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EBioMedicine Dec 2016Artemisinin (ARS) and its derivatives, which are clinically used antimalarial agents, have shown antitumor activities. Their therapeutic potencies, however, are limited...
UNLABELLED
Artemisinin (ARS) and its derivatives, which are clinically used antimalarial agents, have shown antitumor activities. Their therapeutic potencies, however, are limited by their low solubility and poor bioavailability. Here, through a pharmacophore hybridization strategy, we synthesized ARS-drug conjugates, in which the marketed chemotherapeutic agents chlorambucil, melphalan, flutamide, aminoglutethimide, and doxifluridine, were separately bonded to Dihydroartemisinin (DHA) through various linkages. Of these, the artemisinin-melphalan conjugate, ARS4, exhibited most toxicity to human ovarian cancer cells but had low cytotoxicity to normal cells. ARS4 inhibited the growth and proliferation of ovarian cancer cells and resulted in S-phase arrest, apoptosis, and inhibition of migration; these effects were stronger than those of its parent drugs, DHA and melphalan. Furthermore, ARS4 modulated the expression of proteins involved in cell cycle progression, apoptosis, and the epithelial-mesenchymal transition (EMT). Moreover, in mice, ARS4 inhibited growth and intraperitoneal dissemination and metastasis of ovarian cancer cells without observable toxic effects. Our results provide a basis for development of the compound as a chemotherapeutic agent.
RESEARCH IN CONTEXT
Artemisinin compounds have recently received attention as anticancer agents because of their clinical safety profiles and broad efficacy. However, their therapeutic potencies are limited by low solubility and poor bioavailability. Here, we report that ARS4, an artemisinin-melphalan conjugate, possesses marked in-vitro and in-vivo antitumor activity against ovarian cancer, the effects of which are stronger than those for its parent drugs, Dihydroartemisinin and melphalan. In mice, ARS4 inhibits localized growth of ovarian cancer cells and intraperitoneal dissemination and metastasis without appreciable host toxicity. Thus, for patients with ovarian cancer, ARS4 is a promising chemotherapeutic agent.
Topics: Animals; Antineoplastic Agents; Apoptosis; Artemisinins; Cell Cycle; Cell Line, Tumor; Cell Movement; Cell Proliferation; Disease Models, Animal; Drug Combinations; Drug Evaluation, Preclinical; Epithelial-Mesenchymal Transition; Female; Humans; Mice; Neoplasm Metastasis; Neoplasm Staging; Ovarian Neoplasms; Structure-Activity Relationship; Xenograft Model Antitumor Assays
PubMed: 27939426
DOI: 10.1016/j.ebiom.2016.11.026 -
Drug Testing and Analysis Feb 2019Anectodical information suggests that flavonoids may be widely used among athletes for their multiple biochemical and pharmacological effects. We have evaluated in vitro...
Anectodical information suggests that flavonoids may be widely used among athletes for their multiple biochemical and pharmacological effects. We have evaluated in vitro the effects of two synthetic isoflavones, methoxyisoflavone and ipriflavone, on the catalytic activity of human aromatase (CYP19), the enzyme catalyzing the conversion of androgens (ie, testosterone or androstenedione) to estrogens (ie, estradiol and estrone). The potential inhibitory effect was evaluated by measuring the rate of aromatization of testosterone, monitored by gas chromatography-mass spectrometry (GC-MS), both in the presence and in the absence of methoxyisoflavone or ipriflavone, comparing their effects with those of synthetic aromatase inhibitors (formestane, anastrozole, and aminoglutethimide) presently included in the list of prohibited substances and methods, and of natural flavonoids (chrysin, quercetin, and daidzein), that are known inhibitors of CYP19. The preliminary results of our in vitro study show that methoxyisoflavone and ipriflavone act as competitive inhibitors of aromatase, the degree of inhibition measured in vitro being of the same order of magnitude of that of the aromatase inhibitors commonly used in anti-estrogenic therapies. Our preliminary in vitro results indicate that, in principle, a sufficiently large intake of isoflavones could alter the kinetics of the dynamic equilibria between androgens and estrogens, suggesting their monitoring in doping control routine analysis.
