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1.
Molecules ; 29(13)2024 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-38999066

RESUMO

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.


Assuntos
Aldeído Desidrogenase , Inibidores Enzimáticos , Isatina , Simulação de Acoplamento Molecular , Humanos , Isatina/química , Isatina/farmacologia , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/metabolismo , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/síntese química , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/síntese química , Relação Estrutura-Atividade , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Estrutura Molecular
2.
Cell ; 187(13): 3303-3318.e18, 2024 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-38906101

RESUMO

Gamete formation and subsequent offspring development often involve extended phases of suspended cellular development or even dormancy. How cells adapt to recover and resume growth remains poorly understood. Here, we visualized budding yeast cells undergoing meiosis by cryo-electron tomography (cryoET) and discovered elaborate filamentous assemblies decorating the nucleus, cytoplasm, and mitochondria. To determine filament composition, we developed a "filament identification" (FilamentID) workflow that combines multiscale cryoET/cryo-electron microscopy (cryoEM) analyses of partially lysed cells or organelles. FilamentID identified the mitochondrial filaments as being composed of the conserved aldehyde dehydrogenase Ald4ALDH2 and the nucleoplasmic/cytoplasmic filaments as consisting of acetyl-coenzyme A (CoA) synthetase Acs1ACSS2. Structural characterization further revealed the mechanism underlying polymerization and enabled us to genetically perturb filament formation. Acs1 polymerization facilitates the recovery of chronologically aged spores and, more generally, the cell cycle re-entry of starved cells. FilamentID is broadly applicable to characterize filaments of unknown identity in diverse cellular contexts.


Assuntos
Gametogênese , Mitocôndrias , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Aldeído Desidrogenase/metabolismo , Aldeído Desidrogenase/química , Núcleo Celular/metabolismo , Núcleo Celular/ultraestrutura , Coenzima A Ligases/metabolismo , Microscopia Crioeletrônica , Citoplasma/metabolismo , Tomografia com Microscopia Eletrônica , Meiose , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Esporos Fúngicos/metabolismo , Modelos Moleculares , Estrutura Quaternária de Proteína
3.
Theriogenology ; 223: 98-107, 2024 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-38697014

RESUMO

The ALDH1A1 gene encodes a cytoplasmic member of the aldehyde dehydrogenase 1 family, which plays an important role in regulating animal reproductive performance, including estrus cycle and embryonic development. The aim of this study was to characterize ALDH1A1 activity in ovaries of 3-5 year-old yaks and to determine its effects on cell proliferation, apoptosis, and progesterone secretion in luteal cells (LCs). The coding sequence (CDS) of the ALDH1A1 gene was cloned by reverse transcription-PCR and immunohistochemical analysis was used to confirm localization of the ALDH1A1 protein in the ovary. To assess the activity of ALDH1A1 in regulating progesterone secretion, si-ALDH1A1 was transfected into LCs in vitro and progesterone levels in LC supernatants were measured by ELISA. The interference efficiency was assessed by real-time quantitative PCR (RT-qPCR) and immunofluorescence staining, and cell proliferation and apoptosis were evaluated by EdU and TUNEL staining, respectively. The cloned ALDH1A1 sequence contained 1462 bp, encoding 487 amino acids. Immunohistochemical analysis showed that ALDH1A1 protein expression, which was significantly higher in LCs, was mainly found in antral follicles and the corpus luteum (CL). The expression of ALDH1A1 mRNA in LCs was effectively inhibited by si-ALDH1A1transfection, and progesterone secretion was markedly decreased along with the significant down-regulation of progesterone pathway-related genes, STAR, CYP11A1, CYP19A1, CYP17A1, 3ß-HSD, and HSD17B1. Knockdown of ALDH1A1 mRNA expression decreased cell proliferation and increased apoptosis in LCs. The mRNA expression of the proliferation-related genes, PCNA, CCND1, CCNB1 and CDC25A, was significantly down-regulated, while expression of the apoptosis-promoting CASP3 gene was significantly increased. In summary, we characterized the yak ALDH1A1 gene and revealed that ALDH1A1 knockdown promoted apoptosis, repressed cell proliferation, and decreased progesterone secretion by yak LCs, potentially by regulating the mRNA expression of genes related to proliferation, apoptosis, and progesterone synthesis and secretion.


