Synthesis and biological evaluation of 2,3-diaryl isoquinolinone derivatives as anti-breast cancer agents targeting ERα and VEGFR-2

The estrogen receptor α is recognized as important pharmaceutical target for breast cancer therapy, and vascular endothelial growth factor receptors (VEGFRs) play important roles in tumor angiogenesis including breast cancer. A series of 2,3-diaryl isoquinolinone derivatives were designed and synthesized targeting both estrogen receptor α (ERα) and VEGFR-2. Bioactivity evaluation showed that compounds 7c, 7d and 7f exhibited significant anti-proliferative and anti-angiogenesis activities via ERα and VEGFR-2 dependent mechanisms.     

Identification of 10-dehydrooxyglycyuralin E as a selective human estrogen receptor alpha partial agonist

Selective estrogen receptor modulators (SERMs) act as either agonist or antagonist of estrogen receptor (ER) in a tissue selective manner and have been used in several diseases such as breast cancer, postmenopausal syndrome, osteoporosis, and cardiovascular diseases. However, current SERMs may also increase the risk of serious side effects and trigger drug resistance. Herein, a screening program, that was designed to search for novel SERMs, resulted in the identification of a series of 2-arylbenzofuran-containing compounds that are ligands for ERα, when applying the Gaussia-luciferase reporter assay. One of these compounds, 10-dehydrooxyglycyuralin E (T9) was chemically synthesized. T9 showed anti-estrogenic/proliferative activity in ERα-positive breast cancer cells. Pretreatment of T9 prevented the mRNA expression of GREB1, which is an estrogen response gene. Furthermore, by an in silico docking simulation study we demonstrated that T9 showed interactions directly to ERα. Taken together, these results demonstrated that T9 is a candidate of SERMs and a useful seed compound for the foundation of the selective activity of SERMs.     

Synthesis,antiproliferative and pro-apoptotic activity of 2-phenylindoles

Breast cancer is the second most common cancer worldwide after lung cancer with the vast majority of early stage breast cancers being hormone-dependent. One of the major therapeutic advances in the clinical treatment of breast cancer has been the introduction of selective estrogen receptor modulators (SERMs). We describe the design and synthesis of novel SERM type ligands based on the 2-arylindole scaffold to selectively target the estrogen receptor in hormone dependent breast cancers. Some of these novel compounds are designed as bisindole type structures, while others are conjugated to a cytotoxic agent based on combretastatin A4 (CA4) which is a potent inhibitor of tubulin polymerisation. The indole compounds synthesised within this project such as 31 and 86 demonstrate estrogen receptor (ER) binding and strong antiproliferative activity in the ER positive MCF-7 breast cancer cell line with IC₅₀ values of 2.71 μM and 1.86 μM respectively. These active compounds induce apoptotic activity in MCF-7 cells with minimal effects on normal peripheral blood cells. Their strong anti-cancer effect is likely mediated by the presence of two ER binding ligands for 31 and an ER binding ligand combined with a cytotoxic agent for 86.     

Design,synthesis, biological evaluation and molecular docking studies of novel 3-aryl-4-anilino-2H-chromen-2-one derivatives targeting ERα as anti-breast cancer agents

The estrogen receptor (ER) has played an important role in breast cancer development and progression and is a central target for anticancer drug discovery. In order to develop novel selective ERα modulators (SERMs), we designed and synthesized 18 novel 3-aryl-4-anilino-2H-chromen-2-one derivatives based on previously reported lead compounds. The biological results indicated that most of the compounds presented potent ERα binding affinity and possessed better anti-proliferative activities against MCF-7 and Ishikawa cell lines than the positive control tamoxifen. The piperidyl substituted compounds such as 16d and 18d demonstrated strong ERα binding affinities and excellent anti-proliferative activities respectively. Compound 18d displayed the most potent ERα binding affinity with RBA value of 2.83%, while 16d exhibited the best anti-proliferative activity against MCF-7 cells with IC₅₀ value of 4.52 ± 2.47 μM. Further molecular docking studies were also carried out to investigate binding pattern of the newly synthesized compounds with ERα. All these results together with the structure–activity relationships (SARs) indicated that these 3-aryl-4-anilino-2H-chromen-2-one derivatives with basic side chain could serve as promising leads for further optimization as novel SERMs.     

