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1.
Anticancer Res ; 41(9): 4259-4269, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475045

RESUMO

BACKGROUND: Compared to two-dimensional cultures, three-dimensional (3D) cultures have many advantages in cancer studies. Nevertheless, their implementation is unsatisfactory. This study aimed to develop an anchorage-dependent 3D culture model for colorectal cancer research. MATERIALS AND METHODS: Human HCT116, DLD-1 and SW620 colorectal cell lines were cultured in a gelatin sponge, and its applicability for morphological examination was studied. RESULTS: The resulting specimens were suitable for scanning electron microscopy, transmission electron microscopy, and immunohistochemical examination. HCT116 formed smaller structures and migrated through the pores of the sponge. DLD-1 formed larger structures with tight cell-to-cell adhesion. SW620 also formed large structures but small clustered cells tended to attach to the anchorage more favorably. Immunohistochemical staining demonstrated phosphorylated yes-associated protein (YAP) localized near the attachment site in HCT116 cells. CONCLUSION: Because the gelatin sponge provided suitable anchorage and the cultured cells formed distinguishable 3D structures, this method may be useful for further colorectal cancer research.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Técnicas de Cultura de Células/métodos , Neoplasias Colorretais/patologia , Gelatina/química , Tecidos Suporte/química , Fatores de Transcrição/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Neoplasias Colorretais/metabolismo , Células HCT116 , Humanos , Fosforilação
2.
Anticancer Res ; 41(9): 4353-4364, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475055

RESUMO

BACKGROUND/AIM: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a potential anti-tumor agent. However, resistance to TRAIL has been reported in a number of clinical trials. In this study, we investigated the molecular mechanisms by which a novel histone deacetylase (HDAC) inhibitor, CBUD-1001, sensitizes colorectal cancer (CRC) cells to TRAIL-induced apoptosis. MATERIALS AND METHODS: Apoptotic cell death induced by CBUD-1001 and/or TRAIL was assessed on human CRC cells using the MTT assay, FACS analysis and nuclei staining. The involved molecular mechanisms were explored through western blotting analysis. RESULTS: We demonstrated that combined with CBUD-1001, TRAIL significantly enhanced TRAIL-induced apoptosis in CRC cells via mitochondria-mediated pathways. We also found that hyper-acetylation of histone by CBUD-1001 treatment leads to up-regulation of death receptor (DR) 5 in a dose- and time-dependent manner. Furthermore, we identified that enhanced sensitivity to TRAIL by combination with CBUD-1001 depends on the MAPK/CHOP axis, being a key mediator of DR5. CONCLUSION: A novel HDAC inhibitor CBUD-1001 sensitizes TRAIL-induced apoptosis via up-regulation of DR5, and that CBUD-1001 and TRAIL combination treatment offers an effective strategy to overcome TRAIL resistance in CRC cells.


Assuntos
Neoplasias Colorretais/metabolismo , Histona Desacetilase 1/antagonistas & inibidores , Inibidores de Histona Desacetilases/farmacologia , Ligante Indutor de Apoptose Relacionado a TNF/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Regulação para Baixo , Sinergismo Farmacológico , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Humanos
3.
Molecules ; 26(15)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34361570

RESUMO

A novel series of 4-anilinoquinazoline analogues, DW (1-10), were evaluated for anticancer efficacy in human breast cancer (BT-20) and human colorectal cancer (CRC) cell lines (HCT116, HT29, and SW620). The compound, DW-8, had the highest anticancer efficacy and selectivity in the colorectal cancer cell lines, HCT116, HT29, and SW620, with IC50 values of 8.50 ± 2.53 µM, 5.80 ± 0.92 µM, and 6.15 ± 0.37 µM, respectively, compared to the non-cancerous colon cell line, CRL1459, with an IC50 of 14.05 ± 0.37 µM. The selectivity index of DW-8 was >2-fold in colon cancer cells incubated with vehicle. We further determined the mechanisms of cell death induced by DW-8 in SW620 CRC cancer cells. DW-8 (10 and 30 µM) induced apoptosis by (1) producing cell cycle arrest at the G2 phase; (2) activating the intrinsic apoptotic pathway, as indicated by the activation of caspase-9 and the executioner caspases-3 and 7; (3) nuclear fragmentation and (4) increasing the levels of reactive oxygen species (ROS). Overall, our results suggest that DW-8 may represent a suitable lead for developing novel compounds to treat CRC.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias do Colo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células HCT116 , Células HT29 , Humanos
4.
Molecules ; 26(16)2021 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-34443490

