br Results br SERD synthesis
3.1. SERD synthesis and properties
As reported earlier , we designed, synthesized and screened more than 65 new SERD candidates, all of which have the general structure shown in 1, namely 11β-aryloxy estradiols, with a basic amine positioned at the 4-position of the aryl ring (Fig. 1). The basic amine is connected to the aryl unit either directly or via a spacer that varies from 3 to 6 atoms. In some of the more active compounds, we also attached an electron-withdrawing group, e.g. a trifluoromethyl unit or a fluoride atom, in the 3-positon (ortho to the amino chain). Of these compounds, several had activity comparable to fulvestrant 2 but JD128, 4, in par-ticular was more potent than fulvestrant in a number of antitumor as-says as shown below. We note that this new class of steroid-like SERDs lack the prototypical side chain (as in fulvestrant) widely used to design other drugs with ERα antagonism, but these SERD candidates generate
Fig. 1. New substituted estradiols. See text for details.
a full antagonist profile and induce significant ERα down-regulation, likely similar to significant ‘indirect’ receptor antagonism as reported in previous independent studies of 11β−substitutions in ERα [52,53] and other structural changes as in an independent report .
This new series of estradiol analogues, namely 11β-(4-aminoalkyl) aryloxy-estradiols, are expected to bind the ER ligand-binding domain since they are close structural analogues of estradiol (cf. Hansen et al. ). The 11β-aryloxy group, bearing a variable length chain ending in a basic dialkylamino group, would be expected to block the folding of helix-12 by potentially both steric hindrance and a salt bridge forma-tion between the protonated amine and an acidic side chain on helix 12. Thus, these ER antagonists should bind to ER in such a way as to pre-vent the folding of helix-12 and thereby potentially inhibit BC pro-liferation.
The synthesis of the new analogues (Fig. 2) started with estradiol 7 which was converted into the bis (benzyloxy)ketone 8 by a known route [31–34] (protection, benzylic Erteberel to the 9, 11-alkene, hydro-boration-oxidation, and final oxidation to the ketone). Reduction of this protected ketone 8 with sodium borohydride aﬀorded the expected 11β-alcohol 9 by attack of the hydride on the less hindered α–face, away from the hindering 13β–methyl group. Formation of the 11β-alkoxide anion of 9 using potassium hydride in THF/DMF followed by addition of 4-fluoronitrobenzene 10 eﬀected a clean SNAr reaction to aﬀord the 4-nitrophenyl ether 11. Nickel boride reduction [55,56] of the nitro group (sodium borohydride with NiCl2·6H2O in methanol) gave the aminophenyl ether 12 in good yield. Removal of the two benzyl ethers from 12 by catalytic hydrogenolysis using Pd(OH)2 in methanol gave the first analogue, the simple aniline 13 (JD105), namely 11β-(4-aminophenyloxy) estradiol. For nearly all of the other analogues, the crude aniline 12 was not isolated but rather treated directly with an acid chloride. The analogues having a three-atom linker between the aryl ring and the basic amine were all prepared by the same route. Thus, treatment of 12 with chloroacetyl chloride and catalytic DMAP in triethylamine aﬀorded the intermediate chlor-oacetamide, which was immediately reacted with one of four secondary amines, e.g., piperidine, pyrrolidine, morpholine, and dimethylamine,
to give the amides.
Again hydrogenolysis of the benzyl ethers using hydrogen and a palladium catalyst gave the desired analogues, 14a-d (JD101-JD104). After coupling of 12 with the acid chloride to give the amide, hydride reduction aﬀorded the 2-(dialkylamino)ethyl amines, the benzyl ethers of which were hydrogenolyzed to give another set of analogues 15a-d, namely the N-(2-aminoethyl)anilines. In addition the 4-amino group was completely removed to give the simple 11β-phenyl ether 16.
The next set of analogues each had a 3-carbon chain between the aniline and the secondary amine (see Fig. 3). Thus treatment of the crude aniline 12 with 3-chloropropionyl chloride furnished the 3-chloropropanamide and displacement of the chloride with the sec-ondary amines and subsequent hydrogenolysis aﬀorded the analogues with a 5-atom side chain ending in the basic amine, 17a-d (JD106-109).
Likewise using 4-chlorobutanoyl chloride, after displacement of the chloride with the secondary amines and subsequent hydrogenolysis, one obtained the analogues with a 6-atom side chain ending in the basic amine, 18a-d (JD110-112, JD116). Finally, following the same route starting with 5-chloropentanoyl chloride gave the analogues with a 7-atom side chain, 19a-d. Again after coupling of 12 with the 3-carbon acid chloride to give the amide, hydride reduction aﬀorded the 2-(dialkylamino)ethyl amines, the benzyl ethers of which were hydro-genolyzed to give another set of analogues 20a-d, namely the N-(3-aminopropyl) anilines. By substituting the 4-fluoronitrobenzene unit for other aryl fluorides, one could prepare several other sets of analogues (see Fig. 4). Thus, alkylation of the 11β-alcohol 9 with 2, 4-di-fluoronitro-benzene led to the 3-fluoro-4-nitrophenyl ether (which after hydrogenolysis gave the analogue 21). From that compound were prepared the 16 analogues, 23a-d, 24a-d, 25a-d, and 26a-d and the unsubstituted aniline 22. In a similar manner, using 4-fluoro-2-(tri-fluoromethyl) nitrobenzene to alkylate the anion of 12 resulted in the 3-trifluoromethyl-4-nitrophenyl ether (which after hydrogenolysis gave the analogue 27) and thus the 16 additional analogues, 29a-d, 30a-d, 31a-d, and 32a-d and the unsubstituted aniline 28.