Synthesis and structure–activity relationships of novel hybrid ferrocenyl compounds based on a bicyclic core skeleton for breast cancer therapy

Breast cancer is the most frequent cancer in women worldwide, and incidence is increasing year by year. Although current selective estrogen receptor modulators (SERMs) have clear advantages in the treatment of hormone-responsive breast cancer, they are ineffective for ER(−). In this study, we describe the design and synthesis of a series of dual-acting estrogen receptor (ER) and histone deacetylase (HDAC) inhibitors with incorporation of the ferrocenyl moiety, leading to novel hybrid ferrocenyl complexes (FcOBHS–HDACis) for breast cancer therapy. It is worth to note that these ferrocenyl conjugates could not only potently inhibit HDACs and the proliferation of ERα positive (ER(+)) breast cancer cells (MCF-7), but also show significant antiproliferative effect on ER(−) breast cancer cells (MDA-MB-231). Thus, the FcOBHS–HDACi conjugates represent a novel approach to the development of efficiently dual-acting agents for treatment of breast cancer.     

m-Carborane bisphenol structure as a pharmacophore for selective estrogen receptor modulators

A series of m-carborane derivatives was prepared based upon the structures of antiestrogenic drugs and their activities were evaluated by estrogen receptor alpha (ERalpha) binding assay and transactivation assay using human breast cancer cell line, MCF-7 cells. The m-carborane bisphenol 5 exhibited about a thousand times more potent ER agonistic activity than the o-carborane bisphenol 11. The m-carborane bisphenol structure appears to be a favorable hydrophobic pharmacophore for the development of novel selective estrogen receptor modulators (SERMs).     

Unexpected equivalent potency of a constrained chromene enantiomeric pair rationalized by co-crystal structures in complex with estrogen receptor alpha

Despite tremendous progress made in the understanding of the ERα signaling pathway and the approval of many therapeutic agents, ER+?breast cancer continues to be a leading cause of cancer death in women. We set out to discover compounds with a dual mechanism of action in which they not only compete with estradiol for binding with ERα, but also can induce the degradation of the ERα protein itself. We were attracted to the constrained chromenes containing a tetracyclic benzopyranobenzoxepine scaffold, which were reported as potent selective estrogen receptor modulators (SERMs). Incorporation of a fluoromethyl azetidine side chain yielded highly potent and efficacious selective estrogen receptor degraders (SERDs), such as 16aa and surprisingly, also its enantiomeric pair 16ab. Co-crystal structures of the enantiomeric pair 16aa and 16ab in complex with ERα revealed default (mimics the A-D rings of endogenous ligand estradiol) and core-flipped binding modes, rationalizing the equivalent potency observed for these enantiomers in the ERα degradation and MCF-7 anti-proliferation assays.     

Effects of SERM (selective estrogen receptor modulator) treatment on growth and proliferation in the rat uterus

Background  

Selective estrogen receptor modulators (SERMs) have been developed in order to create means to control estrogenic effects on different tissues. A major drawback in treatment of estrogen receptor (ER) positive breast cancer with the antagonist tamoxifen (TAM) is its agonistic effect in the endometrium. Raloxifene (RAL) is the next generation of SERMs where the agonistic effect on the endometrium has been reduced.     

Ligand-selective interdomain conformations of estrogen receptor-alpha

Selective estrogen receptor modulators (SERMs) inhibit estrogen activation of the estrogen receptor (ER) in some tissues but activate ER in other tissues. These tissue-selective actions suggest that SERMs may be identified with tissue specificities that would improve the safety of breast cancer and hormone replacement therapies. The identification of an improved SERM would be aided by understanding the effects of each SERM on the structure and interactions of ER. To date, the inability to obtain structures of the full-length ER has limited our structural characterization of SERM action to their antiestrogenic effects on the isolated ER ligand binding domain. We studied the effects of estradiol and the clinically useful SERMs 4-hydroxytamoxifen and fulvestrant on the conformation of the full-length ERalpha dimer complex by comparing, in living human breast cancer cells, the amounts of energy transfer between fluorophores attached to different domains of ERalpha. Estradiol, 4-hydroxytamoxifen, and fulvestrant all promoted the rapid formation of ERalpha dimers with equivalent interaction kinetics. The amino- and carboxyl-terminal ERalpha domains both contain activation functions differentially affected by these ligands, but the positions of only the carboxyl termini differed upon binding with estradiol, 4-hydroxytamoxifen, or fulvestrant. The association of a specific ERalpha dimer conformation with the binding of ligands of different clinical effect will assist the identification of a SERM with optimal tissue-selective estrogenic and antiestrogenic activities. These studies also provide a roadmap for dissecting important structural and kinetic details for any protein complex from the quantitative analysis of energy transfer.     

