RESUMO
ETS variant transcription factor 6 (ETV6)-neurotrophic receptor tyrosine kinase 3 (NTRK3) (EN) fusions are typically found in rare diseases, such as primary renal fibrosarcoma (only six cases have been reported), secretory carcinoma of the breast and salivary gland (1 case), and AML (4 cases). Few cases have been reported, and expression of the EN gene fusion requires additional clinical data and fundamental research to be supported. The aim of the present study was to determine the inhibitory effect of Andrographis paniculata methanol extract (MeAP) on EN-related cell lines, IMS-M2 and BaF3/EN, as well as evaluate the mechanism of action. Vero cells were used as control cells. Trypan blue staining and MTT were used to evaluate the inhibitory effect of MeAP on tested cells. Western blotting and immunoprecipitation were used to detect the activation of EN after MeAP treatment. The IC50 values of MeAP were found to be 12.38±0.57 µg/ml (IMS-M2) and 13.06±0.49 µg/ml (BaF3/EN). MeAP was observed to inhibit cell proliferation in a time, dose, and cell density-dependent manner. The IC50 value for MeAP in Vero cells was markedly higher, at 109.97±4.24 (µg/ml), indicating a much less sensitive effect. Furthermore, MeAP treatment inhibited EN phosphorylation and induced apoptosis in these cells. Collectively, the present study demonstrated that MeAP has an oncogenic effect on EN fusion-positive cell lines, in particular.
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In recent years, smart lockers have been increasingly utilised in the last-mile delivery stage of e-commerce. Although this technological advance has received significant scientific attention, especially in developed countries, little is known about the general behaviour of Smart Locker users. This study aimed to explore the impact of smart locker use on customer satisfaction with online shopping experience. We used structural equation modelling to analyse data collected from 442 smart locker users in Vietnam. The results revealed that convenience, privacy and security, and reliability attributes significantly impacted the satisfaction of smart locker users through the mediators of perceived value and transaction costs. These findings are expected to reinforce the utilised theories and enrich theoretical research on self-service technologies in last-mile delivery and their influence on consumer behaviour.
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In previous research, the authors demonstrated that the methanol extract of Artemisia vulgaris (AVM) has the ability to inhibit chronic myeloid leukemia (CML) cell proliferation. The aim of the present study was to determine and clarify the mechanism of action of AVM. BCR/ABL activation is present in >90% of CML cases. As a result, cells expressing different forms of BCR/ABL were recruited for the present study, including K562 (human wild-type) or TCCY-T315I (human imatinib-resistant) and the Ba/F3-(T315I/E279K/Y253H) (mouse BCR/ABL point mutation-transfected cells). The results revealed that AVM inhibited the phosphorylation of BCR/ABL and their subsequent molecular signals including AKT and MAPK activation. AVM induced the release of cleaved PARP and cleaved caspase-3 caused apoptosis and inhibited the viability of these cells. Interestingly, AVM appeared to be more sensitive to imatinib-resistant (T315I, Y253H, and E279K) than wild-type BCR/ABL cells, indicating its potential to overcome imatinib-resistant severe issues in CML. Moreover, the effects of various sub-fractions of AVM were then investigated in order to determine the optimal solvent for the identification of anticancer bioactive compounds. The results demonstrated that the ethyl acetate and chloroform fractions were more effective than the n-hexane and water fractions. It is thus concluded that AVM inhibits the activity of BCR/ABL and their subsequent molecular signals, including AKT and MAPK, resulting in cytotoxicity via apoptosis.
RESUMO
We will study the effects of the methanol extract of Sphagneticola trilobata (L.) Pruski (Asteraceae) (MeST) on the growth of leukemia cells that may contain the BCR/ABL gene. This study also clarifies the mechanism of this effect on these cells. For this purpose, the cells harboring wild-type BCR/ABL, imatinib-resistant BCR/ABL (K562 and TCCYT315I), or Ba/F3 cells transfected with wild-type or mutant BCR/ABL genes were used. The results showed that MeST effectively inhibited the viability of leukemia cells in both a dose- and time-dependent manner. The effect of MeST seems to be more sensitive in the cells that carry imatinib-resistant BCR/ABL (especially the T315I BCR/ABL mutation) than those with wild-type BCR/ABL. Furthermore, we have demonstrated that the death caused by MeST is apoptosis and the treatment with MeST could suppress the expression of BCR/ABL, subsequently altering the downstream cascade of BCR/ABL such as AKT and MAPK signaling. In conclusion, MeST has been able to suppress the growth of leukemia cells harboring BCR/ABL. The mechanism of the anti-leukemic effect of MeST on cells harboring imatinib-resistant BCR/ABL mutations could be due to the disruption of the BCR/ABL oncoprotein signaling cascade.
