RESUMO
The ethnopharmacological information gathered over many centuries and the presence of diverse metabolites have made the medicinal plants as the prime source of drugs. Despite the positive attributes of natural products, there are many questions pertaining to their mechanism of actions and molecular targets that impede their development as therapeutic agents. One of the major challenges in cancer research is the toxicity exerted by investigational agents towards the host. An understanding of their molecular targets, underlying mechanisms can reveal their anticancer efficacy, help in optimal therapeutic dose selection, to mitigate their side effects and toxicity towards the host. The purpose of this review is to collate details on natural products that are recently been investigated extensively in the past decade for their anticancer potential. Besides, critical analysis of their molecular targets and underlying mechanisms on multiple cancer cell lines, an in-depth probe of their toxicological screening on rodent models is outlined as well to observe the prevalence of their toxicity towards host. This review can provide valuable insights for researchers in developing methods, strategies during preclinical and clinical evaluation of anticancer candidates.
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Abstract 1'-acetoxychavicol acetate (ACA)-loaded nanostructured lipid carriers (NLCs) were formulated for prostate cancer therapy and to determine the optimal therapeutic dose, we developed a rapid, specific, and accurate reversed-phase high-performance liquid chromatography (RP-HPLC) method to quantify the ACA content in NLCs. The method was validated according to International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines. Chromatographic separation of ACA from the lipid components was performed with an Agilent 1220 Infinity LC system and ultraviolet detector using an Agilent Poroshell C18 column (4.6 x 250.0 mm). The mobile phase consisted of acetonitrile and water (80:20 [v/v]) with a flow rate of 0.8 mL/min in isocratic mode. Linearity of the standard curve was assessed at an ACA concentration range of 5-200 µg/mL, and a 1/x weighted linear regression was adopted for the calibration curve. The calculated limits of detection and quantification were 0.59 µg/mL and 1.79 µg/mL, respectively. The mean percent recovery of ACA was 100.02% (relative SD, 2%), and the coefficients of variation for intraday and interday assays were within the values required by the ICH. We also demonstrated robustness of the method by altering the mobile phase ratio and flow rate. Furthermore, we proved specificity of the method for ACA by comparing chromatograms of the blank NLC and ACA-NLC. Hence, we effectively used this validated method to determine the drug-loading capacity and entrapment efficiency of the NLCs.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Estudo de Validação , Acetatos/agonistas , Neoplasias da Próstata/patologia , Tratamento Farmacológico/classificação , Cromatografia de Fase Reversa/métodosRESUMO
The nucleotide-binding oligomerization domain-like receptor (NLR) family pyrin domain containing (NLRP) 3 inflammasome is a multiprotein complex that triggers Caspase-1-mediated IL-1ß production and pyroptosis, and its dysregulation is associated with the pathogenesis of inflammatory diseases. 1'-Acetoxychavicol acetate (ACA) is a natural compound in the rhizome of tropical ginger Alpinia species with anti-microbial, anti-allergic and anti-cancer properties. In this study, we found that ACA suppressed NLRP3 inflammasome activation in mouse bone marrow-derived macrophages and human THP-1 monocytes. ACA inhibited Caspase-1 activation and IL-1ß production by NLRP3 agonists such as nigericin, monosodium urate (MSU) crystals, and ATP. Moreover, it suppressed oligomerization of the adapter molecule, apoptosis-associated speck-like protein containing a CARD (ASC), and Caspase-1-mediated cleavage of pyroptosis executor Gasdermin D. Mechanistically, ACA inhibited generation of mitochondrial reactive oxygen species (ROS) and prevented release of oxidized mitochondrial DNA, which trigger NLRP3 inflammasome activation. ACA also prevented NLRP3 inflammasome activation in vivo, as evidenced in the MSU crystal-induced peritonitis and dextran sodium sulfate-induced colitis mouse models accompanied by decreased Caspase-1 activation. Thus, ACA is a potent inhibitor of the NLRP3 inflammasome for prevention of NLRP3-associated inflammatory diseases.