RESUMO

Hydroxycamptothecin (SN38) is a natural plant extract isolated from Camptotheca acuminate. It has a broad spectrum of anticancer activity through inhibition of DNA topoisomerase I, which could affect DNA synthesis and lead to DNA damage. Thus, the action of SN38 against cancers could inevitably affect endogenous levels of ribonucleotide (RNs) and deoxyribonucleotide (dRNs) that play critical roles in many biological processes, especially in DNA synthesis and repair. However, the exact impact of SN38 on RNs and dRNs is yet to be fully elucidated. In this study, we evaluated the anticancer effect and associated mechanism of SN38 in human colorectal carcinoma HCT 116 cells. As a result, SN38 could decrease the cell viability and induce DNA damage in a concentration-dependent manner. Furthermore, cell cycle arrest and intracellular nucleotide metabolism were perturbed due to DNA damage response, of which ATP, UTP, dATP, and TTP may be the critical metabolites during the whole process. Combined with the expression of deoxyribonucleoside triphosphates synthesis enzymes, our results demonstrated that the alteration and imbalance of deoxyribonucleoside triphosphates caused by SN38 was mainly due to the de novo nucleotide synthesis at 24 h, and subsequently the salvage pathways at 48 h. The unique features of SN38 suggested that it might be recommended as an effective supplementary drug with an anticancer effect.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Camptotecina/análogos & derivados , Neoplasias Colorretais/metabolismo , DNA/metabolismo , Ribonucleotídeos/metabolismo , Camptotecina/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Células HCT116 , Humanos , Irinotecano/farmacologia , Ribonucleosídeo Difosfato Redutase/metabolismo , Ribonucleotídeo Redutases/metabolismo
5.
Nutrients ; 13(8)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34444911

RESUMO

Plant-derived conjugated linolenic acids (CLnA) have been widely studied for their preventive and therapeutic properties against diverse diseases such as cancer. In particular, punicic acid (PunA), a conjugated linolenic acid isomer (C18:3 c9t11c13) present at up to 83% in pomegranate seed oil, has been shown to exert anti-cancer effects, although the mechanism behind its cytotoxicity remains unclear. Ferroptosis, a cell death triggered by an overwhelming accumulation of lipid peroxides, has recently arisen as a potential mechanism underlying CLnA cytotoxicity. In the present study, we show that PunA is highly cytotoxic to HCT-116 colorectal and FaDu hypopharyngeal carcinoma cells grown either in monolayers or as three-dimensional spheroids. Moreover, our data indicate that PunA triggers ferroptosis in carcinoma cells. It induces significant lipid peroxidation and its effects are prevented by the addition of ferroptosis inhibitors. A combination with docosahexaenoic acid (DHA), a known polyunsaturated fatty acid with anticancer properties, synergistically increases PunA cytotoxicity. Our findings highlight the potential of using PunA as a ferroptosis-sensitizing phytochemical for the prevention and treatment of cancer.


Assuntos
Antineoplásicos/farmacologia , Carcinoma/tratamento farmacológico , Ferroptose/efeitos dos fármacos , Ácidos Linolênicos/farmacologia , Carcinoma/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Ácidos Docosa-Hexaenoicos/metabolismo , Células HCT116 , Humanos , Neoplasias Hipofaríngeas/tratamento farmacológico , Neoplasias Hipofaríngeas/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos
6.
Food Res Int ; 147: 110466, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34399464

RESUMO

Dietary fiber plays an important role in the prevention of colorectal cancer, and arabinoxylans are an important source of this in grains, with some studies reporting the inhibition of cancer cell growth. However, very few studies have been conducted on this, and most previous studies have used oligosaccharides derived from arabinoxylans of specific molecular weight. The aim of this work is to extract, isolate, and analyze arabinoxylans from two different Argentinian genotypes of wheat (hard and soft) and study if they have the capacity to decrease the cellular viability of a colon cancer line (HCT-116). To determine whether the molecular size influences the inhibition of HCT-116 cell viability, specific hydrolysis was performed with endoxylanase, and the cells were exposed to the hydrolyzed arabinoxylans. The arabinoxylans treatment resulted in HCT-116 cell viability of 74% for the soft genotype and 64% for the hard genotype in comparison to nontreated cells. Hydrolyzed-arabinoxylans result in HCT-116 cell viability of 68% for soft and 36% for hard genotypes (the lowest IC50 values) compared to nontreated cells. More importantly, no decrease after the arabinoxylans treatment was observed in the viability of murine noncancer cells known to rapidly respond to polysaccharide presence. The arabinoxylans from hard wheat showed more disubstituted xylose and α-1,2/α-1,3 linkages than the arabinoxylans from soft wheat, the possible cause for showing the best in vitro biological effect. The results showed other beneficial effects than the prebiotic ones and support the use of enzymatic treatment to increase the biological impacts of arabinoxylans.


