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1.
Neoplasma ; 66(6): 946-953, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31607131

RESUMO

The aim of this study was to determine the expression levels of TTK in clear cell renal cell carcinoma (ccRCC) tissues and its possible link with the clinical pathologic characteristics and the prognosis of patients suffering this disease, and to further explore the potential role of TTK in the progression of ccRCC. Immunohistochemical (IHC) assays were performed to detect the expression levels of TTK in 112 samples of ccRCC tissues and corresponding non-tumor tissues. According to the results of IHC assays, patients were divided into TTK high expression and low expression group. Clinical analysis related to the clinical features (age, gender, T stage), and the potential link between TTK expression levels and clinical features were analyzed. In addition, the effects of TTK on the proliferation and invasion of ccRCC cells were detected through colony formation assay and transwell assays, respectively. The possible effects of TTK on tumor growth and metastasis were measured in mice. We found a high expression level of TTK in human ccRCC tissues from patients who received surgical treatment. We also found its expression level was obviously associated with clinical characteristics, such as T stage (p=0.008) and lymphatic metastasis (p=0.023). We further confirmed that knockdown of TTK suppressed cell proliferation and invasion in 2 types of ccRCC cells, HTB-47 and CRL-1932 cells. Furthermore, TTK contributes to tumor growth and metastasis of ccRCC in mice. We found that TTK affected the progression of ccRCC and further mechanically confirmed it could be a novel therapeutic target for ccRCC treatment.


Assuntos
Carcinoma de Células Renais/patologia , Proteínas de Ciclo Celular/metabolismo , Neoplasias Renais/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Tirosina Quinases/metabolismo , Animais , Carcinoma de Células Renais/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Renais/metabolismo , Camundongos , Metástase Neoplásica , Prognóstico
2.
Nat Commun ; 10(1): 4432, 2019 09 30.
Artigo em Inglês | MEDLINE | ID: mdl-31570755

RESUMO

The pathway of homeostatic IgG extravasation is not fully understood, in spite of its importance for the maintenance of host immunity, the management of autoantibody-mediated disorders, and the use of antibody-based biologics. Here we show in a murine model of pemphigus, a prototypic cutaneous autoantibody-mediated disorder, that blood-circulating IgG extravasates into the skin in a time- and dose-dependent manner under homeostatic conditions. This IgG extravasation is unaffected by depletion of Fcγ receptors, but is largely attenuated by specific ablation of dynamin-dependent endocytic vesicle formation in blood endothelial cells (BECs). Among dynamin-dependent endocytic vesicles, IgG co-localizes well with caveolae in cultured BECs. An Abl family tyrosine kinase inhibitor imatinib, which reduces caveolae-mediated endocytosis, impairs IgG extravasation in the skin and attenuates the murine pemphigus manifestations. Our study highlights the kinetics of IgG extravasation in vivo, which might be a clue to understand the pathological mechanism of autoantibody-mediated autoimmune disorders.


Assuntos
Imunoglobulina G/imunologia , Pênfigo/imunologia , Pênfigo/metabolismo , Proteínas Tirosina Quinases/metabolismo , Pele/imunologia , Animais , Autoanticorpos/metabolismo , Doenças Autoimunes/metabolismo , Cavéolas , Modelos Animais de Doenças , Dinaminas/metabolismo , Orelha/patologia , Endocitose , Células Endoteliais/metabolismo , Feminino , Imunoglobulina G/sangue , Cinética , Camundongos , Camundongos Endogâmicos C57BL , Pênfigo/patologia , Pele/patologia , Vesículas Transportadoras/metabolismo
3.
Nat Commun ; 10(1): 4343, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554817

RESUMO

Infant gliomas have paradoxical clinical behavior compared to those in children and adults: low-grade tumors have a higher mortality rate, while high-grade tumors have a better outcome. However, we have little understanding of their biology and therefore cannot explain this behavior nor what constitutes optimal clinical management. Here we report a comprehensive genetic analysis of an international cohort of clinically annotated infant gliomas, revealing 3 clinical subgroups. Group 1 tumors arise in the cerebral hemispheres and harbor alterations in the receptor tyrosine kinases ALK, ROS1, NTRK and MET. These are typically single-events and confer an intermediate outcome. Groups 2 and 3 gliomas harbor RAS/MAPK pathway mutations and arise in the hemispheres and midline, respectively. Group 2 tumors have excellent long-term survival, while group 3 tumors progress rapidly and do not respond well to chemoradiation. We conclude that infant gliomas comprise 3 subgroups, justifying the need for specialized therapeutic strategies.


