RESUMO
Despite many efforts, a comprehensive understanding and clarification of the intricate connections within cancer cell metabolism remain elusive. This might pertain to intracellular dynamics and the complex interplay between cancer cells, and cells with the tumor stroma. Almost a century ago, Otto Warburg found that cancer cells exhibit a glycolytic phenotype, which continues to be a subject of thorough investigation. Past and ongoing investigations have demonstrated intricate mechanisms by which tumors modulate their functionality by utilizing extracellular glucose as a substrate, thereby sustaining the essential proliferation of cancer cells. This concept of "aerobic glycolysis," where cancer cells (even in the presence of enough oxygen) metabolize glucose to produce lactate plays a critical role in cancer progression and is regulated by various signaling pathways. Recent research has revealed that the canonical wingless-related integrated site (WNT) pathway promotes aerobic glycolysis, directly and indirectly, thereby influencing cancer development and progression. The present review seeks to gather knowledge about how the WNT/ß-catenin pathway influences aerobic glycolysis, referring to relevant studies in different types of cancer. Furthermore, we propose the concept of impeding the glycolytic phenotype of tumors by employing specific inhibitors that target WNT/ß-catenin signaling.
Assuntos
Glicólise , Neoplasias , Via de Sinalização Wnt , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Neoplasias/genética , beta Catenina/metabolismo , Efeito Warburg em Oncologia , Animais , Glucose/metabolismoRESUMO
Cancer cell lines play a crucial role as invaluable models in cancer research, facilitating the examination of cancer progression as well as the advancement of diagnostics and treatments. While they may not perfectly replicate the original tumor, they generally exhibit similar characteristics. Low-passage cancer cell lines are generally preferred due to their closer resemblance to the original tumor, as long-term culturing can alter the genetic and molecular profiles of a cell line thereby highlighting the importance of monitoring the passage number (PN). Variations in proliferation, migration, gene expression, and drug sensitivity can be linked to PN differences. PN can also influence DNA methylation levels, metabolic profiles, and the expression of genes/or proteins in cancer cell lines. When conducting research on cancer cell lines, it is crucial for researchers to carefully select the appropriate PN to maintain consistency and reliability of results. Moreover, to ensure dependability and replicability, scientists ought to actively track the growth, migration, and gene/or protein profiles of cancer cell lines at specific PNs. This approach enables the identification of the most suitable range of PNs for experiments, guaranteeing consistent and precise results. Additionally, such efforts serve to minimize disparities and uphold the integrity of research. In this review, we have laid out recommendations for laboratories to overcome these PN discrepancies when working with cancer cell lines.
Assuntos
Neoplasias , Humanos , Reprodutibilidade dos Testes , Neoplasias/genética , Linhagem Celular , Metilação de DNA/genética , Linhagem Celular TumoralRESUMO
Among breast cancer subtypes, the triple negative breast cancer (TNBC) has the worst prognosis. In absence of any permitted targeted therapy, standard chemotherapy is the mainstay for TNBC treatment. Hence, there is a crucial need to identify potential druggable targets in TNBCs for its effective treatment. In recent times, metabolic reprogramming has emerged as cancer cells hallmark, wherein cancer cells display discrete metabolic phenotypes to fuel cell progression and metastasis. Altered glycolysis is one such phenotype, in which even in oxygen abundance majority of cancer cells harvest considerable amount of energy through elevated glycolytic-flux. In the present review, we attempt to summarize the role of key glycolytic enzymes i.e. HK, Hexokinase; PFK, Phosphofructokinase; PKM2, Pyruvate kinase isozyme type 2; and LDH, Lactate dehydrogenase in TNBCs, and possible therapeutic options presently available.