Topics: Aromatase Inhibitors; Doping in Sports; Flavonoids; Humans; In Vitro Techniques; Isoflavones; Testosterone
PubMed: 30118172
DOI: 10.1002/dta.2482 -
Biomaterials Dec 2018The intrinsic characteristics of the tumor microenvironment (TME), including acidic pH and overexpression of hydrolytic enzymes, offer an exciting opportunity for the...
The intrinsic characteristics of the tumor microenvironment (TME), including acidic pH and overexpression of hydrolytic enzymes, offer an exciting opportunity for the rational design of TME-drug delivery systems (DDS). We developed and characterized a pH-responsive biodegradable poly-L-glutamic acid (PGA)-based combination conjugate family with the aim of optimizing anticancer effects. We obtained combination conjugates bearing Doxorubicin (Dox) and aminoglutethimide (AGM) with two Dox loadings and two different hydrazone pH-sensitive linkers that promote the specific release of Dox from the polymeric backbone within the TME. Low Dox loading coupled with a short hydrazone linker yielded optimal effects on primary tumor growth, lung metastasis (∼90% reduction), and toxicological profile in a preclinical metastatic triple-negative breast cancer (TNBC) murine model. The use of transcriptomic analysis helped us to identify the molecular mechanisms responsible for such results including a differential immunomodulation and cell death pathways among the conjugates. This data highlights the advantages of targeting the TME, the therapeutic value of polymer-based combination approaches, and the utility of -omics-based analysis to accelerate anticancer DDS.
Topics: Aminoglutethimide; Animals; Antineoplastic Agents; Cell Line, Tumor; Cell Survival; Doxorubicin; Drug Carriers; Drug Liberation; Female; Heterografts; Humans; Hydrogen-Ion Concentration; Mice, Inbred BALB C; Polyglutamic Acid; Triple Negative Breast Neoplasms; Tumor Microenvironment
PubMed: 30278346
DOI: 10.1016/j.biomaterials.2018.09.023 -
ACS Omega Apr 2022Our present study intended to investigate the encapsulation of DL-AGT within the lipophilic cavity of a β-CD molecule. The consequential inclusion system was...
Synthesis and Characterization of an Inclusion Complex of dl-Aminoglutethimide with β-Cyclodextrin and Its Innovative Application in a Biological System: Computational and Experimental Investigations.
Our present study intended to investigate the encapsulation of DL-AGT within the lipophilic cavity of a β-CD molecule. The consequential inclusion system was characterized by UV-visible spectroscopy and H NMR, PXRD, SEM, and FT-IR studies. Molecular docking was performed for the inclusion complex to discover the most proper orientation, and it was seen that the drug DL-AGT fits into the cavity of β-CD in a 1:1 ratio, which was also confirmed from the Job plot. Furthermore, a comparison was done on the basis of cell viability between the drug and its inclusion complex.
PubMed: 35415366
DOI: 10.1021/acsomega.2c00011 -
Cancer Medicine May 2020Endometrial cancer (EC) is a fatal female reproductive tumor. Bioinformatic tools are increasingly developed to screen out molecular targets related to EC. In this...
