motif contributed significantly to enzyme inhibition. An in silico molecular docking study using the crystal structure of Eg5 further supported the SAR and reasoned the importance of crucial molecular protein-ligand interactions in influencing the inhibition of the ATPase activity of Eg5. The magnitude of the electron-withdrawing functionalities over the hybrids and the critical molecular interactions contributed towards higher in vitro potency of the compounds. The drug-like properties of the synthesized compounds 5a-v were also calculated based on the Lipinski's rule of five and in silico computation of key pharmacokinetic parameters (ADME). Thus, the present work unveils these hybrid molecules as novel Eg5 inhibitors with promising drug-like properties for future development.


Assuntos
Desenho de Drogas , Cinesina/antagonistas & inibidores , Tiadiazóis/farmacologia , Tiazóis/farmacologia , Adenosina Trifosfatases/metabolismo , Relação Dose-Resposta a Droga , Humanos , Cinesina/metabolismo , Microtúbulos/metabolismo , Estrutura Molecular , Relação Estrutura-Atividade , Tiadiazóis/química , Tiazóis/química
20.
Curr Pharm Des ; 24(17): 1873-1878, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29938614

RESUMO

PURPOSE: The purpose of this study was to determine the relationship between Kinesin like protein 6 (KIF6) gene Trp719Arg and major cardiovascular events (MACEs) risk in subjects who received statin therapy. METHODS: PubMed, EmBase, and the Cochrane Library were searched from inception through September 2017. The selected studies evaluated the association of Trp719Arg with MACEs in individuals who received statins. Relative risk (RR) and 95% confidence interval (CI) were used to evaluate the effect of statin therapy on MACEs in subjects carrying polymorphisms, and the odds ratio (OR) and 95% CI were used to evaluate the relationship between Trp719Arg and MACE risk in individuals who received statins, using the random-effects model. RESULTS: Seven studies were included (N=48,885). Overall, we found that statin therapy significantly reduced the risk for MACEs in subjects carrying ArgArg (RR: 0.79; 95% CI: 0.69-0.90; P=0.001), ArgTrp (RR: 0.71; 95% CI: 0.60 -0.83; P<0.001), ArgArg+ArgTrp (RR: 0.71; 95% CI: 0.63 -0.81; P<0.001), and TrpTrp (RR: 0.79; 95% CI: 0.73-0.85; P<0.001). Furthermore, there was no significant difference between subjects carrying ArgArg and those carrying TrpTrp (OR: 1.11; 95% CI: 0.92-1.34; P=0.265). However, ArgTrp (OR: 1.29; 95% CI: 1.07-1.55; P=0.007) and ArgArg+ArgTrp (OR: 1.26; 95% CI: 1.05-1.51; P=0.012) were associated with an increased risk for MACEs when compared with TrpTrp. CONCLUSIONS: Statin therapy significantly reduced the risk for MACEs in subjects carrier specific KIF6 gene Trp719Arg polymorphisms. Further, subjects carrying ArgTrp or ArgArg+ArgTrp had a greater incidence of MACEs as compared with TrpTrp when they received statins.


Assuntos
Arginina/antagonistas & inibidores , Doenças Cardiovasculares/tratamento farmacológico , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Cinesina/antagonistas & inibidores , Arginina/genética , Doenças Cardiovasculares/genética , Variação Genética/efeitos dos fármacos , Variação Genética/genética , Humanos , Cinesina/genética , Polimorfismo Genético/efeitos dos fármacos , Polimorfismo Genético/genética , Fatores de Risco
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