Assuntos
Triticum , Xilanos , Animais , Fibras na Dieta , Células HCT116 , Humanos , Camundongos
7.
Curr Protoc ; 1(8): e219, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34370399

RESUMO

Targeted protein depletion using a conditional degron is a powerful method to probe the role of proteins in living cells because of the speed with which depletion can be induced and its reversibility. The auxin-inducible degron (AID) is one of the most common degron-based technologies used in cell biology. We recently established an improved system, called AID2, which involves expressing a mutant E3 ligase subunit, OsTIR1(F74G), and fusing a protein of interest to the mini-AID (mAID) tag, and that employs a new and more potent ligand, 5-phenyl-indole-3-acetic acid (5-Ph-IAA). The AID2 system overcomes some of the drawbacks associated with the original AID system, i.e., leaky degradation without auxin and the requirement of high auxin doses. With AID2 it is, therefore, now possible to control a degron-fused protein more precisely, enabling target proteins to be degraded with a half-life of 10 to 45 min via the addition of a low dose of 5-Ph-IAA. Importantly, in AID2, it is not necessary to control the expression of OsTIR1(F74G) for suppressing leaky degradation and a parental cell line constitutively expressing OsTIR1(F74G) can be used for the generation of multiple mAID-tagged proteins. Here, we describe a protocol for the tagging of endogenous proteins with mAID in diploid HCT116 cells. Our protocol can be applied to other mammalian cell lines and will enhance the utility of AID2 for studying protein functions in living cells. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Generation of a parental HCT116 cell line expressing OsTIR1(F74G) Basic Protocol 2: Construction of CRISPR and donor plasmids for tagging endogenous genes Basic Protocol 3: Generation of cell lines expressing a protein of interest fused with mAID.


Assuntos
Ácidos Indolacéticos , Proteínas , Animais , Células HCT116 , Humanos , Ácidos Indolacéticos/farmacologia , Proteólise , Ubiquitina-Proteína Ligases/genética
8.
Free Radic Biol Med ; 174: 157-170, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34403740

RESUMO

Tumor hypoxia is a major biological factor that drives resistance to chemotherapy and radiotherapy. We previously demonstrated that the pro-oxidative drug dihydroartemisinin (DHA) efficiently targeted normoxic and hypoxic cancer cells. Although well studied in normoxia, the mechanism behind DHA-mediated cytotoxicity in hypoxia is insufficiently explored. Here, we analyzed the effect of DHA in HCT116 wild type (wt) cells and in HCT116 Bax-/-Baksh cells with a defective intrinsic apoptosis pathway. Normoxic HCT116 wt cells underwent apoptosis shortly after treatment with DHA. Autophagy-associated cell death contributes to short-term cytotoxicity of DHA in normoxia. These cells switched to an apoptosis- and autophagy-independent cell death after treatment with DHA in hypoxia and displayed similar long-term survival in response to DHA in normoxia and hypoxia. In HCT116 Bax-/-Baksh cells, DHA induced cell cycle arrest shortly after treatment irrespective of oxygen levels. Later, HCT116 Bax-/-Baksh cells induced a delayed cell death after treatment with DHA in hypoxia followed by return to normoxia, while treatment with DHA in normoxia was hardly toxic. We identified lower glutathione levels in hypoxic HCT116 cells which correlated with higher lipid peroxidation after treatment with DHA. Moreover, insufficient expression of Bax/Bak counteracted hypoxia-mediated downregulation of mitochondrial function, thereby adding to DHA-induced ROS production and lipid peroxidation in hypoxia. In summary, DHA-mediated cytotoxicity in normoxia depended on Bax/Bak expression, while cytotoxicity after treatment with DHA in hypoxia was regulated independently of Bax/Bak in HCT116 colorectal cancer cells.


