RESUMEN
Aldehyde dehydrogenases (ALDHs) are a family of enzymes that aid in detoxification and are overexpressed in several different malignancies. There is a correlation between increased expression of ALDH and a poor prognosis, stemness, and resistance to several drugs. Several ALDH inhibitors have been generated due to the crucial role that ALDH plays in cancer stem cells. All of these inhibitors, however, are either ineffective, very toxic, or have yet to be subjected to rigorous testing on their effectiveness. Although various drug-like compounds targeting ALDH have been reported in the literature, none have made it to routine use in the oncology clinic. As a result, new potent, non-toxic, bioavailable, and therapeutically effective ALDH inhibitors are still needed. In this study, we designed and synthesized potent multi-ALDH isoform inhibitors based on the isatin and indazole pharmacophore. Molecular docking studies and enzymatic tests revealed that among all of the synthesized analogs, compound 3 is the most potent inhibitor of ALDH1A1, ALDH3A1, and ALDH1A3, exhibiting 51.32%, 51.87%, and 36.65% inhibition, respectively. The ALDEFLUOR assay further revealed that compound 3 acts as an ALDH broad spectrum inhibitor at 500 nM. Compound 3 was also the most cytotoxic to cancer cells, with an IC50 in the range of 2.1 to 3.8 µM for ovarian, colon, and pancreatic cancer cells, compared to normal and embryonic kidney cells (IC50 7.1 to 8.7 µM). Mechanistically, compound 3 increased ROS activity due to potent multi-ALDH isoform inhibition, which increased apoptosis. Taken together, this study identified a potent multi-isoform ALDH inhibitor that could be further developed as a cancer therapeutic.
Asunto(s)
Aldehído Deshidrogenasa , Inhibidores Enzimáticos , Isatina , Simulación del Acoplamiento Molecular , Humanos , Isatina/química , Isatina/farmacología , Aldehído Deshidrogenasa/antagonistas & inhibidores , Aldehído Deshidrogenasa/metabolismo , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/síntesis química , Línea Celular Tumoral , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/síntesis química , Relación Estructura-Actividad , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Estructura MolecularRESUMEN
Current cancer therapy can be effective, but the development of drug resistant disease is the usual outcome. These drugs can eliminate most of the tumor burden but often fail to eliminate the rare, "Drug Tolerant Persister" (DTP) cell subpopulations in residual tumors, which can be referred to as "Persister" cells. Therefore, novel therapeutic agents specifically targeting or preventing the development of drug-resistant tumors mediated by the remaining persister cells subpopulations are needed. Since approximately ninety percent of cancer-related deaths occur because of the eventual development of drug resistance, identifying, and dissecting the biology of the persister cells is essential for the creation of drugs to target them. While there remains uncertainty surrounding all the markers identifying DTP cells in the literature, this review summarizes the drugs and therapeutic approaches that are available to target the persister cell subpopulations expressing the cellular markers ATP-binding cassette sub-family B member 5 (ABCB5), CD133, CD271, Lysine-specific histone demethylase 5 (KDM5), and aldehyde dehydrogenase (ALDH). Persister cells expressing these markers were selected as the focus of this review because they have been found on cells surviving following drug treatments that promote recurrent drug resistant cancer and are associated with stem cell-like properties, including self-renewal, differentiation, and resistance to therapy. The limitations and obstacles facing the development of agents targeting these DTP cell subpopulations are detailed, with discussion of potential solutions and current research areas needing further exploration.
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Antineoplásicos , Resistencia a Antineoplásicos , Neoplasias , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Animales , Resistencia a Antineoplásicos/efectos de los fármacos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Tolerancia a Medicamentos , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/genéticaRESUMEN
The repertoire of modifications to bile acids and related steroidal lipids by host and microbial metabolism remains incompletely characterized. To address this knowledge gap, we created a reusable resource of tandem mass spectrometry (MS/MS) spectra by filtering 1.2 billion publicly available MS/MS spectra for bile-acid-selective ion patterns. Thousands of modifications are distributed throughout animal and human bodies as well as microbial cultures. We employed this MS/MS library to identify polyamine bile amidates, prevalent in carnivores. They are present in humans, and their levels alter with a diet change from a Mediterranean to a typical American diet. This work highlights the existence of many more bile acid modifications than previously recognized and the value of leveraging public large-scale untargeted metabolomics data to discover metabolites. The availability of a modification-centric bile acid MS/MS library will inform future studies investigating bile acid roles in health and disease.