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Patents diagnosed with acute promyelocytic leukemia were treated with Vesanoid® [all-trans retinoic acid (ATRA)]. ATRA promotes the maturation and differentiation of leukemia cells and is therefore capable of reducing the symptoms of leukemia by preventing aggregation of myeloid cells. However, the clinical applications of ATRA are limited by its side effects, including acute retinoid resistance, hypertriglyceridemia, mucocutaneous dryness, nausea, brief recovery time relapse and drug resistance. Therefore, combinations of ATRA and other anticancer drugs are being investigated to overcome these limitations. In our previous study it was shown that in leukemia cells, (-)-epigallocatechin-3-gallate (EGCG) reduced cell proliferation and induced apoptotic cell death. In the present study, an in vitro evaluation of the effects of the combination of EGCG and ATRA on FLT3-mutated cell lines was performed using the isobologram method. The results showed that there was an additive effect in leukemic cells when treated with a combination of ATRA and EGCG. Thus, it was concluded that the cytotoxic effects of EGCG were improved by ATRA.
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FMS-like tyrosine kinase-3 fragments from exon 14 to the end without any mutations or deletions have been reported to fuse to ETV6 (TEL) in a few cases of myeloid/lymphoid neoplasms with eosinophilia carrying a translocation t(12;13)(p13;q12). This fusion protein confers constitutive activation on the FLT3 fragment and induces factor-independent growth in transfected Ba/F3 cells, indicating that it is an oncoprotein. However, the mechanism controlling the stability of this oncoprotein is unknown. In this study, we focus on finding factors controlling the stability of ETV6/FLT3. We have shown that the stability of ETV6/FLT3 is regulated by the Hsp90 chaperone. ETV6/FLT3 fusion protein forms a complex with Hsp90 by coimmunoprecipitation analyses using an Hsp90 antibody. The association between ETV6/FLT3 fusion protein and Hsp90 was impaired after treating ETV6/FLT3 transient transfection cos7 cells with 17-allylamino-17-demethoxygeldanamycin (17-AAG). 17-AAG induced a time- and dose-dependent downregulation of ectopically expressed ETV6/FLT3 protein in cos7 and HeLa-transfected cells. By using cycloheximide to block new protein translation, we found that 17-AAG accelerated the decay of ETV6/FLT3. Our findings could contribute to more understanding of the ETV6/FLT3 regulation through Hsp90 chaperone and open the way to finding effective treatment strategies for this rare disease.
Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas c-ets/metabolismo , Proteínas Repressoras/metabolismo , Tirosina Quinase 3 Semelhante a fms/genética , Animais , Benzoquinonas/farmacologia , Células COS , Catequina/análogos & derivados , Catequina/farmacologia , Chlorocebus aethiops , Células HEK293 , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Células HeLa , Humanos , Lactamas Macrocíclicas/farmacologia , Proteínas de Fusão Oncogênica/antagonistas & inibidores , Proteínas de Fusão Oncogênica/biossíntese , Proteínas de Fusão Oncogênica/genética , Fosforilação , Proteínas Proto-Oncogênicas c-ets/genética , Proteínas Repressoras/genética , Transdução de Sinais , Transfecção , Tirosina Quinase 3 Semelhante a fms/metabolismo , Variante 6 da Proteína do Fator de Translocação ETSRESUMO
Acute myeloid leukemia (AML) is a heterogeneous disease. Numerous molecular abnormalities have been identified in AML and, amongst these, FMSlike tyrosine kinase 3 (FLT3) mutations are one of the most common somatic alterations detected. In the present study, an in vitro investigation was performed to evaluate the effects of alltrans retinoic acid (ATRA) and PKC412, alone and in combination, in FLT3mutated AML cell lines. Trypan blue exclusion test, as well as morphological, western blot and isobologram analyses were conducted. The results indicated that the combined ATRA and PKC412 treatment exhibited additive or synergistic effects in FLT3mutated AML cell lines. These results provided in vitro evidence for the future clinical trials evaluating the effects of a combination treatment using PKC412 and ATRA on AML patients with FLT3mutations.
