European Journal of Medicinal Chemistry 143 (2018) 515e531

Contents lists available at ScienceDirect

European Journal of Medicinal Chemistry

journal homepage: http://www.elsevier.com/locate/ejmech

Review article Tamoxifen a pioneering drug: An update on the therapeutic potential of tamoxifen derivatives

** * Shagufta , Irshad Ahmad

Department of Mathematics and Natural Sciences, School of Arts and Sciences, American University of Ras Al Khaimah, Ras Al Khaimah, United Arab Emirates article info abstract

Article history: Tamoxifen (ICI 46 474), trans-1-(4-b-dimethylaminoethoxyphenyl)-1,2-diphenylbut-1-ene, is the most Received 1 September 2017 commonly used drug for the treatment of estrogen receptor positive breast cancer and has been saving Received in revised form lives worldwide for the past four decades. Tamoxifen is considered a pioneering drug due to its ubiq- 25 October 2017 uitous use in both treatment and chemoprevention of breast cancer and also for research addressing Accepted 20 November 2017 novel selective estrogen receptor modulators (SERMs). Tamoxifen is cost effective, lifesaving, and devoid Available online 24 November 2017 of major side effects in the majority of patients. The discovery of tamoxifen metabolites such as 4- hydroxy tamoxifen, N-desmethyl tamoxifen, and endoxifen has facilitated understanding of tamoxi- Keywords: Tamoxifen fen's and its metabolites' mechanisms of action in breast cancer therapy. Continuous efforts are being Tamoxifen derivatives made by both industry and academia to synthesize novel tamoxifen derivatives in order to better un- Breast cancer derstand the mechanism of this drug's action and to generate new agents with reduced side effects for Tamoxifen metabolites many therapeutic targets. This review article comprises the tamoxifen derivatives reported in the SERMs literature in the last few years and we anticipate that it will assist medicinal chemists in the synthesis of Antiestrogens novel and pharmacologically potent agents for various therapeutic targets. © 2017 Elsevier Masson SAS. All rights reserved.

Contents

1. Introduction ...... 515 2. Therapeutic potential of tamoxifen derivatives ...... 517 2.1. Tamoxifen derivatives with modification on aminoalkyl chain ...... 517 2.2. Tamoxifen derivatives with modification on amino alkyl chain and phenyl rings ...... 517 2.3. Tamoxifen derivatives with modification on amino alkyl and ethyl chain ...... 519 2.4. Tamoxifen derivatives with modification on ethyl chain and phenyl rings ...... 520 2.5. Tamoxifen derivatives with modification on phenyl rings, amino alkyl and ethyl chains ...... 520 2.6. Flexible tamoxifen derivatives ...... 522 2.7. Tamoxifen derivatives with heteroaromatic groups ...... 522 2.8. Tamoxifen conjugates ...... 523 2.9. Derivatives of tamoxifen metabolites ...... 526 3. Conclusion ...... 527 Acknowledgements ...... 527 References...... 527

1. Introduction

* Corresponding author. ** Corresponding author. Tamoxifen (1)(Fig. 1) is considered a groundbreaking drug in E-mail addresses: [email protected] (Shagufta), [email protected]. medical oncology that has saved many lives over the past four ae (I. Ahmad). https://doi.org/10.1016/j.ejmech.2017.11.056 0223-5234/© 2017 Elsevier Masson SAS. All rights reserved. 516 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

N N N O O O N

O O

HO OH HO S I 1 2 3 4

OH N N O O O

HO 5 6 7 Cl Cl

Fig. 1. Chemical structure of drug tamoxifen (1), raloxifene (2) and important tamoxifen derivatives in clinical development trials; droloxifene (3), idoxifene (4), lasofoxifene (5), toremifene (6) and ospemifene (7). decades and progressed to become a significant part of our osteoporosis and prevent breast cancer in high risk post- healthcare [1e4]. The story of the development of tamoxifen as a menopausal women [24e27]. Although the benefits of tamoxifen pioneering medicine for cancer treatment is very fascinating and are prominent, the use of tamoxifen is associated with increased inspiring for researchers worldwide who are working toward the risk of side effects such as hot flashes, menstrual abnormalities, development of novel medicines for various diseases. In the late uterine cancer and thromboembolic phenomena [28,29]. Since 1950s, in the laboratories of Imperial Chemical Industries Ltd. tamoxifen use is associated with an increase in cancer risks and Pharmaceutical division now known as AstraZeneca, a team with unpleasant side effects, it is generally taken for five years followed Dora Richardson (chemist), Michael J. K. Harper (reproductive by different therapeutics depending on the patient's condition; endocrinologist), and Arthur L. Walpole (Head of Reproduction furthermore, its use is not acceptable for all high-risk women. Research) was given the task of developing a post-coital contra- Further results from the Adjuvant Tamoxifen: Longer against ceptive (the morning-after pill). Eventually, tamoxifen (Imperial Shorter (ATLAS) trial suggested that ten years of adjuvant tamox- Chemical Industries (ICI) 46 474, an antiestrogenic trans isomer of a ifen therapy can reduce mortality to greater than five years [30]. substituted triphenyl ethylene, was invented and received mar- Tamoxifen is marketed as a single Z (trans) isomer of p-b- keting approval as a fertility treatment, but the drug never actually dimethylaminoethoxy-1,2-diphenylbut-1-ene and is considered a proved useful in human contraception. Walpole was very inter- lead compound that initiated the SERM development for the ested in exploring tamoxifen's application in cancer research and treatment of various diseases (such as osteoporosis, rheumatoid treatment [5]. In 1972, Walpole collaborated with V.C. Jordan to arthritis) and also for application of the SERM concept for all the conduct scientific research that led to the reinvention of the failed members of the nuclear receptor family [31e38]. Pharmacological ICI 46 474 contraceptive to become tamoxifen, the first targeted studies of tamoxifen in the human body suggested its conversion to agent for the treatment and prevention of breast cancer [6e11]. three active metabolites: 1.) 4-hydroxy tamoxifen (8); 2.) N-des- The “Father of Tamoxifen”, V. C Jordan, introduced the strategy methyltamoxifen (9); and 3.) 4-hydroxy-N-desmethyltamoxifen, of targeting estrogen receptor positive tumors with long term also known as endoxifen (10)(Fig. 2) [39e41]. In humans, N-des- adjuvant tamoxifen therapy that increased the survival rate of methyltamoxifen is the primary metabolite followed by endoxifen hundreds of thousands of breast cancer patients around the world and then 4-hydroxytamoxifen. These metabolites are potent anti- [12,13]. Tamoxifen is inexpensive and readily available to under- estrogens and are used to understand the tamoxifen's mechanism funded healthcare systems and that feature has caused an increase of action [42,43]. Tamoxifen's pharmacological profile indicates in its worldwide popularity as a miracle drug for breast cancer. that it is a prodrug, and its anticancer activity occurs via its active Currently, tamoxifen is used for the treatment of all stages of es- metabolite, 4-hydroxy tamoxifen (8) and its desmethyl analogue trogen receptor (ER) -positive (ERþ) breast cancer in pre- and post- endoxifen (10), which are generated by the action of hepatic CYP menopausal women in addition to hormone treatment for male 2D6 and CYP3A4/3A5 isozymes on tamoxifen after hydroxylation breast cancer [14,15]. In addition, tamoxifen is used for the treat- followed by N-demethylation [44e45]. It has been established that ment of ductal carcinoma in situ and for the prevention of breast patients with variant forms of the gene CYP 2D6 do not receive cancer in women at high risk of developing the disease [16,17]. therapeutic benefits from tamoxifen administration or even suffer Initially, tamoxifen was known as an anti-estrogen that reduced relapses because of slow tamoxifen prodrug metabolism into its estrogen-induced effects by blocking estrogen receptors in breast active metabolites [46e48]. tissues. Later, rigorous pharmacological investigation of tamoxifen Several interesting facts associated with tamoxifen and its me- provided evidence that tamoxifen acts as agonist at estrogen re- tabolites made it a pioneering agent for initiating new therapeutic ceptors in other body tissues such as endometrium, liver, and bone investigation, and its pharmacogenomics is playing a significant and thus, led to the development of a new drug group, the selective role in redefining health care. In recent years, some of the tamox- estrogen receptor modulators (SERMs) [18e23]. One of the ifen analogues; for example, droloxifene (3), idoxifene (4), laso- important examples of a SERM is raloxifene (2)(Fig. 1), a failed foxifene (5), toremifene (6) and ospemifene (7)(Fig. 1) have been breast cancer drug that has been successfully used to treat studied extensively in clinical trials [49]. Literature reports indicate Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 517

35 N O NH O 30 25 20 CYP3A4/3A5

M) 15 μ 10 Tamoxifen (1) N-desmethyltamoxifen (9) GI50 ( 5 0 CYP2D6 CYP2D6 SR M21 HeLa EKVX A498 T-47D HT-29 A2780 UO-31 HCT15 MCF-7 SNB-75 SK-OV3 DU-145 MOLT-4 OVCAR5 LOX IMVI N NH NCI-H460 O O MSTO-21 1H MDA-MB-468 MDA-MB-435 MDA-MB-231

B Fig. 3. GI50 (half minimal growth inhibition concentration) of tamoxifen against CYP3A4/3A5 A different tumor cell lines. Data obtained from Ref [71,81,112,119,122,127]. C HO HO 4-hydroxytamoxifen (8) 4-hydroxy-N-desmethyltamoxifen (10) structural modifications and further discussed their pharmaceu- tical significance. Moreover, tamoxifen metabolites analogues are Fig. 2. Tamoxifen metabolism in the human and related metabolites (8e10). also taken into consideration.

fi that continuous efforts are being made by researchers worldwide 2.1. Tamoxifen derivatives with modi cation on aminoalkyl chain to identify novel tamoxifen derivatives for breast cancer and other therapeutic targets. This review is an effort to provide readers with The presence of a basic aminoalkyl chain on tamoxifen plays a information about recent and continuing development in this major role in its antiestrogenic activity. Replacement of one N- fl research area. methyl group of aminoalkyl side chain by a N-(2,2,2-tri uoroethyl) group has a detrimental effect, and the compound loses its potency to inhibit the growth of the ERþ, MCF-7 cells [68].In2006, 2. Therapeutic potential of tamoxifen derivatives Agouridas et al. reduces the basicity of side chain by synthesizing a series of fluorinated derivatives of tamoxifen and studying their In recent years the biological activity of tamoxifen has been effects on their activities [69]. In addition to examining the effects explored extensively in different therapeutic targets, and the re- of the substituents' chain lengths, the corresponding non- fl sults are very encouraging. In addition to breast cancer, tamoxifen uorinated analogues were also prepared. Analysis of the pre- fi fl has been used to treat infertility, gynecomastia, retroperitoneal pared compound's binding af nity revealed that the uorinated fi e fibrosis, and idiopathic sclerosing mesenteritis [50e52] and exerts compounds' binding af nity (11a c)(Fig. 4) was one order of e beneficial effects on osteoporosis and reduces the incidence of magnitude less than tamoxifen (RBA ~0.2% 0.3%), and the non- fl e cardiovascular diseases [53e55]. This drug has also been studied to ourinated analogues (11d f)(Fig. 4) were as effective as tamox- treat Riedel's thyroiditis, bipolar disorder, and McCune-Albright ifen. Hence it was proposed that decreasing the basicity by fl syndrome [56e58]. Tamoxifen has a wide range of activities, replacing the methyl group of amino alkyl chain with a uorinated including antifungal, antioxidant, and antiviral (especially anti- moiety decreases tamoxifen analogues' capacity to bind to the hepatitis C virus activity) activities, antiangiogenesis properties, ligand binding pocket in addition to reducing the ER-mediated fl induction of intracellular calcium release, stimulation of trans- antagonistic properties. The ourinated compounds displayed forming growth factor beta secretion, alteration of cellular mem- diminished ability to inhibit growth, stabilize ER, and to modulate brane properties, and apoptosis induction [59e66]. Tamoxifen has ethylene response factor (ERF) and activator protein (AP-1) tran- been shown to target a number of proteins, including calmodulin, scriptional activities. In addition, enhancement of agonistic activity protein kinase C, phospholipase C, phosphoinositide kinase, P- on growth was also observed. The efforts to limit metabolic alter- glycoprotein, and swell-induced chloride channels [67]. Tamoxifen ations of the tamoxifen's amino alkyl side chain were not suc- has exhibited very promising anticancer activity against different cessful. It was suggested that additional thoughts and efforts are fi breast and other tumor cell lines (Fig. 3) [71,81,112,119,122,127]. The needed to nd an atom or a group as a substitute for the nitrogen discovery of tamoxifen as an efficient therapeutic agent facilitated atom. the synthesis of various analogues through chemical structure modification of tamoxifen and its metabolites. The reported 2.2. Tamoxifen derivatives with modification on amino alkyl chain tamoxifen derivatives exhibited encouraging biological activities and phenyl rings and provided support in understanding tamoxifen's mechanism of action. Researchers around the world modified tamoxifen's chem- In 2007, Shiina et al. reported the short step synthesis and ical structure by substituting amino alkyl and ethyl chains with cytotoxicity activity of pseudo-symmetrical tamoxifen derivatives several groups and also by introducing substituents on phenyl bearing aminoalkyl chains on two phenyl rings [70]. The three rings. Moreover, in recent years, some reports have highlighted the pseudo-symmetrical tamoxifen derivatives (12aec)(Fig. 4)were importance of incorporation of the additional methylene or heter- synthesized using a three component coupling reaction between oaromatic group in the tamoxifen skeleton in addition to its the aromatic aldehyde, cinnamyltrimethylsilane, and aromatic conjugation with important moieties. In this review, we have made nucleophiles with a Lewis acid catalyst. The antitumor activity of efforts to categorize tamoxifen derivatives on the basis of their these compounds was determined against human promyelocytic 518 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

O N OR O R HN

RO HO 12a;R=CHCH 11a; R=CH2CF3 2 2 N 11b; R=CH2CH2CF3 11c; R=CH2CH2OCF3 OH 12b;R=CHCH 11d; R=CH2CH3 2 2 N 13 11e; R=CH2CH2CH3 11f; R=CH2CH2CH2CH3 12c;R=CH2CH2 N O

H N N N O (CH2)11CH3 O O

N N HO O O 16 14 15

H O N O (CH2)9CH2R HN

C Cl B

A Cl HO O 17a; R=H 17b; R=F 18 17c; R=OH

Fig. 4. Tamoxifen derivatives (11e18) with modifications either on amino alkyl chains or on both amino alkyl chains and phenyl rings.