Assuntos
Álcoois Benzílicos/farmacologia , Inflamassomos/efeitos dos fármacos , Inflamassomos/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Caspase 1/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Peritonite/tratamento farmacológico , Peritonite/metabolismo , Fagocitose/efeitos dos fármacos , Piroptose/efeitos dos fármacos , Células THP-1/efeitos dos fármacos , Células THP-1/metabolismoRESUMO
1'-acetoxychavicol acetate (ACA) extracted from the rhizomes of Alpinia conchigera Griff (Zingiberaceae) has been shown to deregulate the NF-ĸB signaling pathway and induce apoptosis-mediated cell death in many cancer types. However, ACA is a hydrophobic ester, with poor solubility in an aqueous medium, limited bioavailability, and nonspecific targeting in vivo. To address these problems, ACA was encapsulated in a nanostructured lipid carrier (NLC) anchored with plerixafor octahydrochloride (AMD3100) to promote targeted delivery towards C-X-C chemokine receptor type 4 (CXCR4)-expressing prostate cancer cells. The NLC was prepared using the melt and high sheer homogenization method, and it exhibited ideal physico-chemical properties, successful encapsulation and modification, and sustained rate of drug release. Furthermore, it demonstrated time-based and improved cellular uptake, and improved cytotoxic and anti-metastatic properties on PC-3 cells in vitro. Additionally, the in vivo animal tumor model revealed significant anti-tumor efficacy and reduction in pro-tumorigenic markers in comparison to the placebo, without affecting the weight and physiological states of the nude mice. Overall, ACA-loaded NLC with AMD3100 surface modification was successfully prepared with evidence of substantial anti-cancer efficacy. These results suggest the potential use of AMD3100-modified NLCs as a targeting carrier for cytotoxic drugs towards CXCR4-expressing cancer cells.
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Objective: Anoikis is apoptosis that is induced when cells detach from the extracellular matrix and neighboring cells. As anoikis serves as a regulatory barrier, cancer cells often acquire resistance towards anoikis during tumorigenesis to become metastatic. MicroRNAs (miRNAs) are short strand RNA molecules that regulate genes post-transcriptionally by binding to mRNAs and reducing the expression of its target genes. This study aimed to elucidate the role of a novel miRNA, miR-6744-5p, in regulating anoikis in breast cancer and identify its target gene. Methods: An anoikis resistant variant of the luminal A type breast cancer MCF-7 cell line (MCF-7-AR) was generated by selecting and amplifying surviving cells after repeated exposure to growth in suspension. MiRNA microarray analysis identified a list of dysregulated miRNAs from which miR-6744-5p was chosen for overexpression and knockdown studies in MCF-7. Additionally, the miRNA was also overexpressed in a triple-negative breast cancer cell line, MDA-MB-231, to evaluate its ability to impair the metastatic potential of breast cancer cells. Results: This study showed that overexpression and knockdown of miR-6744-5p in MCF-7 increased and decreased anoikis sensitivity, respectively. Similarly, overexpression of miR-6744-5p in MDA-MB-231 increased anoikis and also decreased tumor cell invasion in vitro and in vivo. Furthermore, NAT1 enzyme was identified and validated as the direct target of miR-6744-5p. Conclusions: This study has proven the ability of miR-6744-5p to increase anoikis sensitivity in both luminal A and triple negative breast cancer cell lines, highlighting its therapeutic potential in treating breast cancer.
Assuntos
Arilamina N-Acetiltransferase/genética , Neoplasias da Mama/genética , Regulação Neoplásica da Expressão Gênica , Isoenzimas/genética , MicroRNAs/metabolismo , Animais , Anoikis/genética , Neoplasias da Mama/patologia , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Técnicas de Silenciamento de Genes , Humanos , Células MCF-7 , MicroRNAs/genética , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-ZebraRESUMO
There are innumerable anticancer compounds derived from either natural or synthetic origins. Many of these compounds have been further developed through structural modifications to not only inhibit cancer cell growth but also to exert an antimetastatic effect. This is achieved by attaching different substituents to generate different structure-activity relationships. This review highlights the effectiveness of different functional groups known to have antimigration and antiproliferation activities, such as fluoro, methoxy, methyl, amino, hydroxy, nitro, bromo, chloro, methylamino, ethoxy, carbonyl, iodo, and trifluoromethyl groups. Additionally, the positioning of these functional groups plays an important role in their anticancer activities, which was evident in one of our studies comparing analogues of a natural compound. Thus, this review suggests future recommendations for the design and development of improved anticancer drugs with higher efficacy.