Assuntos
Família Aldeído Desidrogenase 1 , Células Lúteas , Retinal Desidrogenase , Animais , Bovinos/genética , Feminino , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Família Aldeído Desidrogenase 1/genética , Família Aldeído Desidrogenase 1/metabolismo , Apoptose , Proliferação de Células , Regulação da Expressão Gênica/fisiologia , Células Lúteas/metabolismo , Progesterona/metabolismo , Retinal Desidrogenase/genética , Retinal Desidrogenase/metabolismo
4.
Pharmacol Ther ; 259: 108666, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38763322

RESUMO

Acute myocardial infarction (AMI) remains a leading cause of death worldwide. Increased formation of reactive oxygen species (ROS) during the early reperfusion phase is thought to trigger lipid peroxidation and disrupt redox homeostasis, leading to myocardial injury. Whilst the mitochondrial enzyme aldehyde dehydrogenase 2 (ALDH2) is chiefly recognised for its central role in ethanol metabolism, substantial experimental evidence suggests an additional cardioprotective role for ALDH2 independent of alcohol intake, which mitigates myocardial injury by detoxifying breakdown products of lipid peroxidation including the reactive aldehydes, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE). Epidemiological evidence suggests that an ALDH2 mutant variant with reduced activity that is highly prevalent in the East Asian population increases AMI risk. Additional studies have uncovered a strong association between coronary heart disease and this ALDH2 mutant variant. It appears this enzyme polymorphism (in particular, in ALDH2*2/2 carriers) has the potential to have wide-ranging effects on thiol reactivity, redox tone and therefore numerous redox-related signaling processes, resilience of the heart to cope with lifestyle-related and environmental stressors, and the ability of the whole body to achieve redox balance. In this review, we summarize the journey of ALDH2 from a mitochondrial reductase linked to alcohol metabolism, via pre-clinical studies aimed at stimulating ALDH2 activity to reduce myocardial injury to clinical evidence for its protective role in the heart.


Assuntos
Aldeído-Desidrogenase Mitocondrial , Etanol , Infarto do Miocárdio , Oxirredução , Polimorfismo Genético , Humanos , Aldeído-Desidrogenase Mitocondrial/genética , Aldeído-Desidrogenase Mitocondrial/metabolismo , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Animais , Etanol/metabolismo , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Espécies Reativas de Oxigênio/metabolismo
5.
Int J Biol Sci ; 20(7): 2763-2778, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38725845

RESUMO

Dysregulation of the aldehyde dehydrogenase (ALDH) family has been implicated in various pathological conditions, including cancer. However, a systematic evaluation of ALDH alterations and their therapeutic relevance in hepatocellular carcinoma (HCC) remains lacking. Herein, we found that 15 of 19 ALDHs were transcriptionally dysregulated in HCC tissues compared to normal liver tissues. A four gene signature, including ALDH2, ALDH5A1, ALDH6A1, and ALDH8A1, robustly predicted prognosis and defined a high-risk subgroup exhibiting immunosuppressive features like regulatory T cell (Tregs) infiltration. Single-cell profiling revealed selective overexpression of tumor necrosis factor receptor superfamily member 18 (TNFRSF18) on Tregs, upregulated in high-risk HCC patients. We identified ALDH2 as a tumor suppressor in HCC, with three novel phosphorylation sites mediated by protein kinase C zeta that enhanced enzymatic activity. Mechanistically, ALDH2 suppressed Tregs differentiation by inhibiting ß-catenin/TGF-ß1 signaling in HCC. Collectively, our integrated multi-omics analysis defines an ALDH-Tregs-TNFRSF18 axis that contributes to HCC pathogenesis and represents potential therapeutic targets for this aggressive malignancy.


Assuntos
Aldeído-Desidrogenase Mitocondrial , Carcinoma Hepatocelular , Neoplasias Hepáticas , Linfócitos T Reguladores , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/genética , Humanos , Aldeído-Desidrogenase Mitocondrial/metabolismo , Aldeído-Desidrogenase Mitocondrial/genética , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/imunologia , Microambiente Tumoral , Aldeído Desidrogenase/metabolismo , Aldeído Desidrogenase/genética , Animais , Linhagem Celular Tumoral , Masculino , Camundongos , Multiômica
6.
BMC Genomics ; 25(1): 513, 2024 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-38789947