Assuntos
Neoplasias de Mama Triplo Negativas/enzimologia , Neoplasias de Mama Triplo Negativas/genética , Efeito Warburg em Oncologia , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Hexoquinase/genética , Hexoquinase/metabolismo , Humanos , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fosfofrutoquinases/genética , Fosfofrutoquinases/metabolismo , Piruvato Quinase/genética , Piruvato Quinase/metabolismo , Hormônios Tireóideos/genética , Hormônios Tireóideos/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Proteínas de Ligação a Hormônio da TireoideRESUMO
Despite the clinical development of novel adjuvant and neoadjuvant chemotherapeutic drugs, metastatic breast cancer is one of the leading causes of cancer-related death among women. The present review focuses on the relevance, mechanisms, and therapeutic potential of targeting WNT5A as a future anti-metastatic treatment strategy for breast cancer patients by restoring WNT5A signaling as an innovative therapeutic option. WNT5A is an auto- and paracrine ß-catenin-independent ligand that has been shown to induce tumor suppression as well as oncogenic signaling, depending upon cancer type. In breast cancer patients, WNT5A protein expression has been observed to be significantly reduced in between 45 and 75% of the cases and associated with early relapse and reduced disease-free survival. WNT5A triggers various downstream signaling pathways in breast cancer that primarily affect tumor cell migration and invasion. The accumulated in vitro results reveal that treatment of WNT5A-negative breast cancer cells with recombinant WNT5A caused different tumor-suppressive responses and in particular it impaired migration and invasion. The anti-migratory/invasive and anti-metastatic effects of reconstituting WNT5A signaling by the small WNT5A mimicking peptide Foxy5 form the basis for two successful clinical phase 1-studies aiming at determining safety and pharmacokinetics as well as defining dose-level for a subsequent phase 2-study. We conclude that re-installation of WNT5A signaling is an attractive and promising anti-metastatic therapeutic approach for future treatment of WNT5A-negative breast cancer patients.
Assuntos
Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Proteína Wnt-5a/metabolismo , Animais , Materiais Biomiméticos/farmacologia , Neoplasias da Mama/patologia , Humanos , Terapia de Alvo Molecular , Invasividade Neoplásica , Oligopeptídeos/farmacologia , Proteínas Recombinantes/farmacologia , Transdução de Sinais , Proteína Wnt-5a/farmacologiaRESUMO
Cigarette smoking is a key factor for the development and progression of different cancers including mammary tumor in women. Resveratrol (Res) is a promising natural chemotherapeutic agent that regulates many cellular targets including p21, a cip/kip family of cyclin kinase inhibitors involved in DNA damage-induced cell cycle arrest and blocking of DNA replication and repair. We have recently shown that cigarette smoke condensate (CSC) prepared from commercially available Indian cigarette can cause neoplastic transformation of normal breast epithelial MCF-10A cell. Here we studied the mechanism of Res mediated apoptosis in CSC transformed (MCF-10A-Tr) cells in vitro and in vivo. Res mediated apoptosis in MCF-10A-Tr cells was a p21 dependent event. It increased the p21 protein expression in MCF-10A-Tr cells and MCF-10A-Tr cells-mediated tumors in xenograft mice. Res treatment reduced the tumor size(s) and expression of anti-apoptotic proteins (e.g. PI3K, AKT, NFκB) in solid tumor. The expressions of cell cycle regulatory (Cyclins, CDC-2, CDC-6, etc.), BER associated (Pol-ß, Pol-δ, Pol-ε, Pol-η, RPA, Fen-1, DNA-Ligase-I, etc.) proteins and LP-BER activity decreased in MCF-10A-Tr cells but remain significantly unaltered in isogenic p21 null MCF-10A-Tr cells after Res treatment. Interestingly, no significant changes were noted in SP-BER activity in both the cell lines after Res exposure. Finally, it was observed that increased p21 blocks the LP-BER in MCF-10A-Tr cells by increasing its interaction with PCNA via competing with Fen-1 after Res treatment. Thus, Res caused apoptosis in CSC-induced cancer cells by reduction of LP-BER activity and this phenomenon largely depends on p21.