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Antineoplásicos/química , Antineoplásicos/farmacologia , Metástase Neoplásica/tratamento farmacológico , Neoplasias/tratamento farmacológico , Animais , Produtos Biológicos/química , Produtos Biológicos/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Humanos , Metástase Neoplásica/patologia , Neoplasias/patologia , Relação Estrutura-AtividadeRESUMO
Geranylated 4-phenylcoumarins DMDP-1 and DMDP-2 isolated from Mesua elegans were elucidated for their role in inducing caspase-independent programmed cell death (CI-PCD) in prostate cancer cell lines, PC-3 and DU 145, respectively. Cell homeostasis disruption was demonstrated upon treatment, as shown by the increase in calcium ion through colourimetric assay and endoplasmic reticulum (ER) stress markers GRP 78 and p-eIF2α through western blot. Subsequently, cytoplasmic death protease calpain-2 also showed increased activity during DMDP-1 & -2 treatments, while lysosomic death protease cathepsin B activity was significantly increased in PC-3 treated with DMDP-1. Flow cytometry showed a reduction in mitochondrial membrane potential in both cell lines, while western blotting showed translocation of mitochondrial death protease AIF into the cytoplasm in its truncated form. Furthermore, DMDP-1 & -2 treatments caused significant increase in superoxide level and oxidative DNA damage. Concurrent inhibition of calpain-2 and cathepsin B during the treatment showed an attenuation of cell death in both cell lines. Hence, DMDP-1 & -2 induce CI-PCD in prostate cancer cell lines through calpain-2 and cathepsin B.
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Calpaína/metabolismo , Catepsina B/metabolismo , Morte Celular/efeitos dos fármacos , Cumarínicos/farmacologia , Magnoliopsida , Extratos Vegetais/farmacologia , Próstata/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Próstata/metabolismo , Próstata/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologiaRESUMO
Cancer development and progression are extremely complex due to the alteration of various genes and pathways. In most cases, multiple agents are required to control cancer progression. The purpose of this study is to investigate, using a mouse model, the synergistic interactions of anti-cancer agents, 1'-S-1'-acetoxychavicol acetate (ACA), Mycobacterium indicus pranii (MIP), and cisplatin (CDDP) in double and triple combinations to treat chemo-sensitize and immune-sensitize breast cancer. Changes in tumor volume and body weight were monitored. Organs were harvested and stained using hematoxylin-eosin for histopathological assessment. Milliplex enzyme-linked immunosorbent assay (ELISA) was performed to determine cytokine levels, while immunohistochemistry (IHC) was conducted on tumor biopsies to verify systemic drug effects. In vivo mouse models showed tumor regression with maintenance of regular body weight for all the different treatment regimens. IHC results provided conclusive evidence indicating that combination regimens were able to down-regulate nuclear factor kappa-B activation and reduce the expression of its regulated pro-inflammatory proteins. Reduction of pro-inflammatory cytokines (e.g., IL-6, TNF-α, and IFN-É£) levels were observed when using the triple combination, which indicated that the synergistic drug combination was able to significantly control cancer progression. In conclusion, ACA, MIP, and CDDP together serve as promising candidates for further development and for subsequent clinical trials against estrogen-sensitive breast cancer.