Acquired resistance to selective estrogen receptor modulators (SERMs) in clinical practice (tamoxifen & raloxifene) by selection pressure in breast cancer cell populations

Tamoxifen, a pioneering selective estrogen receptor modulator (SERM), has long been a therapeutic choice for all stages of estrogen receptor (ER)-positive breast cancer. The clinical application of long-term adjuvant antihormone therapy for the breast cancer has significantly improved breast cancer survival. However, acquired resistance to SERM remains a significant challenge in breast cancer treatment. The evolution of acquired resistance to SERMs treatment was primarily discovered using MCF-7 tumors transplanted in athymic mice to mimic years of adjuvant treatment in patients. Acquired resistance to tamoxifen is unique because the growth of resistant tumors is dependent on SERMs. It appears that acquired resistance to SERM is initially able to utilize either E₂ or a SERM as the growth stimulus in the SERM-resistant breast tumors. Mechanistic studies reveal that SERMs continuously suppress nuclear ER-target genes even during resistance, whereas they function as agonists to activate multiple membrane-associated molecules to promote cell growth. Laboratory observations in vivo further show that three phases of acquired SERM-resistance exists, depending on the length of SERMs exposure. Tumors with Phase I resistance are stimulated by both SERMs and estrogen. Tumors with Phase II resistance are stimulated by SERMs, but are inhibited by estrogen due to apoptosis. The laboratory models suggest a new treatment strategy, in which limited-duration, low-dose estrogen can be used to purge Phase II-resistant breast cancer cells. This discovery provides an invaluable insight into the evolution of drug resistance to SERMs, and this knowledge is now being used to justify clinical trials of estrogen therapy following long-term antihormone therapy. All of these results suggest that cell populations that have acquired resistance are in constant evolution depending upon selection pressure. The limited availability of growth stimuli in any new environment enhances population plasticity in the trial and error search for survival.     

Selective estrogen receptor modulators (SERMS) and their roles in breast cancer prevention

Tamoxifen has not only proved to be a valuable treatment for estrogen receptor (ER)-positive breast cancer, but is also a pioneering medicine for chemoprevention in high-risk pre- and postmenopausal women. Insights into the pharmacology and toxicology of tamoxifen have led to the recognition of selective ER modulators (SERMs) with estrogen-like actions in maintaining bone density and in lowering circulating cholesterol, but antiestrogenic actions in the breast. Raloxifene, a related SERM, is now available to treat osteoporosis and is also being tested as a preventive for breast cancer and coronary heart disease. Emerging knowledge about the action of SERMs will provide clues for the design of mechanism-based medicines.     

Steroidal oxathiazine inhibitors of estrone sulfatase

The presence of estrone sulfatase in breast tumors and the high levels of circulating estrone sulfate may contribute the major portion of estrogen synthesized locally in breast tissues through conversion of estrone sulfate to estrone by the enzyme. Using inhibitors of estrone sulfatase for the treatment of estrogen-dependent (estrogen receptor positive, ER(+)) breast cancer could be a very effective therapeutic strategy for the treatment of estrogen-dependent breast tumors in postmenopausal women. Therefore, we designed and synthesized several steroidal 2',3'-oxathiazines that inhibit estrone sulfatase and have greatly reduced estrogenic side effects. Our in vitro studies indicate that the oxathiazine compounds have inhibitory activity on estrone sulfatase in MCF-7 human breast cancer cells. These estrone sulfatase inhibitors (ESIs) also inhibit the growth of MCF-7 cells induced by estrone sulfate. In addition, our in vivo experiments demonstrate that our ESIs have moderate antitumor activity against MCF-7 breast cancer xenografts in Balb/c athymic nude mice. The synthesis and biological activity of a number of these unique steroidal ESIs are described.     