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Transformação Celular Neoplásica/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Reparo do DNA/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Glândulas Mamárias Humanas/efeitos dos fármacos , Fumaça/efeitos adversos , Fumar/efeitos adversos , Estilbenos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Transformada , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Inibidor de Quinase Dependente de Ciclina p21/genética , Dano ao DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Endonucleases Flap/metabolismo , Humanos , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Antígeno Nuclear de Célula em Proliferação/metabolismo , Interferência de RNA , Resveratrol , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Transfecção , Regulação para Cima , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cigarette smoking is a crucial factor in the development and progression of multiple cancers including breast. Here, we report that repeated exposure to a fixed, low dose of cigarette smoke condensate (CSC) prepared from Indian cigarettes is capable of transforming normal breast epithelial cells, MCF-10A, and delineate the biochemical basis for cellular transformation. CSC transformed cells (MCF-10A-Tr) were capable of anchorage-independent growth, and their anchorage dependent growth and colony forming ability were higher compared to the non-transformed MCF-10A cells. Increased expression of biomarkers representative of oncogenic transformation (NRP-1, Nectin-4), and anti-apoptotic markers (PI3K, AKT, NFκB) were also noted in the MCF-10A-Tr cells. Short tandem repeat (STR) profiling of MCF-10A and MCF-10A-Tr cells revealed that transformed cells acquired allelic variation during transformation, and had become genetically distinct. MCF-10A-Tr cells formed solid tumors when implanted into the mammary fat pads of Balb/c mice. Data revealed that CSC contained approximately 1.011µg Cd per cigarette equivalent, and Cd (0.0003µg Cd/1×10(7) cells) was also detected in the lysates from MCF-10A cells treated with 25µg/mL CSC. In similar manner to CSC, CdCl2 treatment in MCF-10A cells caused anchorage independent colony growth, higher expression of oncogenic proteins and increased PI3K-AKT-NFκB protein expression. An increase in the expression of PI3K-AKT-NFκB was also noted in the mice xenografts. Interestingly, it was noted that CSC and CdCl2 treatment in MCF-10A cells increased ROS. Collectively, results suggest that heavy metals present in cigarettes of Indian origin may substantially contribute to tumorigenesis by inducing intercellular ROS accumulation and increased expression of PI3K, AKT and NFκB proteins.
Assuntos
Transformação Celular Neoplásica/induzido quimicamente , Células Epiteliais/efeitos dos fármacos , Metais Pesados/toxicidade , NF-kappa B/biossíntese , Fosfatidilinositol 3-Quinase/biossíntese , Proteínas Proto-Oncogênicas c-akt/biossíntese , Fumaça/efeitos adversos , Animais , Mama/efeitos dos fármacos , Mama/metabolismo , Mama/patologia , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Transformada , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Fumar/efeitos adversos , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Per-and polyfluoroalkyl substances (PFAS) are synthetic chemicals that are known for their environmental persistence and adverse health effects. This study comprehensively assessed PFAS contamination in the Kamrup region of Assam, India, focusing on its presence in groundwater and associated health risks. The analysis detected 12 PFAS in groundwater samples from both the Kamrup Metro and Rural regions. In Kamrup Rural, Perfluorohexanoic acid (PFHxA), perfluorononanoic acid (PFNA), and perfluorooctanesulfonic acid (PFOS) were prevalent, whereas in Kamrup Metro, PFNA and PFOS were dominant, based on detection frequencies. These findings are noteworthy, as they demonstrate the widespread presence of PFAS in groundwater, a vital source of drinking water in the region. The assessment of PFAS health risks in India involved hazard quotient calculations for different age groups. Perfluorobutanesulfonic acid (PFBS) posed the highest risk, ranking children > boys > men > girls > women. Overall, ∑PFAS had low hazard (HQ: 0.27-0.41). Further, this study assessed PFBS and PFOS toxicity in human kidney epithelial cell lines (HEK293T) cells, revealing that PFBS was more cytotoxic than PFOS. The study examined the metabolomics of HEK293T cells after PFBS exposure, revealing significant alterations in lipid metabolism, particularly glycerophospholipids, potentially affecting cellular function and health. These findings underscore the importance of monitoring PFAS contamination in drinking water sources, especially in regions such as Kamrup, where groundwater is a primary source. Our metabolomics results show significant health effects at the cellular level, raising concerns about the impact of PFAS exposure on human health. This study highlights PFAS contamination in Kamrup, Assam's groundwater and its health risks, providing valuable insights for policymakers and public health management.