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Antineoplásicos/farmacologia , Álcoois Benzílicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Cisplatino/farmacologia , Mycobacterium/isolamento & purificação , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Álcoois Benzílicos/síntese química , Álcoois Benzílicos/química , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Cisplatino/síntese química , Cisplatino/química , Citocinas/sangue , Combinação de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
The silencing of BclxL in the nonsmall cell lung cancer (NSCLC) cell line, A549, downregulates miR3615p expression. This study aimed to determine the biological effects of miR3615p on NSCLC, and to elucidate the molecular mechanisms through which apoptosis is regulated. MicroRNA (miRNA or miR) functional analyses were performed via transfection of miR3615p mimics and inhibitors, demonstrating that the inhibition of miR3615p induced the apoptosis of NSCLC cells. To elucidate the function of miR3615p in vivo, cells transfected with miR3615p inhibitors were microinjected into zebrafish embryos, and immunostained using antibodies to detect the active form of caspase3. Co-transfection with siBclxL and miR3615p mimics illustrated the association between BclxL, miR3615p and apoptosis; miR3615p mimics blocked the apoptosis initiated by siBclxL. Luciferase reporter assays identified mothers against decapentaplegic homolog 2 (SMAD2) as a novel target of miR3615p and the reduction of its protein level was validated by western blot analysis. To confirm the molecular mechanisms through which apoptosis is regulated, gene rescue experiments revealed that the ectopic expression of SMAD2 attenuated the inhibitory effects on apoptosis induced by miR3615p. In this study, to the best of our knowledge, we provide the first evidence that miR3615p functions as an oncomiR in A549 and SKLU1 cells through the regulation of SMAD2, suggesting that miR3615p may be employed as a potential therapeutic target for the miRNA-based therapy of NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , MicroRNAs/genética , Proteína Smad2/genética , Proteína Smad2/metabolismo , Regiões 3' não Traduzidas , Células A549 , Animais , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/metabolismo , Transfecção , Peixe-Zebra/embriologia , Proteína bcl-X/genéticaRESUMO
BACKGROUND: Pinnatane A from the bark of Walsura pinnata was investigated for its anti-cancer properties by analyzing the cytotoxic activities and cell cycle arrest mechanism induced in two different liver cancer cell lines. METHODS: A 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to analyze the pinnatane A selectivity in inducing cell death in cancer and normal cells. Various biological assays were carried out to analyze the anti-cancer properties of pinnatane A, such as a live/dead assay for cell death microscopic visualization, cell cycle analysis using propidium iodide (PI) to identify the cell cycle arrest phase, annexin V-fluorescein isothiocyanate (annexin V-FITC)/PI flow cytometry assay to measure percentage of cell populations at different stages of apoptosis and necrosis, and DNA fragmentation assay to verify the late stage of apoptosis. RESULTS: The MTT assay identified pinnatane A prominent dose- and time-dependent cytotoxicity effects in Hep3B and HepG2 cells, with minimal effect on normal cells. The live/dead assay showed significant cell death, while cell cycle analysis showed arrest at the G0/G1 phase in both cell lines. Annexin V-FITC/PI flow cytometry and DNA fragmentation assays identified apoptotic cell death in Hep3B and necrotic cell death in HepG2 cell lines. CONCLUSIONS: Pinnatane A has the potential for further development as a chemotherapeutic agent prominently against human liver cells.
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Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Meliaceae/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Clivagem do DNA/efeitos dos fármacos , Humanos , Neoplasias Hepáticas , Estrutura MolecularRESUMO
1'-S-1'-acetoxychavicol acetate (ACA) has been previously reported to reduce tumor volume in nude mice, at an effective dose of 1.56â¯mg/kg body weight. However, the detailed toxicological profile for ACA has not yet been performed. Herein, we investigated the toxicity of intravenous administration of ACA in male and female Sprague-Dawley rats, both acutely (with single doses of 2.00, 4.00 and 6.66â¯mg/kg body weight, for 14â¯days), and sub-acutely (with weekly injections of 0.66, 1.33, and 2.22â¯mg/kg, for 28â¯days). In both toxicity studies, treatment with ACA did not affect behavior, food/water intake or body weight, nor did it induce any changes in clinically relevant hematological and biochemical parameters or mortality, suggesting that the LD50 of ACA was higher than 6.66â¯mg/kg body weight, regardless of sex. Sub-acutely, there was however, mild focal inflammation of kidneys and lobular hepatitis, but these were not associated with significant functional adverse effects. Therefore, the no-observed-adverse-effect level (NOAEL) for intravenous administration of ACA in the present 28-day sub-acute study was 2.22â¯mg/kg body weight, in both male and female rats. These findings provide useful information regarding the safety of ACA use in a healthy, non-tumor-bearing rat model.