Pros and cons of existing treatment modalities in osteoporosis: a comparison between tibolone,SERMs and estrogen (+/-progestogen) treatments

Tibolone, selective estrogen receptor modulators (SERMs) like tamoxifen and raloxifene, and estrogen (±progestogen) treatments prevent bone loss in postmenopausal women. They exert their effects on bone via the estrogen receptor (ER) and the increase in bone mass is due to resorption inhibition. The effect of SERMs on bone mineral density is less than that with the other treatments, but the SERM raloxifene still has a positive effect on vertebral fractures. In contrast to tibolone and estrogens (±progestogen), SERMs do not treat climacteric complaints, whilst estrogen plus progestogen treatments cause a high incidence of bleeding. Estrogen plus progestogen combinations have compromising effects on the breast. Tibolone and SERMs do not stimulate the breast or endometrium. Unlike SERMs, tibolone does not posses antagonistic biological effects via the ER in these tissues. Estrogenic stimulation in these tissues is prevented by local metabolism and inhibition of steroid metabolizing enzymes by tibolone and its metabolites. SERMs and estrogen (±progestogen) treatments increase the risk of venous thromboembolism (VTE), whilst estrogen (±progestogen) combinations have unwanted effects on cardiovascular events. So far, no detrimental effects of tibolone have been observed with respect to VTE or cardiovascular events. The clinical profile of tibolone therefore has advantages over those of other treatment modalities. It is also clear that tibolone is a unique compound with a specific mode of action and that it belongs to a separate class of compounds that can best be described as selective, tissue estrogenic activity regulators (STEARs).     

miR-424对雌激素受体α表达的抑制作用

目的:筛选能够抑制在乳腺癌发生中起关键作用的雌激素受体α(ERα)表达的microRNA(miRNA)分子,并在ERα阳性的乳腺癌细胞株中初步检测其生物学功能。方法:在ERα阳性的乳腺癌细胞ZR75-1中转染多种miRNA的表达载体,Western印迹检测ERα的表达水平,找到可以抑制ERα表达的miRNA分子;将该miRNA的表达载体转染ZR75-1后,在雌激素E2的作用下,检测该miRNA分子对细胞生长的影响。结果:经过筛选,得到能够抑制ERα表达的miRNA分子miR-424;生长曲线结果显示,miR-424能够在不依赖于E2的情况下阻抑ZR75-1的生长。结论:该研究为进一步研究miR-424在ERα信号通路中的生物学功能及研究乳腺癌的发生发展机制奠定了基础。     

Identification of novel proteins induced by estradiol, 4-hydroxytamoxifen and acolbifene in T47D breast cancer cells

Tamoxifen is currently used as adjuvant therapy for estrogen receptor (ER) positive breast cancer patients and as a chemopreventative agent. Although ER is a predictive marker for tamoxifen response, ER status fails to predict tamoxifen response in a significant number of patients highlighting the need to identify new pathways for tamoxifen sensitivity/resistance. To identify novel proteins induced by tamoxifen in breast cancer cells sensitive to tamoxifen growth inhibition, two-dimensional (2D) gel electrophoresis was used to profile proteins in T47D breast cancer cells. Six proteins were identified that were differentially regulated by 17beta-estradiol, 4-hydroxytamoxifen and the pure antagonist acolbifene (EM-652); calreticulin, synapse associated protein 1 (SYAP1), CD2 antigen binding protein 2 (CD2BP2), nucleosome assembly protein 1 like 1 (NAP1L1), d-3-phosphoglycerate dehydrogenase (3-PHGDH) and pyridoxine 5' phosphate oxidase (PNPO). At the mRNA level, these ligands differentially regulated expression of mRNAs encoding the identified proteins in T47D and MCF7 cells but had no effect on mRNA in ERalpha-negative MDA-MB-231 breast cancer cells. These novel SERM-regulated proteins may participate in new or existing pathways for sensitivity or resistance to SERMs.     

Synthesis and evaluation of raloxifene derivatives as a selective estrogen receptor down-regulator

Estrogen receptors (ERs) play a major role in the growth of human breast cancer cells. A selective estrogen receptor down-regulator (SERD) that acts as not only an inhibitor of ligand binding, but also induces the down-regulation of ER, would be useful for the treatment for ER-positive breast cancer. We previously reported that tamoxifen derivatives, which have a long alkyl chain, had the ability to down-regulate ERα. With the aim of expanding range of the currently available SERDs, we designed and synthesized raloxifene derivatives, which had various lengths of the long alkyl chains, and evaluated their SERD activities. All compounds were able to bind ERα, and RC10, which has a decyl group on the amine moiety of raloxifene, was shown to be the most potent compound. Our findings suggest that the ligand core was replaceable, and that the alkyl length was important for controlling SERD activity. Moreover, RC10 showed antagonistic activity and its potency was superior to that of 4,4′-(heptane-4,4-diyl)bis(2-methylphenol) (18), a competitive antagonist of ER without SERD activity. These results provide information that will be useful for the development of promising SERDs candidates.