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Ácidos y Sales Biliares , Microbioma Gastrointestinal , Metabolómica , Espectrometría de Masas en Tándem , Animales , Humanos , Ácidos y Sales Biliares/química , Metabolómica/métodos , Poliaminas , Espectrometría de Masas en Tándem/métodos , Bases de Datos de Compuestos QuímicosRESUMEN
Tobacco smoking and human papillomavirus infection are established etiological agents in the development of head and neck squamous cell carcinoma (HNSCC). The incidence and mortality of HNSCC are higher in men than women. To provide biochemical basis for sex differences, we tested the hypothesis that carcinogen treatment using dibenzo[def,p]chrysene, which is an environmental pollutant and tobacco smoke constituent, in the absence or presence of the mouse papillomavirus infection results in significantly higher levels of DNA damage in the oral cavity in male than in female mice. However, the results of the present investigation do not support our hypothesis since we found that females were more susceptible to carcinogen-induced covalent DNA damage than males independent of the viral infection. Since DNA damage represents only a single-step in the carcinogenesis process, additional factors may contribute to sex differences in humans.
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Bacteria in the gastrointestinal tract produce amino acid bile acid amidates that can affect host-mediated metabolic processes1-6; however, the bacterial gene(s) responsible for their production remain unknown. Herein, we report that bile salt hydrolase (BSH) possesses dual functions in bile acid metabolism. Specifically, we identified a previously unknown role for BSH as an amine N-acyltransferase that conjugates amines to bile acids, thus forming bacterial bile acid amidates (BBAAs). To characterize this amine N-acyltransferase BSH activity, we used pharmacological inhibition of BSH, heterologous expression of bsh and mutants in Escherichia coli and bsh knockout and complementation in Bacteroides fragilis to demonstrate that BSH generates BBAAs. We further show in a human infant cohort that BBAA production is positively correlated with the colonization of bsh-expressing bacteria. Lastly, we report that in cell culture models, BBAAs activate host ligand-activated transcription factors including the pregnane X receptor and the aryl hydrocarbon receptor. These findings enhance our understanding of how gut bacteria, through the promiscuous actions of BSH, have a significant role in regulating the bile acid metabolic network.
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Aciltransferasas , Amidohidrolasas , Aminas , Ácidos y Sales Biliares , Biocatálisis , Microbioma Gastrointestinal , Humanos , Aciltransferasas/metabolismo , Amidohidrolasas/metabolismo , Aminas/química , Aminas/metabolismo , Bacteroides fragilis/enzimología , Bacteroides fragilis/genética , Bacteroides fragilis/metabolismo , Ácidos y Sales Biliares/química , Ácidos y Sales Biliares/metabolismo , Estudios de Cohortes , Escherichia coli/enzimología , Escherichia coli/genética , Escherichia coli/metabolismo , Microbioma Gastrointestinal/fisiología , Ligandos , Receptor X de Pregnano/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Factores de Transcripción/metabolismo , Lactante , Técnicas de Cultivo de CélulaRESUMEN
The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor that plays an important role in gastrointestinal barrier function, tumorigenesis, and is an emerging drug target. The resident microbiota is capable of metabolizing tryptophan to metabolites that are AHR ligands (e.g., indole-3-acetate). Recently, a novel set of mutagenic tryptophan metabolites named indolimines have been identified that are produced by M. morganii in the gastrointestinal tract. Here, we determined that indolimine-200, -214, and -248 are direct AHR ligands that can induce Cyp1a1 transcription and subsequent CYP1A1 enzymatic activity capable of metabolizing the carcinogen benzo(a)pyrene in microsomal assays. In addition, indolimines enhance IL6 expression in a colonic tumor cell line in combination with cytokine treatment. The concentration of indolimine-248 that induces AHR transcriptional activity failed to increase DNA damage. These observations reveal an additional aspect of how indolimines may alter colonic tumorigenesis beyond mutagenic activity.