Assuntos
Antineoplásicos Fitogênicos/toxicidade , Álcoois Benzílicos/toxicidade , Administração Intravenosa , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Comportamento Animal/efeitos dos fármacos , Álcoois Benzílicos/administração & dosagem , Contagem de Células Sanguíneas , Peso Corporal/efeitos dos fármacos , Ingestão de Líquidos/efeitos dos fármacos , Ingestão de Alimentos/efeitos dos fármacos , Feminino , Inflamação/induzido quimicamente , Inflamação/patologia , Dose Letal Mediana , Masculino , Nível de Efeito Adverso não Observado , Ratos , Ratos Sprague-Dawley , Testes de Toxicidade SubcrônicaRESUMO
BACKGROUND: Drug combination therapy to treat cancer is a strategic approach to increase successful treatment rate. Optimizing combination regimens is vital to increase therapeutic efficacy with minimal side effects. MATERIALS AND METHODS: In the present study, we evaluated the in vitro cytotoxicity of double and triple combinations consisting of 1'S-1'-acetoxychavicol acetate (ACA), Mycobacterium indicus pranii (MIP) and cisplatin (CDDP) against 14 various human cancer cell lines to address the need for more effective therapy. Our data show synergistic effects in MCF-7 cells treated with MIP:ACA, MIP:CDDP and MIP:ACA:CDDP combinations. The type of interaction between MIP, ACA and CDDP was evaluated based on combination index being <0.8 for synergistic effect. Identifying the mechanism of cell death based on previous studies involved intrinsic apoptosis and nuclear factor kappa B (NF-κB) and tested in Western blot analysis. Inactivation of NF-κB was confirmed by p65 and IκBα, while intrinsic apoptosis pathway activation was confirmed by caspase-9 and Apaf-1 expression. RESULTS: All combinations confirmed intrinsic apoptosis activation and NF-κB inactivation. CONCLUSION: Double and triple combination regimens that target induction of the same death mechanism with reduced dosage of each drug could potentially be clinically beneficial in reducing dose-related toxicities.
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Antibacterianos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Álcoois Benzílicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Cisplatino/farmacologia , Mycobacterium/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Antibacterianos/química , Protocolos de Quimioterapia Combinada Antineoplásica/química , Álcoois Benzílicos/química , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cisplatino/química , Difusão , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Testes de Sensibilidade Microbiana , NF-kappa B/metabolismo , Relação Estrutura-AtividadeRESUMO
MicroRNAs (miRNAs) have been extensively studied over the decades and have been proposed as potential molecular targets for cancer treatment. Studies have shown that miR-378 participates in numerous biological processes in various cancers; whereas miR-1827 has only been reported in pediatric glioma. The mechanism of how miRNAs modulate lung cancer metastasis remains unclear. Our previous study demonstrated that miR-378 is up-regulated while miR-1827 is down-regulated in high invasive lung cancer sub-cell lines, and their biological functions have been described. Here, we report that miR-378 and miR-1827 modulate lung cancer cell invasion and migration via epithelial-mesenchymal transition (EMT). We also demonstrated that cells treated with miR-378 inhibitors or miR-1827 mimics had reduced number of metastases and ectopic vessels in the zebrafish embryo model. We then showed that miR-378 promoted invasion and miR-1827 suppressed migration by targeting RBX1 and CRKL, respectively. Restored protein expression in miRNA-overexpressed/ miRNA-suppressed cells attenuated the inhibitory/ inducing effect of the miRNA on lung cancer cells. Collectively, our findings highlight that miR-378 and miR-1827 could serve as novel therapeutic targets in lung cancer.
