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1.
Int J Radiat Biol ; 99(12): 1961-1970, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37389464

RESUMO

PURPOSE: PI3K/Akt/mTOR pathway activation causes relapse and resistance after radiotherapy in breast cancer (BC). We aimed to radiosensitize BC cell lines to irradiation (IR) by PKI-402, a dual PI3K/mTOR inhibitor. METHODS: We performed cytotoxicity, clonogenicity, hanging drop, apoptosis and double-strand break detection, and phosphorylation of 16 essential proteins involved in the PI3K/mTOR pathway. RESULTS: Our findings showed that PKI-402 has cytotoxic efficiency in all cell lines. Clonogenic assay results showed that PKI-402 plus IR inhibited the colony formation ability of MCF-7 and breast cancer stem cell lines. Results showed that PKI-402 plus IR causes more apoptotic cell death than IR alone in the MCF-7 cells but did not cause significant changes in the MDA-MB-231. γ-H2AX levels were increased in MDA-MB-231 in PKI-402 plus IR groups, whereas we did not observe any apoptotic and γ-H2AX induction in BCSCs and MCF-10A cells in all treatment groups. Some pivotal phosphorylated proteins of the PI3K/AKT pathway decreased, several proteins increased and others did not change. CONCLUSION: In conclusion, if the combined use of PKI-402 with radiation is supported by in vivo studies, it can contribute to the treatment options and the course of the disease.


Assuntos
Neoplasias da Mama , Fosfatidilinositol 3-Quinases , Humanos , Feminino , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proliferação de Células/efeitos da radiação , Recidiva Local de Neoplasia , Serina-Treonina Quinases TOR/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Neoplasias da Mama/radioterapia , Neoplasias da Mama/patologia , Tolerância a Radiação , Apoptose , Linhagem Celular Tumoral
2.
Eur J Pharmacol ; 910: 174446, 2021 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-34461124

RESUMO

Ponatinib is used for advanced treatment of chronic myeloid leukemia (CML), although low doses to prevent side effects do not suppress survival pathways and eradicate leukemia stem cells (LSCs). We evaluated the potential of ponatinib and PI3K/mTOR dual-inhibitor VS-5584 combination (PoVS) therapy to increase the anti-leukemic effects of ponatinib and investigated the underlying mechanisms at the molecular level. We measured the cytotoxicities of ponatinib, VS-5584, and PoVS (CCK-8 assay), and used the median-effect equation for combination analyses. We investigated the effects of inhibitory concentrations on apoptosis, cell viability and cell-cycle regulation (flow cytometry), protein levels (ELISA, Western blot), transcriptional activities (dual-luciferase reporter assay), gene expressions (qRT-PCR). VS-5584 exerted selective cytotoxic effects against CML and LSC cell lines. VS-5584 inhibited the PI3K/Akt/mTOR pathway, resulting in reduced cell viability, slightly induced caspase-independent apoptosis, prominent G0/G1 cell-cycle blockade that is not a consequence of quiescence. Normal hematopoietic stem cell line was the least affected. Moreover, ponatinib and VS-5584 mediated synergistic anti-leukemic effects on leukemic cells. VS-5584 reduced the ponatinib dose required to target leukemic cells. PoVS treatment inhibited PI3K/Akt/mTOR pathway more consistently than either of the two agents alone through reducing p-Akt, p-mTOR, p-S6K, p-PRAS40, p-S6. The subsequent downstream effects were an increase in C/EBP transcriptional activity and decreases in activities of E2F/DP1, Myc/Max, CREB, STAT3, NFκB, AP-1, Elk-1/SRF. Transcriptional regulation resulted in alterations in the expression levels of target mRNAs. Our results highlight PoVS can be a promising treatment strategy for eliminating CML cells and LSCs selectively, with the reduced ponatinib doses.


Assuntos
Antineoplásicos/farmacologia , Imidazóis/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Purinas/farmacologia , Piridazinas/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , Concentração Inibidora 50 , Janus Quinases/genética , Janus Quinases/metabolismo , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fatores de Transcrição STAT/genética , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição/efeitos dos fármacos
3.
Anticancer Agents Med Chem ; 21(12): 1564-1574, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33106151