Endometrial cancer (EC) is a fatal female reproductive tumor. Bioinformatic tools are increasingly developed to screen out molecular targets related to EC. In this study, GSE17025 and GSE40032 were obtained from Gene Expression Omnibus (GEO). "limma" package and Venn diagram tool were used to identify hub genes. FunRich was used for functional analysis. Retrieval of Interacting Genes Database (STRING) was used to analyze protein-protein interaction (PPI) complex. Cancer Genome Atlas (TCGA), GEPIA, immunohistochemistry staining, and ROC curve analysis were carried out for validation. Univariate and multivariate regression analyses were performed to predict the risk score. Compound muscle action potential (CMap) was used to find potential drugs. GSEA was also done. We retrieved seven oncogenes which were upregulated and hypomethylated and 12 tumor suppressor genes (TSGs) which were downregulated and hypermethylated. The upregulated and hypomethylated genes were strikingly enriched in term "immune response" while the downregulated and hypermethylated genes were mainly focused on term "aromatic compound catabolic process." TCGA and GEPIA were used to screen out EDNRB, CDO1, NDN, PLCD1, ROR2, ESPL1, PRAME, and PTTG1. Among them, ESPL1 and ROR2 were identified by Cox regression analysis and were used to construct prognostic risk model. The result showed that ESPL1 was a negative independent prognostic factor. Cmap identified aminoglutethimide, luteolin, sulfadimethoxine, and maprotiline had correlation with EC. GSEA results showed that "hedgehog signaling pathway" was enriched. This research inferred potential aberrantly methylated DEGs and dysregulated pathways may participate in EC development and firstly reported eight hub genes, including EDNRB, CDO1, NDN, PLCD1, ROR2, ESPL1, PRAME, and PTTG1 that could be used to predict EC prognosis. Aminoglutethimide and luteolin may be used to fight against EC.
Topics: Computational Biology; DNA Methylation; Databases, Genetic; Endometrial Neoplasms; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Hedgehog Proteins; Humans; Prognosis; Proportional Hazards Models; Protein Interaction Maps; Receptor Tyrosine Kinase-like Orphan Receptors; Reproducibility of Results; Separase; Small Molecule Libraries
PubMed: 32170852
DOI: 10.1002/cam4.2956 -
The Journal of Endocrinology Jun 2024Cells actively engaged in de novo steroidogenesis rely on an expansive intracellular network to efficiently transport cholesterol. The final link in the transport chain...
Cells actively engaged in de novo steroidogenesis rely on an expansive intracellular network to efficiently transport cholesterol. The final link in the transport chain is STARD1, which transfers cholesterol to the enzyme complex that initiates steroidogenesis. However, the regulation of ovarian STARD1 is not fully characterized and even less is known for upstream cytosolic cholesterol transporters STARD4 and STARD6. Here, we identified both STARD4 and STARD6 mRNAs in the human ovary but only detected STARD4 protein since the primary STARD6 transcript turned out to be a splice variant. Corpora lutea contained the highest levels of STARD4 and STARD1 mRNA and STARD1 protein, while STARD4 protein was uniformly distributed across ovarian tissues. Cyclic AMP analog (8Br-cAMP) and phorbol ester (PMA) individually increased STARD1 and STARD4 mRNA along with STARD1 protein and its phosphoform in cultured primary human luteinized granulosa cells (hGC). STARD6 transcripts and STARD4 protein were unresponsive to these stimuli. Combining lower doses of PMA and 8Br-cAMP blunted the 8Br-cAMP stimulation of STARD1 protein. Increasing cholesterol levels by blocking its conversion to steroid with aminoglutethimide or by adding LDL reduced the STARD4 mRNA response to stimuli. Sterol depletion reduced the STARD1 mRNA and protein response to PMA. These data support a possible role for STARD4, but not STARD6, in supplying cholesterol for steroidogenesis in the ovary. We demonstrate for the first time how cAMP, PMA and sterol pathways separately and combined differentially regulate STARD4, STARD6 and STARD1 mRNA levels, and STARD1 and STARD4 protein in human primary ovarian cells.
PubMed: 38829257
DOI: 10.1530/JOE-23-0385 -
The Journal of Biological Chemistry Jul 2022Neurosteroids, modulators of neuronal and glial cell functions, are synthesized in the nervous system from cholesterol. In peripheral steroidogenic tissues, cholesterol...