RESUMO

BACKGROUND: Aldehyde dehydrogenases (ALDHs) are a family of enzymes that catalyze the oxidation of aldehyde molecules into the corresponding carboxylic acid, regulate the balance of aldehydes and protect plants from the poisoning caused by excessive accumulation of aldehydes; however, this gene family has rarely been studied in cotton. RESULTS: In the present study, genome-wide identification was performed, and a total of 114 ALDH family members were found in three cotton species, Gossypium hirsutum, Gossypium arboreum and Gossypium raimondii. The ALDH genes were divided into six subgroups by evolutionary analysis. ALDH genes in the same subgroup showed similar gene structures and conserved motifs, but some genes showed significant differences, which may result in functional differences. Chromosomal location analysis and selective pressure analysis revealed that the ALDH gene family had experienced many fragment duplication events. Cis-acting element analysis revealed that this gene family may be involved in the response to various biotic and abiotic stresses. The RT‒qPCR results showed that the expression levels of some members of this gene family were significantly increased under salt stress conditions. Gohir.A11G040800 and Gohir.D06G046200 were subjected to virus-induced gene silencing (VIGS) experiments, and the sensitivity of the silenced plants to salt stress was significantly greater than that of the negative control plants, suggesting that Gohir.A11G040800 and Gohir.D06G046200 may be involved in the response of cotton to salt stress. CONCLUSIONS: In total, 114 ALDH genes were identified in three Gossypium species by a series of bioinformatics analysis. Gene silencing of the ALDH genes of G. hirsutum revealed that ALDH plays an important role in the response of cotton to salt stress.


Assuntos
Aldeído Desidrogenase , Genoma de Planta , Gossypium , Família Multigênica , Filogenia , Gossypium/genética , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Regulação da Expressão Gênica de Plantas , Estresse Fisiológico/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Evolução Molecular , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Inativação Gênica
7.
Biochem Pharmacol ; 224: 116252, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38701866

RESUMO

The mitogen-activated protein kinase (MAPK/ERK) pathway is pivotal in controlling the proliferation and survival of melanoma cells. Several mutations, including those in BRAF, exhibit an oncogenic effect leading to increased cellular proliferation. As a result, the combination therapy of a MEK inhibitor with a BRAF inhibitor demonstrated higher efficacy and lower toxicity than BRAF inhibitor alone. This combination has become the preferred standard of care for tumors driven by BRAF mutations. Aldehyde dehydrogenase 1A1 (ALDH1A1) is a known marker of stemness involved in drug resistance in several type of tumors, including melanoma. This study demonstrates that melanoma cells overexpressing ALDH1A1 displayed resistance to vemurafenib and trametinib through the activation of PI3K/AKT signaling instead of MAPK axis. Inhibition of PI3K/AKT signaling partially rescued sensitivity to the drugs. Consistently, pharmacological inhibition of ALDH1A1 activity downregulated the activation of AKT and partially recovered responsiveness to vemurafenib and trametinib. We propose ALDH1A1 as a new potential target for treating melanoma resistant to MAPK/ERK inhibitors.


Assuntos
Família Aldeído Desidrogenase 1 , Resistencia a Medicamentos Antineoplásicos , Melanoma , Células-Tronco Neoplásicas , Inibidores de Proteínas Quinases , Proteínas Proto-Oncogênicas c-akt , Retinal Desidrogenase , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Melanoma/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/fisiologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Linhagem Celular Tumoral , Família Aldeído Desidrogenase 1/metabolismo , Família Aldeído Desidrogenase 1/genética , Retinal Desidrogenase/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Pirimidinonas/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Piridonas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Vemurafenib/farmacologia , Aldeído Desidrogenase/metabolismo , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/genética , Antineoplásicos/farmacologia , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fenótipo
8.
Cancer Epidemiol Biomarkers Prev ; 33(7): 933-943, 2024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38652503

RESUMO

BACKGROUND: According to the stem cell hypothesis, breast carcinogenesis may be related to the breast stem cell pool size. However, little is known about associations of breast cancer risk factors, such as anthropometric measures, with the expression of stem cell markers in noncancerous breast tissue. METHODS: The analysis included 414 women with biopsy-confirmed benign breast disease in the Nurses' Health Study and Nurses' Health Study II. Birthweight, weight at age 18, current weight, and current height were reported via self-administered questionnaires. IHC staining of stem cell markers (CD44, CD24, and aldehyde dehydrogenase family 1 member A1) in histopathologically normal epithelial and stromal breast tissue was quantified using an automated computational image analysis system. Linear regression was used to examine the associations of early-life and adult anthropometric measures with log-transformed stem cell marker expression, adjusting for potential confounders. RESULTS: Birthweight [≥10.0 vs. <5.5 lbs: ß (95% confidence interval) = 4.29 (1.02, 7.56); P trend = 0.001 in the stroma] and adult height [≥67.0 vs. <63.0 inch: 0.86 (0.14, 1.58); P trend = 0.02 in the epithelium and stroma combined] were positively associated with CD44 expression. Childhood body fatness was inversely associated (P trend = 0.03) whereas adult height was positively associated with CD24 expression in combined stroma and epithelium (P trend = 0.03). CONCLUSIONS: Our data suggest that anthropometric measures, such as birthweight, adult height, and childhood body fatness, may be associated with the stem cell expression among women with benign breast disease. IMPACT: Anthropometric measures, such as birthweight, height, and childhood body fatness, may have long-term impacts on stem cell population in the breast.