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Long-term ligand activation of PPARα in mice causes hepatocarcinogenesis through a mechanism that requires functional PPARα. However, hepatocarcinogenesis is diminished in both Ppara-null and PPARA-humanized mice, yet both lines develop age-related liver cancer independently of treatment with a PPARα agonist. Since PPARα is a master regulator of liver lipid metabolism in the liver, lipidomic analyses were carried out in wild-type, Ppara-null, and PPARA-humanized mice treated with and without the potent agonist GW7647. The levels of hepatic linoleic acid in Ppara-null and PPARA-humanized mice were markedly higher compared to wild-type controls, along with overall fatty liver. The number of liver CD4+ T cells was also lower in Ppara-null and PPARA-humanized mice and was negatively correlated with the elevated linoleic acid. Moreover, more senescent hepatocytes and lower serum TNFα and IFNγ levels were observed in Ppara-null and PPARA-humanized mice with age. These studies suggest a new role for PPARα in age-associated hepatocarcinogenesis due to altered lipid metabolism in Ppara-null and PPARA-humanized mice and the accumulation of linoleic acid as part of an overall fatty liver that is associated with loss of CD4+ T cells in the liver in both transgenic models. Since fatty liver is a known causal risk factor for liver cancer, Ppara-null and PPARA-humanized mice are valuable models for examining the mechanisms of PPARα and age-dependent hepatocarcinogenesis.
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The Rho associated coiled-coil containing protein kinase (ROCK1 and ROCK2) and myotonic dystrophy-related Cdc-42 binding kinases (MRCKα and MRCKß) are critical regulators of cell proliferation and cell plasticity, a process intimately involved in cancer cell migration and invasion. Previously, we reported the discovery of a novel small molecule (DJ4) selective multi-kinase inhibitor of ROCK1/2 and MRCKα/ß. Herein, we further characterized the anti-proliferative and apoptotic effects of DJ4 in non-small cell lung cancer and triple-negative breast cancer cells. To further optimize the ROCK/MRCK inhibitory potency of DJ4, we generated a library of 27 analogs. Among the various structural modifications, we identified four additional active analogs with enhanced ROCK/MRCK inhibitory potency. The anti-proliferative and cell cycle inhibitory effects of the active analogs were examined in non-small cell lung cancer, breast cancer, and melanoma cell lines. The anti-proliferative effectiveness of DJ4 and the active analogs was further demonstrated against a wide array of cancer cell types using the NCI-60 human cancer cell line panel. Lastly, these new analogs were tested for anti-migratory effects in highly invasive MDA-MB-231 breast cancer cells. Together, our results demonstrate that selective inhibitors of ROCK1/2 (DJE4, DJ-Allyl) inhibited cell proliferation and induced cell cycle arrest at G2/M but were less effective in cell death induction compared with dual ROCK1/2 and MRCKα/ß (DJ4 and DJ110).
RESUMEN
The aryl hydrocarbon receptor (AHR) is a ligand activated transcription factor that plays an integral role in homeostatic maintenance by regulating cellular functions such as cellular differentiation, metabolism, barrier function, and immune response. An important but poorly understood class of AHR activators are compounds derived from host and bacterial metabolism of tryptophan. The commensal bacteria of the gut microbiome are major producers of tryptophan metabolites known to activate the AHR, while the host also produces AHR activators through tryptophan metabolism. We used targeted mass spectrometry-based metabolite profiling to determine the presence and metabolic source of these metabolites in the sera of conventional mice, germ-free mice, and humans. Surprisingly, sera concentrations of many tryptophan metabolites are comparable between germ-free and conventional mice. Therefore, many major AHR-activating tryptophan metabolites in mouse sera are produced by the host, despite their presence in feces and mouse cecal contents. Here we present an investigation of AHR activation using a complex mixture of tryptophan metabolites to examine the biological relevance of circulating tryptophan metabolites. AHR activation is rarely studied in the context of a mixture at relevant concentrations, as we present here. The AHR activation potentials of individual and pooled metabolites were explored using cell-based assays, while ligand binding competition assays and ligand docking simulations were used to assess the detected metabolites as AHR agonists. The physiological and biomedical relevance of the identified metabolites was investigated in the context of a cell-based model for rheumatoid arthritis. We present data that reframe AHR biology to include the presence of a mixture of ubiquitous tryptophan metabolites, improving our understanding of homeostatic AHR activity and models of AHR-linked diseases.