Assuntos
Neoplasias Encefálicas/genética , Metilação de DNA , Epigenômica/métodos , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Receptores Proteína Tirosina Quinases/genética , Quinase do Linfoma Anaplásico/genética , Quinase do Linfoma Anaplásico/metabolismo , Neoplasias Encefálicas/classificação , Neoplasias Encefálicas/metabolismo , Feminino , Glioma/classificação , Glioma/metabolismo , Humanos , Lactente , Recém-Nascido , Masculino , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptor trkA/genética , Receptor trkA/metabolismo , Análise de Sobrevida , Sequenciamento Completo do Exoma/métodos
4.
Eur J Med Chem ; 182: 111664, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31494475

RESUMO

As an important source of drugs, natural products play an important role in the discovery and development of new drugs. More than 60% of anti-tumor drugs are closely related to natural products. At the same time, as the main cause of tumors, the abnormal activity of tyrosine kinase has become an important target for clinical treatment. Although, small molecule targeted drugs dominate the cancer treatment. Natural active products are driving the development of new tyrosine kinase inhibitors with their unique mode of action and molecular structure diversity. Obtaining new chemical entities with tyrosine kinase inhibitory activity from natural active products will bring new breakthroughs in the research of anticancer drugs. In this paper, different tyrosine kinases are mainly classified as targets, and natural products and derivatives which have been found to inhibit tyrosine kinase activity have been described. It is hoped that by analyzing the different aspects of the source, structural characteristics, mechanism of action and biological activity of these natural products, we will find new members that can be developed into drugs and promote the development of anti-tumor drugs.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Produtos Biológicos/farmacologia , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Antineoplásicos Fitogênicos/química , Produtos Biológicos/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Inibidores de Proteínas Quinases/química , Proteínas Tirosina Quinases/metabolismo
5.
Zhonghua Shao Shang Za Zhi ; 35(8): 580-586, 2019 Aug 20.
Artigo em Chinês | MEDLINE | ID: mdl-31474037

RESUMO

Objective: To investigate the role and mechanism of nonreceptor tyrosine kinase Tec in the production of pro-inflammatory cytokine interleukin-8 (IL-8) induced by endotoxin/lipopolysaccharide (LPS) in human alveolar epithelial cells A549. Methods: Human alveolar epithelial cells A549 were routinely cultured and passaged in Roswell Park Memorial Institute-1640 medium containing 10% fetal bovine serum. The second or third passage of cells were collected for subsequent experiments. (1) Cells were collected and divided into 6 groups with 4 wells in each group according to the random number table. Cells in blank control group were routinely cultured for 2 h. Cells in simple LPS group were routinely cultured for 1 h and then stimulated by 1 µg/mL LPS for 1 h. Cells in simple LFM-A13 group were cultured with conventional culture medium adding 75 µmol/L LFM-A13 for 1 h and then cultured with replaced conventional culture medium for 1 h. Cells in 25 µmol/L LFM-A13+ LPS group, 75 µmol/L LFM-A13+ LPS group, and 100 µmol/L LFM-A13+ LPS group were cultured with conventional culture medium adding 25, 75, and 100 µmol/L LFM-A13 respectively for 1 h and then all stimulated by 1 µg/mL LPS added into the replaced conventional culture medium for 1 h. The protein expression of Tec in cells of each group was detected by Western blotting, and the content of IL-8 in cell culture supernatant of each group was determined by enzyme-linked immunosorbent assay. (2) Cells were collected and divided into 5 groups with 4 wells in each group according to the random number table. Cells in blank control group were routinely cultured for 2 h. Cells in small interfering RNA (siRNA) control+ LPS group were transfected with empty lentivirus for 10 h and then stimulated by 1 µg/mL LPS added into the conventional culture medium for 2 h. Cells in Tec mus-298 RNA interference (RNAi)+ LPS group, Tec mus-299 RNAi+ LPS group, and Tec mus-300 RNAi+ LPS group were transfected with lentivirus loaded with Tec mus-298 RNAi, Tec mus-299 RNAi, and Tec mus-300 RNAi respectively for 10 h and then stimulated by 1 µg/mL LPS added into the conventional culture medium for 2 h. The protein expression of Tec in cells of each group was detected by Western blotting to screen Tec-siRNA with the best silencing effect on Tec gene. (3) Cells were collected and divided into 4 groups with 4 wells in each group according to the random number table. Cells in blank control group were routinely cultured for 2 h. Cells in virus control group were transfected with empty lentivirus for 10 h and then routinely cultured for 2 h. Cells in simple LPS group were stimulated by 1 µg/mL LPS added into the conventional culture medium for 2 h. Cells in Tec-siRNA+ LPS group were transfected with lentivirus loaded with Tec-siRNA with the best silencing effect on Tec gene for 10 h and then stimulated by 1 µg/mL LPS added into the conventional culture medium for 2 h. The protein expressions of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) MAPK of cells in each group were detected by Western blotting. Data were processed with one-way analysis of variance and the least significant difference-t test. Results: (1) Compared with that of blank control group, the protein expression of Tec of cells in simple LPS group was obviously increased (t=9.72, P<0.05), but the protein expression of Tec of cells in simple LFM-A13 group was not obviously changed (t=4.31, P=0.05). Compared with that of simple LPS group, the protein expression of Tec of cells in 25 µmol/L LFM-A13+ LPS group, 75 µmol/L LFM-A13+ LPS group, or 100 µmol/L LFM-A13+ LPS group was obviously decreased (t=9.72, 9.07, 16.33, P<0.05 or P<0.01). Compared with (189±22) pg/mL of blank control group, the content of IL-8 in culture supernatant of cells in simple LPS group was obviously increased [(214±10) pg/mL, t=2.18, P<0.05], but the content of IL-8 in culture supernatant of cells in simple LFM-A13 group was not obviously changed [(173±43) pg/mL, t=0.64, P>0.05]. Compared with that of simple LPS group, the content of IL-8 in culture supernatant of cells in 25 µmol/L LFM-A13+ LPS group was not obviously changed [(204±38) pg/mL, t=0.54, P>0.05], but the content of IL-8 in culture supernatant of cells in 75 µmol/L LFM-A13+ LPS group and 100 µmol/L LFM-A13+ LPS group was obviously decreased [(144±44), (137±51) pg/mL, t=3.63, 2.55, P<0.05 or P<0.01]. (2) Compared with that of blank control group, the protein expression of Tec of cells in siRNA control+ LPS group was obviously increased (t=14.24, P<0.01). Compared with that of siRNA control+ LPS group, the protein expression of Tec of cells in Tec mus-298 RNAi+ LPS group or Tec mus-299 RNAi+ LPS group was obviously decreased (t=36.03, 18.23, P<0.01), but the protein expression of Tec of cells in Tec mus-300 RNAi+ LPS group was not obviously changed (t=4.08, P>0.05). The protein expression of Tec was the lowest in cells of Tec mus-298 RNAi+ LPS group, so Tec mus-298 RNAi was used in subsequent experiment. (3) Compared with 1.16±0.16 and 0.78±0.11 of blank control group, the protein expressions of p38 MAPK and ERK MAPK of cells in virus control group were not obviously changed (1.66±0.13, 0.89±0.11, t=11.09, 3.60, P>0.05), but the protein expressions of p38 MAPK and ERK MAPK of cells in simple LPS group were obviously increased (2.83±0.29, 1.86±0.37, t=9.70, 7.23, P<0.05). Compared with those of simple LPS group, the protein expression of p38 MAPK and protein expression of ERK MAPK of cells in Tec-siRNA+ LPS group were obviously decreased (0.69±0.16, 1.03±0.24, t=13.78, 4.12, P<0.05 or P<0.01). Conclusions: Tec may mediate the production and release of pro-inflammatory cytokine IL-8 from human alveolar epithelial cells A549 induced by LPS via the p38 MAPK and ERK MAPK signal pathways.