leukemia (HL-60) cancer cells using MTT assay in which the effi- derivatives were synthesized using McMurry coupling reaction and ciency of pseudo-symmetrical tamoxifen derivatives were deter- the characterization and structural assignment of E, Z isomers were mined by measuring their capacity to decrease cell viability. It was assisted by 2D-NOESY experiments. The compounds' in vitro anti- observed that compounds 12a and b, bearing pyrrolidine and proliferative activities were performed on three different cell lines: piperidine side chain respectively, showed better cytotoxic activ- 1.) human breast cancer cell line (MCF-7), which overexpress the ities and induced apoptosis in the HL-60 cancer cells. In contrast, estrogen receptor; 2.) HeLa, an estrogen-independent human tu- compound 12c with a morpholine side chain was inactive when mor cell line (cervix adenocarcinoma); and 3.) MSTO-211H, another used in the same cell line. 12a and b effectively reduced cell estrogen-independen ttumor cell line (biphasic mesothelioma). viability in a dose-dependent manner. 12a, at final concentrations With respect to tamoxifen, most of the compounds in the series of 5, 7.5, and 10 mM and a 6-h incubation period inhibited cell showed comparable or even higher antiproliferative activity on viability by >90%. Similarly, compound 12 b at final concentrations estrogen sensitive MCF-7 cells, whereas when compared with 4- of 7.5 and 10 mM and a 6-h incubation period inhibited cell viability hydroxytamoxifen, the compounds in the series were less effec- by >80%. In order to further confirm that 12a- or b-induced cell tive. Also, the Z isomers exhibited slightly more activity toward death was caused by apoptosis or necrosis, agarose gel electro- MCF-7 cell lines compared to E isomers. With respect to anti- phoresis for DNA cleavage was performed with HL-60 cells and proliferative activity on MCF-7 cell lines, the most active compound 9 mM final concentration of the tested compound. The result was of the series was 13 (Fig. 4), which showed a GI50 ¼ 2.9 mM that is 4 positive, and DNA fragmentation was observed, which suggested times higher than the reference drug, tamoxifen (GI50 ¼ 12.0 mM). that the pseudo-symmetrical tamoxifen derivatives a12 and b As expected, tamoxifen's antiproliferative effects on two estrogen- induced cell death through apoptosis. independent cell lines (HeLa and MSTO-211H) was much reduced Christodoulou et al. (2013) designed and synthesized novel (GI50 ¼ 32.6 mM and 23.3 mM, respectively). In comparison, except derivatives of tamoxifen by substituting tamoxifen's aminoalkyl for 13 (HeLa, GI50 ¼ 7.4 mM and MSTO-211H, GI50 ¼ 8.4 mM) most of group with an amide side chain [71]. Although in all the compound the derivatives exhibited GI50 values for HeLa and MSTO-211H of the series triarylethylene skeleton of tamoxifen having hydroxyl similar to the values obtained for MCF-7 cells. These results sug- group in position 4 of the phenyl moiety was sustained. The gested that novel tamoxifen derivatives' mechanisms of action with Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 519 respect to antiproliferative activity were independent from those ERa-positive and ERb-negative breast cancer cells via different actions on the estrogen receptor. To further study the possible pathways; for example, pathways involving production of oxidative molecular targets responsible for the cytotoxicity, the effects on stress, induction of mitochondrial permeability transition, cer- relaxation activities of DNA topoisomerase I and II were assayed. amide generation or changes in cell membrane fluidity may be These novel derivatives were able to inhibit topoisomerase II- involved [79]. Tamoxifen was also shown to inhibit DNA top- mediated relaxation activity; therefore, it was suggested that po- oisomerases and be taken up by hepatic mitochondria in order to tential intracellular targets could be nuclear enzymes. attain high concentrations with the purpose of inhibiting both b- Furthermore, Ridaifen-B (12a)(Fig. 4), a tamoxifen derivative, oxidation and respiration [71,80]. Based on these results Christo- has shown higher potency than tamoxifen with respect to inducing doulouet et al. (2016) synthesized a few tamoxifen analogues that ER-cell apoptosis via mitochondrial membrane potential pertur- showed significant antiproliferative activity on both estrogen- bation [72] Thus, Hosegawa et al. (2014), using a series of tamoxifen dependent (MCF-7) and -independent (HeLa) human tumor cell derivatives that consisted of different Ridaifen forms, identified lines. This revealed involvement of a molecular target different nonpeptide and noncovalent inhibitors of the human 20S protea- from the ER [81]. The most promising compounds of the series some [73]. The most active derivative was 14 (Fig. 4), which, at were syn isomers with an isobutyramide moiety as side chain. submicromolar levels (IC50 ¼ 0.64, 0.34, and 0.43 mM) inhibited the These compounds exhibited 4-fold more potent cytotoxicity than three different enzymatic activities (CT-L, T-L and PGPH, respec- the reference drug on both estrogen-dependent and -independent tively) of the 20s proteasome. A series of 14 analogues were pre- human cell lines. Additionally, these compounds displayed the pared and then examined in order to study structure-activity ability to inhibit topoisomerase II-mediated relaxation activity of relationships. The smallest analogue of 14 was compound 15 supercoiled pBR322 DNA. Further, a continuing novel series of (Fig. 4), which was found to inhibit proteasome activity with po- tamoxifen analogues having triaryl skeletons substituted with the tency similar to 14. These derivatives induced apoptotic cell deaths isobutyramide moiety was prepared. The antiproliferative activity via the proteasome inhibition within the cells. Kinetic analyses of of these compounds was evaluated on two human tumor cell lines the inhibition and docking simulations suggested that the analogue (MCF-7 and HeLa) and two human ovarian cancer cell lines (A2780 of 14 interacted with the protease subunit in a different manner. and OVCAR5). These compounds' biological activities suggested The selective estrogen receptor down regulators (SERDs) are a that the presence of an OH group or Cl moieties in the aromatic ring class of antagonists that not only inhibit the binding of estrogens B favors cytotoxic activity, whereas a methoxy or hydroxyl sub- such as 17b-estradiol (E2) to the ER but also can induce rapid ER stituent in the aromatic ring A has a detrimental effect on cytotoxic down-regulation [74,75]. They have no agonistic activity in many activity. Most of the compounds were more active in MCF-7 and tissues and have been considered useful for ER þ or tamoxifen HeLa cell lines compared to A2780 and OVCAR5 cell lines. Com- resistant breast cancer cells [76]. Shoda et al. (2014) designed and pound 18 (Fig. 4) showed the lowest GI50 values in MCF-7 and HeLa synthesized tamoxifen derivatives by introducing a long alkyl chain cell lines (GI50 ¼ 6.6 mM and 2.2 mM, respectively). Additional (such as hexyl, dodecyl, and octadecyl) on 4-hydroxytamoxifen's studies with 18 suggested that it was able to induce apoptosis and amine moiety and then evaluated its ability to facilitate ER degra- that topoisomerase II was a possible intracellular target. dation in MCF-7 cells and also examine its binding affinity for ER [77]. It was expected that a long alkyl chain on the amine moiety 2.3. Tamoxifen derivatives with modification on amino alkyl and may destabilize the ER by protruding from the ligand binding ethyl chain pocket and thus inhibiting helix 12 interactions and coactivator binding. These results suggested that the presence of a secondary Nonsteroidal anti-inflammatory drugs (NSAIDs) are commonly amine is essential for ERa down-regulation. The most promising used to treat rheumatoid arthritis and osteoarthritis, and these compound in the series was 16 (Fig. 4), which, at 10 mM caused a agents mostly act through the cyclooxygenase (COX) inhibitory reduction in the ERa protein levels in cells that were treated with pathway. Several reports have suggested that selective COX-2 in- this compound. In addition, a decrease in the ERa level in MCF- hibitors are associated with fewer side effects compared to 7 cells was observed in a concentration-dependent manner at nonselective one [82,83] Selective COX-2 inhibitors are also useful doses of compound 16 ranging from 1 to 30 mM after a 6-h incu- in the treatment of a wide variety of cancer and neurodegenerative bation period. It was observed that compound 16-induced reduc- disorders [84]. In 2004, Uddin et al. designed and synthesized tion in ERa was caused by cellular proteasomal degradation. acyclic triaryl olefinic compounds, which were structurally similar Later, in order to obtain more potent SERDs and to explain the to tamoxifen, as selective COX-2 inhibitors [85,86]. The presence of mechanism of ER down-regulation, six new tamoxifen derivatives identical C-1 phenyl substituents excluded the possibility of (E)- that had various length and terminal groups (octyl, decyl, tetra- and (Z)- stereoisomers. The most active compound in this series decyl, hexadecyl, 10-hydroxydecyl, and 10-fluorodecyl) of the long was 1,1-diphenyl-2-(4-methylsulfonylphenyl)hex-1-ene (19) alkyl side chain were synthesized [78]. Western blotting results (Fig. 5), which exhibited high potency (IC50 ¼ 0.014 mM) and se- showed that C10-bearing decyl group on the amine moiety of 4- lective COX-2 inhibitory activity (selectivity index >7142). OHT were most potent among the compounds having simple In the literature, sulfoximines, monoaza sulfone analogues, are alkyl chains on the amine moiety; thus, alkyl chain length appears considered potentially useful groups for drug development to play a significant role in the ER down-regulation. ER binding [87e89]. Considering the significance of triaryl alkenes and sul- affinity studies revealed that compound 17a's binding affinity foximines in medicinal chemistry, Chen et al. (2012) used sulfonyl- (Fig. 4) for ERa (IC50 ¼ 3.6 nM) was better than 4-hydroxy tamox- substituted triaryl olefin 19 as a starting point to synthesized ifen (IC50 ¼ 5.6 nM). The introduction of the fluoro group on the sulfoximine-based analogues 20 and 21 (Fig. 5) [90]. Initially, the alkyl chain terminus (17b) maintained high ERa binding COX inhibitory activity of the synthesized compounds was (IC50 ¼ 3.4 nM) and increased the potency of SERD activity. How- checked; however, the compounds showed very low COX inhibitory ever, introduction of hydrophilic hydroxyl group on alkyl chain activity. Furthermore, sulfoximines' (20 and 21) estrogen binding terminus (17c) decreased binding affinity for ERa (IC50 ¼ 210 nM) in affinities were evaluated using human recombinant enzymes, and addition to reducing the ability to down-regulate the ERa protein sulfone 19 was used as a reference compound. Sulfone 19 was se- levels. lective and moderately active for ER b (77%) whereas sulfoximines It has been reported that tamoxifen can induce apoptosis in both (20 and 21) were basically ERa and ERb unselective and exhibited 520 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