Assuntos
Família Aldeído Desidrogenase 1 , Antígeno CD24 , Receptores de Hialuronatos , Retinal Desidrogenase , Humanos , Feminino , Adulto , Antígeno CD24/metabolismo , Família Aldeído Desidrogenase 1/metabolismo , Receptores de Hialuronatos/metabolismo , Retinal Desidrogenase/metabolismo , Pessoa de Meia-Idade , Biópsia , Neoplasias da Mama/patologia , Neoplasias da Mama/metabolismo , Mama/patologia , Antropometria/métodos , Células-Tronco/metabolismo , Células-Tronco/patologia , Aldeído Desidrogenase/metabolismo
9.
Biochemistry ; 63(9): 1075-1088, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38602394

RESUMO

Aldehyde dehydrogenase enzymes (ALDHs) are widely studied for their roles in disease propagation and cell metabolism. Their use in biocatalysis applications, for the conversion of aldehydes to carboxylic acids, has also been recognized. Understanding the structural features and functions of both prokaryotic and eukaryotic ALDHs is key to uncovering novel applications of the enzyme and probing its role in disease propagation. The thermostable enzyme ALDHTt originating fromThermus thermophilus, strain HB27, possesses a unique extension of its C-terminus, which has been evolutionarily excluded from mesophilic counterparts and other thermophilic enzymes in the same genus. In this work, the thermophilic adaptation is studied by the expression and optimized purification of mutant ALDHTt-508, with a 22-amino acid truncation of the C-terminus. The mutant shows increased activity throughout production compared to native ALDHTt, indicating an opening of the active site upon C-terminus truncation and giving rationale into the evolutionary exclusion of the C-terminal extension from similar thermophilic and mesophilic ALDH proteins. Additionally, the C-terminus is shown to play a role in controlling substrate specificity of native ALDH, particularly in excluding catalysis of certain large and certain aromatic ortho-substituted aldehydes, as well as modulating the protein's pH tolerance by increasing surface charge. Dynamic light scattering and size-exclusion HPLC methods are used to show the role of the C-terminus in ALDHTt oligomeric stability at the cost of catalytic efficiency. Studying the aggregation rate of ALDHTt with and without a C-terminal extension leads to the conclusion that ALDHTt follows a monomolecular reaction aggregation mechanism.


Assuntos
Aldeído Desidrogenase , Estabilidade Enzimática , Thermus thermophilus , Thermus thermophilus/enzimologia , Aldeído Desidrogenase/química , Aldeído Desidrogenase/metabolismo , Aldeído Desidrogenase/genética , Especificidade por Substrato , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Multimerização Proteica , Cinética , Domínio Catalítico , Sequência de Aminoácidos
10.
Food Chem ; 450: 139323, 2024 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-38636386

RESUMO

Radix puerariae thomsonii (RPT) contains many phenolics and exhibits various health benefits. Although the free phenolics in RPT have been identified, the composition and content of bound phenolics, which account for approximately 20% of the total phenolic content, remain unknown. In this study, 12 compounds were isolated and identified from RPT-bound phenolic extracts, of which 2 were novel and 6 were reported first in RPT. ORAC and PSC antioxidant activities of 12 compounds, as well as their effects on alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), α-glucosidase, and α-amylase were evaluated. Genistein exhibited the highest ORAC activity, while daidzin demonstrated superior PSC activity. Five compounds, including two new compounds, exhibited the ability to activate both ADH and ALDH. All the compounds except 4-hydroxyphenylacetic acid methyl ester and 2,4,4'-trihydroxydeoxybenzoin demonstrated inhibitory effects on α-glucosidase and α-amylase. Alkaline hydrolysis and stepwise enzymatic hydrolysis revealed that bound phenolics in RPT mainly exist within starch.