Assuntos
Apoptose , Neoplasias Colorretais , Artemisininas , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/genética , Células HCT116 , Humanos , Hipóxia , Proteína X Associada a bcl-2/genética
9.
Molecules ; 26(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34443432

RESUMO

BACKGROUND: It is well known that more than 90% of cancer deaths are due to metastases. However, the entire tumorigenesis process is not fully understood, and it is evident that cells spreading from the primary tumor play a key role in initiating the metastatic process. Tumor proliferation and invasion also elevate the concentration of regular and irregular metabolites in the serum, which may alter the normal function of the entire human homeostasis and possibly causes cancer metabolism syndrome, also referred to as cachexia. METHODS: We report on the modification of commercially available hemodialysis membranes to selectively capture circulating tumor cells from the blood stream by means of immobilized human anti-EpCAM antibodies on the inner surface of the fibers. All critical steps are described that required in situ addition of the immuno-affinity feature to hemodialyzer cartridges in order to capture EpCAM positive circulating tumor cells, which represents ~80% of cancer cell types. RESULTS: The cell capture efficiency of the suggested technology was demonstrated by spiking HCT116 cancer cells both into buffer solution and whole blood and run through on the modified cartridge. Flow cytometry was used to quantitatively evaluate the cell clearance performance of the approach. CONCLUSIONS: The suggested modification has no significant effect on the porous structure of the hemodialysis membranes; it keeps its cytokine removal capability, addressing cachexia simultaneously with CTC removal.


Assuntos
Neoplasias/patologia , Neoplasias/terapia , Células Neoplásicas Circulantes/patologia , Diálise Renal , Citometria de Fluxo , Fluorescência , Células HCT116 , Humanos , Membranas , Polímeros/química , Sulfonas/química
10.
Molecules ; 26(15)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34361576

RESUMO

Prunus mahaleb L. fruit has long been used in the production of traditional liqueurs. The fruit also displayed scavenging and reducing activity, in vitro. The present study focused on unravelling peripheral and central protective effects, antimicrobial but also anti-COVID-19 properties exerted by the water extract of P. mahaleb. Anti-inflammatory effects were studied in isolated mouse colons exposed to lipopolysaccharide. Neuroprotection, measured as a blunting effect on hydrogen-peroxide-induced dopamine turnover, was investigated in hypothalamic HypoE22 cells. Antimicrobial effects were tested against different Gram+ and Gram- bacterial strains. Whereas anti-COVID-19 activity was studied in lung adenocarcinoma H1299 cells, where the gene expression of ACE2 and TMPRSS2 was measured after extract treatment. The bacteriostatic effects induced on Gram+ and Gram- strains, together with the inhibition of COX-2, TNFα, HIF1α, and VEGFA in the colon, suggest the potential of P. mahaleb water extract in contrasting the clinical symptoms related to ulcerative colitis. The inhibition of the hydrogen peroxide-induced DOPAC/DA ratio indicates promising neuroprotective effects. Finally, the downregulation of the gene expression of ACE2 and TMPRSS2 in H1299 cells, suggests the potential to inhibit SARS-CoV-2 virus entry in the human host. Overall, the results support the valorization of the local cultivation of P. mahaleb.


Assuntos
Bactérias/efeitos dos fármacos , Colo/efeitos dos fármacos , Neuroproteção , Extratos Vegetais/farmacologia , SARS-CoV-2/efeitos dos fármacos , Enzima de Conversão de Angiotensina 2/metabolismo , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Antivirais/química , Antivirais/farmacologia , COVID-19 , Linhagem Celular , Colite Ulcerativa/tratamento farmacológico , Citocinas/genética , Citocinas/metabolismo , Dopamina/metabolismo , Frutas/química , Regulação da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Humanos , Inflamação/tratamento farmacológico , Masculino , Camundongos , Extratos Vegetais/química , Prunus/química , Serina Endopeptidases/metabolismo
11.
Int J Mol Sci ; 22(15)2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34360807

RESUMO

This study investigated the roles of low-molecular-weight fucoidan (LMWF) in enhancing the anti-cancer effects of fluoropyrimidine-based chemotherapy. HCT116 and Caco-2 cells were treated with LMWF and 5-FU. Cell viability, cell cycle, apoptosis, and migration were analyzed in both cell types. Potential mechanisms underlying how LMWF enhances the anti-cancer effects of fluoropyrimidine-based chemotherapy were also explored. The cell viability of HCT116 and Caco-2 cells was significantly reduced after treatment with a LMWF--5FU combination. In HCT116 cells, LMWF enhanced the suppressive effects of 5-FU on cell viability through the (1) induction of cell cycle arrest in the S phase and (2) late apoptosis mediated by the Jun-N-terminal kinase (JNK) signaling pathway. In Caco-2 cells, LMWF enhanced the suppressive effects of 5-FU on cell viability through both the c-mesenchymal-epithelial transition (MET)/Kirsten rat sarcoma virus (KRAS)/extracellular signal-regulated kinase (ERK) and the c-MET/phosphatidyl-inositol 3-kinases (PI3K)/protein kinase B (AKT) signaling pathways. Moreover, LMWF enhanced the suppressive effects of 5-FU on tumor cell migration through the c-MET/matrix metalloproteinase (MMP)-2 signaling pathway in both HCT116 and Caco-2 cells. Our results demonstrated that LMWF is a potential complementary therapy for enhancing the efficacies of fluoropyrimidine-based chemotherapy in colorectal cancers (CRCs) with the wild-type or mutated KRAS gene through different mechanisms. However, in vivo studies and in clinical trials are required in order to validate the results of the present study.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias Colorretais , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Proteínas de Neoplasias/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Células CACO-2 , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Fluoruracila/farmacologia , Células HCT116 , Humanos , Polissacarídeos/farmacologia
12.
Molecules ; 26(16)2021 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-34443472