Assuntos
Células Epiteliais Alveolares/metabolismo , Interleucina-8/metabolismo , Proteínas Tirosina Quinases/metabolismo , Células A549 , Humanos , Lipopolissacarídeos , Transdução de Sinais
6.
Eur J Med Chem ; 181: 111545, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31400706

RESUMO

Abnormal activation of B-cell receptor (BCR) signaling plays a key role in the development of lymphoid malignancies, and could be reverted by the simultaneous inhibition of Lyn, Fyn and Blk, three members of the Src family kinase (SFK). Fyn and Blk are also promising targets for the treatment of some forms of T-cell non-Hodgkin lymphoma which point to the druggability of SFKs for the treatment of these cancers. We recently identified Si308 as a potent Fyn inhibitor, while preliminary data showed that it might also inhibit Lyn and Blk. Here, molecular modelling studies were coupled with enzymatic assays to further investigate the effect of Si308 on Lyn and Blk. A small library of pyrazolo[3,4-d]pyrimidines structurally related to Si308 was synthesized and tested on human lymphoma cell lines. Compound 2h emerged as a new multitarget inhibitor of Lyn, Fyn and Blk endowed with remarkable antiproliferative effects on human B and T lymphoma cell lines. Its favorable ADME properties make the compound suitable for further developments.


Assuntos
Antineoplásicos/farmacologia , Linfoma de Células T/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-fyn/antagonistas & inibidores , Pirazóis/farmacologia , Pirimidinas/farmacologia , Quinases da Família src/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Linfoma de Células T/metabolismo , Linfoma de Células T/patologia , Estrutura Molecular , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Pirazóis/síntese química , Pirazóis/química , Pirimidinas/síntese química , Pirimidinas/química , Relação Estrutura-Atividade , Células Tumorais Cultivadas , Quinases da Família src/metabolismo
7.
Biochem Soc Trans ; 47(4): 1101-1116, 2019 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-31395755

RESUMO

The SRC, Abelson murine leukemia viral oncogene homolog 1, TEC and C-terminal SRC Kinase families of non-receptor tyrosine kinases (collectively the Src module kinases) mediate an array of cellular signaling processes and are therapeutic targets in many disease states. Crystal structures of Src modules kinases provide valuable insights into the regulatory mechanisms that control activation and generate a framework from which drug discovery can advance. The conformational ensembles visited by these multidomain kinases in solution are also key features of the regulatory machinery controlling catalytic activity. Measurement of dynamic motions within kinases substantially augments information derived from crystal structures. In this review, we focus on a body of work that has transformed our understanding of non-receptor tyrosine kinase regulation from a static view to one that incorporates how fluctuations in conformational ensembles and dynamic motions influence activation status. Regulatory dynamic networks are often shared across and between kinase families while specific dynamic behavior distinguishes unique regulatory mechanisms for select kinases. Moreover, intrinsically dynamic regions of kinases likely play important regulatory roles that have only been partially explored. Since there is clear precedence that kinase inhibitors can exploit specific dynamic features, continued efforts to define conformational ensembles and dynamic allostery will be key to combating drug resistance and devising alternate treatments for kinase-associated diseases.