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The aryl hydrocarbon receptor (AHR) mediates intestinal barrier homeostasis. Many AHR ligands are also CYP1A1/1B1 substrates, which can result in rapid clearance within the intestinal tract, limiting systemic exposure and subsequent AHR activation. This led us to the hypothesis that there are dietary substrates of CYP1A1/1B1 that functionally increase the half-life of potent AHR ligands. We examined the potential of urolithin A (UroA), a gut bacterial metabolite of ellagitannins, as a CYP1A1/1B1 substrate to enhance AHR activity in vivo. UroA is a competitive substrate for CYP1A1/1B1 in an in vitro competition assay. A broccoli-containing diet promotes the gastric formation of the potent hydrophobic AHR ligand and CYP1A1/1B1 substrate, 5,11-dihydroindolo[3,2-b]carbazole (ICZ). In mice, dietary exposure to UroA in a 10% broccoli diet led to a coordinated increase in duodenal, cardiac, and pulmonary AHR activity, but no increase in activity in the liver. Thus, CYP1A1 dietary competitive substrates can lead to enhanced systemic AHR ligand distribution from the gut, likely through the lymphatic system, increasing AHR activation in key barrier tissues. Finally, this report will lead to a reassessment of the dynamics of distribution of other hydrophobic chemicals present in the diet.
Asunto(s)
Citocromo P-450 CYP1A1 , Tracto Gastrointestinal , Pulmón , Receptores de Hidrocarburo de Aril , Animales , Ratones , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Ligandos , Hígado/metabolismo , Pulmón/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Dieta , Tracto Gastrointestinal/metabolismoRESUMEN
In the face of mechanical, chemical, microbial, and immunologic pressure, intestinal homeostasis is maintained through balanced cellular turnover, proliferation, differentiation, and self-renewal. Here, we present evidence supporting the role of the aryl hydrocarbon receptor (AHR) in the adaptive reprogramming of small intestinal gene expression, leading to altered proliferation, lineage commitment, and remodeling of the cellular repertoire that comprises the intestinal epithelium to promote intestinal resilience. Ahr gene/protein expression and transcriptional activity exhibit marked proximalHI to distalLO and cryptHI to villiLO gradients. Genetic ablation of Ahr impairs commitment/differentiation of the secretory Paneth and goblet cell lineages and associated mucin production, restricts expression of secretory/enterocyte differentiation markers, and increases crypt-associated proliferation and villi-associated enterocyte luminal exfoliation. Ahr-/- mice display a decrease in intestinal barrier function. Ahr+/+ mice that maintain a diet devoid of AHR ligands intestinally phenocopy Ahr-/- mice. In contrast, Ahr+/+ mice exposed to AHR ligands reverse these phenotypes. Ligand-induced AHR transcriptional activity positively correlates with gene expression (Math1, Klf4, Tff3) associated with differentiation of the goblet cell secretory lineage. Math1 was identified as a direct target gene of AHR, a transcription factor critical to the development of goblet cells. These data suggest that dietary cues, relayed through the transcriptional activity of AHR, can reshape the cellular repertoire of the gastrointestinal tract.
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Células Epiteliales , Receptores de Hidrocarburo de Aril , Animales , Ratones , Diferenciación Celular , Células Epiteliales/metabolismo , Mucosa Intestinal/metabolismo , Intestinos , Ligandos , Receptores de Hidrocarburo de Aril/genética , Receptores de Hidrocarburo de Aril/metabolismoRESUMEN
The aryl hydrocarbon receptor (AHR) mediates intestinal barrier homeostasis. Many AHR ligands are also CYP1A1/1B1 substrates, which can result in the rapid clearance within the intestinal tract, limiting AHR activation. This led us to the hypothesis that there are dietary substrates of CYP1A1/1B1 that increase the half-life of potent AHR ligands. We examined the potential of urolithin A (UroA) as a CYP1A1/1B1 substrate to enhance AHR activity in vivo. UroA is a competitive substrate for CYP1A1/1B1 in an in vitro competition assay. A broccoli-containing diet promotes the gastric formation of the potent hydrophobic AHR ligand and CYP1A1/1B1 substrate, 5,11-dihydroindolo[3,2-b]carbazole (ICZ). Dietary exposure to UroA in a broccoli diet led to a coordinated increase in duodenal, cardiac, and pulmonary AHR activity, but no increase in activity in liver. Thus, CYP1A1 dietary competitive substrates can lead to intestinal escape, likely through the lymphatic system, increasing AHR activation in key barrier tissues.