O O O S S S O N N CN H

R R

20a; R=CH CH CH CH 21a; R=CH CH CH CH 19 2 2 2 3 2 2 2 3 20b;R=CH3 21b;R=CH3

N O O O

O HO NH2 HN 22 23 R 24 OH O H H N N 23a; R= 23b; R=

Fig. 5. Tamoxifen derivatives (19e24) with modifications on amino alkyl and ethyl chain/ethyl chain and phenyl ring. almost equal affinity for both ERs. Compound 21a (10 mM) showed 2.4. Tamoxifen derivatives with modification on ethyl chain and maximum inhibition of up to 91% for ER a and 80% for ER b. phenyl rings Literature reports have recommended that the tamoxifen's tri- arylethylene framework acts as an estrogen agonist, and the two In the human genome, orphan estrogen-related receptors exist alkyl chains attached to it are responsible for full or partial antag- as three subtypes (specifically ERRa, ERRb and ERRg) [95]. The onist behavior. Tamoxifen acts as a partial anti-estrogen agent after sequencing of these three orphan nuclear receptors sequencing is alkyl chain lengthening, and the presence of alkyl chains of similar to classic estrogen receptors (ERa and ERb), but they do not different lengths on its amine moiety increase its antagonist effect bind to estradiol or other related steroidal estrogens. The fasting- in MCF-7 cells. Furthermore replacement of chloroethyl by an induced cofactor PGC-1a has been reported as an endogenous aminoethyl group in ospemifene (7), a triaryl ethylene SERM, protein ligand for the ERRs [96e98]. ERRg is expressed in the spinal resulted in substantial improvement in its anti-breast cancer ac- cord and CNS, and it has been identified that the potent estrogen tivity [91e93]. In view of all of these factors, Kaur et al. (2016) receptor antagonist 4-hydroxytamoxifen acts as an inverse agonist designed and synthesized a new series of triarylethylenes by of ERRg [99]. With the aim of developing ERRg inverse agonist with introducing polar amino-/amidoethyl groups instead of chloro selectivity against the classical estrogen receptors, Chao et al. ethyl groups and replacing O-dimethylaminoethyl chains with (2006) selected 4-hydroxytamoxifen for further modifications and short O-methyl chains [94]. To further examine the effect of studies [100]. 4-hydroxytamoxifen analogues with extended ethyl structural modifications on tamoxifen, these compounds were side chain and polar functionality for ERRg interactions and unfa- evaluated for their anti-breast cancer activities against MCF-7 vorable interactions with ERa were designed and synthesized. The (ERþ) and MDA-MB-231 (ER-) cell lines. Two forms of compound binding affinities of these compounds toward ERRg and ERa sug- 22 (Fig. 5) (MCF-7 and MDA-MB-231 IC50 ¼ 16.9 and 11.4 mM, gested that analogues with hydroxyalkyl groups were more selec- respectively) and 23a (Fig. 5) (MCF-7 and MDA-MB-231; IC50 ¼ 12.1 tive in binding to ERRg when compared with that of 4- and 12.2 mM, respectively) showed promising activity against both hydroxytamoxifen. With analogue 24 (Fig. 5), a 25-fold improve- the cell lines whereas compound 23b (Fig. 5) was selectively active ment in binding selectivity for ERRg over ERa was observed (ERRg, only against MDA-MB-231 (MCF-7, IC50 50 mM; MDA-MB-231, IC50 ¼ 0.079 mM; ERa,IC50 ¼ 0.32 mM). IC50 ¼ 11.5 mM). Structure activity relationship studies suggested that the replacement of longer O-dimethylaminoethyl chains with 2.5. Tamoxifen derivatives with modification on phenyl rings, fi O-methyl chains did not have any signi cant effects on its activities, amino alkyl and ethyl chains whereas the introduction of an amino or oxalamido substitution on O-methyl analogues increased the potency in addition to making The literature indicates that the presence of tamoxifen's ami- þ these analogues more effective against both ER and ER-cell lines. noalkoxy group is essential for its receptor binding affinity, and Furthermore, western blotting and scratch assays were performed significant reductions in binding interactions were observed with a to evaluate two paraemters: 1.) the expression of proteins associ- decrease in protonated amino groups' basicity, whereas ethyl group ated with adhesion, migration and metastasis and 2.) migration of replacement with a methyl group in tamoxifen showed no change human breast cancer MDA-MB-231 (ER-) cells. In both studies, in antiproliferative activity [101e103]. These results motivated compound 22 was most effective and it also showed anti- Abdellatif et al. (2013) to design and synthesize tamoxifen ana- metastasis properties by suppressing wound healing and inva- logues by replacing the dimethyl amino moiety in tamoxifen with sion. Experimental toxicity against MCF-7 cell lines was supported secondary amines such as piperidino, piperazino, and/or N-meth- through in silico molecular docking studies. Representative com- ylpiperazino [104]. Additionally, the ethyl group was replaced by fi pounds 22 and 23a exhibited good binding af nity with ERs. methyl group and the para position of phenyl ring was substituted Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 521 with a fluorine atom. The antiproliferative activity of these com- analogues [112]. The fluorinated compounds' antiproliferative ac- pounds was examined using an MTT assay for ER-positive and ER- tivities were evaluated using four human tumor cell lines (HT-29 negative cell lines (MCF-7 and MDA-MB-231 cell lines respectively). colon carcinoma cells, M21 skin melanoma, MCF-7 estrogen- The most active compounds were 25a (Fig. 6) (MCF-7 and NDA-MB- dependent breast adenocarcinoma, and MDA-MB-231 estrogen- 231; IC50 ¼ 6.75 and 10.53 mM, respectively) and 25b (Fig. 6) (MCF-7 independent breast adenocarcinoma). Most of the compounds and MDA-MB-231; IC50 ¼ 5.58 and 13.04 mM, respectively), which showed similar or better activities than tamoxifen, and the most showed better activity compared to tamoxifen (MCF-7 and MDA- active compound in the MCF-7 cell line was compound 27 (Fig. 6) MB-231; IC50 ¼ 27.96 and 64.85 mM, respectively) in both the cell (GI50 ¼ 3.6 mM). The 4-hydroxytamoxifen is one of the main lines; however, the rest of the compounds showed activities similar metabolite of tamoxifen and is well known to have better activity to tamoxifen. Docking studies revealed that the synthesized com- than the parent compound [113,114]. Similarly, in this study, the 4- pounds have a low docking score energy with ERa. In addition, hydroxy derivatives were more potent than their parent com- compounds 25a and b (the most active ones) showed hydrogen pounds. The alkene geometry plays a very important role in bond interactions with the Asp-831 amino acid. Compounds 25a tamoxifen activity; subsequently Z-tamoxifen acts as an antagonist, and b were capable of inhibiting MDA-MB-231 proliferation and whereas E-tamoxifen behaves as an agonist. In contrast, this series indicated that the increased anticancer activities were ER- of fluorinated compound exhibited similar activities for both independent. Further tests were performed with 25a and b in or- geometrical isomers, and it was suggested that this behavior might der to explain their ability to induce ER-independent cell death. The be due to the compounds' in vitro isomerization. results revealed that both the compounds were highly capable of Furthermore, to identify the structural features of tamoxifen triggering classic caspase-dependent apoptosis and exhibited high that confer selective binding properties to the ER relative to other caspase 3/7 activities in MCF-7 treated cells compared to tamox- targets such as protein kinase C (PKC), extensive structure activity ifen. Structure activity relationship studies suggested that relationship studies have been done with the tamoxifen frame- replacement of tamoxifen's dimethyl amino group with more basic work. Tamoxifen's high affinity for estrogen and low affinity for PKC groups such as piperazino and N-methyl piperazino and introduc- compromise its utility to selectively target PKC for brain disorders. tion of a fluorine atom on the p-position of phenyl provided more Carpenter et al. (2016) used tamoxifen's triphenylethylene core as a active compounds than tamoxifen. framework to design and synthesize molecules with increased af- One of the common strategies to fine tune the physico-chemical finity for PKC and decreased affinity for the ERs [115]. Based on the properties of a bioactive compound is the introduction of fluorine previous study of Bigon et al., compound 28a (Fig. 6), bearing a atom and this led to the development of panomifene (26)(Fig. 6) diethyl amino side chain, was selected as the lead compound for [105e109]. Mainly to prevent the incidence of new cancers the 26 is further modification [116]. Compound 28a exhibited reasonable considered superior to tamoxifen [110,111]. Following the same PKC inhibition (12% at 3 mM) and lower ER binding affinity strategy Forest et al. (2013) replaced the ethyl substituent with a (IC50 ¼ > 10 000 nM) relative to tamoxifen (PKC inhibition ¼ 27% at fluorine atom and then synthesized fluorinated tamoxifen 3 mM and ER binding affinity, IC50 ¼ 222 nM). A series of novel triarylacrylonitrile analogues were synthesized and screened for PKC inhibition and ER binding affinity. The most potent compound X of the series was 28b, containing a more basic (4-methylpiperazin- H fi N 1-yl)ethoxy side chain, signi cantly inhibited PKC at a concentra- N N O O O OH tion of 3 mM (83%) and caused a decrease in ER binding affinity (IC50 ¼ >10 000 nM) compared to tamoxifen. Cytochrome P450 (CYP450), mainly CYP2D6 and CYP3A4 en- R zymes, are involved in tamoxifen metabolism and activation to the more active 4-hydroxy tamoxifen (8) and endoxifen (10). High F CF3 HO variations in tamoxifen's clinical outcomes can be observed as a 27 result of genetic polymorphisms in the CYP2D6 genes [117,118].In 25a; R=H,X=NCH3 26 25b; R=F,X=NH 2016, Ahmed et al. designed and synthesized tamoxifen analogues by retaining tamoxifen's pharmacophoric features and following O R1 possible metabolic pathways that do not involve the CYP2D6 N O R2 O C9H19 enzyme [119]. These analogues were evaluated for their anti- proliferative activity on MCF-7 breast cancer cell lines and binding affinity for ER-a and ER-b receptors. All of the compounds showed better antiproliferative activity than tamoxifen on MCF-7 cells. The most promising compound was 29 (Fig. 6), with a GI ¼ 0.005 mM R1 50 N CN N and a 1000 times more potency than tamoxifen (GI50 ¼ 1.58 mM). In O R2 O addition, after incubation of compound 29 in human liver micro- 29 28a; NR1R2 = N somes (HLM) and human hepatocytes (hHEP), the active hydroxyl metabolite was detected; this suggests other enzymes' involvement in its metabolism. 28b; NR1R2 = N N Combretastatin A-4 (CA-4), a natural product having an aryl- O ethylene moiety, is a potent tubulin assembly inhibitor O (IC50 ¼ 1.2 mM) in addition to possessing strong cytotoxic activity HO O against selected human cell lines such as DU-145 prostate cancer cells (GI50 ¼ 2 nM) [120,121]. In comparison to CA-4, tamoxifen has O no significant effects on tubulin polymerization (IC50 > 40 mM). 30 Consequently, Tanpure et al. (2009) designed and synthesized Fig. 6. Tamoxifen derivatives (25e30) with modification on phenyl rings, amino alkyl, tetra-substituted alkenes by combining structural and electronic and ethyl chain. components of tamoxifen and combretastatin A-4 using McMurry 522 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 coupling [122]. The compounds were evaluated for their inhibition the phenyl ring and ethylene group. Additionally, several analogues of tubulin polymerization and cell growth in selected human can- of tamoxifen bearing dimethylaminoethyloxy, pyrrolidinylethyloxy, cer cell lines. Altogether the compounds were less potent than CA- or piperidinylethyloxy side chains were synthesized, and the effects 4, and none of them were able to significantly inhibit tubulin as- of cyclization, size of the cyclic structure, and the effects of altering sembly. Of all the series, compound 30 (Fig. 6) was more cytotoxic the nitrogen's basicity have been studied. The tamoxifen analogues than tamoxifen against the three cell lines, especially against the were evaluated for their antiproliferative activity on MCF-7 cell human ovarian cancer cell line SK-OV-3 (GI50 ¼ 0.6 mM). Similar to lines and their binding affinities for ERa and ERb. All of the tamoxifen, compound 30 was not able to inhibit tubulin assembly tamoxifen analogues showed better antiproliferative activity than (IC50 > 40 mM); therefore, it was assumed that compound 30's tamoxifen, and the most active compound (32) contained pyrroli- cytotoxicity occurred via different mechanisms. dinylethyloxy side chain (Fig. 7) exhibiting IC50 < 0.25 mM. Com- pound 32 also exhibited 80-times more ERa binding than tamoxifen and 900 times more selectivity towards ERa than 2.6. Flexible tamoxifen derivatives tamoxifen. The mode of compound 32's binding with ERa was examined with a computational docking study using crystal Meegan and coworkers studied several flexible and non- structure obtained from the co-crystallization of ERa with 4-OH- isomerisable estrogen receptor modulator analogues of tamox- TAM [126]. As anticipated, the results showed that introduction of ifen, and the most potent lead compound was identified as 31 the additional methylene group allow the compound to easily (Fig. 7), which contained an additional methylene group positioned adopt the required arrangement for binding in an established between the aryl ring C and vinylic carbon and had an IC50 12.5 mM antiestrogenic mode. against MCF-7 cell line [123,124]. Furthermore to improve tamox- ifen analogues' antiproliferative and receptor binding activities, a 2.7. Tamoxifen derivatives with heteroaromatic groups second generation derivatives of compound 31 was synthesized by introducing a hydroxyl group on ring B [125]. The effects of halo- Considering the ER binding affinity and antiestrogenic activity of gens and other oxygen containing species (such as esters and car- heteroatom containing tamoxifen analogues [128,129], Wenckens bamates) on the phenyl ring in addition to the effects of different et al. (2003) designed and synthesized a series of tamoxifen ana- amino alkyl chains were examined. Most of the second generation logues by replacing the 1Z-alkoxyphenyl group by N-alkoxypyr- forms of compound 31 exhibited high antiproliferative activity azole and by introducing the functionalized phenyl or against the MCF-7 human breast cancer cell lines. The cytotoxic heteroaromatic groups at the 2Z-position of tamoxifen [130]. Also, a assessment results suggested that the compounds exhibited low few analogues of 4-hydroxytamoxifen were prepared in which the cytotoxicity, indicating their mode of action is cytostatic rather than 1E-4-hydroxyphenyl group was replaced with a 1-hydroxypyrazol- cytotoxic. The most active compound was 32 (Fig. 7), which showed 4-yl group. The binding affinity of the synthesized N-alkoxypyr- high ER binding affinity (IC50 ¼ 20 nM) with up to 12 fold ERa/b azole analogues to the estrogen receptor (ER ) were determined selectivity. The compound also displayed antiestrogenic effects at a and compared with tamoxifen. Substantial differences in binding 40 nM with little estrogenic stimulation when evaluated in the affinities to the ER were observed between the 40 and 50 pyrazolyl Ishikawa cell lines and also promoted apoptosis in MCF-7 cells in a a analogues. Most of the compounds of the 4'pyrazolyl series FACS based assay. The docking study of compound 32 was per- exhibited good binding to the ER and were comparable to formed using the crystal structure obtained from the cocrystalli- a tamoxifen (IC ¼ 0.1 mM). The compounds' cytostatic properties zation of ERa with 4-hydroxytamoxifen as found in the PDB 50 were examined in the MCF-7 cell lines and showed high affinity for database (PDB ID 3ERT) [126]. The results suggest that compound ER . The activity results suggest that replacement of the phenoxy 32 binds in an antiestrogenc manner with some modifications in its a group in tamoxifen with 1-pyrazoloxy group does not have signif- benzylic ring C orientation. icant effects on its antiestrogenic activity. Compound 33 (Fig. 8), The genetic polymorphisms in CYP2D6 are considered respon- which is a close tamoxifen analogue, showed growth inhibition sible for variation in tamoxifen's clinical outcomes. The poly- morphism may result in the formation of inactive proteins devoid of enzymatic activity or may lead to an enzyme with reduced ac- tivity. Bearing in mind the antiproliferative activity of flexible N N OH tamoxifen analogues, in 2016, Elghazawy et al. synthesized a series O O of flexible tamoxifen analogues to overcome the genetic poly- N N N N morphism of the CYP2D6 enzymes [127]. The site for metabolic para-hydroxylation was blocked by introducing either a hydroxyl or fl O ester group, and exibility to the rigid triphenylethylene backbone O was provided by introducing a benzylic methylene spacer between O 33 34 35a

N N N O O O O A C

O B O E/Z O S 35b 31 HO 32 36

Fig. 7. Flexible tamoxifen derivatives (31 and 32). Fig. 8. Tamoxifen derivatives with heteroaromatic groups (33e36). Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 523

(IC50 ¼ 3 mM) similar to tamoxifen (IC50 ¼ 2.7 mM) on MCF-7 cell ring of 4-hydroxytamoxifen with a ferrocenyl unit. The biological lines. Moreover, the introduction of substituents in the 2Z-or1Z- activity results of ferrocifens (37) demonstrated that these com- phenyl group or thienyl group does not display significant changes pounds had strong antiproliferative effects on both hormone- in the potency. The most potent compound of the series was dependent (ERþ) and -independent (ER-) breast cancer cells compound 34 (Fig. 8), which was unable to isomerize around the [139,140]. Motivated by these results in 2007, the group synthe- double bond and inhibited the growth of the MCF-7 cell line at sized compound 38 (Fig. 9), in which the amino alkyl chain of 4- IC50 ¼ 1.0 mM. hydroxytamoxifen was replaced by a stable and a lipophilic ferro- 5 Taking into consideration the abundance of the structural cenyl group eOCH2CO-[(h -C5H4)FeCp] [141]. The relative binding component quinone and dienone in several cytotoxic agents affinity test with estrogen receptors showed positive results for [131e134], Srivastava and his co-workers (2015) designed and these compounds. As expected, the (Z) isomer of 38 was well synthesized a series of constrained tamoxifen look alikes contain- recognized and showed good affinity for both the ERa and b es- ing the spirodienone moiety [135]. The series was synthesized us- trogen receptor isoforms (13.9% and 12.8%, respectively), and the ing iodine-catalyzed ipso-cyclization followed by Suzuki coupling. (E) isomer exhibited a drop in affinity with both isomers (ERa and b The molecular docking results of the series suggest that these [1.2% and 1.6%, respectively]). Next, to understand the binding af- compounds fit properly in the binding pocket and have similar finities the molecular modeling on (E)-38 and (Z)-38 on the anti- binding mode in ER binding site as that of the co-crystallized ligand estrogenic form of the estrogen receptor was investigated. The 4-hydroxytamoxifen. The compound's series was also evaluated study suggested that carbonylferrocenyl group substitution has in vitro against ERþ, MCF-7 and ER-, MDA-MB-231 breast cancer minimal effects on the interactions of ligand to the ERa, and the cell lines. Most of the compounds were active against MCF-7 cells [(Z)-38]-cavity complex was found to be twice as stable as that of with IC50 values < 6.5 mM. Compound 35a (Fig. 8), containing a 4- the [(E)-38]-cavity complex. The antiproliferative activity of both hydoxy substituent on the phenyl ring, showed maximum activity diastereomers was examined on hormone-dependent MCF-7 against both cell lines (MCF-7 and MDA-MB-231; IC50 ¼ 5.76 and breast cancer cells and hormone-independent PC-3 prostate cancer 8.85 mM, respectively), and compound 35b (Fig. 8), containing a 4- cells. Both diastereomers (E)-38 and (Z)-38 displayed similar anti- methoxy group on the phenyl ring, was most selective against MCF- proliferative activity, with an average of 10.4 mM on MCF-7 cells and 7 cells (MCF-7 and MDA-MB-231; IC50 ¼ 5.86 and 64.0 mM, 8.9 mM on PC-3 cells, which is not as strong as that of the ferrocifens respectively). 37 (n ¼ 3; IC50 ¼ 0.5 mM on MCF-7). It was anticipated that the Pharmaceutical limitations associated with tamoxifen, existing antiproliferative activity of 38 could occur either through an anti- as two geometrical stereoisomers with opposite actions, leads to hormonal mechanism or was due to the cytotoxic character of the the development of nonisomerizable antiestrogens such as nafox- ferrocenyl group. idine and ring-fused analogues such as benzocycloheptene, ben- Subsequently, a series of cobaltifens, organometallic analogues zoxepine, and benzothiepines [136e137]. The problem of of tamoxifen in which a phenyl ring has been replaced by an isomerization was solved with the discovery of these non- organo-cobalt sandwich moiety, was synthesized [142]. The pres- isomerizable compounds; however, these compounds were not ence of the organo-cobalt sandwich supports multiple functional- potent for antiproliferative or antiestrogenic activity. Consequently, izations and allows modification of its electronic characters, redox Ansari et al. (2015) designed and synthesized a novel series of properties, and steric parameters [143]. The compounds were substituted dibenzo[b,f]thiepines and dibenzo[b,f]oxepines as anti- screened for their ERa binding affinity and antiproliferative activ- breast cancer agents [138]. The compounds were tamoxifen ana- ities against hormone-dependent (MCF-7) and hormone- logues and contained planar tricyclic cores with pendant phenyl independent (MDA-MB-231) breast cancer cells. The ERa binding rings as attachments. The desired antiestrogenic conformation was affinities for cobaltifens were quite low (below 1%) and significantly achieved by the presence of basic tert-aminoalkoxy group on the lower than ferrocenyl derivatives (RBA 10e15%). It was suggested phenyl ring with perpendicular orientation. The synthesized com- that this could be because of the size of the organometallic cobalt pounds were evaluated on ERþ and ER-breast cancer cell lines and units with four phenyl groups which is significantly more bulky the results showed that most of the compounds possessed prom- than the unsubstituted cyclopentadienyl ring of ferrocene. Cellular ising in vitro antiproliferative activity. The dibenzo[b.f]thiepine proliferation results revealed that the dihydroxycobaltifens showed analogues with the sulfur atom were more active as anti-breast estrogenic actvity against MCF-7 cells but not on MDA-MB- cancer agents compared to oxygen containing dibenzo[b,f]oxefine 231 cells, whereas, the aminoalkyl-hydroxycobaltifens were analogues. The most active compound of the series was 36 (Fig. 8) weakly cytotoxic toward both cell lines. Surprisingly the bis- that inhibited both the breast cancer cell lines in the micromolar (dimethylamino-ethoxy)cobaltifens such as 39a and b (Fig. 9) range (MCF-7 and MDA-MB-231; IC50 ¼ 1.33 and 5 mM, respec- exhibited high cytotoxicity toward both cell lines, specifically MDA- tively). Also, it lacked any cytotoxic effects at 50 mM on the normal MB-231 cells (IC50 ¼ 3.8 and 2.5 mM, respectively). human embryonic kidney (HEK-293) cells, which was much better Later, considering the significance of ferrocenyl derivatives of than tamoxifen at 18 mM. Further analysis of compound 36 on cell hydroxytamoxifen for antiproliferative activity against both cycle distribution and apoptosis of MCF-7 cells followed by an LDH estrogen-responsive and estrogen-refractory breast cancer cells release assay suggested that 36 inhibited cellular proliferation via [144,145], a series of tetrasubstituted olefins bearing a ferrocenyl G0/G1 arrest in MCF-7 cells and was primarily due to apoptosis, not group were designed and synthesized [146]. Earlier reports indi- necrosis. As anticipated, molecular docking studies showed better cated the antiproliferative studies were done only on breast and binding of compound 36 with estrogen receptors in comparison to prostate cancer cell lines; hence to obtain a broader view, using the 4-hydroxytamoxifen binding to the same receptors. MTT test, the synthesized compounds were evaluated on four tu- mor types, including SF-295 (human glioblastoma), HCT-8 (human 2.8. Tamoxifen conjugates colon cancer), MDA-MB-435 (human melanoma), and HL-60 (hu- man promyelocytic leukemia). More than one third of the com- Introduction of the organometallic substituent on the tamoxifen pounds exhibited IC50 values <2 mM for one or more cell lines. The skeleton and its effects on cytotoxicity were first studied by the Top most encouraging compound of the series was 40 (Fig. 9), which and Jaouen group. Initially, some ferrocenyl derivatives of hydrox- was a 2-ferrocenyl-1.1-diphenyl-but-1-ene that showed an inter- ytamoxifen (37)(Fig. 9) were prepared by substituting the b-phenyl esting combination of growth inhibition and low hemolytic activity 524 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