RESUMO
We previously reported that quinacrine (QC) has anticancer activity against breast cancer cells. Here, we examine the mechanism of action of QC and its ability to inhibit Wnt-TCF signaling in two independent breast cancer cell lines. QC altered Wnt-TCF signaling components by increasing the levels of adenomatous polyposis coli (APC), DAB2, GSK-3ß and axin and decreasing the levels of ß-catenin, p-GSK3ß (ser 9) and CK1. QC also reduced the activity of the Wnt transcription factor TCF/LEF and its downstream targets cyclin D1 and c-MYC. Using a luciferase-based Wnt-TCF transcription factor assay, it was shown that APC levels were inversely associated with TCF/LEF activity. Induction of apoptosis and DNA damage was observed after treatment with QC, which was associated with increased expression of APC. The effects induced by QC depend on APC because the inhibition of Wnt-TCF signaling by QC is lost in APC-knockdown cells, and consequently, the extent of apoptosis and DNA damage caused by QC is reduced compared with parental cells. Because we previously showed that QC inhibits topoisomerase, we examined the effect of another topoisomerase inhibitor, etoposide, on Wnt signaling. Interestingly, etoposide treatment also reduced TCF/LEF activity, ß-catenin and cyclin D1 levels commensurate with induction of DNA damage and apoptosis. Lycopene, a plant-derived antioxidant, synergistically increased QC activity and inhibited Wnt-TCF signaling in cancer cells without affecting the MCF-10A normal breast cell line. Collectively, the data suggest that QC-mediated Wnt-TCF signal inhibition depends on APC and that the addition of lycopene synergistically increases QC anticancer activity.
Assuntos
Proteína da Polipose Adenomatosa do Colo/metabolismo , Neoplasias da Mama/tratamento farmacológico , Carotenoides/farmacologia , Quinacrina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Fator 1 de Transcrição de Linfócitos T/antagonistas & inibidores , Proteínas Wnt/antagonistas & inibidores , Proteína da Polipose Adenomatosa do Colo/antagonistas & inibidores , Proteína da Polipose Adenomatosa do Colo/genética , Anticarcinógenos/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Apoptose , Western Blotting , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclo Celular , Proliferação de Células , Ensaio Cometa , Ciclina D1/metabolismo , Sinergismo Farmacológico , Etoposídeo/farmacologia , Feminino , Citometria de Fluxo , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Humanos , Licopeno , Regiões Promotoras Genéticas , RNA Interferente Pequeno/genética , Fator 1 de Transcrição de Linfócitos T/metabolismo , Fatores de Transcrição TCF , Transativadores/metabolismo , Transcrição Gênica , Células Tumorais Cultivadas , Ensaio Tumoral de Célula-Tronco , Proteínas Wnt/metabolismo , beta Catenina/metabolismoRESUMO
Based on known heterocyclic topoisomerase II inhibitors and anticancer agents, various indenoindolone derivatives were predicted as potential topoisomerase II-inhibiting anticancer agents. They are hydrazones, (thio)semicarbazones, and oximes of indenoindolones, and indenoindolols. These derivatives with suitable substitutions exhibited potent specific inhibition of human DNA TopoIIα while not showing inhibition of topoisomerase I and DNA intercalation, despite the fact that parent indenoindolones are known poor/moderate inhibitors of topoisomerase II. The potent topoisomerase II inhibitor indenoindolone derivatives exhibited good anticancer activities compared to etoposide and 5-fluorouracil, and relatively low toxicity to normal cells. These derivatizations of indenoindolones were found to result in enhancement of anticancer activities.
Assuntos
Antineoplásicos/farmacologia , Indóis/química , Indóis/farmacologia , Neoplasias/tratamento farmacológico , Inibidores da Topoisomerase II/farmacologia , Antineoplásicos/química , Antineoplásicos/uso terapêutico , DNA Topoisomerases Tipo II/química , DNA Topoisomerases Tipo II/metabolismo , Humanos , Indóis/uso terapêutico , Modelos Moleculares , Neoplasias/enzimologia , Inibidores da Topoisomerase II/química , Inibidores da Topoisomerase II/uso terapêuticoRESUMO
Integrin-mediated cellular delivery was attempted to optimize practical applications of hydrophobic ionophores. The potent ionophore preferentially transports H+/Cl- across the lipid bilayers following a symport mechanism. The RGD-peptide-appended tag was stimulated by glutathione to generate the active ionophore, prompting the transport of Cl- under the cellular environment.