Neurosteroids, modulators of neuronal and glial cell functions, are synthesized in the nervous system from cholesterol. In peripheral steroidogenic tissues, cholesterol is converted to the major steroid precursor pregnenolone by the CYP11A1 enzyme. Although pregnenolone is one of the most abundant neurosteroids in the brain, expression of CYP11A1 is difficult to detect. We found that human glial cells produced pregnenolone, detectable by mass spectrometry and ELISA, despite the absence of observable immunoreactive CYP11A1 protein. Unlike testicular and adrenal cortical cells, pregnenolone production in glial cells was not inhibited by CYP11A1 inhibitors DL-aminoglutethimide and ketoconazole. Furthermore, addition of hydroxycholesterols increased pregnenolone synthesis, suggesting desmolase activity that was not blocked by DL-aminoglutethimide or ketoconazole. We explored three different possibilities for an alternative pathway for glial cell pregnenolone synthesis: (1) regulation by reactive oxygen species, (2) metabolism via a different CYP11A1 isoform, and (3) metabolism via another CYP450 enzyme. First, we found oxidants and antioxidants had no significant effects on pregnenolone synthesis, suggesting it is not regulated by reactive oxygen species. Second, overexpression of CYP11A1 isoform b did not alter synthesis, indicating use of another CYP11A1 isoform is unlikely. Finally, we show nitric oxide and iron chelators deferoxamine and deferiprone significantly inhibited pregnenolone production, indicating involvement of another CYP450 enzyme. Ultimately, knockdown of endoplasmic reticulum cofactor NADPH-cytochrome P450 reductase had no effect, while knockdown of mitochondrial CYP450 cofactor ferredoxin reductase inhibited pregnenolone production. These data suggest that pregnenolone is synthesized by a mitochondrial cytochrome P450 enzyme other than CYP11A1 in human glial cells.
Topics: Aminoglutethimide; Cholesterol; Cholesterol Side-Chain Cleavage Enzyme; Humans; Ketoconazole; Neuroglia; Neurosteroids; Pregnenolone; Reactive Oxygen Species
PubMed: 35688208
DOI: 10.1016/j.jbc.2022.102110 -
Turkish Journal of Pharmaceutical... Dec 2022Aromatase is an enzyme that catalyzes the conversion of androgens to estrogens. While inhibition of aromatase is a useful approach for treating breast cancer, it may...
OBJECTIVES
Aromatase is an enzyme that catalyzes the conversion of androgens to estrogens. While inhibition of aromatase is a useful approach for treating breast cancer, it may also have toxicological consequences due to its endocrine disrupting/modulating effect. In this study, sensitivity and performance of two assays -a cell free and a cell-based- for evaluating aromatase activity were investigated by testing known aromatase inhibitors and partial validation of the methods was performed. Advantages and disadvantages of these methods are also discussed.
MATERIALS AND METHODS
Aromatase activity was evaluated two models; direct measurement with a cell-free assay using a fluorescent substrate and recombinant human enzyme and indirect evaluation with a cell-based assay where cell proliferation was determined in estrogen receptor positive human breast cancer cells (MCF-7 BUS) in the absence of estrogen and the presence of testosterone.
RESULTS
In the cell-free direct measurement assay, reference compounds ketoconazole and aminoglutethimide have been shown to inhibit the aromatase enzyme with half-maximal inhibitory concentration (IC) values concordant with literature. In cell-based indirect measurement assay, only ketoconazole dose-dependently inhibited cell proliferation with 3.47 x 10 M IC. Inter-assay and intra-assay reproducibility of both methods was found to be within acceptable deviation levels.
CONCLUSION
Both methods can be successfully applied. However, to evaluate the potential aromatase activity of the novel compounds , it seems better to perform both the cell-based and the cell-free assays that allows low-moderate biotransformation and eliminate cytotoxicity potential, respectively.
PubMed: 36544280
DOI: 10.4274/tjps.galenos.2021.85530 -
Journal of Chromatography. A Oct 2016The separation of aminoglutethimide enantiomers by the continuous multicolumn chromatographic processes were investigated experimentally and theoretically, where the...