Assuntos
Proteínas Tirosina Quinases/metabolismo , Animais , Catálise , Domínio Catalítico , Descoberta de Drogas , Ativação Enzimática , Humanos , Fosforilação , Conformação Proteica , Proteínas Tirosina Quinases/química , Transdução de Sinais , Domínios de Homologia de src
8.
Eur J Med Chem ; 181: 111512, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31404861

RESUMO

Curcumin is a natural substance known for ages, exhibiting a multidirectional effect in cancer prevention and adjuvant cancer therapies. The great advantage of using nutraceuticals of vegetable origin in comparison to popular cytostatic drugs is the minimized side effect and reduced toxicity. The targets in oncological therapy are, among others, tyrosine kinases, important mediators of signaling pathways whose impaired expression is observed in many types of cancer. Unfortunately, the hydrophobic nature of the curcumin molecule often limits its bioavailability, which is why many studies focus on the chemical modification of this compound. Current research is aimed at modifying structures that improve the pharmacokinetic parameters of curcumin, e.g. the formation of nanoparticles, complexes with metals or the synthesis of curcumin derivatives with functional substituents that allow tumor targeting. The article is a review and analysis of current literature on the properties of curcumin and its derivatives in the treatment of cancers directed to signaling pathways of tyrosine kinases and confronts the problem of low assimilation of curcumin with potential therapeutic effects.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Curcumina/farmacologia , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Proliferação de Células/efeitos dos fármacos , Curcumina/química , Curcumina/isolamento & purificação , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/isolamento & purificação , Proteínas Tirosina Quinases/metabolismo
9.
Neurochem Res ; 44(9): 2044-2056, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31278631

RESUMO

Depression is a highly debilitating and life-threatening psychiatric disorder. The classical antidepressants are still not adequate due to undesirable side effects. Therefore, the development of new drugs for depression treatment is an urgent strategic to achieving clinical needs. Licorisoflavan A is a bioactive ingredient isolated from Glycyrrhizae Radix and has been recently reported for neuroprotective effects. In this study, the antidepressant-like effect and neural mechanism of licorisoflavan A were explored. In the mice behavioral despair test, we observed that licorisoflavan A exhibited powerful antidepressant-like effect in forced swimming test (FST), tail suspension test (TST), without affecting locomotor activity in open field test (OFT). Additionally, licorisoflavan A administration significantly restored Chronic mild stress (CMS)-induced changes in sucrose preference test (SPT), FST, and TST, without altering the locomotion in OFT. In chronical-stimulated mice, the licorisoflavan A treatment effectively attenuated the expressions of Brain-derived neurotrophic factor (BDNF), tyrosine kinase B (TrkB), the phosphorylations of cAMP response element binding protein (CREB), extracellular signal-regulated kinase (ERK)-1/2, eukaryotic elongation factor 2 (eEF2), mammalian target of rapamycin (mTOR), initiation factor 4E-binding protein 1 (4E-BP-1), and p70 ribosomal protein S6 kinase (p70S6K) in hippocampus of CMS-induced mice. Additionally, licorisoflavan A could reverse the decreases in synaptic proteins post-synaptic density protein 95 (PSD-95) and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor subunit glutamate receptor 1 (GluR1) caused by CMS, and its antidepressant-like effect was blocked by the AMPA receptor antagonist NBQX. These findings served as preclinical evidence that licorisoflavan A exerted potent antidepressant-like effects involving BDNF-TrkB pathway and AMPA receptors. Licorisoflavan A might be used as a potential medicine against depression-like disorder.


Assuntos
Antidepressivos/uso terapêutico , Benzopiranos/uso terapêutico , Depressão/tratamento farmacológico , Receptores de AMPA/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Antidepressivos/isolamento & purificação , Comportamento Animal/efeitos dos fármacos , Benzopiranos/isolamento & purificação , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Hipocampo/metabolismo , Locomoção/efeitos dos fármacos , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Fosforilação/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Tirosina Quinases/metabolismo
10.
Artif Cells Nanomed Biotechnol ; 47(1): 2754-2763, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31286799

RESUMO

Colon cancer is one of the most malignant cancers. Histone modification is closely related to tumour development. Our study explored the functions of anti-silencing function 1A (ASF1A) on H4Y72ph in colon cancer cells. Colon cancer cell lines and clinical specimens were obtained and/or transfected with full length ASF1A or interference mRNA to mimic or silence of ASF1A expression. Immunoprecipitation and GST pull down was used to target targeting ASF1A or H4Y72ph. Cells were transfected with H4WT- or H4Y72F-expressing. An in vitro kinase activity assay was set to determine whether ASF1A could phosphorylate H4. The severity of autophagy was measured by detecting number of autophagosomes, number of EGFP-LC3, LC3-II/I, percentage of degradation and expression of autophagy associated gene (ATG). ASF1A positively regulated H4Y72ph; Immunoprecipitation assay and GST pull down results showed that ASF1A interacted directly with H4. In addition, ASF1A silence inhibited autophagosomes number, EGFP-LC3 number, LC3-II/I, percentage of degradation and ATG expression. Moreover, H4Y72F impaired the promoting autophagy effects of ASF1A. The ASF1A-H4Y72ph axis promoted colon cancer autophagy via transcriptional regulation of ATG genes. ASF1A regulated H4Y72ph and promotes autophagy in colon cancer cells via a kinase activity through regulation of ATG.