RESUMEN
The aryl hydrocarbon receptor (AHR) is a ligand activated transcription factor that plays an integral role in homeostatic maintenance by regulating cellular functions such as cellular differentiation, metabolism, barrier function, and immune response. An important but poorly understood class of AHR activators are compounds derived from host and bacterial metabolism of tryptophan. The commensal bacteria of the gut microbiome are major producers of tryptophan metabolites known to activate the AHR, while the host also produces AHR activators through tryptophan metabolism. We used targeted mass spectrometry-based metabolite profiling to determine the presence and metabolic source of these metabolites in the sera of conventional mice, germ-free mice, and humans. Surprisingly, sera concentrations of many tryptophan metabolites are comparable between germ-free and conventional mice. Therefore, many major AHR-activating tryptophan metabolites in mouse sera are produced by the host, despite their presence in feces and mouse cecal contents. AHR activation is rarely studied in the context of a mixture at relevant concentrations, as we present here. The AHR activation potentials of individual and pooled metabolites were explored using cell-based assays, while ligand binding competition assays and ligand docking simulations were used to assess the detected metabolites as AHR agonists. The physiological and biomedical relevance of the identified metabolites was investigated in the context of cell-based models for cancer and rheumatoid arthritis. We present data here that reframe AHR biology to include the presence of ubiquitous tryptophan metabolites, improving our understanding of homeostatic AHR activity and models of AHR-linked diseases.
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Therapeutic aryl hydrocarbon receptor (AHR) modulating agents gained attention in dermatology as non-steroidal anti-inflammatory drugs that improve skin barrier properties. By exploiting AHR's known ligand promiscuity, we generated novel AHR modulating agents by lead optimization of a selective AHR modulator (SAhRM; SGA360). Twenty-two newly synthesized compounds were screened yielding two novel derivatives, SGA360f and SGA388, in which agonist activity led to enhanced keratinocyte terminal differentiation. SGA388 showed the highest agonist activity with potent normalization of keratinocyte hyperproliferation, restored expression of skin barrier proteins and dampening of chemokine expression by keratinocytes upon Th2-mediated inflammation in vitro. The topical application of SGA360f and SGA388 reduced acute skin inflammation in vivo by reducing cyclooxygenase levels, resulting in less neutrophilic dermal infiltrates. The minimal induction of cytochrome P450 enzyme activity, lack of cellular toxicity and mutagenicity classifies SGA360f and SGA388 as novel potential therapeutic AHR ligands and illustrates the potential of medicinal chemistry to fine-tune AHR signaling for the development of targeted therapies in dermatology and beyond.
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Receptores de Hidrocarburo de Aril , Enfermedades de la Piel , Humanos , Receptores de Hidrocarburo de Aril/metabolismo , Ligandos , Piel/metabolismo , Queratinocitos/metabolismo , Inflamación/metabolismo , Enfermedades de la Piel/tratamiento farmacológicoRESUMEN
Ulcerative colitis (UC) is an idiopathic, chronic and relapsing colonic inflammatory disease. Despite the involvement of diverse intricate mechanisms, COX mediated inflammatory pathway is crucial in the pathophysiology of colitis. Thus, COX inhibition is imperative for managing colitis-associated inflammation. However, the use of COX inhibitory classical non-steroidal anti-inflammatory drugs (NSAIDs) for inflammation resolution has been linked to sudden increased flare-ups. Therefore, considering the anti-inflammatory and pro-resolution effects of antioxidant and essential trace element Selenium (Se), a Seleno-derivative of Celecoxib called Selenocoxib-3 was characterized and evaluated for its favourable pharmacokinetics, safety margins and anti-inflammatory therapeutic potential in DSS-induced experimental colitis. The serum pharmacokinetic profiling [elimination rate constant (K) and clearance (Cl) and toxicity profiling suggested enhanced efficacy, therapeutic potential and lesser toxicity of Selenocoxib-3 as compared to its parent NSAID Celecoxib. In vivo studies demonstrated that Selenocoxib-3 efficiently resolves the gross morphological signs of DSS-induced colitis such as diarrhoea, bloody stools, weight loss and colon shortening. Further, intestinal damage evaluated by H & E staining and MPO activity suggested of histopathological disruptions, such as neutrophil infiltration, mucodepletion and cryptitis, by Selenocoxib-3. The expression profiles of COX-1/2 demonstrated mitigation of pro-inflammatory mediators thereby promoting anti-inflammatory efficacy of Selenocoxib-3 when compared with Celecoxib. The current study suggests translational applicability of Se-containing novel class of COX inhibitors for efficiently managing inflammatory disorders such as UC.