Assuntos
Fenóis , Extratos Vegetais , Pueraria , alfa-Amilases , alfa-Glucosidases , Pueraria/química , Fenóis/química , Fenóis/farmacologia , alfa-Amilases/química , alfa-Amilases/metabolismo , alfa-Amilases/antagonistas & inibidores , alfa-Glucosidases/química , alfa-Glucosidases/metabolismo , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Sítios de Ligação , Álcool Desidrogenase/química , Álcool Desidrogenase/metabolismo , Aldeído Desidrogenase/química , Aldeído Desidrogenase/metabolismo , Antioxidantes/química , Antioxidantes/farmacologia , Raízes de Plantas/química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Estrutura Molecular , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia
11.
Chem Biol Interact ; 394: 110993, 2024 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-38604394

RESUMO

Aldehyde dehydrogenase 7A1 (ALDH7A1) catalyzes a step of lysine catabolism. Certain missense mutations in the ALDH7A1 gene cause pyridoxine dependent epilepsy (PDE), a rare autosomal neurometabolic disorder with recessive inheritance that affects almost 1:65,000 live births and is classically characterized by recurrent seizures from the neonatal period. We report a biochemical, structural, and computational study of two novel ALDH7A1 missense mutations that were identified in a child with rare recurrent seizures from the third month of life. The mutations affect two residues in the oligomer interfaces of ALDH7A1, Arg134 and Arg441 (Arg162 and Arg469 in the HGVS nomenclature). The corresponding enzyme variants R134S and R441C (p.Arg162Ser and p.Arg469Cys in the HGVS nomenclature) were expressed in Escherichia coli and purified. R134S and R441C have 10,000- and 50-fold lower catalytic efficiency than wild-type ALDH7A1, respectively. Sedimentation velocity analytical ultracentrifugation shows that R134S is defective in tetramerization, remaining locked in a dimeric state even in the presence of the tetramer-inducing coenzyme NAD+. Because the tetramer is the active form of ALDH7A1, the defect in oligomerization explains the very low catalytic activity of R134S. In contrast, R441C exhibits wild-type oligomerization behavior, and the 2.0 Å resolution crystal structure of R441C complexed with NAD+ revealed no obvious structural perturbations when compared to the wild-type enzyme structure. Molecular dynamics simulations suggest that the mutation of Arg441 to Cys may increase intersubunit ion pairs and alter the dynamics of the active site gate. Our biochemical, structural, and computational data on two novel clinical variants of ALDH7A1 add to the complexity of the molecular determinants underlying pyridoxine dependent epilepsy.


Assuntos
Aldeído Desidrogenase , Mutação de Sentido Incorreto , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/química , Aldeído Desidrogenase/metabolismo , Humanos , Simulação de Dinâmica Molecular , Cristalografia por Raios X , Modelos Moleculares , Epilepsia/genética , Lactente , Masculino
12.
Int J Biol Macromol ; 265(Pt 2): 131091, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38521319

RESUMO

Acetaldehyde dehydrogenase 2 (ALDH2) is a crucial enzyme in alcohol metabolism, and oral administration of ALDH2 is a promising method for alcohol detoxification. However, recombinant ALDH2 is susceptible to hydrolysis by digestive enzymes in the gastrointestinal tract and is expressed as inactive inclusion bodies in E. coli. In this study, we performed three rounds of rational design to address these issues. Specifically, the surface digestive sites of pepsin and trypsin were replaced with other polar amino acids, while hydrophobic amino acids were incorporated to reshape the catalytic cavity of ALDH2. The resulting mutant DE2-852 exhibited a 45-fold increase in soluble expression levels, while its stability against trypsin and pepsin increased by eightfold and twofold, respectively. Its catalytic efficiency (kcat/Km) at pH 7.2 and 3.2 improved by more than four and five times, respectively, with increased Vmax and decreased Km values. The enhanced properties of DE2-852 were attributed to the D457Y mutation, which created a more compact protein structure and facilitated a faster collision between the substrate and catalytic residues. These results laid the foundation for the oral administration and mass preparation of highly active ALDH2 and offered insights into the oral application of other proteins.


Assuntos
Aldeído Desidrogenase , Pepsina A , Humanos , Aldeído-Desidrogenase Mitocondrial/genética , Aldeído-Desidrogenase Mitocondrial/química , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Tripsina , Escherichia coli/genética , Escherichia coli/metabolismo , Aminoácidos
13.
Methods Mol Biol ; 2777: 83-89, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38478337

RESUMO

Cancer stem cells (CSCs) are a small tumor cell subpopulation, driving cancer initiation, progression, multidrug resistance, and metastasis. Several methods are used to detect and isolate CSCs by flow cytometry. Among these, measurement of aldehyde dehydrogenase (ALDH) activity within the cell is an assay widely used to identify and isolate CSCs from different types of solid tumors. The aldehyde dehydrogenase (ALDH) is a polymorphic enzyme responsible for the oxidation of aldehydes to carboxylic acids, overexpressed both in normal and cancer stem cells. In this chapter, it is described how CSCs are detected and isolated by using ALDH activity assay.