OH OH O

O Fe

Fe

O n HO O HO OH N (E)-38 37 (Z)-38 O n= 2, 3, 4, 5 Fe

N O N SH O O 7 Alk = AuNP Fe Ph Co Ph O 40 N 41 TAM-PEG-SH Ph Ph

39a; Alk =CH3 39b; Alk = CH2CH3 N O O O 7 O

=

42

N O S O O O N O 43 F2HB N

O O

O n O N HN

HN N O

44a;n=2 44b;n=4 OR O

HN NH O H H H N O S O RO

45a;R=CH2CH2 N

45b;R=CH2CH2N(CH3)2

Fig. 9. Tamoxifen conjugates (37e45). Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 525 and was then selected for further examinations. This compound uptake into the cancerous cell in comparison to healthy cells. exhibited promising activities on SF-295 (IC50 ¼ 1.0 mM), HCT-8 Following this approach Hawco et al. (2013) planned to conjugate (IC50 ¼ 0.9 mM), and HL-60 (IC50 ¼ 1.04 mM) cell lines and moder- prodigiosenes [164,165], a class of tripyrrolic compound with sig- ate activity on the MDA-MB-231 breast cancer cell line nificant anticancer activity, to tumor selective ligands in order to (IC50 ¼ 16 mM). deliver a drug to a specific site. Several ER ligands such as tamoxifen Selective targeting and delivery of gold nanoparticles func- [166], fulvestrant [167], and hematoporphyrin [168] were selected tionalized with ligands of cell surface receptors overexpressed by to target ER positive breast cancers. The tripyrrolic skeleton was malignant cells has been well documented [147e150]. Moreover, attached to the selected ligands via ester linkages with several ER isoforms are located both intracellularly and on the cell mem- hydrocarbon chain lengths [169]. The synthesized conjugates were brane [151,152]. After considering these facts, Dreaden et al. (2009) screened for their in vitro biological activities on different cancer synthesized gold nanoparticle tamoxifen analogues as selective and cell lines. The results revealed that the tamoxifen conjugates 44a potent agents for breast cancer treatment [153]. The thiol- and b (Fig. 9) were the most potent against MCF-7 cells with poly(ethylene glycol) tamoxifen (PEG-SH-TAM) derivative was GI50 ¼ 30 and 50 nM, respectively. The porphyrin conjugate was synthesized and subsequently conjugated with gold nanoparticle inactive, whereas the estrone conjugate exhibited moderate activ- (AuNP). Tamoxifenegold nanoparticle conjugate (TAM-PEG-SH- ity on both ER negative and positive cell lines. The structure activity AuNP; 41)(Fig. 9) exhibited drug potency 2.7 times greater than relationship studies suggest that conjugates with shorter chain free tamoxifen; this was due to the conjugate's selective intracel- length conjugates, which are linked with 2 and 4 carbon atoms, are lular delivery to ER(þ) breast cancer cells, caused by both receptor- more active than the conjugates with a longer chain length (linker and ligand-dependence in vitro. These results suggest that plasma with 8 carbon atoms). membrane-localized ERa may facilitate selective uptake and Ridaifen B (RID-B) 12a, a tamoxifen derivative, has been shown retention of this and other therapeutic nanoparticle conjugates. to be more active on cell proliferation compared to tamoxifen. RID- Following a similar synthetic approach Nelson et al. (2011) syn- B is equally active on ERþ and ER-cell lines; its mode of action is thesized single walled carbon nano tube (SWCNT) tamoxifen con- different from the existing cancer drugs, including tamoxifen, jugates (42)(Fig. 9) and characterized them by using different suggesting an ER-independent mode of action [170].In2013, analytical techniques, including NMR [154]. It was anticipated that Sugawara and his co-workers investigated RID-B binding proteins the conjugate comprising both SWCNT and tamoxifen linked by via a T7 phage display screen and binding analysis with synthesized octa(ethyleneglycol) (OEG) could be used in breast cancer treat- biotinylated RID-B derivative (Bio-RID-B) 45a (Fig. 9) [171]. The ment due to the advantages of SWCNTs in drug delivery systems study identified Grb 10 interacting GYF protein 2 (GIGYF2) as an and photothermal therapy in addition to tamoxifen's recognition RID-B binding protein, which is involved in the PI3K/Akt signaling properties as a selective targeting agent and potent endocrine pathway and is up-regulated in breast cancer cells in which Akt is treatment drug. highly phosphorylated. RID-B binds directly to GIGYF2 and reduces One of the important strategies for understanding the mecha- Akt's phosphorylation level, suggesting an ER-independent anti- nisms of estrogen signaling and the specific physiological responses cancer activity for RID-B's mechanism of action. Subsequently, a associated with this signaling is to develop new tamoxifen-based biotinylated RID-G derivative (Bio-RID-G) 45b (Fig. 9) was synthe- chemical probes. Only a few examples of fluorescent tamoxifen sized and a chemical genetic approach was used to identify the probes have been reported in the literature [155,156]. The probes target proteins of RID-G, another tamoxifen analogue containing a having direct attachment of dyes to tamoxifen or metabolite were dimethyl amino alkyl chain on two phenyl rings and having high having good binding affinity and selectivity nevertheless they were growth inhibitory activity against various cancer cell lines [172]. not helpful to solve the contentious membrane receptor problems. Using this approach, a phage display screen was combined with a The tamoxifen probe developed specifically for membrane associ- statistical analysis using drug potency and gene expression profiles ated ER studies also found to have poor affinity and loss of speci- in thirty nine cancer cell lines. This approach assisted in under- ficity compared to parent compound. These results inspired Ho standing the physiological relevance of the direct association [173]. et al. (2016) to design and synthesize new selective tamoxifen- The study identified three proteins, calmodulin (CaM), heteroge- based fluorescent probes [157]. Tamoxifen was selected (instead neous nuclear ribonucleoproteins A2/B1 (hnRNP A2/B1), and zinc of its metabolite 4-hydroxy tamoxifen) to tether the fluorophore finger protein 638 (ZNF638) as targets of RID-G as part of its growth since it is commercially available and tamoxifen-based probes are inhibitory activity. known to have sufficient binding compared to the metabolite- Tamoxifen and its main metabolite (4-hydroxytamoxifen) are based probes. Dye attachment was done on the basic alkylami- known to form adducts with DNA, and in animal models hepatic noethoxy side chain with a short ethylene glycol linker. The BOD- toxicity has been observed in the presence of these two agents ® IPY FL fluorophore was selected for the study due to its well- [174]. In 2014, Tajmir-Riahi and coworkers used chemical and characterized cell permeability and unusual cytoplasmic localiza- molecular modeling approaches to examine the binding affinities tion [158]. The cellular localization of the synthesized fluorescent of tamoxifen (1) and its metabolites for DNA with the aim of ® BODIPY FL ethylene glycol-linked tamoxifen conjugate (43)(Fig. 9) establishing the tamoxifen and its metabolites' mechanism of was visualized by fluorescent confocal microscopy in ER-positive binding to DNA [175]. The results suggested that tamoxifen and its MCF-7 and ER-negative MDA 231 breast cell lines. Results metabolites bind to DNA via both hydrophobic and hydrophilic ® revealed that the BODIPY FL conjugate (43) was internalized in the interactions. In the DNA duplex, tamoxifen and its metabolites ER-positive cell via a receptor-mediated mechanism of uptake; showed different binding sites, and the order of binding was 4- however, no internalization of 43 was observed in ER-negative hydroxytamoxifen (8) > tamoxifen (1) > endoxifen (10), indi- cells. Additional increases in concentrations of 43 exhibited no cating the formation of a more stable complex with 4- change in the degree of uptake or localization, and also no locali- hydroxytamoxifen. Next, considering the fact that loading of zation of 43 in the cytoplasm was observed. tamoxifen and its metabolites with serum protein increases the In the literature, several reports have described the significance solubility of the drug, improves its tissue-specific targeting, and of the targeted drug design approach to be used to overcome multi facilitates constant release of the drug, a comparative study on the drug resistance [159e163]. An anticancer agent can be attached to a binding affinity of serum proteins with tamoxifen and its metab- compound known to accumulate in cancer cells, which increases its olites was performed [176]. In this study, both human and bovine 526 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 serum albumin (HSA and BSA, respectively) were used, and the modulatory activities. Norendoxifen, a human metabolite of results of multiple spectroscopic methods and docking studies tamoxifen with high potency and selectivity as an AI [194],was were considered. The results revealed that tamoxifen and its me- selected as a lead compound for further development. It was ex- tabolites bind serum proteins through hydrophobic, hydrophilic, pected that close structural similarity of nornedoxifen with and/or H-bonding interactions and that HAS conjugates were more tamoxifen would preserve the dual aromatase inhibitory and ER stable than BSA conjugates. 4-hydroxytamoxifen forms a stronger modulatory activities. The aromatase inhibitory activity should bond than tamoxifen and endoxifen. Additionally, major drug inhibit tumor growth by blocking estrogen biosynthesis in the conjugation-induced perturbations in serum protein conformation breast, and its ER modulatory activity should reduce the side effects were observed, and 4-hydroxytamoxifen formed a stronger bond in bones and other tissues caused by estrogen reduction. Initially than tamoxifen and endoxifen. Later, encouraged by polyamido- (E)-norendoxifen (46a), (Z)-norendoxifen (46b), and (E, Z)-nor- amine (PAMAM) significance as drug delivery tools, either through endoxifen isomers (46c)(Fig. 10), were synthesized and their aro- physical interactions or chemical bonding, the loading efficacies of matase inhibitory and estrogen receptor affinities were determined antitumor drugs (doxorubicin and tamoxifen) with PAMAM-G4 [195]. Compound 46c displayed strong aromatase inhibitory ac- were reviewed, and correlations between drug interactions and tivity with IC50 ¼ 102 nM and good binding affinity to both ERa and polymer morphology were established [177]. The results indicated b with EC50 ¼ 27 nM and 35 nM, respectively. It was observed that that hydrophilic, hydrophobic, and H-bonding contacts were 46a was a more potent aromatase inhibitor (IC50 ¼ 76.8 nM) than responsible for drug-polymer conjugation and that doxorubicin 46b (IC50 ¼ 1029 nM). In contrast, 46b displayed a higher binding formed more stable conjugates with PAMAM-G4 than did tamox- affinity toward ERa (EC50 ¼ 17 nM) and b (EC50 ¼ 28 nM) than di ifen. The drug loading efficacy was 40%e50%, and PAMAM nano- 46a (EC50 ¼ 59 nM and 79 nM, respectively). particles were observed to be efficient for both drugs' in vitro Next, a series of structurally related norendoxifen analogues transport. were designed by molecular modeling using a structure-based drug design approach. The designed analogues were synthesized and 2.9. Derivatives of tamoxifen metabolites then pharmacologically evaluated with a view to optimizing the efficacy against both aromatase and ER, improve the aromatase PKC is an important enzyme in cell signaling pathways. It is selectivity versus other cytochrome P450 enzymes, and to further involved in numerous brain diseases such as Parkinson's, Alz- explore the structure activity relationships [196]. Most of the ana- heimers, and bipolar diseases (BPD) in addition to substance abuse logues of the series exhibited promising aromatase inhibitory ac- disorders [178e181]. PKC is considered a promising therapeutic tivities and ER binding affinities. The most potent compound was target for these diseases; however in vivo validation has been 40-hydroxynorendoxifen (47)(Fig. 10) with high aromatase inhibi- difficult due to PKC inhibitor impermeability to the central nervous tory potency (IC50 ¼ 45 nM) and ER binding affinity (ERa and b: system [182,183]. Tamoxifen is the only PKC inhibitor that can EC50 ¼ 15 and 9.5 nM, respectively). In comparison to norendoxifen, penetrate the blood brain barrier and inhibit cellular PKC activity analogue 47 was a more potent antagonist than norendoxifen of [184]. Preclinical and clinical studies with the relatively selective estradiol-stimulated progesterone receptor mRNA expression in PKC inhibitor Z-tamoxifen support the significance of PKC as a MCF-7 cells. Also, selectivity of 47 for aromatase versus other cy- target for treatment of BPD [185]. BPD is a chronic, debilitating tochrome P450 enzyme was much better than norendoxifen (10). illness characterized by drastic swings in mood, energy, and func- Furthermore, in solution, E/Z isomerization was observed for tional ability, and it mostly affects the adult population [186]. most of the norendoxifen analogues that were similar to 4- Conversely, capriciousness was observed in tamoxifen bioavail- hydroxytamoxifen and was possibly due to the presence of a ability and function due to CYP2D6 genetic polymorphism, which phenolic hydroxyl group in one of the para positions [197]. An in- extensively metabolizes tamoxifen into its active metabolites, 4- crease in the number of para phenolic hydroxyl groups increased hydroxytamoxifen (8) and endoxifen (10). In 2010, Ali et al. syn- the isomerization rate and was also dependent on the temperature thesized endoxifen, an active metabolite of tamoxifen, studied its and solvent [198]. E/Z isomerization can affect the synthesis of pure PKC inhibitory activity in vitro, and then compared it with the E and Z norendoxifen isomers in addition to influencing the accu- known PKC inhibitor, tamoxifen [187]. In comparison to tamoxifen, racy of biological tests for pure E and Z isomers. Therefore, to endoxifen significantly inhibited PKC in a concentration-dependent overcome the problems associated with E/Z isomerization, a series manner and was found to be four fold more potent. At the con- of triphenylethylene bisphenol analogues was designed and syn- centration of 0.2 mM, endoxifen exhibited 78% PKC inhibition, thesized for use as dual AI/SERM agents [199]. In these analogues, whereas the tamoxifen showed only 25%. the possibility of E/Z isomerization was solved by eliminating nor- In postmenopausal women, third generation aromatase in- endoxifen's aminoethoxy side chain. The hydrogen bond donor hibitors (AIs) such as anastrozole, exemestane, and letrozole have groups (such as hydroxyl or amino groups) were introduced at the mainly replaced tamoxifen as the preferred treatment for hormone meta or para positions of the phenyl ring as a substitute to the receptor-positive breast cancer. Clinical studies have suggested that aminoethoxy side chain. In addition, iron-coordinating groups AIs are much more superior and effective than tamoxifen in the (such as nitrile, imidazole, or triazole groups) were used as sub- treatment of ER-positive breast cancer in postmenopausal women, stitutes for the ethyl group. The synthesized compounds were and these drugs have demonstrated enhanced safety profiles, evaluated for their aromatase inhibitory activities, ER-a and -b tolerability, and disease free survival rates [188e190]. On the other binding affinities, and abilities to antagonize b-estradiol-stimulated hand, the use of AIs as anti-breast cancer drugs also involves transcriptional activity in MCF-7 human breast cancer cells. The various side effects such as reduction of bone density, severe biological activity results suggest that replacement of the ethyl musculoskeletal pain, and increased frequency of cardiovascular group with an imidazole group was favorable for aromatase and thromboembolic events due to the overall estrogen reduction inhibitory activity and ER binding affinities. The most potent [191e193]. Continuous efforts are being made by researchers compound of the synthesized series was the imidazole-containing worldwide to develop novel AIs with novel mechanisms that can compound 48 (Fig. 10). Compound 48 displayed a very high aro- cause fewer side effects. Incidentally, Cushman and coworkers matase inhibitory activity (IC50 ¼ 4.77 nM) in addition to ER (2013) thought to develop new breast cancer chemotherapeutic binding affinities (ERa and b;EC50 ¼ 27 nM and 41 nM, agents with dual aromatase inhibitory and estrogen receptor respectively). Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 527

