17 research outputs found
A novel aspirin prodrug inhibits NFκB activity and breast cancer stem cell properties
INTRODUCTION: Activation of cyclooxygenase (COX)/prostaglandin and nuclear factor κB (NFκB) pathways can promote breast tumor initiation, growth, and progression to drug resistance and metastasis. Thus, anti-inflammatory drugs have been widely explored as chemopreventive and antineoplastic agents. Aspirin (ASA), in particular, is associated with reduced breast cancer incidence but gastrointestinal toxicity has limited its usefulness. To improve potency and minimize toxicity, ASA ester prodrugs have been developed, in which the carboxylic acid of ASA is masked and ancillary pharmacophores can be incorporated. To date, the effects of ASA and ASA prodrugs have been largely attributed to COX inhibition and reduced prostaglandin production. However, ASA has also been reported to inhibit the NFκB pathway at very high doses. Whether ASA prodrugs can inhibit NFκB signaling remains relatively unexplored. METHODS: A library of ASA prodrugs was synthesized and screened for inhibition of NFκB activity and cancer stem-like cell (CSC) properties, an important PGE2-and NFκB-dependent phenotype of aggressive breast cancers. Inhibition of NFκB activity was determined by dual luciferase assay, RT-QPCR, p65 DNA binding activity and Western blots. Inhibition of CSC properties was determined by mammosphere growth, CD44(+)CD24(−)immunophenotype and tumorigenicity at limiting dilution. RESULTS: While we identified multiple ASA prodrugs that are capable of inhibiting the NFκB pathway, several were associated with cytotoxicity. Of particular interest was GTCpFE, an ASA prodrug with fumarate as the ancillary pharmacophore. This prodrug potently inhibits NFκB activity without innate cytotoxicity. In addition, GTCpFE exhibited selective anti-CSC activity by reducing mammosphere growth and the CD44(+)CD24(−)immunophenotype. Moreover, GTCpFE pre-treated cells were less tumorigenic and, when tumors did form, latency was increased and growth rate was reduced. Structure-activity relationships for GTCpFE indicate that fumarate, within the context of an ASA prodrug, is essential for anti-NFκB activity, whereas both the ASA and fumarate moieties contributed to attenuated mammosphere growth. CONCLUSIONS: These results establish GTCpFE as a prototype for novel ASA-and fumarate-based anti-inflammatory drugs that: (i) are capable of targeting CSCs, and (ii) may be developed as chemopreventive or therapeutic agents in breast cancer. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12885-015-1868-7) contains supplementary material, which is available to authorized users
Mechanisms of estrogen carcinogenesis in human breast epithelial cells.
Mechanisms of estrogen carcinogenesis in human breast epithelial cells
Photochemical reactions of thiols with organic nitrates \u2014 Oxygen atom transfer via a thionitrate
Nitroglycerin is an organic nitrate that has been used in the clinical treatment of angina for 130 years, yet important details of its mechanism of action remain unanswered. The biological activity of nitrates suggests that they are bioactivated to NO via a three-electron reduction. The involvement of free or bound protein thiols in this reduction has often been proposed. To examine the involvement of thiyl radicals in such a process, the photochemical generation of benzenethiyl radical from thiol and disulfide precursors was studied in the presence of isopropyl nitrate. Analysis of reaction products and kinetics led to the conclusion that photolysis of the nitrate to NO2 dominated the observed photochemistry. Formation of sulfonothioate and NO as products, and trapping of NO2 by 4-chlorophenol, indicated a mechanism involving oxygen atom transfer from N to S via a thionitrate intermediate. The results of the study did not indicate a rapid reaction between thiyl radical and organic nitrate. Despite weak nitrate absorption of light >300 nm and a relatively high BDE for homolysis to give NO2, the photochemistry under thiyl-generating conditions was driven by nitrate photolysis to NO2. A novel nitrate, containing a phenyl disulfanyl group linked to nitrate groups, did not undergo photolysis to NO2 or generate sulfonothioate, but did yield NO. These observations suggest that reaction between thiyl radicals and nitrates leading to NO release is a viable pathway, but it is subservient to other competing reactions, such as photolysis, in the case of IPN, and reaction with thiolate, in the case of the novel nitrate.NRC publication: Ye
Estrogen-Induced Apoptosis of Breast Epithelial Cells Is Blocked by NO/cGMP and Mediated by Extranuclear Estrogen Receptors
Estrogen action, via both nuclear and extranuclear estrogen receptors (ERs), induces a variety of cellular signals that are prosurvival or proliferative, whereas nitric oxide (NO) can inhibit apoptosis
via caspase S-nitrosylation and via activation of soluble guanylyl cyclase to produce cGMP. The action of 17β-estradiol (E2) at ER is known to elicit NO signaling via activation of NO synthase (NOS) in many tissues. The MCF-10A nontumorigenic, mammary epithelial cell line is genetically stable
and insensitive to estrogenic proliferation. In this cell line, estrogens or NOS inhibitors alone had no significant effect, whereas in combination, apoptosis was induced rapidly in the absence of serum; the presence of inducible NOS was confirmed by proteomic analysis. The application of pharmacological agents determined that apoptosis was dependent upon NO/cGMP signaling via
cyclic GMP (cGMP)-dependent protein kinase and could be replicated by inhibition of the phosphatidylinositol 3 kinase/serine-threonine kinase pathway prior to addition of E2. Apoptosis was confirmed by nuclear staining and increased caspase-3 activity in E2 NOS inhibitor-treated cells.