Assuntos
Neoplasias , Células-Tronco Neoplásicas , Linhagem Celular Tumoral , Células-Tronco Neoplásicas/metabolismo , Aldeído Desidrogenase/metabolismo , Citometria de Fluxo , Neoplasias/patologia
14.
Int J Mol Sci ; 25(6)2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38542325

RESUMO

The cancer stem cell (CSC) hypothesis postulates that heterogeneous human cancers harbor a population of stem-like cells which are resistant to cytotoxic therapies, thus providing a reservoir of relapse following conventional therapies like chemotherapy and radiation (RT). CSCs have been observed in multiple human cancers, and their presence has been correlated with worse clinical outcomes. Here, we sought to evaluate the impact of drug dosing of the multi-tyrosine kinase inhibitor, sorafenib, on CSC and non-CSCs in soft tissue sarcoma (STS) models, hypothesizing differential effects of sorafenib based on dose and target cell population. In vitro, human cancer cell lines and primary STS from surgical specimens were exposed to escalating doses of sorafenib to determine cell viability and expression of CSC marker aldehyde dehydrogenase (ALDH). In vivo, ALDHbright CSCs were isolated, exposed to sorafenib, and xenograft growth and survival analyses were performed. We observed that sarcoma CSCs appear to paradoxically respond to the tyrosine kinase inhibitor sorafenib at low doses with increased proliferation and stem-like function of CSCs, whereas anti-viability effects dominated at higher doses. Importantly, STS patients receiving neoadjuvant sorafenib and RT on a clinical trial (NCT00864032) showed increased CSCs post therapy, and higher ALDH scores post therapy were associated with worse metastasis-free survival. These data suggest that low-dose sorafenib may promote the CSC phenotype in STS with clinically significant effects, including increased tumor growth and higher rates of metastasis formation in sarcoma patients.


Assuntos
Sarcoma , Humanos , Sorafenibe/farmacologia , Sorafenibe/uso terapêutico , Sorafenibe/metabolismo , Aldeído Desidrogenase/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Inibidores de Proteínas Quinases/metabolismo , Sarcoma/patologia , Células-Tronco Neoplásicas/metabolismo , Linhagem Celular Tumoral
15.
Oncogene ; 43(14): 1007-1018, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38361046

RESUMO

One-third of pediatric patients with osteosarcoma (OS) develop lung metastases (LM), which is the primary predictor of mortality. While current treatments of patients with localized bone disease have been successful in producing 5-year survival rates of 65-70%, patients with LM experience poor survival rates of only 19-30%. Unacceptably, this situation that has remained unchanged for 30 years. Thus, there is an urgent need to elucidate the mechanisms of metastatic spread in OS and to identify targetable molecular pathways that enable more effective treatments for patients with LM. We aimed to identify OS-specific gene alterations using RNA-sequencing of extremity and LM human tissues. Samples of extremity and LM tumors, including 4 matched sets, were obtained from patients with OS. Our data demonstrate aberrant regulation of the androgen receptor (AR) pathway in LM and predicts aldehyde dehydrogenase 1A1 (ALDH1A1) as a downstream target. Identification of AR pathway upregulation in human LM tissue samples may provide a target for novel therapeutics for patients with LM resistant to conventional chemotherapy.


Assuntos
Neoplasias Ósseas , Neoplasias Pulmonares , Osteossarcoma , Humanos , Criança , Aldeído Desidrogenase/metabolismo , Receptores Androgênicos/genética , Neoplasias Pulmonares/patologia , Osteossarcoma/patologia , Neoplasias Ósseas/patologia , RNA
16.
J Membr Biol ; 257(1-2): 3-16, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38356054