Assuntos
Bicamadas Lipídicas , Oligopeptídeos , Ionóforos/química , Transporte de Íons , Bicamadas Lipídicas/química , Transporte Biológico , Oligopeptídeos/químicaRESUMO
Light-based three-dimensional (3D) printing has been under use extensively to fabricate complex geometrical constructs which find a vast application in the fields of drug delivery and tissue engineering fields due to its ability to recapitulate the intricate biological architecture and thus provides avenues to achieve previously unachievable biomedical devices. The inherent problem associated with light-based 3D printing (from a biomedical perspective) is that of light scattering causing inaccurate and defective prints which results in erroneous drug loading in 3D printed dosage forms and can also render the environment of the polymers toxic for the biological cells and tissues. In this regard, an innovative additive comprising of a nature-derived drug-cum-photoabsorber (curcumin) entrapped in naturally derived protein (bovine serum albumin) is envisaged to act as a photoabsorbing system that can improve the printing quality of 3D printed drug delivery formulations (macroporous pills) as well as provide stimuli-responsive release of the same upon oral ingestion. The delivery system was designed to endure the chemically and mechanically hostile gastric environment and deliver the drug in the small intestine to improve absorption. A 3 × 3 grid macroporous pill was designed (specifically to withstand the mechanically hostile gastric environment) and 3D printed using Stereolithography comprising of a resin system including acrylic Acid, PEGDA and PEG 400 along with curcumin loaded BSA nanoparticles (Cu-BSA NPs) as a multifunctional additive and TPO as the photoinitiator. The 3D printed macroporous pills were found to show excellent fidelity to CAD design as evident from the resolution studies. The mechanical performance of the macroporous pills was found to be extremely superior to monolithic pills. The pills found to release curcumin in pH responsive manner with slower release at acidic pH but faster release at intestinal pH due to its similar swelling behavior. Finally, the pills were found to be cytocompatible to mammalian kidney and colon cell lines.
Assuntos
Curcumina , Nanopartículas , Curcumina/farmacologia , Curcumina/uso terapêutico , Impressão Tridimensional , Estereolitografia , PolímerosRESUMO
The small molecule Quinacrine (QC, a derivative of 9-aminoacridine), an anti-malaria drug, displays activity against cancer cell lines and can simultaneously suppress nuclear factor-κB (NF-κB) and activate p53 signaling. In this study, we investigated the anticancer mechanism underlying these drug activities in breast cancer cell lines. QC caused a dose-dependent decrease of both anchorage dependent and independent growth of breast cancer cells (MCF-7 and MDA-MB-231) without affecting normal breast epithelial cells (MCF-10A), as evident from clonogenic cell survival, [3-(4,5-dimethylthiazol-2yl-)-2,5-diphenyl tetrazolium bromide] viability, wound healing and soft agar growth. QC activated the proapoptotic marker Bax, PARP cleavage, p53 and its downstream target, p21 (Cip1/Waf1) and downregulated the antiapoptotic marker Bcl-xL and relative luciferase activity of NF-κB in MCF-7 cells. Results of DAPI nuclear staining and FACS analysis show that QC increased apoptosis in a dose-dependent manner. QC caused apoptosis by increasing the cell population in S-phase and simultaneously decreasing the G1 and G2/M populations. A dose-dependent increase of DNA damage as measured by the comet assay was seen in MCF-7 cells after exposure to QC. With regards to the mechanism of DNA damage, we found that QC inhibited topoisomerase activity in MCF-7 cells by increasing the unwinding of supercoiled DNA. Collectively, the results demonstrate that QC has efficient anticancer potential against breast cancer cells via not only an induction of p53 and p21 but also an induction of S phase arrest, DNA damage and inhibition of topoisomerase activity.