H N NH H2N 2 HO O 2 O OH O OH E-46a Z-46b EZ-46c

N HO O2N N A

B C NH2 HO O HO OH H2N NH2 48 49 47

Fig. 10. Derivatives of tamoxifen metabolites (46e49).

Later, to optimize the aromatase inhibition and ER binding af- dynasty through perseverance and not by sudden action” (14th finity of norendoxifen analogues the structural features of a third Century Arab Historian Ibn Khaldun) and “No advances occur in generation AIs letrozole [200], which was a symmetrically isolation; they build on the work of previous generations and by substituted diphenylmethane fragment, was taken into consider- collegial interaction” (Father of Tamoxifen, V C Jordan). ation. The new series of norendoxifen (10) analogues were syn- thesized by eliminating the aminoethoxyl side chain, substituting Acknowledgements the para position of ‘A’ ring with a nitro/amino group and para position of ‘B’ and ‘C’ ring with hydroxyl or amino group [201]. The The authors, Dr. Shagufta and Dr. Irshad Ahmad are thankful to resulting analogues were devoid of geometrical isomerization and the School of Graduate Studies and Research, American University possessed hydrogen bond acceptors similar to letrozole. The bio- of Ras Al Khaimah for their support. logical activity results exhibited that the analogues bearing para- ‘ ’ amino group in the A ring of triphenylethylene scaffold was potent References for aromatase inhibitor activity. The most promising compound of the series was 49 (Fig. 10) with significant aromatase inhibitory [1] V.C. Jordan, Tamoxifen: a most unlikely pioneering medicine, Nat. Rev. Drug Discov. 2 (2003) 205e213. activity ((IC50 ¼ 62.2 nM) and binding activity to both ERa and b ¼ [2] B. Fisher, J.P. Costantino, D.L. Wickerham, R.S. Cecchini, W.M. Cronin, (EC50 72.1 and 70.8 nM, respectively). The structure activity A. Robidoux, T.B. Bevers, M.T. Kavanah, J.N. Atkins, R.G. Margolese, relationship studies suggest that the presence of amino groups in C.D. Runowicz, J.M. James, L.G. Ford, N. Wolmark, Tamoxifen for the pre- the para position of ‘A’ and ‘B’ was favorable for aromatase inhibi- vention of breast cancer: current status of the national surgical adjuvant breast and bowel project P-1 study, J. Natl. Cancer Inst. 97 (2005) tory activity, whereas replacement of ethyl side chain with a methyl 1652e1662. group had detrimental effects on aromatase inhibition and ER [3] K. Dhingra, Antiestrogensetamoxifen, SERMs and beyond, Invest New Drugs binding affiniy. Replacement of the unsymmetrical diphenyl- 17 (3) (1999) 285e311. methylene substructure of norendoxifen with symmetrical diphe- [4] D.J. Grainger, J.C. Metcalfe, Tamoxifen: teaching an old drug new tricks? Nat. Med. 2 (1996) 381e385. nylmethylene substructure preserved the activity and eliminated [5] V.C. Jordan, The development of tamoxifen for breast cancer therapy: a the triphenylethylene's E/Z isomerization. Additionally, in contrast tribute to the late Arthur L, Walpole. Breast Cancer Res. Treat. 11 (3) (1998) e to previous reports [202,203], it was observed that the amino- 197 209. fi [6] V.C. Jordan, S. Koerner, Tamoxifen (ICI 46, 474) and the human carcinoma 8S ethoxyl side chain in triphenylalkene derivatives was insigni cant oestrogen receptor, Eur. J. Cancer 11 (1975) 205e206. for both aromatase inhibitory and ER binding affinity. [7] V.C. Jordan, Effect of tamoxifen (ICI 46, 474) on initiation and growth of DMBA-induced rat mammary carcino, Eur. J. Cancer 12 (1976) 419e424. [8] V.C. Jordan, K.E. Allen, Evaluation of the antitumour activity of the non- 3. Conclusion steroidal antioestrogen monohydroxytamoxifen in the DMBA-induced rat mammary carcinoma model, Eur. J. Cancer 16 (1980) 239e251. In conclusion, we can certainly mention that tamoxifen is one of [9] V.C. Jordan, Tamoxifen (ICI 46, 474) as a targeted therapy to treat and pre- vent breast cancer, Br. J. Pharmacol. 147 (2006) S269eS276. the important drug templates explored by researchers worldwide [10] V.C. Jordan, Tamoxifen the first targeted long-term adjuvant therapy for for cancer and other therapeutic targets. Tamoxifen has immensely breast cancer, Endocrine-Related cancer 21 (2014) R235eR246. decreased the death rates from breast cancer around the world, and [11] V.C. Jordan, Tamoxifen: catalyst for the change to targeted therapy, Eur. J. Cancer 44 (2008) 30e38. patients are living longer, recurrence-free lives with less morbidity. [12] EBCTCG, Tamoxifen for early breast cancer: an overview of the randomized Since the discovery of tamoxifen in the 1970s as a failed contra- trials, Lancet 354 (1998) 1451e1467. ceptive to the promising breast cancer drug, the research on the [13] EBCTCG, Effect of chemotherapy and hormonal therapy for early breast tamoxifen template is still ongoing. It has been well-documented cancer on recurrence and 15-years survival: an overview of the randomized trials, Lancet 365 (2005) 1687e1717. that tamoxifen tremendously supports the discovery of several [14] L. Hughes-Davies, C. Caldas, G.C. Wishart, Tamoxifen: the drug that came in significant compounds with promising biological activities, and in from the cold, Br. J. Cancer 101 (2009) 875e878. the future, discovery of more promising compounds is expected. [15] T.G. Hayes, Pharmacologic treatment of male breast cancer, Exp. Opin. Pharmacother. 10 (2009) 2499e2510. This review article is an attempt to make available researchers the [16] B. Fisher, J. Dignam, N. Wolmark, D.L. Wickerham, E.R. Fisher, E. Mamounas, thorough progressions made in the last few years in the tamoxifen R. Smith, M. Begovic, N.V. Dimitrov, R.G. Margolese, C.G. Kardinal, research domain and it's our firm believe that it will support M.T. Kavanah, L. Fehrenbacher, R.H. Oishi, Tamoxifen in treatment of intra- ductal breast cancer: national Surgical Adjuvant Breast and Bowel Project B- readers with their ongoing research. In the end, we would like to 24 randomised controlled trial, Lancet 353 (9169) (1999) 1993e2000. conclude with these lines: “A new dynasty gives over the ruling [17] J. Cuzick, T. Powles, U. Veronesi, J. Forbes, R. Edwards, S. Ashley, P. Boyle, 528 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