RESUMO

Feruloylacetone (FER) is a natural degradant of curcumin after heating, which structurally reserves some functional groups of curcumin. It is not as widely discussed as its original counterpart has been previously; and in this study, its anticancer efficacy is investigated. This study focuses on the suppressive effect of FER on colon cancer, as the efficacious effect of curcumin on this typical cancer type has been well evidenced. In addition, demethoxy-feruloylacetone (DFER) was applied to compare the effect that might be brought on by the structural differences of the methoxy group. It was revealed that both FER and DFER inhibited the proliferation of HCT116 cells, possibly via suppression of the phosphorylated mTOR/STAT3 pathway. Notably, FER could significantly repress both the STAT3 phosphorylation and protein levels. Furthermore, both samples showed capability of arresting HCT116 cells at the G2/M phase via the activation of p53/p21 and the upregulation of cyclin-B. In addition, ROS elevation and changes in mitochondrial membrane potential were revealed, as indicated by p-atm elevation. The apoptotic rate rose to 36.9 and 32.2% after being treated by FER and DFER, respectively. In summary, both compounds exhibited an anticancer effect, and FER showed a greater proapoptotic effect, possibly due to the presence of the methoxy group on the aromatic ring.


Assuntos
Antineoplásicos/farmacologia , Antioxidantes/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/patologia , Curcumina/farmacologia , Mitocôndrias/efeitos dos fármacos , Estirenos/farmacologia , Antineoplásicos/química , Antioxidantes/química , Apoptose/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Curcumina/química , Curcumina/metabolismo , Ciclina B1/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/agonistas , Fase G2/efeitos dos fármacos , Células HCT116 , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/metabolismo , Fenol/química , Fenol/farmacologia , Fosforilação/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/metabolismo , Estirenos/química , Serina-Treonina Quinases TOR/metabolismo , Proteína Supressora de Tumor p53/agonistas
13.
Nat Commun ; 12(1): 4814, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376668

RESUMO

Glutamoptosis is the induction of apoptotic cell death as a consequence of the aberrant activation of glutaminolysis and mTORC1 signaling during nutritional imbalance in proliferating cells. The role of the bioenergetic sensor AMPK during glutamoptosis is not defined yet. Here, we show that AMPK reactivation blocks both the glutamine-dependent activation of mTORC1 and glutamoptosis in vitro and in vivo. We also show that glutamine is used for asparagine synthesis and the GABA shunt to produce ATP and to inhibit AMPK, independently of glutaminolysis. Overall, our results indicate that glutamine metabolism is connected with mTORC1 activation through two parallel pathways: an acute alpha-ketoglutarate-dependent pathway; and a secondary ATP/AMPK-dependent pathway. This dual metabolic connection between glutamine and mTORC1 must be considered for the future design of therapeutic strategies to prevent cell growth in diseases such as cancer.


Assuntos
Apoptose/fisiologia , Proliferação de Células/fisiologia , Glutamina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Transdução de Sinais/fisiologia , Proteínas Quinases Ativadas por AMP/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Células HCT116 , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Sirolimo/análogos & derivados , Sirolimo/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
14.
Int J Mol Sci ; 22(12)2021 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-34208547