Assuntos
Autofagia , Proteínas de Ciclo Celular/metabolismo , Neoplasias do Colo/patologia , Histonas/metabolismo , Chaperonas Moleculares/metabolismo , Proteínas Tirosina Quinases/metabolismo , Adulto , Proteínas Relacionadas à Autofagia/genética , Proteínas de Ciclo Celular/deficiência , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Neoplasias do Colo/enzimologia , Neoplasias do Colo/metabolismo , Feminino , Inativação Gênica , Humanos , Masculino , Chaperonas Moleculares/genética , Fosforilação , Transcrição Genética
11.
Eur J Med Chem ; 179: 358-375, 2019 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-31260890

RESUMO

ALK and ROS1 kinases have become promising therapeutic targets since Crizotinib was used to treat non-small-cell lung cancer clinically. Aiming to explore new potent inhibitors, a series of 2-amino-4-(1-piperidine) pyridine derivatives that stabilized a novel DFG-shifted conformation in the kinase domain of ALK were designed and synthesized on the base of lead compound A. Biological evaluation highlighted that most of these new compounds could also potently inhibit ROS1 kinase, leading to the promising inhibitors against both ROS1 and ALK. Among them, the representative compound 2e stood out potent anti-proliferative activity against ALK-addicted H3122 and ROS1-addicted HCC78 cell lines (IC50 = 6.27 µM and 10.71 µM, respectively), which were comparable to that of Crizotinib. Moreover, 2e showed impressive enzyme activity against clinically Crizotinib-resistant ALKL1196M with an IC50 value of 41.3 nM, which was about 2-fold more potent than that of Crizotinib. 2e also showed potent inhibitory activity in about 6-fold superior to Crizotinib (IC50: 104.7 nM vs. 643.5 nM) in Ba/F3 cell line harboring ROS1G2032R. Furthermore, molecular modeling disclosed that all the representative inhibitors could dock into the active site of ALK and ROS1, which gave a probable explanation of anti Crizotinib-resistant mutants. These results indicated that our work has established a path forward for the generation of anti Crizotinib-resistant ALK/ROS1 dual inhibitors.


Assuntos
Quinase do Linfoma Anaplásico/antagonistas & inibidores , Antineoplásicos/farmacologia , Crizotinibe/farmacologia , Piperidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Piridinas/farmacologia , Células A549 , Quinase do Linfoma Anaplásico/metabolismo , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Crizotinibe/química , Relação Dose-Resposta a Droga , Desenho de Drogas , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Estrutura Molecular , Piperidinas/química , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Piridinas/química , Relação Estrutura-Atividade
12.
Anticancer Res ; 39(7): 3341-3345, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31262854

RESUMO

In early 2000, the term 'targeted therapy' became popular and was used to indicate all types of tyrosine kinase inhibitors (TKI). However, the term targeted therapy had been used much earlier. Targeting tumor metabolism was already considered as targeted therapy, with methotrexate and 5-fluorouracil as the most successful examples. Hormone therapy is another successful type of targeted therapy. Imatinib was the first TKI for the fusion protein BCR-ABL and represented a breakthrough in the treatment of chronic myeloid leukemia. Many other TKIs have been introduced into the clinic, but most were less specific and had multiple targets, and therefore, by definition, not targeted. However, with the introduction of TKIs developed specifically against mutations in the active site of a TK, more truly targeted TKI have been approved, such as new anaplastic lymphoma kinase - echinoderm microtubule-associated protein-like 4 (ALK-EML4) inhibitors and the epidermal growth factor-T790M-targeted osimertinib. This article summarizes the content of the Burger-Kelland award lecture given by the Author in February 2019 during the 40th EORTC-PAMM Group meeting in Verona, Italy and reviews the development of various targeted agents.