Overview of the main outcomes in breast-cancer prevention trials, Lancet Pharmacol. Ther. 25 (2) (1984) 127e205. 361 (2003) 296e300. [45] B.S. Katzenellenbogen, M.J. Norman, R.L. Eckert, S.W. Peltz, W.F. Mangel, [18] V.C. Jordan, Chemosuppression of breast cancer with tamoxifen: laboratory Bioactivities, estrogen receptor interactions, and plasminogen activator- evidence and future clinical investigations, Cancer Invest 6 (5) (1998) inducing activities of tamoxifen and hydroxy-tamoxifen isomers in MCF-7 589e595. human breast cancer cells, Cancer Res. 44 (1) (1984) 112e119. [19] V.C. Jordan, Selective estrogen receptor modulation: a personal perspective, [46] J.N. Beverage, T.M. Sissung, A.M. Sion, R. Danesi, W.D. Figg, CYP2D6 poly- Cancer Res. 61 (2001) 5683e5687. morphisms and the impact on tamoxifen therapy, J. Pharm. Sci. 96 (9) (2007) [20] V.C. Jordan, B.M. O'Malley, Selective estrogen receptor modulators and 2224e2231. antihormonal resistance in breast cancer, J. Clin. Oncol. 25 (36) (2007) [47] W. Schroth, M.P. Goetz, U. Hamann, P.A. Fasching, M. Schmidt, S. Winter, 5815e5824. P. Fritz, W. Simon, V.J. Suman, M.M. Ames, S.L. Safgren, M.J. Kuffel, [21] Shagufta, A.K. Srivastava, R. Sharma, R. Mishra, A.K. Balapure, P.S.R. Murthy, H.U. Ulmer, J. Bolander,€ R. Strick, M.W. Beckmann, H. Koelbl, G. Panda, Substituted phenanthrenes with basic amino side chains: a new R.M. Weinshilboum, J.N. Ingle, M. Eichelbaum, M. Schwab, H. Brauch, Asso- series of anti-breast cancer agents, Bioorg. Med. Chem. 14 (2006) ciation between CYP2D6 polymorphisms and outcomes among women with 1497e1505. early stage breast cancer treated with tamoxifen, JAMA 302 (13) (2009) [22] I. Ahmad, Shagufta. Recent developments in steroidal and nonsteroidal 1429e1436. aromatase inhibitors for the chemoprevention of estrogen-dependent breast [48] V.O. Dezentje, H.J. Guchelaar, J.W.R. Nortier, C.J.H. van de Velde, cancer, Eur. J. Med. Chem. 102 (2015) 375e386. H. Gelderblom, Clinical implications of CYP2D6 genotyping in tamoxifen [23] W.C. Park, V.C. Jordan, Selective estrogen receptor modulators (SERMS) and treatment for breast cancer, Clin. Cancer Res. 15 (1) (2009) 15e21. their roles in breast cancer prevention, Trends Mol. Med. 8 (2) (2002) 82e88. [49] F. Marin, M.C. Barbancho, Clinical pharmacology of selective estrogen re- [24] S.R. Cummings, S. Eckert, K.A. Krueger, et al., The effect of raloxifene on risk ceptor modulators (SERMs), in: A. Cano, J. Calaf, I. Alsina, J.L. Duenas-Diez~ of breast cancer in postmenopausal women: results from the MORE ran- (Eds.), Selective Estrogen Receptor Modulators. New Brand of Multitarget domized trial, JAMA 281 (1999) 2189e2197. Drugs vol. 2006, Springer, Berlin Heidelberg New York, 2006, pp. 49e69. [25] V.C. Jordan, Optimising endocrine approaches for the chemoprevention of [50] A.Z. Steiner, M. Terplan, R.J. Paulson, Comparison of tamoxifen and clomi- breast cancer. Beyond the study of Tamoxifen and Raloxifene (STAR) trial, phene citrate for ovulation induction: a meta-analysis, Hum. Reprod. 20 (6) Eur. J. Cancer 42 (2006) 2909e2913. (2005) 1511e1515. [26] V.G. Vogel, J.P. Costantino, D.L. Wickerham, et al., Effects of tamoxifen vs [51] E.F. van Bommel, T.R. Hendriksz, A.W. Huiskes, A.G. Zeegers, Brief commu- raloxifene on the risk of developing invasive breast cancer and other disease nication: tamoxifen therapy for nonmalignant retroperitoneal fibrosis, Ann. outcomes: the NSABP Study of Tamoxifen and Raloxifene (STAR) P-2 trial, Intern. Med. 144 (2) (2006) 101e106. JAMA 295 (23) (2006) 2727e2741. [52] S. Akram, D.S. Pardi, J.A. Schaffner, T.C. Smyrk, Sclerosing mesenteritis: [27] V.C. Jordan, Tamoxifen or Raloxifene for breast cancer chemoprevention: a clinical features, treatment, and outcome in ninety-two patients, Clin. tale of two choices, Cancer Epidemiol. Biomar Prev. 16 (2007) 2207e2209. Gastroenterology Hepatology 5 (5) (2007) 589e596. [28] C.J. Fabian, B.F. Kimler, Chemoprevention for high risk women: tamoxifen [53] S. Turken, E. Siris, D. Seldin, E. Flaster, G. Hyman, R. Lindsay, Effects of and beyond, Breast J. 7 (5) (2001) 311e320. tamoxifen on spinal bone density in women with breast cancer, J. Natl. [29] S. Mikelman, N. Mardirossian, M.E. Gnegy, Tamoxifen and amphetamine Cancer Inst. 81 (1989) 1086e1088. abuse: are there therapeutic possibilities, J. Chem. Neuroanat. 83e84 (2017) [54] R.R. Love, R.B. Mazess, H.S. Barden, S. Epstein, P.A. Newcomb, V.C. Jordan, 50e58. P.P. Carbone, D.L. DeMets, Effects of tamoxifen on bone mineral density in [30] C. Davies, H. Pan, J. Godwin, R. Gray, R. Arriagada, V. Raina, M. Abraham, postmenopausal women with breast cancer, N. Engl. J. Med. 326 (1992) V.H. Medeiros Alencar, A. Badran, K. Bonfill, Long-term effects of con- 852e856. tinuining adjuvant tamoxifen to 10 years versus stopping at 5 years after [55] C.C. McDonald, F.E. Alexander, B.W. Whyte, A.P. Forrest, H.J. Stewart, Cardiac diagnosis of oestrogen receptor-positive breast cancer: ATLAS, a randomized and vascular morbidity in women receiving adjuvant tamoxifen for breast trial, Lancet 381 (2013) 805e816. cancer in a randomised trial. The Scottish Cancer Trials Breast Group, BMJ [31] L. Gennari, D. Merlotti, R. Nuti, Selective estrogen receptor modulator (SERM) 311 (1995) 977e980. for the treatment of osteoporosis in postmenopausal women: focus on [56] N. Dabelic, T. Jukic, Z. Labar, S.A. Novosel, N. Matesa, Z. Kusic, Riedel's lasofoxifen, Clin. Interv. Aging 5 (2010) 19e29. thyroiditis treated with tamoxifen, Croat. Med. J. 44 (2) (2003) 239e241. [32] R.J. Steffan, E. Matelan, M.A. Ashwell, et al., Synthesis and activity of [57] A. Yildiz, S. Guleryuz, D.P. Ankerst, D. Ongür, P.F. Renshaw, Protein kinase C substituted 4-(indazol-3-yl)phenols as pathway-selective estrogen receptor inhibition in the treatment of mania: a double-blind, placebo-controlled trial ligands useful in the treatment of rheumatoid arthritis, J. Med. Chem. 47 (26) of tamoxifen, Archives General Psychiatry 65 (3) (2008) 255e263. (2004) 6435e6438. [58] E.A. Eugster, S.D. Rubin, E.O. Reiter, P. Plourde, H.C. Jou, O.H. Pescovitz, [33] C. Jochems, U. Islander, A. Kallkopf, M. Lagerquist, C. Ohlsson, H. Carlsten, Tamoxifen treatment for precocious puberty in McCune-Albright syndrome: Role of raloxifene as a potent inhibitor of experimental postmenopausal a multicenter trial, J. Pediatr. 143 (1) (2003) 60e66. polyarthritis and osteoporosis, Arthritis & Rheumatism 56 (10) (2007) [59] K. Dolan, S. Montgomery, B. Buchheit, L. DiDone, M. Wellington, D.J. Krysan, 3261e3270. Antifungal activity of tamoxifen: in vitro and in vivo activities and mecha- [34] Y.-P. Xiao, F.-M. Tian, M.-W. Dai, W.-Y. Wang, L.-T. Shao, L. Zhang, Are nistic characterization, Antimicrob. Agents Chemother. 53 (8) (2009) estrogen-related drugs new alternatives for the management of osteoar- 3337e3346. thritis? Arthritis Res. Ther. 18 (2016) 151e159. [60] H. Wiseman, M. Cannon, H.R.V. Arnstein, B. Halliwell, Enhancement by [35] A. Andersson, A.I. Bernardi, A. Stubelius, M. Nurkkala-Karlsson, C. Ohlsson, tamoxifen of the membrane antioxidant action of the yeast membrane sterol H. Carlsten, U. Islander, Selective oestrogen receptor modulators lasofoxifene ergosterol: relevance to the antiyeast and anticancer action of tamoxifen, and bazedoxifene inhibit joint inflammation and osteoporosis in ovariec- Biochim. Biophys. Acta 1181 (1993) 201e206. tomised mice with collagen-induced arthritis, Rheumatol. Oxf. 55 (3) (2016) [61] H. Wiseman, P. Quinn, B. Halliwell, Tamoxifen and related compounds 553e563. decrease membrane fluidity in liposomes. Mechanism for the antioxidant [36] T. Thomas, M.A. Gallo, T.J. Thomas, Estrogen receptors as targets for drug action of tamoxifen and relevance to cardioprotective actions? FEBS Lett. 330 development for breast cancer, osteoporosis and cardiovascular diseases, (1993) 53e56. Curr. Cancer Drug Targets 4 (6) (2004) 483e499. [62] K. Watashi, D. Inoue, M. Hijikata, K. Goto, H.H. Aly, K. Shimotohno, Anti- [37] V.C. Jordan, Antiestrogens and selective estrogen receptor modulators as hepatitis C virus activity of tamoxifen reveals the functional association of multifunctional medicines. 2. Clinical considerations and new agents, J. Med. estrogen receptor with viral RNA polymerase NS5B, J. Biol. Chem. 282 (2007) Chem. 46 (7) (2003) 1081e1111. 32765e32772. [38] V.C. Jordan, Antiestrogens and selective estrogen receptor modulators as [63] E.M. Wagner, S.J. Gallagher, S. Reddy, W. Mitzner, Effects of tamoxifen on multifunctional medicines. 1. Receptor interactions, J. Med. Chem. 46 (6) ischemia-induced angiogenesis in the mouse lung, Angiogenesis 6 (2003) (2003) 883e908. 65e71. [39] V.C. Jordan, New insights into the metabolism of tamoxifen and its role in the [64] U.W. Nilsson, J.A. Jonsson, C. Dabrosin, Tamoxifen decreases extracellular treatment and prevention of breast cancer, Steroids 72 (2007) 829e842. TGF-beta1 secreted by breast cancer cells-a posttranslational regulation [40] J.M. Hoskins, L.A. Carey, H.L. McLeod, CYP2D6 and tamoxifen: DNA matters in involving matrix metalloproteinase activity, Exp. Cell Res. 315 (2009) 1e9. breast cancer, Nat. Rev. Cancer 9 (2009) 576e586. [65] S.Z. Khan, C.L. Longland, F. Michelangeli, The effects of phenothiazines and [41] M.J. Bijl, R.H. van Schaik, L.A. Lammers, A. Hofman, A.G. Vulto, T. van Gelder, other calmodulin antagonists on the sarcoplasmic and endoplasmic reticu- B.H. Sticker, L.E. Visser, The CYP2D6*4 polymorphism affects breast cancer lum Ca (2þ) pumps, Biochem. Pharmacol. 15 (2000) 1797e1806. survival in tamoxifen users, Breast Cancer Res. Treat. 118 (2009) 125e130. [66] E. Baral, E. Nagy, I. Berczi, Modulation of natural killer cell mediated cyto- [42] X. Wu, J.R. Hawse, M. Subramaniam, M.P. Goetz, J.N. Ingle, T.C. Spelsberg, The toxicity by tamoxifen and estradiol, Cancer 75 (1995) 591e599. tamoxifen metabolite, endoxifen, is a potent antiestrogen that targets es- [67] Z.Y. Friedman, Recent advance in the molecular mechanisms of tamoxifen trogen receptor alpha for degradation in breast cancer cells, Cancer Res. 69 action, Cancer Investig. 16 (1998) 391e396. (2009) 1722e1727. [68] A.B. Foster, R. McCague, A. Seago, G. Leclercq, S. Stoessel, F. Roy, Modification [43] H. Wiseman, G. Paganga, C. Rice-Evans, B. Halliwell, Protective actions of of the basic side chain in tamoxifen: effects on microsomal metabolism and tamoxifen and 4-hydroxytamoxifen against oxidative damage to human in vitro biological activity, Anti-Cancer Drug Des. (1986) 245e257. low-density lipoproteins: a mechanism accounting for the cardioprotective [69] V. Agouridas, I. Laı€os, A. Cleeren, E. Kizilian, E. Magnier, J.-C. Blazejewskia, action of tamoxifen? Biochem. J. 292 (1993) 635e638. G. Leclercq, Loss of antagonistic activity of tamoxifen by replacement of one [44] B.J. Furr, V.C. Jordan, The pharmacology and clinical uses of tamoxifen, N-methyl of its side chain by fluorinated residues, Bioorg Med Chem 14 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 529

(2006) 7531e7538. [97] J.M. Huss, R.P. Kopp, D.P. Kelly, Peroxisome proliferator-activated receptor [70] I. Shiina, Y. Sano, K. Nakata, T. Kikuchi, A. Sasaki, M. Ikekita, Y. Hasome, coactivator-1alpha (PGC-1alpha) coactivates the cardiac-enriched nuclear Synthesis of the new pseudo-symmetrical tamoxifen derivatives and their receptors estrogen-related receptor-alpha and -gamma. Identification of anti-tumor activity, Bioorg Med. Chem. Lett. 17 (2007) 2421e2424. novel leucine-rich interaction motif within PGC-1alpha, J. Biol. Chem. 277 [71] M.S. Christodoulou, N. Fokialakis, D. Passarella, A.N. García-Argaez, O.M. Gia, (2002) 40265e40274. I. Pongratz, L.D. Via, S.A. Haroutounian, Synthesis and biological evaluation of [98] Y. Kamei, H. Ohizumi, Y. Fujitani, T. Nemoto, T. Tanaka, N. Takahashi, novel tamoxifen analogues, Bioorg Med. Chem. 21 (2013) 4120e4131. T. Kawada, M. Miyoshi, O. Ezaki, A. Kakizuka, PPARgamma coactivator 1beta/ [72] Y. Nagahara, I. Shiina, K. Nakata, A. Sasaki, T. Miyamoto, M. Ikekita, Induction ERR ligand 1 is an ERR protein ligand, whose expression induces a high- of mitochondria-involved apoptosis in estrogen receptor-negative cells by a energy expenditure and antagonizes obesity, Proc. Natl. Acad. Sci. U. S. A. novel tamoxifen derivative, ridaifen-B, Cancer Sci. 99 (2008) 608e614. 100 (2003) 12378e12383. [73] M. Hasegawa, Y. Yasuda, M. Tanaka, K. Nakata, E. Umeda, Y. Wang, [99] P. Coward, D. Lee, M.V. Hull, J.M. Lehmann, 4-Hydroxytamoxifen binds to and C. Watanabe, S. Uetake, T. Kunoh, M. Shionyu, R. Sasaki, I. Shiina, deactivates the estrogen-related receptor gamma, Proc. Natl. Acad. Sci. U. S. T. Mizukami, A novel tamoxifen derivative, ridaifen-F, is a nonpeptidic small- A. 98 (2001) 8880e8884. molecule proteasome inhibitor, Eur. J. Med. Chem. 71 (2014) 290e305. [100] E.Y.H. Chao, J.L. Collins, S. Gaillard, A.B. Miller, L. Wang, L.A. Orband-Miller, [74] S.J. Howell, S.R. Johnston, A. Howell, The use of selective estrogen receptor R.T. Nolte, D.P. McDonnel, T.M. Willson, W.J. Zuercher, Structure-guided modulators and selective estrogen receptor down-regulators in breast can- synthesis of tamoxifen analogs with improved selectivity for the orphan cer, Best. Pract. Res. Clin. Endocrinol. Metab. 18 (2004) 47e66. ERRg, Bioorg Med. Chem. Lett. 16 (2006) 821e824. [75] C.K. Baumann, M. Castiglione-Gertsch, Estrogen receptor modulators and [101] K.R. Abdellatif, C.A. Velazquez, Z. Huang, M.A. Chowdhury, E.E. Knaus, Triaryl down regulators: optimal use in postmenopausal women with breast cancer, (Z)-olefins suitable for radiolabeling with iodine-124 or fluorine-18 radio- Drugs 67 (2007) 2335e2353. nuclides for positron emission tomography imaging of estrogen positive [76] C.E. Connor, J.D. Norris, G. Broadwater, T.M. Willson, M.M. Gottardis, breast tumors, Bioorg Med. Chem. Lett. 21 (2011) 1195e1198. M.W. Dewhirst, D.P. McDonnell, Circumventing tamoxifen resistance in [102] R.J. Baumann, T.L. Bush, D.E. Cross-Doersen, E.A. Cashman, P.S. Wright, breast cancers using antiestrogens that induce unique conformational J.H. Zwolshen, G.F. Davis, D.P. Matthews, D.M. Bender, A.J. Bitonti, Clomi- changes in the estrogen receptor, Cancer Res. 61 (2001) 2917e2922. phene analogs with activity in vitro and in vivo against human breast cancer [77] T. Shoda, K. Okuhira, M. Kato, Y. Demizu, H. Inoue, M. Naito, M. Kurihara, cells, Biochem. Pharmacol. 55 (1998) 841e851. Design and synthesis of tamoxifen derivatives as a selective estrogen re- [103] L. Zheng, Q. Wei, B. Zhou, L. Yang, Z.L. Liu, Synthesis of 1,1,2- ceptor down-regulator, Bioorg Med. Chem. Lett. 24 (2014) 87e89. triphenylethylenes and their antiproliferative effect on human cancer cell [78] T. Shoda, M. Kato, R. Harada, T. Fujisato, K. Okuhira, Y. Demizu, H. Inoue, lines, Anticancer Drugs 18 (2007) 1039e1055. M. Naito, M. Kurihara, Synthesis and evaluation of tamoxifen derivatives [104] K.R.A. Abdellatif, A. Belal, H.A. Omar, Design, synthesis and biological eval- with a long alkyl side chain as selective estrogen receptor down-regulators, uation of novel triaryl (Z)-olefins as tamoxifen analogues, Bioorg Med. Chem. Bioorg Med. Chem. 23 (2015) 3091e3096. Lett. 23 (2013) 4960e4963. [79] S. Mandlekar, A.N. Kong, Mechanisms of tamoxifen-induced apoptosis, [105] K. Park, N.R. Kiteringham, P.M. O'Neill, Annu. Metabolism of fluorine- Apoptosis 6 (2001) 469e477. containing drugs, Rev. Pharmacol. Toxicol. 41 (2001) 443e470. [80] I. Larosche, P. Letteron, B. Fromenty, N. Vadrot, A. Abbey-Toby, G. Feldmann, [106] S. Purser, P.R. Moore, S. Swallow, V. Gouverneur, Fluorine in medicinal D. Pessayre, A. Mansouri, Tamoxifen inhibits topoisomerases, depletes chemistry, Chem. Soc. Rev. 37 (2008) 320e330. mitochondrial DNA, and triggers steatosis in mouse liver, J. Pharm. Exp. Ther. [107] K.L. Kirk, Fluorination in medicinal chemistry: methods, strategies, and 321 (2007) 526e535. recent developments, Org. Process Res. Dev. 12 (2008) 305e321. [81] M.S. Christodoulou, M. Zarate, F. Ricci, G. Damia, S. Pieraccini, F. Dapiaggi, [108] E.P. Gillis, K.J. Eastman, M.D. Hill, D.J. Donnelly, N.A. Meanwell, Applications M. Sironi, L.L. Presti, A.N. García-Arg_aez, L.D. Via, Passarella D. 4-(1,2- of fluorine in medicinal chemistry, J. Med. Chem. 58 (21) (2015) 8315e8359. diarylbut-1-en-1-yl)isobutyranilide derivatives as inhibitors of topoisomer- [109] V. Erdelyi-Toth, F. Gyergyay, I. Szamel, E. Pap, J. Kralovanszky, E. Bojti, ase II, Eur. J. Med. Chem. 118 (2016) 79e89. M. Csorgo,€ S. Drabant, I. Klebovich, Pharmacokinetics of panomifene in [82] F.E. Silverstein, G. Faich, J.L. Goldstein, L.S. Simon, T. Pincus, A. Whelton, healthy volunteers at phase I/a study, Anti-Cancer Drugs 8 (1997) 603e609. R. Makuch, G. Eisen, N.M. Agrawal, W.F. Stenson, A.M. Burr, W.W. Zhao, [110] K. Monostory, K. Jemnitz, L. Vereczkey, G. Czira, Species differences in J.D. Kent, J.B. Lefkowith, K.M. Verburg, G.S. Geis, Gastrointestinal toxicity metabolism of panomifene, an analogue of tamoxifen, Drug Metab. Dispos. with celecoxib vs nonsteroidal anti-inflammatory drugs for osteoarthritis 25 (1997) 1370e1378. and rheumatoid arthritis: the CLASS study: a randomized controlled trial. [111] T. Konno, T. Daitoh, A. Noiri, J. Chae, T. Ishihara, H.A. Yamanaka, Highly regio- Celecoxib Long-term Arthritis Safety Study, JAMA 284 (2000) 1247e1255. and stereoselective carbocupration of fluoroalkylated internal Alkynes: a [83] C. Bombardier, L. Laine, A. Reicin, D. Shapiro, R. Burgos-Vargas, B. Davis, short total synthesis of the antiestrogenic drug panomifene, Org. Lett. 6 R. Day, M.B. Ferraz, C.J. Hawkey, M.C. Hochberg, T.K. Kvien, T.J. Schnitzer, (2004) 933e936. Comparison of upper gastrointestinal toxicity of rofecoxib and naproxen in [112] B. Malo-Forest, G. Landelle, J.-A. Roy, J. Lacroix, R.C. Gaudreault, J.-F. Paquin, patients with rheumatoid arthritis, N. Engl. J. Med. 343 (2000) 1520e1528. Synthesis and growth inhibition activity of fluorinated derivatives of [84] M. Katori, M. Majima, Cyclooxygenase-2: its rich diversity of roles and tamoxifen, Bioorg Med. Chem. Lett. 23 (2013) 1712e1715. possible application of its selective inhibitors, Inflamm. Res. 49 (2000) [113] P. De Medina, G. Favre, M. Poirot, Multiple targeting by the antitumor drug 367e392. tamoxifen: a structure-activity study, Curr. Med. Chem. Anticancer Agents 4 [85] M.J. Uddin, P.N.P. Rao, E.E. Knaus, Design of acyclic triaryl olefins: a new class (2004) 491e508. of potent and selective cyclooxygenase-2 (COX-2) inhibitors, Bioorg Med. [114] M.D. Johnson, H. Zuo, K.-H. Lee, J.P. Trebley, J.M. Rae, R.V. Weatherman, Chem. Lett. 14 (2004) 1953e1956. Z. Desta, D.A. Flockhart, T.C. Skaar, Pharmacological characterization of 4- [86] M.J. Uddin, P.N.P. Rao, E.E. Knaus, Design and synthesis of acyclic triaryl (Z)- hydroxy-N-desmethyl tamoxifen, a novel active metabolite of tamoxifen, olefins: a novel class of cyclooxygenase-2 (COX-2) inhibitors, Bioorg Med. Breast Cancer Res. Treat. 85 (2004) 151e159. Chem. 12 (2004) 5929e5940. [115] C. Carpenter, R.J. Sorenson, Y. Jin, S. Klossowski, T. Cierpicki, M. Gnegy, [87] U. Lucking, Sulfoximines: a neglected opportunity in medicinal chemistry, H.D. Showalter, Design and synthesis of triarylacrylonitrile analogues of Angew. Chem. Int. Ed. Engl. 52 (36) (2013) 9399e9408. tamoxifen with improved binding selectivity to protein kinase C, Bioorg [88] C. Worch, A.C. Mayer, C. Bolm, in: T. Toru, C. Bolm (Eds.), Organosulfur Med. Chem. 24 (2016) 5495e5504. Chemistry in Asymmetric Synthesis, Wiley/VCH, Weinheim, 2008. [116] E. Bignon, M. Pons, J.-C. Dore, J. Gilbert, T. Ojasoo, J.-F. Miquel, J.P. Raynaud, [89] I. Ahmad, Shagufta, Sulfones: an important class of organic compounds with A.C. Paulet, Influence of di- and tri-phenylethylene estrogen/antiestrogen diverse biological activities, Int. J. Pharm. Pharm. Sci. 7 (3) (2015) 19e27. structure on the mechanisms of protein kinase C inhibition and activation as [90] X.Y. Chen, S.J. Park, H. Buschmann, M.D. Rosa, C. Bolm, Syntheses and bio- revealed by a multivariate analysis, Biochem. Pharmacol. 42 (1991) logical activities of sulfoximine-based acyclic triaryl olefins, Bioorg Med. 1373e1383. Chem. Letts 22 (2012) 4307e4309. [117] A.J. Ellis, V.M. Hendrick, R. Williams, B.S. Komm, Selective estrogen receptor [91] S. Ray, Sangita, The potent triarylethylene pharmacophore, Drugs Future 29 modulators in clinical practice: a safety overview, Expert Opin. Drug Saf. 14 (2004) 185e203. (2015) 921e934. [92] M.R. Schneider, R.W. Hartmann, F. Sinowatz, W. Amselgruber, Nonsteroidal [118] Z. Desta, B.A. Ward, N.V. Soukhova, D.A. Flockhart, Comprehensive evalua- antiestrogens and partial estrogens with prostatic tumor inhibiting activity, tion of tamoxifen sequential biotransformation by the human cytochrome J. Cancer Res. Clin. 112 (1986) 258e265. P450 system in vitro: prominent roles for CYP3A and CYP2D6, J. Pharmacol. [93] G. Kaur, M.P. Mahajan, M.K. Pandey, P. Singh, S.R. Ramisetti, A.K. Sharma, Exp. Ther. 310 (2004) 1062e1075. Design, synthesis and evaluation of Ospemifene analogs as anti-breast cancer [119] N.S. Ahmed, N.H. Elghazawy, A.K. ElHady, M. Engel, R.W. Hartmann, agents, Eur. J. Med. Chem. 86 (2014) 211e218. A.H. Abadi, Design and synthesis of novel tamoxifen analogues that avoid [94] G. Kaur, M.P. Mahajan, M.K. Pandey, P. Singh, S R b Ramisetti, A.K. Sharma, CYP2D6 metabolism, Eur. J. Med. Chem. 112 (2016) 171e179. Design, synthesis, and anti-breast cancer evaluation of new triarylethylene [120] R. Siles, J.F. Ackley, M.B. Hadimani, J.J. Hall, B.E. Mugabe, R. Guddneppanavar, analogs bearing short alkyl- and polar amino-/amido-ethyl chains, Bioorg K.A. Monk, J.C. Chapuis, G.R. Pettit, D.J. Chaplin, K. Edvardsen, M.L. Trawick, Med. Chem. Letts 26 (2016) 1963e1969. C.M. Garner, K.G. Pinney, Combretastatin dinitrogen-substituted stilbene [95] V. Giguere, To ERR in the estrogen pathway, Trends Endocr. Metab. 13 (2002) analogues as tubulin-binding and vascular-disrupting agents, J. Nat. Prod. 71 220e225. (2008) 313e320. [96] B. Horard, J.M. Vanacker, Estrogen receptor-related receptors: orphan re- [121] G.R. Pettit, M.P. Grealish, D.L. Herald, M.R. Boyd, E. Hamel, R.K. Pettit, Anti- ceptors desperately seeking a ligand, J. Mol. Endocrinol. 31 (2003) 349e357. neoplastic agents. 443. Synthesis of the cancer cell growth inhibitor 530 Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531