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The innate immune system serves as the first line of defense to protect the host from pathogen infection. As a first step, the pattern recognition receptors (PRRs) recognize pathogen-associated molecular patterns (PAMPs), such as non-self DNA derived from pathogens, and damage-associated molecular patterns (DAMPs), such as self DNA released from damaged or injured cells. Sensing of such DNAs elicits innate immune responses through the production of type I interferons (IFNs) and proinflammatory cytokines resulting from the activation of interferon regulatory factor 3 (IRF3) and nuclear factor kappa B (NF-κB), respectively. These cytokines are key players in interlinking innate and adaptive immune responses. However, defects in DNA sensors and their signaling cascades lead to dysregulation of immune responses, autoimmune diseases, and cancer progression. Here we provide an update on DNA signaling pathways in response to pathogen infection and cell injury, and on the roles of regulators in governing the immune system and maintaining host homeostasis. We also discuss the evasion of immunosurveillance by pathogens.
Assuntos
DNA/imunologia , Imunidade Inata/imunologia , Receptores de Reconhecimento de Padrão/imunologia , Transdução de Sinais/imunologia , Animais , Citocinas/biossíntese , Citocinas/imunologia , Humanos , Interferon Tipo I/biossíntese , Interferon Tipo I/imunologiaRESUMO
Nine analogs of 1'S-1'-acetoxychavicol acetate (ACA) were hemi-synthesized and evaluated for their anticancer activities against seven human cancer cell lines. The aim of this study was to investigate the anti-proliferative, apoptotic, and anti-migration effects of these compounds and to explore the plausible underlying mechanisms of action. We found that ACA and all nine analogs were non toxic to human mammary epithelial cells (HMECs) used as normal control cells, and only ACA, 1'-acetoxyeugenol acetate (AEA), and 1'-acetoxy-3,5-dimethoxychavicol acetate (AMCA) inhibited the growth of MDA-MB-231 breast cancer cells with a half-maximal inhibitory concentration (IC50) value of <30.0 µM based on 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay results, and were selected for further investigation. DNA fragmentation assays showed that these three compounds markedly induced apoptosis of MDA-MB-231 cells. Western blot analysis revealed increased expression levels of cleaved PARP, p53, and Bax, while decreased expression levels of Bcl-2 and Bcl-xL were seen after treatment, indicating that apoptosis was induced via the mitochondrial pathway. Moreover, ACA, AEA, and AMCA effectively inhibited the migration of MDA-MB-231 cells. They also downregulated the expression levels of pFAK/FAK and pAkt/Akt via the integrin ß1-mediated signaling pathway. Collectively, ACA and its hemi-synthetic analogs, AEA and AMCA are seen as potential anticancer agents following their abilities to suppress growth, induce apoptosis, and inhibit migration of breast cancer cells.
Assuntos
Antineoplásicos/farmacologia , Álcoois Benzílicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias/tratamento farmacológico , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Álcoois Benzílicos/síntese química , Álcoois Benzílicos/química , Western Blotting , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Fragmentação do DNA/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Humanos , Concentração Inibidora 50 , Neoplasias/patologiaRESUMO
Lung cancer remains a major health problem with a low 5-year survival rate of patients. Recent studies have shown that dysregulation of microRNAs (miRNAs) are prevalent in lung cancer and these aberrations play a significant role in the progression of tumour progression. In the present study, bioinformatics analyses was employed to predict potential miR-608 targets, which are associated with signaling pathways involved in cancer. Luciferase reporter assay identified AKT2 as a novel target of miR-608, and suppression of its protein levels was validated through western blot analysis. Zebrafish embryos were microinjected with cells transfected with miR-608 to elucidate the role of miR-608 in vivo, and immunostained with antibodies to detect activated caspase-3. We present the first evidence that miR-608 behaves as a tumour suppressor in A549 and SK-LU-1 cells through the regulation of AKT2, suggesting that selective targeting of AKT2 via miR-608 may be developed as a potential therapeutic strategy for miRNA-based non-small cell lung cancer (NSCLC) therapy.