Assuntos
Antineoplásicos/uso terapêutico , Terapia de Alvo Molecular , Neoplasias/terapia , Humanos , Neoplasias/metabolismo , Proteínas Tirosina Quinases/metabolismo
13.
Cell Prolif ; 52(5): e12656, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31264309

RESUMO

OBJECTIVES: Cell migration has a key role in cancer metastasis, which contributes to drug resistance and tumour recurrence. Better understanding of the mechanisms involved in this process will potentially reveal new drug targets for cancer therapy. Fer is a non-receptor protein tyrosine kinase aberrantly expressed in various human cancers, whereas its role in tumour progression remains elusive. MATERIALS AND METHODS: Transgenic flies and epigenetic analysis were employed to investigate the role of Drosophila Fer (FER) in cell migration and underlying mechanisms. Co-immunoprecipitation assay was used to monitor the interaction between FER and Drosophila JNK (Bsk). The conservation of Fer in regulating JNK signalling was explored in mammalian cancer and non-cancer cells. RESULTS: Overexpression of FER triggered cell migration and activated JNK signalling in the Drosophila wing disc. Upregulation and downregulation in the basal activity of Bsk exacerbated and eliminated FER-mediated migration, respectively. In addition, loss of FER blocked signal transduction of the JNK pathway. Specifically, FER interacted with and promoted the activity of Bsk, which required both the kinase domain and the C-terminal of Bsk. Lastly, Fer regulated JNK activities in mammalian cells. CONCLUSIONS: Our study reveals FER as a positive regulator of JNK-mediated cell migration and suggests its potential role as a therapeutic target for cancer metastasis.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Proteínas Tirosina Quinases/metabolismo , Animais , Animais Geneticamente Modificados/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proteínas de Drosophila/química , Matriz Extracelular/metabolismo , Regulação da Expressão Gênica , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/química , Metaloproteinase 1 da Matriz/metabolismo , Domínios Proteicos , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/genética , Interferência de RNA , RNA Interferente Pequeno , Transdução de Sinais , Asas de Animais/metabolismo
14.
Int J Mol Sci ; 20(14)2019 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-31336846

RESUMO

Acute myeloid leukemia (AML) is a myeloid malignancy carrying a heterogeneous molecular panel of mutations participating in the blockade of differentiation and the increased proliferation of myeloid hematopoietic stem and progenitor cells. The historical "3 + 7" treatment (cytarabine and daunorubicin) is currently challenged by new therapeutic strategies, including drugs depending on the molecular landscape of AML. This panel of mutations makes it possible to combine some of these new treatments with conventional chemotherapy. For example, the FLT3 receptor is overexpressed or mutated in 80% or 30% of AML, respectively. Such anomalies have led to the development of targeted therapies using tyrosine kinase inhibitors (TKIs). In this review, we document the history of TKI targeting, FLT3 and several other tyrosine kinases involved in dysregulated signaling pathways.


Assuntos
Antineoplásicos/uso terapêutico , Biomarcadores Tumorais , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Terapia de Alvo Molecular , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Tirosina Quinases/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Ensaios Clínicos como Assunto , Regulação Leucêmica da Expressão Gênica , Humanos , Leucemia Mieloide Aguda/etiologia , Leucemia Mieloide Aguda/patologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Proteoma , Transdução de Sinais , Transcriptoma , Resultado do Tratamento
15.
Oxid Med Cell Longev ; 2019: 1754593, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31285782

RESUMO

Chemiluminescence (CL) is one of the most useful methods for detecting reactive oxygen species (ROS). Although fluorescence dyes or genetically encoded biosensors have been developed, CL is still used due to its high sensitivity, ease of use, and low cost. While initially established and used to measure high levels of ROS in phagocytic cells, CL assays are not ideal for measuring low levels of ROS. Here, we developed a newly modified CL assay using a chemiluminescent imaging system for measuring low concentrations of ROS in nonphagocytic cells. We found that dissolving luminol in NaOH, rather than DMSO, increased the H2O2-induced CL signal and that the addition of 4-iodophenylboronic acid (4IPBA) further increased CL intensity. Our new system also increased the rate and intensity of the CL signal in phorbol 12-myristate 13-acetate- (PMA-) treated HT-29 colon cancer cells compared to those in luminol only. We were able to quantify ROS levels from both cells and media in parallel using an H2O2 standard. A significant benefit to our system is that we can easily measure stimulus-induced ROS formation in a real-time manner and also investigate intracellular signaling pathways from a single sample simultaneously. We found that PMA induced tyrosine phosphorylation of protein tyrosine kinases (PTKs), such as focal adhesion kinase (FAK), protein tyrosine kinase 2 (Pyk2), and Src, and increased actin stress fiber formation in a ROS-dependent manner. Interestingly, treatment with either N-acetyl-L-cysteine (NAC) or diphenyleneiodonium (DPI) reduced the PMA-stimulated phosphorylation of these PTKs, implicating a potential role in cellular ROS signaling. Thus, our newly optimized CL assay using 4IPBA and a chemiluminescent imaging method provides a simple, real-time, and low-cost method for the quantification of low levels of ROS.