RESUMO

BACKGROUND: Dysregulation of the cell cycle is one of the main causes of melanomagenesis. Genomewide studies showed that the expression of Aurora -A and -B significantly has been upregulated in melanoma. However, there is no FDA approved drug targeting aurora kinases in the treatment of melanoma. In addition, the development of resistance to chemotherapeutic agents in the treatment of melanoma and, as a result, the relapse due to heterogeneous cell groups in patients is a second phenomenon that causes treatment failure. Therefore, there is an urgent need for therapeutic alternatives targeting both melanoma and Melanoma Cancer Stem Cells (MCSCs) in treatments. At this stage, cell cycle regulators become promising targets. OBJECTIVE: In this study, we aimed to identify the effects of Aurora kinase inhibitor CCT137690 on the cytotoxicity, apoptosis, cell cycle, migration, and colony formation and expression changes of genes related to proliferation, cell death and cell cycle in melanoma and melanoma cancer stem cell. In addition, we investigated the apoptotic and cytostatic effects of CCT137690 in normal fibroblast cells. METHODS: We evaluated the cytotoxic effect of CCT137690 in MCSCs, NM2C5 referring as melanoma model cells and WI-38 cells by using the WST-1 test. The effect of CCT137690 on apoptosis was detected via Annexin V and JC-1 method; on cell cycle progression by cell cycle test; on gene expression by using RT-PCR, on migration activity by wound healing assay and clonal growth by clonogenic assay in NM2C5 cells and MCSCs. The effects of CCT137690 in WI-38, referring as healthy fibroblast cell, were assessed through Annexin V and cell cycle method. RESULTS: CCT137690 was determined to have a cytotoxic and apoptotic effect in MCSCs and melanoma. It caused polyploidy and cell cycle arrest at the G2/M phase in MCSCs and melanoma cells. The significant decrease in the expression of MMP2, MMP7, MMP10, CCNB1, IRAK1, PLK2 genes, and the increase in the expression of PTEN, CASP7, p53 genes were detected. CONCLUSION: Aurora kinases inhibitor CCT137690 displays promising anticancer activity in melanoma and especially melanoma cancer stem cells. The effect of CCT137690 on melanoma and MCSC may provide a new approach to treatment protocols.


Assuntos
Antineoplásicos/farmacologia , Aurora Quinase A/antagonistas & inibidores , Imidazóis/farmacologia , Melanoma/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Aurora Quinase A/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Imidazóis/química , Melanoma/metabolismo , Melanoma/patologia , Inibidores de Proteínas Quinases/química , Piridinas/química , Relação Estrutura-Atividade , Células Tumorais Cultivadas
4.
J Cell Biochem ; 120(1): 380-395, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30145828

RESUMO

The effect of Wnt pathway in head and neck cancer could not be elucidated, even though the aberrant Wnt signaling plays a key role in the development of many types of cancer. The inhibitor of ß-catenin responsive transcription (ICRT-3) blocks the Wnt signaling pathway by binding to ß-catenin, which is a coactivator of the Wnt signaling pathway and a promising agent for inhibiting aberrant signaling. In our study, we aimed to evaluate the effect of ICRT-3 on the cytotoxicity, apoptosis, cell cycle progression, migration, and gene expressions in head and neck cancer stem cell (HNCSC) and hypopharynx cancer. The effect of this compound on cytotoxicity and cell viability in FaDu and HNCSC line was assessed by using the water-soluble tetrazolium salt-1 method. The effect of ICRT-3 on apoptosis was detected by using Annexin V and caspase-3, caspase-9 kit, on cell cycle progression by cycle test plus DNA reagent kit, on gene expression by dual luciferase reporter assay, and on migration activity by wound healing assay in both cell lines. ICRT-3 was determined to have cytotoxic and apoptotic effect in both cell lines. In addition, it was also found that the administration of ICRT-3 caused cell cycle arrest and significant decrease in gene expression level and migration ability of the cells.


Assuntos
Citotoxinas/farmacologia , Neoplasias de Cabeça e Pescoço/patologia , Células-Tronco Neoplásicas/metabolismo , Oxazóis/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Carcinoma de Células Escamosas/patologia , Caspase 3/metabolismo , Caspase 9/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citotoxinas/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hipofaríngeas/patologia , Concentração Inibidora 50 , Fator 1 de Ligação ao Facilitador Linfoide/metabolismo , Oxazóis/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição 4/metabolismo , beta Catenina/antagonistas & inibidores , beta Catenina/metabolismo
5.
J Cell Biochem ; 120(2): 1282-1293, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30368861

RESUMO

It is emphasized that cancer stem cells (CSCs) forming the subpopulation of tumour cells are responsible for tumour growth, metastasis, and cancer drug resistance. Inadequate response to conventional therapy in breast cancer leads researchers to find new treatment methods and literature surveys that support CSC studies. A selective anticancer agent BIBR1532 inhibits the telomerase enzyme. Many of the chemotherapeutic drugs used in clinical trials have harmful effects, but the advantage of telomerase-based inhibitors is that they are less toxic to healthy tissues. The phosphoinositide 3-kinase (PI3K)/serine/threonine kinase (Akt)/mammalian target of rapamycin (mTOR) pathway is common in breast cancer, and the interaction between the mTOR pathway and human telomerase reverse transcriptase (hTERT) is essential for the survival of cancer cells. In our study, we treated MCF-7, breast cancer stem cell (BCSC) and normal breast epithelial cell MCF10A with the BIBR1532 inhibitor. The IC 50 doses for the 48th hour of BIBR1532 treatment were detected as 34.59 µM in MCF-7, 29.91 µM in BCSCs, and 29.07 µM in MCF10A. It has been observed that this agent induces apoptosis in the BCSC and MCF-7 cell lines. According to the results of cell cycle analysis, G 2 /M phase accumulation was observed in BCSC and MCF-7 cell lines. It has also been shown that BIBR1532 suppresses telomerase activity in BCSC and MCF-7. The effect of BIBR1532 on the mTOR signalling pathway has been investigated for the first time in this study. It is thought that the telomerase inhibitor may bring a new approach to the treatment and it may be useful in the treatment of CSCs.

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