Assuntos
Adenocarcinoma/enzimologia , Adenocarcinoma/genética , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Regiões 3' não Traduzidas , Células A549 , Adenocarcinoma de Pulmão , Animais , Apoptose/fisiologia , Caspase 3/metabolismo , Inativação Gênica , Xenoenxertos , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transfecção , Peixe-ZebraRESUMO
BACKGROUND: The compound, 1'-S-1'-acetoxychavicol acetate (ACA), isolated from the rhizomes of a Malaysian ethno-medicinal plant, Alpinia conchigera Griff. (Zingiberaceae), was previously shown to have potential in vivo antitumour activities. In the development of a new drug entity, potential interactions of the compound with the cytochrome P450 superfamily metabolizing enzymes need to be ascertain. PURPOSE: The concomitant use of therapeutic drugs may cause potential drug-drug interactions by decreasing or increasing plasma levels of the administered drugs, leading to a suboptimal clinical efficacy or a higher risk of toxicity. Thus, evaluating the inhibitory potential of a new chemical entity, and to clarify the mechanism of inhibition and kinetics in the various CYP enzymes is an important step to predict drug-drug interactions. STUDY DESIGN: This study was designed to assess the potential inhibitory effects of Alpinia conchigera Griff. rhizomes extract and its active constituent, ACA, on nine c-DNA expressed human cytochrome P450s (CYPs) enzymes using fluorescent CYP inhibition assay. METHODS/RESULTS: The half maximal inhibitory concentration (IC50) of Alpinia conchigera Griff. rhizomes extract and ACA was determined for CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C19, CYP2D6, CYP2E1, CYP3A4 and CYP3A5. A. conchigera extract only moderately inhibits on CYP3A4 (IC50 = 6.76 ± 1.88µg/ml) whereas ACA moderately inhibits the activities of CYP1A2 (IC50 = 4.50 ± 0.10µM), CYP2D6 (IC50 = 7.50 ± 0.17µM) and CYP3A4 (IC50 = 9.50 ± 0.57µM) while other isoenzymes are weakly inhibited. In addition, mechanism-based inhibition studies reveal that CYP1A2 and CYP3A4 exhibited non-mechanism based inhibition whereas CYP2D6 showed mechanism-based inhibition. Lineweaver-Burk plots depict that ACA competitively inhibited both CYP1A2 and CYP3A4, with a Ki values of 2.36 ± 0.03 µM and 5.55 ± 0.06µM, respectively, and mixed inhibition towards CYP2D6 with a Ki value of 4.50 ± 0.08µM. Further, molecular docking studies show that ACA is bound to a few key amino acid residues in the active sites of CYP1A2 and CYP3A4, while one amino residue of CYP2D6 through predominantly Pi-Pi interactions. CONCLUSION: Overall, ACA may demonstrate drug-drug interactions when co-administered with other therapeutic drugs that are metabolized by CYP1A2, CYP2D6 or CYP3A4 enzymes. Further in vivo studies, however, are needed to evaluate the clinical significance of these interactions.
Assuntos
Alpinia/química , Álcoois Benzílicos/farmacologia , Inibidores das Enzimas do Citocromo P-450/farmacologia , Álcoois Benzílicos/química , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Inibidores das Enzimas do Citocromo P-450/química , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Interações Medicamentosas , Humanos , Concentração Inibidora 50 , Simulação de Acoplamento Molecular , Extratos Vegetais/química , Extratos Vegetais/farmacologiaRESUMO
MicroRNAs (miRNAs) are short non-coding RNAs that regulate genes posttranscriptionally. Past studies have reported that miR-210 is up-regulated in many cancers including cervical cancer, and plays a pleiotropic role in carcinogenesis. However, its role in regulating response towards anti-cancer agents has not been fully elucidated. We have previously reported that the natural compound 1'S-1'-acetoxychavicol acetate (ACA) is able to induce cytotoxicity in various cancer cells including cervical cancer cells. Hence, this study aims to investigate the mechanistic role of miR-210 in regulating response towards ACA in cervical cancer cells. In the present study, we found that ACA down-regulated miR-210 expression in cervical cancer cells, and suppression of miR-210 expression enhanced sensitivity towards ACA by inhibiting cell proliferation and promoting apoptosis. Western blot analysis showed increased expression of mothers against decapentaplegic homolog 4 (SMAD4), which was predicted as a target of miR-210 by target prediction programs, following treatment with ACA. Luciferase reporter assay confirmed that miR-210 binds to sequences in 3'UTR of SMAD4. Furthermore, decreased in SMAD4 protein expression was observed when miR-210 was overexpressed. Conversely, SMAD4 protein expression increased when miR-210 expression was suppressed. Lastly, we demonstrated that overexpression of SMAD4 augmented the anti-proliferative and apoptosis-inducing effects of ACA. Taken together, our results demonstrated that down-regulation of miR-210 conferred sensitivity towards ACA in cervical cancer cells by targeting SMAD4. These findings suggest that combination of miRNAs and natural compounds could provide new strategies in treating cervical cancer.