Apoptosis was partially inhibited by a pure ER antagonist and replicated by agonists selective for extranuclear ER. Cells were rescued from E2-induced apoptosis after NOS blockade, by NO-donors and cGMP pathway agonists; preincubation with NO donors was required. The NOS and ER status of breast cancer tissues is significant in etiology, prognosis, and therapy. In this study, apoptosis of preneoplastic mammary epithelial cells was triggered by estrogens via a rapid, extranuclear ER-mediated response, after removal of an antiapoptotic NO/cGMP/cGMP-dependent protein kinase signal. (Endocrinology 151: 5602–5616, 2010
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17 novembre 19401940/11/17 (A69).Appartient à l’ensemble documentaire : PoitouCh
Additional file 3: Figure S2. of A novel aspirin prodrug inhibits NFκB activity and breast cancer stem cell properties
GTCpFE inhibits cytokine-induced NFκB target gene expression in breast cancer cells. MCF-7 cells were pretreated for 2 hours with increasing concentrations of GTCpFE followed by treatment with IL-1β (10 ng/ml) for another 2 hours. Expression of NFκB target genes, ICAM1 and CCL2 was measured by RT-QPCR. Drug inhibitory activity is plotted as % of IL-1β alone. (PPTX 72 kb
Structural Modulation of Oxidative Metabolism in Design of Improved Benzothiophene Selective Estrogen Receptor Modulators
Raloxifene and arzoxifene are benzothiophene selective estrogen receptor
modulators (SERMs) of clinical use in postmenopausal osteoporosis and
treatment of breast cancer and potentially in hormone replacement therapy. The
benefits of arzoxifene are attributed to improved bioavailability over
raloxifene, whereas the arzoxifene metabolite, desmethylarzoxifene (DMA) is a
more potent antiestrogen. As polyaromatic phenolics, benzothiophene SERMs
undergo oxidative metabolism to electrophilic quinoids. The long-term clinical
use of SERMs demands increased understanding of correlations between structure
and toxicity, with metabolism being a key component. A homologous series of
4′-substituted 4′-desmethoxyarzoxifene derivatives was developed,
and metabolism was studied in liver and intestinal microsomes. Formation of
glutathione conjugates was assayed in rat liver microsomes and novel adducts
were characterized by liquid chromatography-tandem mass spectrometry.
Formation of glucuronide conjugates was assayed in human intestine and liver
microsomes, demonstrating formation of glucuronides ranging from 5 to 100% for
the benzothiophene SERMs: this trend was inversely correlated with the loss of
parent SERM in rat liver microsomal incubations. Molecular orbital
calculations generated thermodynamic parameters for oxidation that correlated
with Hammett substituent constants; however, metabolism in liver microsomes
correlated with a combination of both Hammett and Hansch lipophilicity
parameters. The results demonstrate a rich oxidative chemistry for the
benzothiophene SERMs, the amplitude of which can be powerfully modulated, in a
predictable manner, by structural tuning of the 4′-substituent. The
predicted extensive metabolism of DMA was confirmed in vivo and compared with
the relatively stable arzoxifene and F-DMA
Benzothiophene Selective Estrogen Receptor Modulators with Modulated Oxidative Activity and Receptor Affinity
Raloxifene and Desmethylarzoxifene Block Estrogen- Induced Malignant Transformation of Human Breast Epithelial Cells
There is association between exposure to estrogens and the development and progression of hormone-dependent gynecological cancers. Chemical carcinogenesis by catechol estrogens derived from oxidative metabolism is thought to
contribute to breast cancer, yet exact mechanisms remain elusive. Malignant transformation was studied in MCF-10A
human mammary epithelial cells, since estrogens are not proliferative in this cell line. The human and equine estrogen
components of estrogen replacement therapy (ERT) and their catechol metabolites were studied, along with the influence of co-administration of selective estrogen receptor modulators (SERMs), raloxifene and desmethyl-arzoxifene (DMA), and
histone deacetylase inhibitors. Transformation was induced by human estrogens, and selectively by the 4-OH catechol metabolite, and to a lesser extent by an equine estrogen metabolite. The observed estrogen-induced upregulation of
CYP450 1B1 in estrogen receptor negative MCF-10A cells, was compatible with a causal role for 4-OH catechol estrogens, as was attenuated transformation by CYP450 inhibitors. Estrogen-induced malignant transformation was blocked by SERMs
correlating with a reduction in formation of nucleobase catechol estrogen (NCE) adducts and formation of 8-oxo-dG. NCE
adducts can be formed consequent to DNA abasic site formation, but NCE adducts were also observed on incubation of
estrogen quinones with free nucleotides. These results suggest that NCE adducts may be a biomarker for cellular electrophilic stress, which together with 8-oxo-dG as a biomarker of oxidative stress correlate with malignant transformation
induced by estrogen oxidative metabolites. The observed attenuation of transformation by SERMs correlated with these biomarkers and may also be of clinical significance in breast cancer chemoprevention