RESUMO

Cancer is one of the main causes of death among humans, second only to cardiovascular diseases. In recent years, numerous studies have been conducted on the pathophysiology of cancer, and it has been established that this disease is developed by a group of stem cells known as cancer stem cells (CSCs). Thus, cancer is considered a stem cell disease; however, there is no comprehensive consensus about the characteristics of these cells. Several different signaling pathways including Notch, Hedgehog, transforming growth factor-ß (TGF-ß), and WNT/ß-catenin pathways cause the self-renewal of CSCs. CSCs change their metabolic pathways in order to access easy energy. Therefore, one of the key objectives of researchers in cancer treatment is to destroy CSCs. Nuclear factor erythroid 2-related factor 2 (Nrf2) plays an essential role in the protection of CSCs from reactive oxygen species (ROS) and chemotherapeutic agents by regulating antioxidants and detoxification enzymes. Human epidermal growth factor receptor 2 (HER2) is a member of the tyrosine kinase receptor family, which contributes to the protection of cancer cells against treatment and implicated in the invasion, epithelial-mesenchymal transition (EMT), and tumorigenesis. Aldehyde dehydrogenases (ALDHs) are highly active in CSCs and protect the cells against damage caused by active aldehydes through the regulation of aldehyde metabolism. On the other hand, ALDHs promote the formation and maintenance of tumor cells and lead to drug resistance in tumors through the activation of various signaling pathways, such as the ALDH1A1/HIF-1α/VEGF axis and Wnt/ß-catenin, as well as changing the intracellular pH value. Given the growing body of information in this field, in the present narrative review, we attempted to shed light on the function of Nrf2, HER2, and ALDH in CSCs.


Assuntos
Aldeído Desidrogenase , Fator 2 Relacionado a NF-E2 , Células-Tronco Neoplásicas , Receptor ErbB-2 , beta Catenina , Humanos , Aldeído Desidrogenase/metabolismo , Aldeídos/metabolismo , beta Catenina/metabolismo , Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Receptor ErbB-2/metabolismo
17.
Chem Biol Interact ; 391: 110910, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38364885

RESUMO

Aldehyde dehydrogenase 1A (ALDH1A) isoforms may be a useful target for overcoming chemotherapy resistance in high-grade serous ovarian cancer (HGSOC) and other solid tumor cancers. However, as different cancers express different ALDH1A isoforms, isoform selective inhibitors may have a limited therapeutic scope. Furthermore, resistance to an ALDH1A isoform selective inhibitor could arise via induction of expression of other ALDH1A isoforms. As such, we have focused on the development of pan-ALDH1A inhibitors, rather than on ALDH1A isoform selective compounds. Herein, we report the development of a new group of pan-ALDH1A inhibitors to assess whether broad spectrum ALDH1A inhibition is an effective adjunct to chemotherapy in HGSOC. Optimization of the CM10 scaffold, aided by ALDH1A1 crystal structures, led to improved biochemical potencies, improved cellular efficacy as demonstrated by reduction in ALDEFLUOR signal in HGSOC cells, and substantial improvements in liver microsomal stability. Based on this work we identified two compounds 17 and 25 suitable for future in vivo proof of concept experiments.


Assuntos
Isoenzimas , Neoplasias , Humanos , Aldeído Desidrogenase/metabolismo , Retinal Desidrogenase/metabolismo , Aldeído Oxirredutases/metabolismo
18.
Nat Commun ; 15(1): 1032, 2024 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-38310110

RESUMO

Glutarate is a key monomer in polyester and polyamide production. The low efficiency of the current biosynthetic pathways hampers its production by microbial cell factories. Herein, through metabolic simulation, a lysine-overproducing E. coli strain Lys5 is engineered, achieving titer, yield, and productivity of 195.9 g/L, 0.67 g/g glucose, and 5.4 g/L·h, respectively. Subsequently, the pathway involving aromatic aldehyde synthase, monoamine oxidase, and aldehyde dehydrogenase (AMA pathway) is introduced into E. coli Lys5 to produce glutarate from glucose. To enhance the pathway's efficiency, rational mutagenesis on the aldehyde dehydrogenase is performed, resulting in the development of variant Mu5 with a 50-fold increase in catalytic efficiency. Finally, a glutarate tolerance gene cbpA is identified and genomically overexpressed to enhance glutarate productivity. With enzyme expression optimization, the glutarate titer, yield, and productivity of E. coli AMA06 reach 88.4 g/L, 0.42 g/g glucose, and 1.8 g/L·h, respectively. These findings hold implications for improving glutarate biosynthesis efficiency in microbial cell factories.