Assuntos
Antineoplásicos/farmacologia , Álcoois Benzílicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , MicroRNAs/metabolismo , Proteína Smad4/metabolismo , Neoplasias do Colo do Útero/metabolismo , Regiões 3' não Traduzidas , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Proteína Smad4/genética , Regulação para CimaRESUMO
BACKGROUND: Cervical cancer is the fourth most frequent malignancy affecting women worldwide, but drug resistance and toxicities remain a major challenge in chemotherapy. The use of natural compounds is promising because they are less toxic and able to target multiple signaling pathways. The 1'S-1'-acetoxychavicol acetate (ACA), a natural compound isolated from wild ginger Alpinia conchigera, induced cytotoxicity on various cancer cells including cervical cancer. MicroRNAs (miRNAs) are short noncoding RNAs that regulate numerous biological processes, such as apoptosis and chemosensitivity. Past studies reported that miR-629 is upregulated in many cancers, and its expression was altered in ACA-treated cervical cancer cells. However, the role of miR-629 in regulating sensitivity toward ACA or other anticancer agents has not been reported. Hence, this study aims to investigate the role of miR-629 in regulating response toward ACA on cervical cancer cells. METHODS: The miR-629 expression following transfection with miR-629 hairpin inhibitor and hairpin inhibitor negative control was measured using quantitative real-time polymerase chain reaction (RT-qPCR). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to investigate sensitivity toward ACA. Apoptosis was detected using Annexin V/propidium iodide and Caspase 3/7 assays. The gene target for miR-629 was identified using miRNA target prediction programs, luciferase reporter assay and Western blots. Gene overexpression studies were performed to evaluate its role in regulating response toward ACA. RESULTS: Transfection with miR-629 hairpin inhibitor downregulated its expression in both cervical cancer cell lines. Suppression of miR-629 increased sensitivity toward ACA by reducing cell proliferation and inducing apoptosis. Luciferase reporter assay confirmed RSU1 as a direct target of miR-629. Overexpression of miR-629 decreased RSU1 protein expression, while inhibition of miR-629 increased RSU1 protein expression. Overexpression of RSU1 augmented antiproliferative and apoptosis-inducing effects of ACA. CONCLUSION: Our findings showed that combination of ACA with miR-629 and RSU1 may provide a potential strategy in treating cervical cancer.
RESUMO
Mycobacterium indicus pranii (MIP) is a non-pathogenic mycobacterium, which has been tested on several cancer types like lung and bladder where tumour regression and complete recovery was observed. In discovering the potential cytotoxic elements, a preliminary test was carried out using four different fractions consisting of live bacteria, culture supernatant, heat killed bacteria and heat killed culture supernatant of MIP against two human cancer cells A549 and CaSki by 3-(4,5-dimethyl thiazol)-2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis was investigated in MCF-7 and ORL-115 cancer cells by poly-(ADP-ribose) polymerase (PARP) and DNA fragmentation assays. Among four MIP fractions, only heat killed MIP fraction (HKB) showed significant cytotoxicity in various cancer cells with inhibitory concentration, IC50 in the range 5.6-35.0 µl/(1.0 × 10(6) MIP cells/ml), while cytotoxicity effects were not observed in the remaining fractions. HKB did not show cytotoxic effects on non-cancerous cells contrary to cancerous cells, suggesting its safe usage and ability to differentially recognize between these cells. Evaluation on PARP assay further suggested that cytotoxicity in cancer cells were potentially induced via caspase-mediated apoptosis. The cytotoxic and apoptotic effects of MIP HKB have indicated that this fraction can be a good candidate to further identify effective anti-cancer agents.