Assuntos
Anticarcinógenos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/enzimologia , DNA Topoisomerases/metabolismo , Quinacrina/farmacologia , Inibidores da Topoisomerase/farmacologia , Adulto , Apoptose/efeitos dos fármacos , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Dano ao DNA/efeitos dos fármacos , DNA Super-Helicoidal/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/enzimologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , NF-kappa B/metabolismo , Poli(ADP-Ribose) Polimerases/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteína X Associada a bcl-2/metabolismoRESUMO
We previously showed that quinacrine (QC), a small molecule antimalarial agent, also presented anticancer activity in breast cancer cells through activation of p53, p21, and inhibition of topoisomerase activity. Here we have systematically studied the detailed cell death mechanism of this drug using three colon cancer cell lines (HCT-116 parental, isogenic HCT-116 p53-/-, and HCT-116 p21-/- sublines). QC caused a dose-dependent reduction in cell viability in all three cell lines. However, the parental cells were more susceptible to QC-mediated cell death, suggesting that p53- and p21-dependent processes were involved. QC-mediated cell death was measured with the following endpoints: the Bax/Bcl-xL ratio, cleaved PARP, apoptotic nuclei visualized by DAPI staining, and COMET formation. In addition, markers of autophagy were measured. Acridine orange staining revealed increased accumulation of autophagic vacuoles (AVs) after QC treatment in a dose-dependent manner in parental cells, and decreased staining in isogenic HCT-116 p53-/- and HCT-116 p21-/- cells. Immunofluorescence of LC3B was significantly lowered in QC-treated cells lacking p53 or p21, compared to the parental cells. Interestingly, the expression of the autophagy marker LC3B-II after exposure to QC was decreased in either p53 or p21 null cells compared to parental cells. After deletion of p21 in HCT-116 p53-/- cells, no change in LC3B-II expression was noted following QC treatment. Collectively, the results suggest that QC-mediated autophagy and apoptosis dependent on p53 and p21.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Quinacrina/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Western Blotting , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Humanos , Células Tumorais CultivadasRESUMO
Scaffold hybridization of several natural and synthetic anticancer leads led to the consideration of indenoindolones as potential novel anticancer agents. A series of these compounds were prepared by a diversity-feasible synthetic method. They were found to possess anticancer activities with higher potency compared to etoposide and 5-fluorouracil in kidney cancer cells (HEK 293) and low toxicity to corresponding normal cells (Vero). They exerted apoptotic effect with blocking of cell cycle at G2/M phase.
Assuntos
Antineoplásicos/síntese química , Apoptose/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Indenos/síntese química , Indóis/síntese química , Animais , Antineoplásicos/farmacologia , Biomarcadores/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Etoposídeo/farmacologia , Citometria de Fluxo , Fluoruracila/farmacologia , Células HEK293 , Humanos , Indenos/farmacologia , Indóis/farmacologia , Concentração Inibidora 50 , Células VeroRESUMO
Treatment of melanoma with a BRAF inhibitor (BRAFi) frequently initiates development of BRAFi resistance, leading to increased tumor progression and metastasis. Previously, we showed that combined inhibition of elevated WNT5A and IL-6 signaling reduced the invasion and migration of BRAFi-resistant (BRAFi-R) melanoma cells. However, the use of a combined approach per se and the need for high inhibitor concentrations to achieve this effect indicate a need for an alternative and single target. One such target could be myristoylated alanine-rich C-kinase substrate (MARCKS), a downstream target of WNT5A in BRAFi-sensitive melanoma cells. Our results revealed that MARCKS protein expression and activity are significantly elevated in PLX4032 and PLX4720 BRAFi-R A375 and HTB63 melanoma cells. Surprisingly, neither WNT5A nor IL-6 contributed to the increases in MARCKS expression and activity in BRAFi-R melanoma cells, unlike in BRAFi-sensitive melanoma cells. However, despite the above findings, our functional validation experiments revealed that MARCKS is essential for the increased metastatic behavior of BRAFi-R melanoma cells. Knockdown of MARCKS in BRAFi-R melanoma cells caused reductions in the F-actin content and the number of filopodia-like protrusions, explaining the impaired migration, invasion and metastasis of these cells observed in vitro and in an in vivo zebrafish model. In our search for an alternative explanation for the increased activity of MARCKS in BRAFi-R melanoma cells, we found elevated basal activities of PKCα, PKCε, PKCι, and RhoA. Interestingly, combined inhibition of basal PKC and RhoA effectively impaired MARCKS activity in BRAFi-R melanoma cells. Our results reveal that MARCKS is an attractive single antimetastatic target in BRAFi-R melanoma cells.