RESUMO

BACKGROUND: Granulin is a secreted, glycosylated peptide-originated by cleavage from a precursor protein-which is involved in cell growth, tumor invasion and angiogenesis. However, the specific prognostic impact of granulin in human colorectal cancer has only been studied to a limited extent. Thus, we wanted to assess the expression of granulin in colorectal cancer patients to evaluate its potential as a prognostic biomarker. METHODS: Expressional differences of granulin in colorectal carcinoma tissue (n = 94) and corresponding healthy colon mucosa were assessed using qRT-PCR. Immunohistochemistry was performed in colorectal cancer specimens (n = 97), corresponding healthy mucosa (n = 47) and colorectal adenomas (n = 19). Subsequently, the results were correlated with histopathological and clinical patients' data. HCT-116 cells were transfected with siRNA for invasion and migration assays. RESULTS: Immunohistochemistry and qRT-PCR revealed tumoral over expression of granulin in colorectal cancer specimens compared to corresponding healthy colon mucosa and adenomas. Tumoral overexpression of granulin was associated with a significantly impaired overall survival. Moreover, downregulation of granulin by siRNA significantly diminished the invasive capacities of HCT-116 cells in vitro. CONCLUSION: Expression of granulin differs in colorectal cancer tissue, adenomas and healthy colon mucosa. Furthermore, granulin features invasive and migrative capabilities and overexpression of granulin correlates with a dismal prognosis. This reveals its potential as a prognostic biomarker and granulin could be a worthwhile molecular target for individualized anticancer therapy.


Assuntos
Biomarcadores Tumorais , Neoplasias Colorretais/diagnóstico , Neoplasias Colorretais/mortalidade , Granulinas/metabolismo , Idoso , Idoso de 80 Anos ou mais , Neoplasias Colorretais/etiologia , Neoplasias Colorretais/metabolismo , Feminino , Expressão Gênica , Granulinas/genética , Células HCT116 , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Estadiamento de Neoplasias , Prognóstico , Análise de Sobrevida
15.
Int J Mol Sci ; 22(13)2021 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-34203267

RESUMO

Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) is an RNA-binding protein and serves as a post-transcriptional fine-tuner regulating the expression of mRNA targets. However, the clinicopathological roles of IGF2BP1 in colorectal cancer (CRC) remains limited. Thus, we aimed to elucidate the clinical significance and biomarker potentials of IGF2BP1 in CRC. A total of 266 specimens from two sets of CRC patients were collected. IGF2BP1 expression was studied by immunohistochemical (IHC) staining. The Kaplan-Meier survival plot and a log-rank test were used for survival analysis. The Cox proportional hazards model was applied to determine the survival impact of IGF2BP1. Public datasets sets from The Cancer Genome Atlas (TCGA) and Human Cancer Metastasis Database (HCMDB), receiver operating characteristic (ROC) plotter, and two CRC cell lines, HCT-116 and DLD-1, were used for validating our findings. We showed that IGF2BP1 was overexpressed in tumor specimens compared to 13 paired normal parts by examining the immunoreactivity of IGF2BP1 (p = 0.045). The increased expression of IGF2BP1 in primary tumor parts was observed regardless of metastatic status (p < 0.001) in HCMDB analysis. IGF2BP1 expression was significantly associated with young age (59.6% vs. 46.7%, p-value = 0.043) and advanced stage (61.3% vs. 40.0%, p-value = 0.001). After controlling for confounding factors, IGF2BP1 remained an independent prognostic factor (HR = 1.705, p-value = 0.005). TCGA datasets analysis indicated that high IGF2BP1 expression showed a lower 5-year survival rate (58% vs. 65%) in CRC patients. The increased expression of IGF2BP1 in chemotherapy non-responder rectal cancer patients was observed using a ROC plotter. Overexpression of IGF2BP1 promoted the colony-forming capacity and 5-fluorouracil and etoposide resistance in CRC cells. Here, IGF2BP1 was an independent poor prognostic marker in CRC patients and contributed to aggressive phenotypes in CRC cell lines.


Assuntos
Biomarcadores/metabolismo , Neoplasias Colorretais/metabolismo , Proteínas de Ligação a RNA/metabolismo , Biomarcadores/química , Neoplasias Colorretais/genética , Células HCT116 , Humanos , Estimativa de Kaplan-Meier , Prognóstico , Modelos de Riscos Proporcionais , Proteínas de Ligação a RNA/genética , Curva ROC
16.
Int J Mol Sci ; 22(13)2021 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-34208885