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Colitis Ulcerosa , Colitis , Animales , Celecoxib/efectos adversos , Antiinflamatorios/farmacología , Colitis/metabolismo , Colitis Ulcerosa/inducido químicamente , Colitis Ulcerosa/tratamiento farmacológico , Colitis Ulcerosa/patología , Antiinflamatorios no Esteroideos/farmacología , Colon , Inflamación/metabolismo , Ciclooxigenasa 2/metabolismo , Sulfato de Dextran/farmacología , Modelos Animales de EnfermedadRESUMEN
(1) Background: Recently, a number of side chain length variants for tetrahydrocannabinol and cannabidiol have been identified in cannabis; however, the precursor to these molecules would be based upon cannabigerol (CBG). Because CBG, and its side chain variants, are rapidly converted to other cannabinoids in the plant, there are typically only small amounts in plant extracts, thus prohibiting investigations related to CBG and CBG variant therapeutic effects. (2) Methods: To overcome this, we developed an efficient synthesis of corresponding resorcinol fragments using the Wittig reaction which, under acid catalyzed coupling with geraniol, produced the desired side chain variants of CBG. These compounds were then tested in an animal model of chemotherapeutic-induced neuropathic pain and to reduce colorectal cancer cell viability. (3) Results: We found that all side-chain variants were similarly capable of reducing neuropathic pain in mice at a dose of 10 mg/kg. However, the molecules with shorter side chains (i.e., CBGV and CBGB) were better at reducing colorectal cancer cell viability. (4) Conclusions: The novel synthesis method developed here will be of utility for studying other side chain derivatives of minor cannabinoids such as cannabichromene, cannabinol, and cannabielsoin.
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Cannabinoides , Cannabis , Neoplasias Colorrectales , Neuralgia , Ratones , Animales , Cannabinoides/farmacología , Cannabis/química , Dronabinol , Neuralgia/tratamiento farmacológicoRESUMEN
In a series of previous studies we reported that black raspberry (BRB) powder inhibits dibenzo[a,l]pyrene (DBP)-induced DNA damage, mutagenesis, and oral squamous cell carcinoma (OSCC) development in mice. In the present study, using human oral leukoplakia (MSK-Leuk1) and squamous cell carcinoma (SCC1483) cells, we tested the hypothesis that BRB extract (BRBE) will enhance the synthesis of glutathione (GSH) and in turn increase GSH conjugation of the fjord-region DBP diol epoxide (DBPDE) derived from DBP leading to inhibition of DBP-induced DNA damage. The syntheses of DBPDE-GSH conjugate, DBPDE-dA adduct, and the corresponding isotope-labeled internal standards were performed; LC-MS/MS methods were used for their quantification. BRBE significantly (p < 0.05) increased cellular GSH by 31% and 13% at 6 and 24 h, respectively, in OSCC cells; in MSK-LeuK1 cells, the levels of GSH significantly (p < 0.05) increased by 55% and 22%, at 1 and 6 h. Since BRBE significantly enhanced the synthesis of GSH in both cell types, subsequent experiments were performed in MSK-Leuk1 cells. Western blot analysis was performed to determine the types of proteins involved in the synthesis of GSH. BRBE significantly (p < 0.05) increased the protein expression (2.5-fold) of the glutamate-cysteine ligase catalytic subunit (GCLC) but had no effect on the glutamate-cysteine ligase modifier subunit (GCLM) and glutathione synthetase (GSS). LC-MS/MS analysis showed that pretreatment of cells with BRBE followed by DBPDE significantly (p < 0.05) increased the levels of DBPDE-GSH conjugate (2.5-fold) and decreased DNA damage by 74% measured by assessing levels of DBPDE-dA adduct formation. Collectively, the results of this in vitro study clearly support our hypothesis, and the LC-MS/MS methods developed in the present study will be highly useful in testing the same hypothesis initially in our mouse model and ultimately in smokers.