RESUMO
Oxidative damage and accumulation of extracellular matrix (ECM) components play a crucial role in the adverse outcome of cardiac hypertrophy. Evidence suggests that nuclear factor erythroid-derived factor 2 related factor 2 (Nrf2) can modulate oxidative damage and adverse myocardial remodeling. Daphnetin (Daph) is a coumarin obtained from the plant genus Daphne species that exerts anti-oxidative and anti-inflammatory properties. Herein, we investigated the roles of Daph in transverse aortic constriction (TAC)-induced cardiac hypertrophy and fibrosis in mice. TAC-induced alterations in cardiac hypertrophy markers, histopathological changes, and cardiac function were markedly ameliorated by oral administration of Daph in mice. We found that Daph significantly reduced the reactive oxygen species (ROS) generation, increased the nuclear translocation of Nrf2, and consequently, reinstated the protein levels of NAD(P)H quinone dehydrogenase1 (NQO1), heme oxygenase-1 (HO-1), and other antioxidants in the heart. Besides, Daph significantly inhibited the TAC-induced accumulation of ECM components, including α-smooth muscle actin (α-SMA), collagen I, collagen III, and fibronectin, and interfered with the TGF-ß1/Smad2/3 signaling axis. Further studies revealed that TAC-induced terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) positive nuclei and the protein levels of Bax/Bcl2 ratio and cleaved caspase 3 were substantially decreased by Daph treatment. We further characterized the effect of Daph on angiotensin II (Ang-II)-stimulated H9c2 cardiomyoblast cells and observed that Daph markedly decreased the Ang-II induced increase in cell size, production of ROS, and proteins associated with apoptosis and fibrosis. Mechanistically, Daph alone treatment enhanced the protein levels of Nrf2, NQO1, and HO-1 in H9c2 cells. The inhibition of this axis by Si-Nrf2 transfection abolished the protective effect of Daph in H9c2 cells. Taken together, Daph effectively counteracted the TAC-induced cardiac hypertrophy and fibrosis by improving the Nrf2/HO-1 axis and inhibiting the TGF-ß1/Smad2/3 signaling axis.
Assuntos
Heme Oxigenase-1 , Proteínas de Membrana , Fator 2 Relacionado a NF-E2 , Proteína Smad2 , Proteína Smad3 , Fator de Crescimento Transformador beta1 , Umbeliferonas , Angiotensina II/metabolismo , Animais , Cardiomegalia/tratamento farmacológico , Cardiomegalia/metabolismo , Colágeno/metabolismo , Heme Oxigenase-1/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Fator 2 Relacionado a NF-E2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Smad2/antagonistas & inibidores , Proteína Smad2/metabolismo , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Fator de Crescimento Transformador beta1/metabolismo , Umbeliferonas/farmacologia , Regulação para Cima , Remodelação Ventricular/efeitos dos fármacosRESUMO
Resveratrol (Res) can modulate multiple cellular pathways relevant for tumorigenesis but is less effective in colon cancer compared to breast cancer. To increase the chemopreventive potential of Res in combination with 5-fluorouracil (5-FU), a systematic study was carried out in colon cancer cells. HCT-116 cells were treated with Res and 5-FU and several cell-based assays, such as MTT, clonogenic, wound healing, DAPI, comet assay, and Western blot, were performed. A significant inhibition of cell proliferation, migration, and increased apoptosis were observed when moderate concentration of Res (15 microM) was associated with very low concentration of 5-FU (0.5 microM). This combination caused apoptosis by blocking the cells at S phase and enhanced the DNA damage. Expression levels of p-JNK and p-p38 were increased without affecting pERK. 5-FU could be used as a therapeutic modality to improve efficacy of Res-based chemotherapy against colon cancer.
Assuntos
Anticarcinógenos/farmacologia , Antimetabólitos Antineoplásicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Dano ao DNA , Fluoruracila/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Estilbenos/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células HCT116 , Humanos , ResveratrolRESUMO
WNT5A is a well-known mediator of melanoma cell invasion and metastasis via its ability to activate protein kinase C (PKC), which is monitored by phosphorylation of the endogenous PKC substrate myristoylated alanine-rich c-kinase substrate (MARCKS). However, a possible direct contribution of MARCKS in WNT5A-mediated melanoma cell invasion has not been investigated. Analyses of melanoma patient databases suggested that similar to WNT5A expression, MARCKS expression appears to be associated with increased metastasis. A relationship between the two is suggested by the findings that recombinant WNT5A (rWNT5A) induces both increased expression and phosphorylation of MARCKS, whereas WNT5A silencing does the opposite. Moreover, WNT5A-induced invasion of melanoma cells was blocked by siRNA targeting MARCKS, indicating a crucial role of MARCKS expression and/or its phosphorylation. Next, we employed a peptide inhibitor of MARCKS phosphorylation that did not affect MARCKS expression and found that it abolished WNT5A-induced melanoma cell invasion. Similarly, rWNT5A induced the accumulation of phosphorylated MARCKS in membrane protrusions at the leading edge of melanoma cells. Our results demonstrate that WNT5A-induced phosphorylation of MARCKS is not only an indicator of PKC activity but also a crucial regulator of the metastatic behavior of melanoma and therefore an attractive future antimetastatic target in melanoma patients.