Assuntos
Espécies Reativas de Oxigênio/metabolismo , Acetilcisteína/farmacologia , Compostos de Boro/farmacologia , Quinase 1 de Adesão Focal/metabolismo , Células HT29 , Humanos , Immunoblotting , Iodobenzenos/farmacologia , Oniocompostos/farmacologia , Fosforilação/efeitos dos fármacos , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos
16.
Immunohorizons ; 3(1): 28-36, 2019 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-31356174

RESUMO

In addition to functioning as a ligand to engage TCRs and drive TCR signaling, MHC class II molecules are signaling molecules that generate a number of signals within APCs, such as B lymphocytes. Moreover, MHC class II signaling is critical for B cell activation and development of a robust humoral immune response. Murine class II molecules exist in two distinct conformational states, based primarily on the differential pairing of transmembrane domain GxxxG dimerization motifs (i.e., M1- and M2-paired class II). This laboratory has previously reported that the binding of a multimerized form of an anti-class II mAb that selectively recognizes M1-paired I-Ak class II drives intracellular calcium signaling in resting murine B cells and that this signaling is dependent on both src and Syk protein tyrosine kinase activity. In contrast, multimerized forms of two different anti-I-Ak mAbs that bind both M1- and M2-paired class II fail to elicit a response. In this report, a flow cytometry-based calcium flux assay is used to demonstrate that coligation of M1- and M2-paired I-Ak class II results in the active and selective inhibition of M1-paired I-Ak class II B cell calcium signaling by M2-paired class II molecules. Because M1- and M2-paired class II can be loaded with different sets of peptides derived from Ags acquired by distinct pathways of endocytosis, these findings suggest an MHC class II signaling-based mechanism by which CD4 T cells of differing specificities can either enhance or suppress B cell activation.


Assuntos
Linfócitos B/imunologia , Linfócitos B/metabolismo , Sinalização do Cálcio/imunologia , Antígenos de Histocompatibilidade Classe II/metabolismo , Ativação Linfocitária/imunologia , Animais , Anticorpos Monoclonais/metabolismo , Células Apresentadoras de Antígenos/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Comunicação Celular/imunologia , Reações Cruzadas/imunologia , Epitopos de Linfócito B/metabolismo , Antígenos de Histocompatibilidade Classe II/imunologia , Imunidade Humoral , Motivo de Ativação do Imunorreceptor Baseado em Tirosina , Motivo de Inibição do Imunorreceptor Baseado em Tirosina , Camundongos , Peptídeos/imunologia , Proteínas Tirosina Fosfatases/metabolismo , Proteínas Tirosina Quinases/metabolismo , Baço/citologia
17.
Life Sci ; 232: 116597, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31238052

RESUMO

LncRNA SNHG3 (SNHG3) is involved in tumor development and progression, but little is known about how SNHG3 functions in laryngeal carcinoma (LC). Real time-PCR (RT-PCR) was used to estimate the expression of SNHG3 in LC tissues and cell lines TU212, TU686, and Hep-2. Cell viability, migration, and invasion were evaluated. Our results showed increased SNHG3 in LC tissues and cell lines. Loss of function of SNHG3 reduced cell viability, migration, and invasion of TU212 and TU686 cells. Western blot analyses demonstrated that the protein levels of MMP2 and MMP9 decreased after SNHG3 silencing. Additionally, bioinformatics software predicted that SNHG3 could sponge miR-384 at the 3'-UTR with complementary binding sites, which was validated by a dual-luciferase reporter assay. RT-PCR analysis revealed that knockdown of SNHG3 upregulated miR-384 expression and that overexpression of miR-384 decreased SNHG3. Furthermore, a dual-luciferase reporter assay showed that miR-384 could bind to the 3'-UTR of WEE1, and inhibition of miR-384 markedly increased WEE1 expression. The mRNA and protein levels of WEE1 were downregulated upon deletion of SNGH3. Suppression of WEE1 partly abolished the tumorigenic migration and invasion potential of the miR-384 inhibitor in migration and invasion. Inhibition of miR-384 partially reversed the biological activities of SNHG3 in TU212 and TU686 cells. Collectively, our results indicate that SNHG3 regulated LC cell migration and invasion via the miR-384/WEE1 axis.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/metabolismo , MicroRNAs/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Tirosina Quinases/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Regiões 3' não Traduzidas , Carcinogênese , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Sobrevivência Celular/fisiologia , Humanos , Neoplasias Laríngeas/patologia , MicroRNAs/genética , Invasividade Neoplásica , Proteínas Nucleares/genética , Proteínas Tirosina Quinases/genética
18.
Psychopharmacology (Berl) ; 236(11): 3301-3315, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31197433