hydroxyphenstatin and its sodium diphosphate prodrug, J. Med. Chem. 43 G. Jaouen, Synthesis, biochemical properties and molecular modeling studies (2000) 2731e2737. of organometallic specific estrogen receptor modulators (SERMs), the fer- [122] R.P. Tanpure, A.R. Harkrider, T.E. Strecker, E. Hamel, M.L. Trawick, rocifens and hydroxyferrocifens: evidence for an antiproliferative effect of K.G. Pinney, Application of the McMurry coupling reaction in the synthesis of hydroxyferrocifens on both hormone-dependent and hormone-independent tri- and tetra-arylethylene analogues as potential cancer chemotherapeutic breast cancer cell lines, Chem. Eur. J. 9 (2003) 5223e5236. agents, Bioorg Med. Chem. 17 (2009) 6993e7001. [146] A. Cabral de Oliveira, E.A. Hillard, P. Pigeon, D.D. Rocha, F.A.R. Rodrigues, [123] M.J. Meegan, R.B. Hughes, D.G. Lloyd, D.C. Williams, D.M. Zisterer, Flexible R.C. Montenegro, L.V. Costa-Lotufo, M.O.F. Goulart, G. Jaouen, Biological estrogen receptor modulators: design, synthesis, and antagonistic effects in evaluation of twenty-eight ferrocenyl tetrasubstituted olefins: cancer cell human MCF-7 breast cancer cells, J. Med. Chem. 44 (2001) 1072e1084. growth inhibition, ROS production and hemolytic activity, Eur. J. Med. Chem. [124] D.G. Lloyd, R.B. Hughes, D.M. Zisterer, D.C. Williams, C. Fattorusso, 46 (2011) 3778e3787. B. Catalanotti, G. Campiani, M.J. Meegan, Benzoxepin-Derived estrogen re- [147] X. Huang, I.H. El-Sayed, M.A. El-Sayed, Cancer cell imaging and photothermal ceptor Modulators: a novel molecular scaffold for the estrogen receptor, therapy in the near-infrared region by using gold nanorods, J. Am. Chem. So J. Med. Chem. 47 (2004) 5612e5615. 128 (2006) 2115e2120. [125] D.G. Lloyd, H.M. Smith, T. O'Sullivan, D.M. Zisterer, M.J. Meegan, Synthesis, [148] V. Dixit, J. Van den Bossche, D.M. Sherman, D.H. Thompson, R.P. Andres, structure-activity relationship and antiestrogenic effects in human MCF-7 Synthesis and grafting of thioctic acid-PEG-folate conjugates onto Au breast cancer cells of flexible estrogen receptor modulators, Med. Chem. 1 nanoparticles for selective targeting of folate receptor-positive tumor cells, (4) (2005) 335e353. Bioconjugate Chem. 17 (2006) 603e609. [126] A.K. Shiau, D. Barstad, P.M. Loria, L. Cheng, P.J. Kushner, D.A. Agard, [149] T.B. Huff, L. Tong, Y. Zhao, M.N. Hansen, J.X. Cheng, A. Wei, Hyperthermic G.L. Greene, The structural basis of estrogen receptor/coactivators recogni- effects of gold nanorods on tumor cells, Nanomedicine 2 (2007) 125e132. tion and the antagonism of this interaction by tamoxifen, Cell 95 (7) (1998) [150] L. Tong, Y. Zhao, T.B. Huff, M.N. Hansen, A. Wei, J.X. Cheng, Gold nanorods 927e937. mediate tumor cell death by compromising membrane integrity, Adv. Mater [127] N.H. Elghazawy, M. Engel, R.W. Hartmann, M.M. Hamed, N.S. Ahmed, 2007 (19) (2007) 3136e3141. A.H. Abadi, Design and synthesis of novel flexible ester-containing analogues [151] E.R. Levin, Integration of the extranuclear and nuclear actions of estrogen, of tamoxifen and their evaluation as anticancer agents, Future Med. Chem. 8 Mol. Endocrinol. 19 (2005) 1951e1959. (3) (2016) 249e256. [152] C.H. Campbell, N. Bulayeva, D.B. Brown, B. Gametchu, C.S. Watson, Regula- [128] S.A. Haroutounian, A.W. Scribner, J.A. Katzenellenbogen, Derivatives of 4- tion of the membrane estrogen receptor-alpha: role of cell density, serum, styrylpyridines: synthesis, estrogen receptor binding affinity, and photo- cell passage number, and estradiol, FASEB J. 16 (2002) 1917e1927. physical properties, Steroids 60 (1995) 636e645. [153] E.C. Dreaden, S.C. Mwakwari, Q.H. Sodji, A.K. Oyelere, M.A. El-Sayed, [129] S. Auger, Y. Merand, J.D. Pelletier, D. Poirier, F. Labrie, Synthesis and bio- Tamoxifen-Poly(ethylene glycol)-thiol gold nanoparticle conjugates: logical activities of thioether derivatives related to the antiestrogens enhanced potency and selective delivery for breast cancer treatment, Bio- tamoxifen and ICI 164384, J. Steroid Biochem. Mol. Biol. 52 (1995) 547e565. conjugate Chem. 20 (2009) 2247e2253. [130] M. Wenckens, P. Jakobsen, P. Vedsø, P.O. Huusfeldt, B. Gissel, M. Barfoed, [154] D.J. Nelson, R. Shagufta Kumar, Characterization of a tamoxifen-tethered B.L. Brockdorff, A.E. Lykkesfeldt, M. Begtrup, N-alkoxypyrazoles as bio- single-walled carbon nanotube conjugate by using NMR spectroscopy, mimetics for the alkoxyphenyl group in tamoxifen, Bioorg Med. Chem. 11 Anal. Bioanal. Chem. 404 (2012) 771e776. (2003) 1883e1899. [155] E.L. Rickert, S. Oriana, C. Hartman-Frey, X. Long, T.T. Webb, K.P. Nephew, [131] G. Daletos, N.J. de Voogd, W.E.G. Muller, V. Wray, W.H. Lin, D. Feger, R.V. Weatherman, Synthesis and characterization of fluorescent 4- M. Kubbutat, A.H. Aly, P. Proksch, Cytotoxic and protein kinase inhibiting hydroxytamoxifen conjugates with unique antiestrogenic properties, Bio- nakijiquinones and nakijiquinols from the sponge Dactylospongia meta- conjugate Chem. 21 (2010) 903e910. chromia, J. Nat. Prod. 77 (2014) 218e226. [156] F. Abendroth, M. Solleder, D. Mangoldt, P. Welker, K. Licha, M. Weber, [132] S.T. Duggan, G.M. Keating, Pegylated liposomal doxorubicin: a review of its O. Seitz, High affinity fluorescent ligands for the estrogen receptor, Eur. J. use in metastatic breast cancer, ovarian cancer, multiple myeloma and AIDS- Org. Chem. 10 (2015) 2157e2166. related Kaposi's sarcoma, Drugs 71 (2011) 2531e2558. [157] L.A. Ho, E. Thomas, R.A. McLaughlin, G.R. Flematti, R.O. Fuller, A new selective [133] N.M. Kogan, M. Schlesinger, E. Priel, R. Rabinowitz, E. Berenshtein, fluorescent probe based on tamoxifen, Bioorg Med. Chem. Lett. 26 (2016) M. Chevion, R. Mechoulam, HU-331, a novel cannabinoid-based anticancer 4879e4883. topoisomerase II inhibitor, Mol. Cancer Ther. 6 (2007) 173e183. [158] B. Yu, Z. Qin, G.T. Wijewickrama, P. Edirisinghe, J.L. Bolton, G.R.J. Thatcher, [134] N. Tabata, T. Sunazuka, H. Tomoda, T. Nagamitsu, Y. Iwai, S. Omura, Dio- Comparative methods for analysis of protein covalent modification by lmycins, new anticoccidial agents produced by Streptomyces sp. II. Structure electrophilic quinoids formed from xenobiotics, Bioconjugate Chem. 20 elucidation of diolmycins A1, A2, B1 and B2, and synthesis of diolmycin A1, (2009) 728e741. J. Antibiot. 46 (1993) 762e769. [159] K. Cyrus, M. Wehenkel, E.-Y. Choi, H. Lee, H. Swanson, K.-B. Kim, Jostling for [135] S. Archana, R. Geesala, N.B. Rao, S. Satpati, G. Puroshottam, A. Panasa, A. Dixit, position: optimizing linker location in the design of estrogen receptor- A. Das, A.K. Srivastava, Development of constrained tamoxifen mimics and targeting PROTACs, Chem. Med. Chem. 5 (2010) 979e985. their antiproliferative properties against breast cancer cells, Bioorg Med. [160] D. Zhang, S.-H. Baek, A. Ho, K. Kim, Degradation of target protein in living Chem. Lett. 25 (2015) 680e684. cells by small-molecule proteolysis inducer, Bioorg Med. Chem. Lett. 14 [136] R. McCague, R. Kuroda, G. Leclercq, S. Stoessel, Synthesis and estrogen re- (2004) 645e648. ceptor binding of 6,7-dihydro-8-phenyl-9-[4-[2-(dimethylamino)ethoxy] [161] A.F. Gacio, C. Fernandez-Marcos, N. Swamy, D. Dunn, R. Ray, Photodynamic phenyl]-5H-benzocycloheptene, a nonisomerizable analog of tamoxifen. X- cell-kill analysis of breast tumor cells with a tamoxifen-pyropheophorbide ray crystallographic studies, J. Med. Chem. 29 (1986) 2053e2059. conjugate, J. Cell Biochem. 99 (2006) 665e670. [137] M.J. Meegan, I. Barrett, J. Zimmermann, A.J.S. Knox, M. Daniela, A.M. Zistere, [162] R. Schobert, G. Bernhardt, B. Biersack, S. Bollwein, M. Fallahi, A. Grotemeier, D.G. Lloyd, Benzothiepin-derived molecular scaffolds for estrogen receptor G.L. Hammond, Steroid conjugates of dichloro(6-aminomethylnicotinate) modulators: synthesis and antagonistic effects in breast cancer cells, J. Enzy platinum(II): effects on DNA, sex hormone binding globulin, the estrogen Inhib. Med. Chem. 22 (2007) 655e666. receptor, and various breast cancer cell lines, Chem. Med. Chem. 2 (2007) [138] M.I. Ansari, M.K. Hussain, A. Arun, B. Chakravarti, R. Konwar, K. Hajela, 333e342. Synthesis of targeted dibenzo[b,f]thiepines and dibenzo[b,f]oxepines as po- [163] P.J. Burke, T.H.J. Koch, Design, synthesis, and biological evaluation of Dox- tential lead molecules with promising anti-breast cancer activity, Eur. J. Med. orubicinFormaldehyde conjugates targeted to breast cancer cells, J. Med. Chem. 99 (2015) 113e124. Chem. 47 (2004) 1193e1206. [139] G. Jaouen, S. Top, A. Vessieres, G. Leclercq, M.J. McGlinchey, The first [164] R. Pandey, R. Chander, K.B. Sainis, Prodigiosins as anti cancer agents: living organometallic selective estrogen receptor modulators (SERMs) and their upto their name, Curr. Pharm. Des. 15 (2009) 732e741. relevance to breast cancer, Curr. Med. Chem. 11 (2004) 2505e2517. [165] N.R. Williamson, P.C. Fineran, T. Gristwood, S.R. Chawrai, F.J. Leeper, [140] E. Hillard, A. Vessieres, F. Le Bideau, D. Plazuk, D. Spera, M. Huche, G. Jaouen, G.P.C. Salmond, Anticancer and immunosuppressive properties of bacterial A series of unconjugated ferrocenyl phenols: prospects as anticancer agents, prodiginines, Future Microbiol. 2 (2007) 605e618. Chem. Med. Chem. 1 (2006) 551e559. [166] J. Peng, S. Sengupta, V.C. Jordan, Potential of selective estrogen receptor [141] A. Nguyen, S. Top, A. Vessieres, P. Pigeon, M. Huche, E.A. Hillard, G. Jaouen, modulators as treatments and preventives of breast cancer, Anticancer Organometallic analogues of tamoxifen: effect of the amino side-chain Agents Med. Chem. 9 (2009) 481e499. replacement by a carbonyl ferrocenyl moiety in hydroxytamoxifen, [167] M. Dowsett, R. Nicholson, R. Pietras, Biological characteristics of the pure J. Organomet. Chem. 692 (2007) 1219e1225. antiestrogen fulvestrant: overcoming endocrine resistance, Breast Cancer [142] K. Nikitin, Y. Ortin, H. Müller-Bunz, M.-A. Plamont, G. Jaouen, A. Vessieres, Res. Treat. 93 (2005) 11e18. M.J. McGlinchey, Organometallic SERMs (selective estrogen receptor mod- [168] T. Ishikawa, H. Nakagawa, Y. Hagiya, N. Nonoguchi, S.-I. Miyatake, ulators): cobaltifens, the (cyclobutadiene)cobalt analogues of hydrox- T. Kuroiwa, Key role of human ABC transporter ABCG2 in photodynamic ytamoxifen, J. Organomet. Chem. 695 (2010) 595e608. therapy and photodynamic diagnosis, Adv. Pharmacol. Sci. (2010). Article ID [143] A.R. Kudinov, E.V. Mutsenek, D.A. Loginov, (Tetramethylcyclobutadiene)co- 587306. balt chemistry, Coord. Chem. Rev. 248 (2004) 571e585. [169] C.L.A. Hawco, E. Marchal, M.I. Uddin, A.E.G. Baker, D.P. Corkery, G. Dellaire, [144] S. Top, J. Tang, A. Vessieres, D. Carrez, C. Provot, G. Jaouen, Ferrocenyl A. Thompson, Synthesis and biological evaluation of prodigiosene conjugates hydroxytamoxifen: a prototype for a new range of estradiol receptor site- of porphyrin, estrone and 4-hydroxytamoxifen, Bioorg Med. Chem. 21 directed cytotoxics, Chem. Commun. 8 (1996) 955e956. (2013) 5995e6002. [145] S. Top, A. Vessieres, G. Leclercq, J. Quivy, J. Tang, J. Vaisserman, M. Huche, [170] I. Shiina, Y. Sano, K. Nakata, T. Kikuchi, A. Sasaki, M. Ikekita, Y. Nagahara, Shagufta, I. Ahmad / European Journal of Medicinal Chemistry 143 (2018) 515e531 531