RESUMO

We previously demonstrated that flavonoid metabolites inhibit cancer cell proliferation through both CDK-dependent and -independent mechanisms. The existing evidence suggests that gut microbiota is capable of flavonoid biotransformation to generate bioactive metabolites including 2,4,6-trihydroxybenzoic acid (2,4,6-THBA), 3,4-dihydroxybenzoic acid (3,4-DHBA), 3,4,5-trihyroxybenzoic acid (3,4,5-THBA) and 3,4-dihydroxyphenylacetic acid (DOPAC). In this study, we screened 94 human gut bacterial species for their ability to biotransform flavonoid quercetin into different metabolites. We demonstrated that five of these species were able to degrade quercetin including Bacillus glycinifermentans, Flavonifractor plautii, Bacteroides eggerthii, Olsenella scatoligenes and Eubacterium eligens. Additional studies showed that B. glycinifermentans could generate 2,4,6-THBA and 3,4-DHBA from quercetin while F. plautii generates DOPAC. In addition to the differences in the metabolites produced, we also observed that the kinetics of quercetin degradation was different between B. glycinifermentans and F. plautii, suggesting that the pathways of degradation are likely different between these strains. Similar to the antiproliferative effects of 2,4,6-THBA and 3,4-DHBA demonstrated previously, DOPAC also inhibited colony formation ex vivo in the HCT-116 colon cancer cell line. Consistent with this, the bacterial culture supernatant of F. plautii also inhibited colony formation in this cell line. Thus, as F. plautii and B. glycinifermentans generate metabolites possessing antiproliferative activity, we suggest that these strains have the potential to be developed into probiotics to improve human gut health.


Assuntos
Ácido 3,4-Di-Hidroxifenilacético/farmacologia , Antineoplásicos/farmacologia , Bactérias/classificação , Bromobenzoatos/farmacologia , Ácido Gálico/farmacologia , Hidroxibenzoatos/farmacologia , Quercetina/química , Ácido 3,4-Di-Hidroxifenilacético/química , Actinobacteria/genética , Actinobacteria/isolamento & purificação , Actinobacteria/metabolismo , Antineoplásicos/química , Bacillus/genética , Bacillus/isolamento & purificação , Bacillus/metabolismo , Bactérias/genética , Bactérias/isolamento & purificação , Bactérias/metabolismo , Proteínas de Bactérias , Bacteroides/genética , Bacteroides/isolamento & purificação , Bacteroides/metabolismo , Bromobenzoatos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Clostridiales/genética , Clostridiales/isolamento & purificação , Clostridiales/metabolismo , Eubacterium/genética , Eubacterium/isolamento & purificação , Eubacterium/metabolismo , Ácido Gálico/química , Microbioma Gastrointestinal , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Células HCT116 , Humanos , Hidroxibenzoatos/química , Filogenia , Análise de Sequência de RNA
17.
Int J Mol Sci ; 22(13)2021 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-34208896

RESUMO

In this paper, we report our investigations on five T30175 analogues, prepared by replacing sequence thymidines with abasic sites (S) one at a time, in comparison to their natural counterpart in order to evaluate their antiproliferative potential and the involvement of the residues not belonging to the central core of stacked guanosines in biological activity. The collected NMR (Nuclear Magnetic Resonance), CD (Circular Dichroism), and PAGE (Polyacrylamide Gel Electrophoresis) data strongly suggest that all of them adopt G-quadruplex (G4) structures strictly similar to that of the parent aptamer with the ability to fold into a dimeric structure composed of two identical G-quadruplexes, each characterized by parallel strands, three all-anti-G-tetrads and four one-thymidine loops (one bulge and three propeller loops). Furthermore, their antiproliferative (MTT assay) and anti-motility (wound healing assay) properties against lung and colorectal cancer cells were tested. Although all of the oligodeoxynucleotides (ODNs) investigated here exhibited anti-proliferative activity, the unmodified T30175 aptamer showed the greatest effect on cell growth, suggesting that both its characteristic folding in dimeric form and its presence in the sequence of all thymidines are crucial elements for antiproliferative activity. This straightforward approach is suitable for understanding the critical requirements of the G-quadruplex structures that affect antiproliferative potential and suggests its application as a starting point to facilitate the reasonable development of G-quadruplexes with improved anticancer properties.


Assuntos
Antineoplásicos/química , Aptâmeros de Nucleotídeos/química , Neoplasias Colorretais/genética , Neoplasias Pulmonares/genética , Timidina/genética , Substituição de Aminoácidos , Antineoplásicos/farmacologia , Aptâmeros de Nucleotídeos/genética , Aptâmeros de Nucleotídeos/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Dicroísmo Circular , Neoplasias Colorretais/tratamento farmacológico , Ensaios de Seleção de Medicamentos Antitumorais , Quadruplex G , Células HCT116 , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Espectroscopia de Ressonância Magnética
18.
Molecules ; 26(12)2021 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-34201326