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Carcinoma de Células Escamosas , Neoplasias de la Boca , Rubus , Humanos , Ratones , Animales , Carcinógenos , Crisenos , Benzopirenos/metabolismo , Compuestos Epoxi , Nicotiana/metabolismo , Glutamato-Cisteína Ligasa , Aductos de ADN , Cromatografía Liquida , Estuarios , Neoplasias de la Boca/inducido químicamente , Espectrometría de Masas en Tándem , Glutatión/metabolismo , Extractos Vegetales/farmacologíaRESUMEN
Nonalcoholic steatohepatitis (NASH) is a rapidly developing pathology around the world, with limited treatment options available. Some farnesoid X receptor (FXR) agonists have been applied in clinical trials for NASH, but side effects such as pruritus and low-density lipoprotein elevation have been reported. Intestinal FXR is recognized as a promising therapeutic target for metabolic diseases. Glycine-ß-muricholic acid (Gly-MCA) is an intestine-specific FXR antagonist previously shown to have favorable metabolic effects on obesity and insulin resistance. Herein, we identify a role for Gly-MCA in the pathogenesis of NASH, and explore the underlying molecular mechanism. Gly-MCA improved lipid accumulation, inflammatory response, and collagen deposition in two different NASH models. Mechanistically, Gly-MCA decreased intestine-derived ceramides by suppressing ceramide synthesis-related genes via decreasing intestinal FXR signaling, leading to lower liver endoplasmic reticulum (ER) stress and proinflammatory cytokine production. The role of bile acid metabolism and adiposity was excluded in the suppression of NASH by Gly-MCA, and a correlation was found between intestine-derived ceramides and NASH severity. This study revealed that Gly-MCA, an intestine-specific FXR antagonist, has beneficial effects on NASH by reducing ceramide levels circulating to liver via lowering intestinal FXR signaling, and ceramide production, followed by decreased liver ER stress and NASH progression. Intestinal FXR is a promising drug target and Gly-MCA a novel agent for the prevention and treatment of NASH.
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Enfermedad del Hígado Graso no Alcohólico , Ratones , Animales , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Ceramidas/metabolismo , Glicina/farmacología , Receptores Citoplasmáticos y Nucleares/farmacología , Intestinos , Obesidad/tratamiento farmacológicoRESUMEN
Despite recent improvements in multiple myeloma (MM) treatment, MM remains an incurable disease and most patients experience a relapse. The major reason for myeloma recurrence is the persistent stem cell-like population. It has been demonstrated that overexpression of Bruton's tyrosine kinase (BTK) in MM stem cell-like cells is correlated with drug resistance and poor prognosis. We have developed a novel small BTK inhibitor, KS151, which is unique compared to other BTK inhibitors. Unlike ibrutinib, and the other BTK inhibitors such as acalabrutinib, orelabrutinib, and zanubrutinib that covalently bind to the C481 residue in the BTK kinase domain, KS151 can inhibit BTK activities without binding to C481. This feature of KS151 is important because C481 becomes mutated in many patients and causes drug resistance. We demonstrated that KS151 inhibits in vitro BTK kinase activities and is more potent than ibrutinib. Furthermore, by performing a semi-quantitative, sandwich-based array for 71-tyrosine kinase phosphorylation, we found that KS151 specifically inhibits BTK. Our western blotting data showed that KS151 inhibits BTK signaling pathways and is effective against bortezomib-resistant cells as well as MM stem cell-like cells. Moreover, KS151 potentiates the apoptotic response of bortezomib, lenalidomide, and panobinostat in both MM and stem cell-like cells. Interestingly, KS151 inhibits stemness markers and is efficient in inhibiting Nanog and Gli1 stemness markers even when MM cells were co-cultured with bone marrow stromal cells (BMSCs). Overall, our results show that we have developed a novel BTK inhibitor effective against the stem cell-like population, and potentiates the response of chemotherapeutic agents.
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A series of seleno-biotin analogs were synthesized and their anticancer activity and mode of action were assessed using ovarian cancer cells. Compound 2, out of the other analogs, in direct comparison to biotin alone, more effectively reduced the cell viability and induced apoptosis in ovarian cancer cell lines in a dose dependent manner as demonstrated by the cell viability assay, trypan blue dye exclusion assay, Annexin V/7-AAD, and Caspase 3/7 apoptosis assays. Furthermore, compound 2 showed efficacy better than 5-fluorouracil (5-FU) and similar to cisplatin, in vitro; notably it was more cytotoxic to drug-resistant Hey A8 cells than cisplatin. The cytotoxicity of compound 2 was primarily mediated by reactive oxygen species (ROS) as demonstrated by DCFDA based ROS estimation. Biotin receptors (BR) saturation and the use of a BR negative cell line showed a significant decline in the cytotoxic ativity of the compound 2, confirming that its activity is BR-mediated. These experiments demonstrated that selenium modified biotin which contains an ester linked redox cycling selenocyanate group has the potential for human therapeutic applications against ovarian and other cancers over-expressing BR.