RESUMO
The identification of novel antimetastatic therapeutic targets is necessary for improved treatment of patients with acquired BRAF inhibitor-resistant (BRAFi-R) melanoma, in whom metastasis is a major concern. Our present study focused on the identification of such targets to explore novel antimetastatic therapeutic options for BRAFi-R melanoma patients. We confirmed the development of BRAFi resistance in our BRAFi-treated melanoma cell lines by demonstrating reduced sensitivity to BRAF inhibitors, increased ERK1/2 activity and increased WNT5A expression. Here, we demonstrated for the first time that high secretion of interleukin-6 (IL-6) was associated with increased invasive migration of BRAFi-R melanoma cells. This finding could be readily explained by the increased expression of WNT5A in BRAFi-R melanoma cells and the presence of an IL-6/WNT5A positive feedback loop in parental melanoma cells. Surprisingly, however, we found that the IL-6/WNT5A positive feedback loop present in parental melanoma cells was lost during the development of acquired BRAFi resistance, meaning that IL-6 and WNT5A signalling were independent events in BRAFi-R melanoma cells. Despite the absence of an IL-6/WNT5A loop, we found that both an IL-6 blocking antibody and the WNT5A antagonist Box5 alone impaired the elevated invasive migration of BRAFi-R melanoma cells, but combined use of the two was more effective. This impaired invasive migration of BRAFi-R melanoma cells correlated well with the reduction in Cdc42-GTPase activity and alterations of the actin cytoskeleton in these cells. In summary, our novel identification of IL-6 as a key independent promoter of the invasive migration of BRAFi-R melanoma cells stresses that a combination of a blocking IL-6 antibody and administration of the WNT5A antagonist Box5 might be an attractive antimetastatic approach for future treatment of BRAFi-R melanoma patients.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Movimento Celular , Resistencia a Medicamentos Antineoplásicos , Interleucina-6/metabolismo , Melanoma/tratamento farmacológico , Melanoma/patologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Actinas/metabolismo , Anticorpos Bloqueadores/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Humanos , Mutação/genética , Invasividade Neoplásica , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Wnt-5a , Proteína cdc42 de Ligação ao GTP/metabolismoRESUMO
Increased expression and signalling of WNT5A and interleukin-6 (IL-6) have both been shown to promote melanoma progression. Here, we investigated the proposed existence of a WNT5A-IL-6 positive feedback loop that drives melanoma migration and invasion. First, the HOPP algorithm revealed that the invasive phenotype of cultured melanoma cells was significantly correlated with increased expression of WNT5A or IL-6. In three invasive melanoma cell lines, endogenous WNT5A protein expression was related to IL-6 protein secretion. Knockdown with anti-IL-6 siRNAs or treating WM852 melanoma cells with a neutralising anti-IL-6 antibody reduced WNT5A protein expression. Conversely, the silencing of WNT5A expression by WNT5A siRNAs or treating WM852 melanoma cells with Box5 (a WNT5A antagonist) significantly reduced IL-6 secretion. Interestingly, these effects occurred at the protein level but not at the transcriptional levels. Functionally, we demonstrated that combined siRNA knockdown of WNT5A and IL-6 expression or the simultaneous inhibition of WNT5A and IL-6 signalling inhibited melanoma cell invasion more effectively than suppressing each factor individually. Together, our results demonstrate that WNT5A and IL-6 are connected through a positive feedback loop in melanoma cells and that the combined targeting of both molecules could serve as an effective therapeutic means to reduce melanoma metastasis.