Assuntos
Escherichia coli , Glutaratos , Escherichia coli/genética , Escherichia coli/metabolismo , Glutaratos/metabolismo , Glucose/metabolismo , Engenharia Metabólica/métodos , Aldeído Desidrogenase/metabolismo
19.
J Microbiol Biotechnol ; 34(4): 838-845, 2024 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-38247212

RESUMO

Excessive alcohol consumption can have serious negative consequences on health, including addiction, liver damage, and other long-term effects. The causes of hangovers include dehydration, alcohol and alcohol metabolite toxicity, and nutrient deficiency due to absorption disorders. Additionally, alcohol consumption can slow reaction times, making it more difficult to rapidly respond to situations that require quick thinking. Exposure to a large amount of ethanol can also negatively affect a person's righting reflex and balance. In this study, we evaluated the potential of lactic acid bacteria (LAB) to alleviate alcohol-induced effects and behavioral responses. Two LAB strains isolated from kimchi, Levilactobacillus brevis WiKim0168 and Leuconostoc mesenteroides WiKim0172, were selected for their ethanol tolerance and potential to alleviate hangover symptoms. Enzyme activity assays for alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) were then conducted to evaluate the role of these bacteria in alcohol metabolism. Through in vitro and in vivo studies, these strains were assessed for their ability to reduce blood alcohol concentrations and protect against alcohol-induced liver damage. The results indicated that these LAB strains possess significant ethanol tolerance and elevate ADH and ALDH activities. LAB administration remarkably reduced blood alcohol levels in rats after excessive alcohol consumption. Moreover, the LAB strains showed hepatoprotective effects and enhanced behavioral outcomes, highlighting their potential as probiotics for counteracting the adverse effects of alcohol consumption. These findings support the development of functional foods incorporating LAB strains that can mediate behavioral improvements following alcohol intake.


Assuntos
Álcool Desidrogenase , Aldeído Oxirredutases , Etanol , Lactobacillales , Probióticos , Animais , Etanol/metabolismo , Álcool Desidrogenase/metabolismo , Ratos , Masculino , Probióticos/administração & dosagem , Lactobacillales/metabolismo , Concentração Alcoólica no Sangue , Fígado/metabolismo , Fígado/efeitos dos fármacos , Administração Oral , Leuconostoc mesenteroides , Aldeído Desidrogenase/metabolismo , Levilactobacillus brevis/metabolismo , Ratos Sprague-Dawley , Alimentos Fermentados/microbiologia
20.
Theranostics ; 14(2): 714-737, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38169509

RESUMO

Rationale: Current therapies for metastatic osseous disease frequently fail to provide a durable treatment response. To date, there are only limited therapeutic options for metastatic prostate cancer, the mechanisms that drive the survival of metastasis-initiating cells are poorly characterized, and reliable prognostic markers are missing. A high aldehyde dehydrogenase (ALDH) activity has been long considered a marker of cancer stem cells (CSC). Our study characterized a differential role of ALDH1A1 and ALDH1A3 genes as regulators of prostate cancer progression and metastatic growth. Methods: By genetic silencing of ALDH1A1 and ALDH1A3 in vitro, in xenografted zebrafish and murine models, and by comparative immunohistochemical analyses of benign, primary tumor, and metastatic specimens from patients with prostate cancer, we demonstrated that ALDH1A1 and ALDH1A3 maintain the CSC phenotype and radioresistance and regulate bone metastasis-initiating cells. We have validated ALDH1A1 and ALDH1A3 as potential biomarkers of clinical outcomes in the independent cohorts of patients with PCa. Furthermore, by RNAseq, chromatin immunoprecipitation (ChIP), and biostatistics analyses, we suggested the molecular mechanisms explaining the role of ALDH1A1 in PCa progression. Results: We found that aldehyde dehydrogenase protein ALDH1A1 positively regulates tumor cell survival in circulation, extravasation, and metastatic dissemination, whereas ALDH1A3 plays the opposite role. ALDH1A1 and ALDH1A3 are differentially expressed in metastatic tumors of patients with prostate cancer, and their expression levels oppositely correlate with clinical outcomes. Prostate cancer progression is associated with the increasing interplay of ALDH1A1 with androgen receptor (AR) and retinoid receptor (RAR) transcriptional programs. Polo-like kinase 3 (PLK3) was identified as a transcriptional target oppositely regulated by ALDH1A1 and ALDH1A3 genes in RAR and AR-dependent manner. PLK3 contributes to the control of prostate cancer cell proliferation, migration, DNA repair, and radioresistance. ALDH1A1 gain in prostate cancer bone metastases is associated with high PLK3 expression. Conclusion: This report provides the first evidence that ALDH1A1 and PLK3 could serve as biomarkers to predict metastatic dissemination and radiotherapy resistance in patients with prostate cancer and could be potential therapeutic targets to eliminate metastasis-initiating and radioresistant tumor cell populations.


Assuntos
Neoplasias da Próstata , Receptores Androgênicos , Masculino , Humanos , Animais , Camundongos , Peixe-Zebra/metabolismo , Linhagem Celular Tumoral , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Neoplasias da Próstata/genética , Biomarcadores , Família Aldeído Desidrogenase 1 , Retinal Desidrogenase
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