The separation of aminoglutethimide enantiomers by the continuous multicolumn chromatographic processes were investigated experimentally and theoretically, where the columns were packed with cellulose tris 3,5-dimethylphenyl-carbamate stationary phase (brand name Chiralcel OD) and mobile phase was a mixture of n-hexane and ethanol with monoethanolamine additive. The continuous enantioseparation processes included a synchronous shifting process (SMB) and an asynchronous shifting process (VARICOL), which allowed reducing the column number (here from six-column SMB to five-column VARICOL process). Transport-dispersive model with the consideration of both intraparticle mass transfer resistance and axial dispersion was adopted to design and optimize the operation conditions for the separation of aminoglutethimide enantiomers by SMB process and VARICOL process. According to the optimized operation conditions, experiments were carried out on VARICOL-Micro unit using five-column VARICOL process with 1/1.5/1.5/1 configuration and six-column SMB process with 1/2/2/1 configuration. Products of R-aminoglutethimide (R-AG) enantiomer and S-aminoglutethimide (S-AG) enantiomer with more than 99.0% purity were obtained continuously from extract stream and raffinate stream, respectively. Furthermore, the experiemntal data obtained from five-column VARICOL process were compared with that from six-column SMB process, the feasibility and efficiency for the separation of guaifenesin enantiomers by VARICOL processes were evaluated.
Topics: Aminoglutethimide; Cellulose; Chromatography; Ethanol; Guaifenesin; Hexanes; Indicators and Reagents; Organophosphates; Phenylcarbamates; Stereoisomerism
PubMed: 27544751
DOI: 10.1016/j.chroma.2016.08.031 -
Medicine Jul 2017Steroid profiling was introduced to determine the endogenous steroid misuse in sports. Thus, screening for the exogenous use of these prohibited substances can be... (Randomized Controlled Trial)
Randomized Controlled Trial
Steroid profiling was introduced to determine the endogenous steroid misuse in sports. Thus, screening for the exogenous use of these prohibited substances can be established by monitoring a range of endogenous steroids, which constitute the steroid profile and evaluate their concentrations and ratios against reference values. The steroid profiling is currently based on population statistics. As large interindividual variations exist, athlete biological passport (ABP) analysis is ongoing. This study aimed to identify new biomarker(s) for aromatase inhibitor detection in sports using statistical analysis and adapt the model into ABP analysis.Forty-one Chinese nonathlete volunteers (21 males and 20 females) were administered 3 nonsteroidal aromatase inhibitors (aminoglutethimide, letrozole, and anastrozole) independently. Statistical analysis was performed on 16 steroid profile parameters.After administration, the concentrations of endogenous androgen biomarkers including testosterone (T), epitestosterone, androsterone (AN), etiocholanolone (ETIO), 5α-diol, 5β-diol, and dehydroepiandrosterone were increased, while the level of estrogen was decreased. These biomarkers returned to the baselines levels within 1 month. In females, the concentrations of endogenous biomarkers were affected by nonsteroidal aromatase inhibitors, without a common trend. Three new endogenous biomarkers (AN/estrone, ETIO/estrone, and T/estrone) elevated significantly after treatment. The 3 new models were more sensitive than the World Anti-Doping Agency ratio biomarkers. They were also effective in exponentially weighted moving average chart analysis.Verification experiment demonstrated that the biomarker T/estrone was valid in judging the steroidal aromatase inhibitor abuse. The screening of these new endogenous biomarkers can provide additional parameters to support ABP monitoring and specific information regarding the administered steroids.
Topics: Aminoglutethimide; Anastrozole; Aromatase Inhibitors; Biomarkers, Pharmacological; Doping in Sports; Female; Hormones; Humans; Letrozole; Male; Models, Biological; Nitriles; Single-Blind Method; Steroids; Substance-Related Disorders; Triazoles; Young Adult
PubMed: 28700478
DOI: 10.1097/MD.0000000000007411