Assuntos
Antineoplásicos/farmacologia , Leucemia Mieloide Aguda/genética , Mutação , Inibidores de Proteínas Quinases/farmacologia , Estaurosporina/análogos & derivados , Tretinoína/farmacologia , Tirosina Quinase 3 Semelhante a fms/genética , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Estaurosporina/farmacologiaRESUMO
OBJECTIVE: To investigate the inhibited effect of epigallocatechin-3-gallate (EGCG) on the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations. METHODS: Cell proliferation assay was performed to test the effects of EGCG on cell growth of IMS-M2 cells harboring the NPM1 mutations. Western blot analysis were performed to test the protein expression of NPM1, AKT, those associated with apoptosis. RESULTS: EGCG can down-regulate the expression of NPM1 in IMS-M2 cells harboring the NPM1 mutations. Moreover, EGCG also suppressed the cell proliferation and induced apoptosis in IMS-M2 cells. CONCLUSIONS: The results suggested that EGCG could be considered as a reagent for treatment of AML patients with NPM1 mutations.
RESUMO
Acute myeloid leukemia (AML) is a heterogeneous disease characterized by a block in differentiation and uncontrolled proliferation. FLT3 is a commonly mutated gene found in AML patients. In clinical trials, the presence of a FLT3-ITD mutation significantly correlates with an increased risk of relapse and dismal overall survival. Therefore, activated FLT3 is a promising molecular target for AML therapies. In this study, we have shown that green tea polyphenols including (-)-epigallocatechin-3-gallate (EGCG), (-)-epigallocatechin (EGC), and (-)-epicatechin-3-gallate (ECG) suppress the proliferation of AML cells. Interestingly, EGCG, EGC and ECG showed the inhibition of FLT3 expression in cell lines harboring FLT3 mutations. In the THP-1 cells harboring FLT3 wild-type, EGCG showed the suppression of cell proliferation but did not suppress the expression of FLT3 even at the concentration that suppress 100% cell proliferation. Moreover, EGCG-, EGC-and ECG-treated cells showed the suppression of MAPK, AKT and STAT5 phosphorylation. Altogether, we suggest that green tea polyphenols could serve as reagents for treatment or prevention of leukemia harboring FLT3 mutations.
Assuntos
Antineoplásicos/farmacologia , Catequina/análogos & derivados , Leucemia Mieloide Aguda/tratamento farmacológico , Tirosina Quinase 3 Semelhante a fms/genética , Catequina/farmacologia , Linhagem Celular Tumoral/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Lactente , Leucemia Mieloide Aguda/genética , Masculino , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Mutagênese Insercional , Fosforilação , Mutação Puntual , Processamento de Proteína Pós-Traducional , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT5/metabolismo , Estaurosporina/análogos & derivados , Estaurosporina/farmacologia , Tirosina Quinase 3 Semelhante a fms/metabolismoRESUMO
The ETV6-NTRK3 (EN) fusion gene which encodes a chimeric tyrosine kinase was first identified by cloning of the t(12;15)(p13;q25) translocation in congenital fibrosarcoma (CFS). Since then, EN has been also found in congenital mesoblastic nephroma (CMN), secretory breast carcinoma (SBC) and acute myelogenous leukemia (AML). Using IMS-M2 and M0-91 cell lines harboring the EN fusion gene, and Ba/F3 cells stably transfected with EN, we demonstrated that PKC412, also known as midostaurin, is an inhibitor of EN. Inhibition of EN activity by PKC412 suppressed the activity of it downstream molecules leading to inhibition of cell proliferation and induction of apoptosis. Our data for the first time suggested that PKC412 could serve as therapeutic drug for treatment of patients with this fusion.