RESUMO

The development of cancer treatments requires continuous exploration and improvement, in which the discovery of new drugs for the treatment of cancer is still an important pathway. In this study, based on the molecular hybridization strategy, a new structural framework with an N-aryl-N'-arylmethylurea scaffold was designed, and 16 new target compounds were synthesized and evaluated for their antiproliferative activities against four different cancer cell lines A549, MCF7, HCT116, PC3, and human liver normal cell line HL7702. The results have shown seven compounds with 1-methylpiperidin-4-yl groups having excellent activities against all four cancer cell lines, and they exhibited scarcely any activities against HL7702. Among them, compound 9b and 9d showed greatly excellent activity against the four kinds of cells, and the IC50 for MCF7 and PC3 cell lines were even less than 3 µM.


Assuntos
Antineoplásicos/química , Antineoplásicos/síntese química , Ureia/química , Ureia/síntese química , Células A549 , Antineoplásicos/farmacologia , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Células HCT116 , Humanos , Células PC-3 , Relação Estrutura-Atividade , Ureia/farmacologia
19.
Molecules ; 26(13)2021 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-34203232

RESUMO

Colorectal cancer is one of the life-threatening ailments causing high mortality and morbidity worldwide. Despite the innovation in medical genetics, the prognosis for metastatic colorectal cancer in patients remains unsatisfactory. Recently, lichens have attracted the attention of researchers in the search for targets to fight against cancer. Lichens are considered mines of thousands of metabolites. Researchers have reported that lichen-derived metabolites demonstrated biological effects, such as anticancer, antiviral, anti-inflammatory, antibacterial, analgesic, antipyretic, antiproliferative, and cytotoxic, on various cell lines. However, the exploration of the biological activities of lichens' metabolites is limited. Thus, the main objective of our study was to evaluate the anticancer effect of secondary metabolites isolated from lichen (Usnea barbata 2017-KL-10) on the human colorectal cancer cell line HCT116. In this study, 2OCAA exhibited concentration-dependent anticancer activities by suppressing antiapoptotic genes, such as MCL-1, and inducing apoptotic genes, such as BAX, TP53, and CDKN1A(p21). Moreover, 2OCAA inhibited the migration and invasion of colorectal cancer cells in a concentration-dependent manner. Taken together, these data suggest that 2OCAA is a better therapeutic candidate for colorectal cancer.


Assuntos
Antineoplásicos , Apoptose/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Triterpenos , Usnea/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Neoplasias Colorretais/metabolismo , Células HCT116 , Humanos , Triterpenos/química , Triterpenos/farmacologia
20.
Molecules ; 26(13)2021 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-34202689

RESUMO

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancers worldwide. One of its subtypes is associated with defective mismatch repair (dMMR) genes. Saffron has many potentially protective roles against colon malignancy. However, these roles in the context of dMMR tumors have not been explored. In this study, we aimed to investigate the effects of saffron and its constituents in CRC cell lines with dMMR. METHODS: Saffron crude extracts and specific compounds (safranal and crocin) were used in the human colorectal cancer cell lines HCT116, HCT116+3 (inserted MLH1), HCT116+5 (inserted MSH3), and HCT116+3+5 (inserted MLH1 and MSH3). CDC25b, p-H2AX, TPDP1, and GAPDH were analyzed by Western blot. Proliferation and cytotoxicity were analyzed by MTT. The scratch wound assay was also performed. RESULTS: Saffron crude extracts restricted (up to 70%) the proliferation in colon cells with deficient MMR (HCT116) compared to proficient MMR. The wound healing assay indicates that deficient MMR cells are doing better (up to 90%) than proficient MMR cells when treated with saffron. CDC25b and TDP1 downregulated (up to 20-fold) in proficient MMR cells compared to deficient MMR cells, while p.H2AX was significantly upregulated in both cell types, particularly at >10 mg/mL saffron in a concentration-dependent manner. The reduction in cellular proliferation was accompanied with upregulation of caspase 3 and 7. The major active saffron compounds, safranal and crocin reproduced most of the saffron crude extracts' effects. CONCLUSIONS: Saffron's anti-proliferative effect is significant in cells with deficient MMR. This novel effect may have therapeutic implications and benefits for MSI CRC patients who are generally not recommended for the 5-fluorouracil-based treatment.


Assuntos
Neoplasias do Colo/tratamento farmacológico , Crocus/química , Reparo de Erro de Pareamento de DNA/efeitos dos fármacos , Instabilidade de Microssatélites/efeitos dos fármacos , Extratos Vegetais/farmacologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células HCT116 , Humanos , Extratos Vegetais/química
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