Y. Hasome, T. Yamori, K. Yamazaki, Synthesis and pharmacological evalua- M. von Euler, T. Sahmoud, A. Webster, M. Steinberg, Arimidex writing C, tion of the novel pseudo-symmetrical tamoxifen derivatives as anti-tumor investigators comm M. Anastrozole is superior to tamoxifen as first-line agents, Biochem. Pharmacol. 75 (2008) 1014e1026. therapy in hormone receptor positive advanced breast carcinoma-results [171] S. Tsukuda, T. Kusayanagi, E. Umeda, C. Watanabe, Y-t Tosaki, S. Kamisuki, of two randomized trials designed for combined analysis, Cancer 92 (2001) T. Takeuchi, Y. Takakusagi, I. Shiina, F. Sugawara, B. Ridaifen, A tamoxifen 2247e2258. derivative, directly binds to Grb10 interacting GYF protein 2, Bioorg Med. [189] J.M. Nabholtz, J. Bonneterre, A. Buzdar, J.F.R. Robertson, B. Thurlimann, Ari- Chem. 21 (2013) 311e320. midex writing comm behalf, I. Anastrozole (arimidex (Tm)) versus tamoxifen [172] W.Z. Guo, Y. Wang, E. Umeda, I. Shiina, S. Dan, T. Yamori, Search for novel as first-line therapy for advanced breast cancer in postmenopausal women: anti-tumor agents from ridaifens using JFCR39, a panel of human cancer cell survival analysis and updated safety results, Eur. J. Cancer 39 (2003) lines, Biol. Pharm. Bull. 36 (2013) 1008e1016. 1684e1689. [173] K. Ikeda, S. Kamisuki, S. Uetake, A. Mizusawa, N. Ota, T. Sasaki, S. Tsukuda, [190] A. Howell, J. Cuzick, M. Baum, A. Buzdar, M. Dowsett, J.F. Forbes, G. Hoctin- T. Kusayanagi, Y. Takakusagi, K. Morohashi, T. Yamori, S. Dan, I. Shiina, Boes, I. Houghton, G.Y. Locker, J.S. Tobias, A.T. Grp, Results of the ATAC F. Sugawara, G. Ridaifen, Tamoxifen analog, is a potent anticancer drug (arimidex, tamoxifen, alone or in combination) trial after completion of 5 working through a combinatorial association with multiple cellular factors, Years' adjuvant treatment for breast cancer, Lancet 365 (2005) 60e62. Bioorg Med. Chem. 23 (2015) 6118e6124. [191] M.S. Ewer, S. Gluck, A Woman's heart the impact of adjuvant endocrine [174] M. Lazzeroni, D. Serrano, B.K. Dunn, B.M. Heckman Stoddard, O. Lee, S. Khan, therapy on cardiovascular health, Cancer 115 (2009) 1813e1826. A. Decensi, Oral low dose and topical tamoxifen for breast cancer prevention: [192] B. Bird, S.M. Swain, Cardiac toxicity in breast cancer survivors: review of modern approaches for an old drug, Breast Cancer Res. 14 (2012) 214. potential CardiacProblems, Clin. Cancer Res. 14 (2008) 14e24. [175] P. Bourassa, T.J. Thomas, H.A. Tajmir-Riahi, Locating the binding sites of [193] N. Bundred, J. The effects of aromatase inhibitors on lipids and thrombosis, antitumor drug tamoxifen and its metabolites with DNA, J. Pharm. Biomed. Br. J. Cancer 93 (2005) S23eS27. Anal. 95 (2014) 193e199. [194] W.J. Lu, C. Xu, Z.F. Pei, A.S. Mayhoub, M. Cushman, D.A. Flockhart, The [176] P. Bourassa, T.J. Thomas, H.A. Tajmir-Riahi, A short review on the delivery of tamoxifen metabolite norendoxifen is a potent and selective inhibitor of breast anticancer drug tamoxifen and its metabolites by serum proteins, aromatase (Cyp19) and a potential lead compound for novel therapeutic J. Nanomed Res. 4 (2) (2016) 00080. agents, Breast Cancer Res. Treat. 133 (2012) 99e109. [177] P. Chanphai, L. Bekale, S. Sanyakamdhorn, D. Agudelo, G. Berube, T.J. Thomas, [195] W. Lv, J. Liu, D. Lu, D.A. Flockhart, M. Cushman, Synthesis of mixed (E,Z)-, (E)- H.A. Tajmir-Riahi, PAMAM dendrimers in drug delivery: loading efficacy and , and (Z)-norendoxifen with dual aromatase inhibitory and estrogen receptor polymer morphology, Can. J. Chem. 95 (2017) 891e896. modulatory activities, J. Med. Chem. 56 (11) (2013) 4611e4618. [178] D. Zhang, V. Anantharam, A. Kanthasamy, A.G. Kanthasamy, Neuroprotective [196] W. Lv, J. Liu, T.C. Skaar, D.A. Flockhart, M. Cushman, Design and synthesis of effect of protein kinase C delta inhibitor rottlerin in cell culture and animal norendoxifen analogues with dual aromatase inhibitory and estrogen re- models of Parkinson's disease, J. Pharmacol. Exp. Ther. 322 (2007) 913e922. ceptor modulatory activities, J. Med. Chem. 58 (6) (2015) 2623e2648. [179] J.L. Garrido, J.A. Godoy, A. Alvarez, M. Bronfman, N.C. Inestrosa, Protein ki- [197] V.W. Winkler, M.A. Nyman, R.S. Egan, Diethylstilbestrol cis-trans isomeri- nase C inhibits amyloid beta peptide neurotoxicity by acting on members of zation and estrogen activity of diethylstilbestrol isomers, Steroids 17 (1971) the Wnt pathway, FASEB J. 2002 (1982-1984) 16. 197e207. [180] H.K. Manji, R.H. Lenox, Signaling: cellular insights into the pathophysiology [198] J.A. Katzenellenbogen, K.E. Carlson, B.S. Katzenellenbogen, Facile geometric of bipolar disorder, Biol. Psychiatry 48 (2000) 518e530. isomerization of phenolic non-steroidal estrogens and antiestrogens: limi- [181] K.C. Schmitt, M.E. Reith, Regulation of the dopamine transporter: aspects tations to the interpretation of experiments characterizing the activity of relevant to psychostimulant drugs of abuse, Ann. N. Y. Acad. Sci. 1187 (2010) individual isomers, J. Steroid Biochem. Mol. Biol. 22 (1985) 589e596. 316e340. [199] W. Lv, J. Liu, T.C. Skaar, E. O'Neill, G. Yu, D.A. Flockhart, M. Cushman, Syn- [182] F. Battaini, Protein kinase C isoforms as therapeutic targets in nervous sys- thesis of triphenylethylene bisphenols as aromatase inhibitors that also tem disease states, Pharmacol. Res. 44 (2001) 353. modulate estrogen receptors, J. Med. Chem. 59 (1) (2016) 157e170. [183] D. Mochly-Rosen, K. Das, K.V. Grimes, Protein kinase C, an elusive thera- [200] B.P. Haynes, M. Dowsett, W.R. Miller, J.M. Dixon, A.S. Bhatnagar, The phar- peutic target? Nat. Rev. Drug Disc 11 (2012) 937e957. macology of letrozole, J. Steroid Biochem. Mol. Biol. 87 (2003) 35e45. [184] C.A.J. Zarate, J.B. Singh, P.J. Carlson, J. Quiroz, L. Jolkovsky, D.A. Luckenbaugh, [201] L.-M. Zhao, H.-S. Jin, J. Liu, T.C. Skaar, J. Ipe, W. Lv, D.A. Flockhart, H.K. Manji, Efficacy of a protein kinase C inhibitor (tamoxifen) in the treat- M. Cushman, A new Suzuki synthesis of triphenylethylenes that inhibit ment of acute mania: a pilot study, Bipolar Disord. 9 (2007) 561e570. aromatase and bind to estrogen receptors a and b, Bioorg Med. Chem. 24 [185] H. Einat, H.K. Manji, Cellular plasticity cascades: genes-to-behavior pathways (2016) 5400e5409. in animal models of bipolar disorder, Biol. Psychiatry 59 (2006) 1160e1171. [202] P. Arsenyan, E. Paegle, I. Domracheva, A. Gulbe, I. Kanepe-Lapsa, [186] M.M. Weissman, P.J. Leaf, G.L. Tishler, D.G. Blazer, M. Karno, M. Bruce, I. Shestakova, Selenium analogues of raloxifene as promising anti- L.P. Florio, Affective disorders in five United States communities, Psychol. proliferative agents in treatment of breast cancer, Eur. J. Med. Chem. 87 Med. 18 (1988) 141e153. (2014) 471e483. [187] S.M. Ali, A. Ahmad, A. Shahabuddin, m U. Ahmad, S. Shieikh, I. Ahmad, [203] K. Ohta, Y. Chiba, A. Kaise, Y. Endo, Structure-activity relationship study of Endoxifen is a new otent inhibitor of PKC: a potential therapeutic ageny for diphenylamine-based estrogen receptor (ER) antagonists, Bioorg Med. Chem. bipolar disorder, Biorg Med. Chem. Lett. 20 (2010) 2665e2667. 23 (2015) 861e867. [188] J. Bonneterre, A. Buzdar, J.M.A. Nabholtz, J.F.R. Robertson, B. Thurlimann,