RESUMO

RATIONALE: The volatile anesthetic isoflurane is suggested to produce a rapid and robust antidepressive effect in preliminary clinical trials. Recently, isoflurane was found to activate the tropomyosin receptor kinase B (TrkB) signaling which is the underlying mechanism of the rapid antidepressant ketamine. OBJECTIVE: Our study investigated the effect of isoflurane anesthesia on chronic unpredictable mild stressed (CUMS) model in mice and verified the role of brain-derived neurotrophic factor (BDNF)/TrkB/ the mammalian target of rapamycin (mTOR) signaling in the antidepressant effect of isoflurane. METHODS: We employed the CUMS model of depression to assess the rapid antidepressant effect of isoflurane by the forced swimming test (FST), the sucrose preference test (SPT), and the novelty suppressed feeding test (NSFT). The protein expression of BDNF and TrkB/protein kinase B (PKB or Akt)/mTOR was determined through Western blot. The dendritic spine density in the hippocampus and medial prefrontal cortex (PFC) was measured by the Golgi staining. RESULTS: A brief burst-suppressing isoflurane anesthesia rapidly reversed the behavioral deficits caused by CUMS procedure, normalized the expression of BDNF and further activated the TrkB signaling pathway in CUMS-induced stressed mice in both prefrontal cortex (PFC) and hippocampus (HC). All of those behavioral and proteomic effects were blocked by K252a, a selective receptor inhibitor of TrkB. Isoflurane significantly promoted the formation of dendritic spines in both medial prefrontal cortex (mPFC), CA1, CA3, and DG of the hippocampus. CONCLUSION: Our study indicates that isoflurane exerts a rapid antidepressant-like effect in CUMS depression animal model, and the activation of BDNF/TrkB signaling pathway plays an indispensable role in the biological and behavioral antidepressant effects of isoflurane. A single exposure to isoflurane could repair synaptic damage caused by chronic stimulation.


Assuntos
Antidepressivos/farmacologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Depressão/metabolismo , Isoflurano/farmacologia , Glicoproteínas de Membrana/metabolismo , Proteínas Tirosina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Anestésicos Inalatórios/farmacologia , Anestésicos Inalatórios/uso terapêutico , Animais , Antidepressivos/uso terapêutico , Depressão/tratamento farmacológico , Depressão/psicologia , Relação Dose-Resposta a Droga , Isoflurano/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Transdução de Sinais/fisiologia , Estresse Psicológico/metabolismo
19.
Nat Microbiol ; 4(10): 1661-1670, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31182798

RESUMO

Chromosome segregation in bacteria is poorly understood outside some prominent model strains1-5 and even less is known about how it is coordinated with other cellular processes. This is the case for the opportunistic human pathogen Streptococcus pneumoniae (the pneumococcus)6, which lacks the Min and the nucleoid occlusion systems7, and possesses only an incomplete chromosome partitioning Par(A)BS system, in which ParA is absent8. The bacterial tyrosine kinase9 CpsD, which is required for capsule production, was previously found to interfere with chromosome segregation10. Here, we identify a protein of unknown function that interacts with CpsD and drives chromosome segregation. RocS (Regulator of Chromosome Segregation) is a membrane-bound protein that interacts with both DNA and the chromosome partitioning protein ParB to properly segregate the origin of replication region to new daughter cells. In addition, we show that RocS interacts with the cell division protein FtsZ and hinders cell division. Altogether, this work reveals that RocS is the cornerstone of a nucleoid protection system ensuring proper chromosome segregation and cell division in coordination with the biogenesis of the protective capsular layer.


Assuntos
Proteínas de Bactérias/metabolismo , Segregação de Cromossomos , Proteínas de Ligação a DNA/metabolismo , Streptococcus pneumoniae/citologia , Cápsulas Bacterianas/metabolismo , Proteínas de Bactérias/genética , Divisão Celular , Proteínas do Citoesqueleto/metabolismo , Proteínas de Ligação a DNA/genética , Deleção de Genes , Modelos Biológicos , Complexo de Reconhecimento de Origem/genética , Complexo de Reconhecimento de Origem/metabolismo , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/metabolismo
20.
Diagn Pathol ; 14(1): 62, 2019 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-31221175

RESUMO

The use of immunohistochemistry (IHC) as a companion diagnostic is an increasingly important part of the case workup by pathologists and is often central to clinical decision making. New predictive molecular markers are constantly sought for to improve treatment stratification parallel to drug development. Unfortunately, official biomarker guidelines lag behind, and pathologists are often left hesitating when medical oncologists request off-labelled biomarker testing. We performed a literature review of five commonly requested off-label IHC predictive biomarkers in gastrointestinal tract (GIT) malignancies: HER2, mismatch repair (MMR), PD-L1, BRAF V600E and ROS1. We found that HER2 amplification is rare and poorly associated to IHC overexpression in extracolonic and extragastric GIT cancers; however in KRAS wild type colorectal cancers, which fail conventional treatment, HER2 IHC may be useful and should be considered. For MMR testing, more evidence is needed to recommend reflex testing in GIT cancers for treatment purposes. MMR testing should not be discouraged in patients considered for second line checkpoint inhibitor therapy. With the exception of gastric tumors, PD-L1 IHC is a weak predictor of checkpoint inhibitor response in the GIT and should be replaced by MMR in this context. BRAF inhibitors showed activity in BRAF V600E mutated cholangiocarcinomas and pancreatic carcinomas in non-first line settings. ROS1 translocation is extremely rare and poorly correlated to ROS1 IHC expression in the GIT; currently there is no role for ROS1 IHC testing in GIT cancers. Overall, the predictive biomarker literature has grown exponentially, and official guidelines need to be updated more regularly to support pathologists' testing decisions.


Assuntos
Biomarcadores Tumorais/análise , Neoplasias Colorretais/terapia , Reparo de Erro de Pareamento de DNA/genética , Uso Off-Label , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/patologia , Feminino , Humanos , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas/análise , Proteínas Proto-Oncogênicas/genética
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