Inhibition of the enzyme catechol O-methyltransferase (COMT) represents a viable strategy for regulation of the catabolism of catecholamine neurotransmitters or their precursors, and is of considerable interest in the therapy of Parkinsons disease. Herein, we report the development of a new generation of potent bisubstrate inhibitors of COMT derived from nitro-substituted ligand 1 (K i = 28 nM, Table 1), which achieve high biological activity despite the lack of a NO 2 substituent on the catechol moiety. Their synthesis takes advantage of a convergent approach, in which a series of functionalized catechol intermediates is prepared (Schemes 2 -7) and coupled to a common adenosine-derived allylic amine building block (Scheme 8). Biological activities of the newly synthesized inhibitors, determined by in vitro enzymatic assay and kinetic studies, clearly demonstrate that high inhibitory potency of the bisubstrate inhibitors is not correlated with the pK a of the catechol OH groups. Aromatic residues, connected to the catechol via a biaryl-type linkage, were found to maximally benefit from additional favorable hydrophobic interactions with the enzyme and thus to be preferred replacements of the NO 2 group in 1. A competitive kinetic inhibition mechanism (Fig. 2) with respect to the cofactor binding site was confirmed in all cases, supporting a bisubstrate inhibition mode for inhibitors 2 -19.
nation with MeOLi/MeOH and careful drying of the resulting lithium carboxylate under high vacuum, followed by Br/Li exchange (t-BuLi/THF) and subsequent quenching with the appropriate electrophile, gave rise to acylated building blocks 29 and 32, and secondary alcohol 45 in moderate yields (25 -38%; Scheme 3). Intermediate 45 was easily oxidized to give (pyridin-4-yl)carbonyl derivative 30 by treatment with ortho-iodoxybenzoic acid (IBX; 95%). Alternatively, deprotonation of 25 with LiH/ THF, followed by Br/Li exchange (t-BuLi/THF), avoided the change of the solvent medium, furnishing carbamoyl derivative 37 upon quenching with N,N-dimethylcarbamoyl chloride (21%). Interestingly, attempts to generate the respective dianionic intermediate by treatment of 25 with 2 equiv. or an excess of alkyllithium reagent led to exclusive formation of the corresponding 5-unfunctionalized catechol 38 (replacement of Br by H) upon reaction with electrophiles and aqueous workup.
Starting from intermediate 39, a variety of catechol building blocks could be accessed using Pd 0 -catalyzed cross-coupling reactions. Thus, Suzuki cross-coupling of 39 with (4-fluorophenyl)-or (4-methylphenyl)boronic acid furnished the corresponding biaryl carboxylates 46 and 47 (78 and 79%, resp.; Scheme 4), which, after hydrolysis, provided carboxylic acids 21 and 22 in high yields (89 and 93%, resp.). Heterocyclic and benzylic residues could be introduced by one-pot Pd 0 -catalyzed conversion of Br derivative 39 to the corresponding boronate, followed by Suzuki cross-coupling with the desired aryl or benzyl (Bn) bromide to give biaryls 48 -50 and 5-Bn carboxylate 51 (40 -80% overall). Again, the methyl esters of these intermediates were subjected to basic hydrolysis to furnish the corresponding carboxylic acids 23, 24, 26, and 35 (92 -99%). Furthermore, (N,N-dimethylcarbamoyl)ethenyl-substituted intermediate 52 was obtained by Heck cross-coupling of 39 with N,N-dimethylacrylamide, using Ph 3 A C H T U N G T R E N N U N G P as Pd ligand (44%). Addition of Bu 4 A C H T U N G T R E N N U N G NBr to the reaction mixture significantly accelerated the conversion of 39 to 52. Subsequent hydrolysis provided carboxylic acid 33 (77%).
Cross-coupling methodology was also used for the synthesis of 5-CN-functionalized catechol building blocks 27 and 53 starting from 5-Br intermediates 39 and 54, respectively [9d]. In this case, protection of the catechol OH groups as the corresponding bis(4-methoxyphenyl)methyldioxy moieties was introduced to improve the outcome Scheme 2. Synthesis of Catechol Building Blocks 25 and 31 a) SOCl 2 , MeOH, D, 12 h; 80 -88%. b) Ph 2 CCl 2 (neat), 1608, 40 min; 52 -76%. c) LiOH, THF/H 2 O 1 : 1, D, 4 h; 82 -91%. Scheme 4. Synthesis of Catechol Building Blocks 21 -24, 26, 27, 33, 35, and 53 a) (4-Fluorophenyl)-or (4-methylphenyl)boronic acid, [Pd(PPh 3 ) 4 ], Na 2 CO 3 , benzene/EtOH/H 2 O, D, 12 h; 78 -79%. b) LiOH, THF/H 2 O 1:1, D, 4 h; 77 -99%. c) KCN, [Pd(PPh 3 ) 4 ], 18-crown-6, toluene/ DMF 3 : 1, 1008, 16 h; 48 -76%. d) LiOH, MeOH, 208, 16 h; 92 -93%. e) N,N-Dimethylacrylamide, Pd(OAc) 2 , Na 2 CO 3 , Bu 4 A C H T U N G T R E N N U N G NBr, P(OPh) 3 , N,N-dimethylacetamide (DMA), 1408, 72 h; 44%. f) Pinacol diboron, [Pd(PPh 3 ) 4 ], AcOK, toluene, D; 4 h. g) Aryl or benzyl bromide, [Pd(PPh 3 ) 4 ], Na 2 CO 3 , toluene/EtOH/H 2 O, D, 16 h; 40 -80% (2 steps).
General Procedure 4 (GP 4) for the Synthesis of Functionalized Catechol-carboxylic Acids Starting from 6-Bromo-2,2-diphenyl-1,3-benzodioxole-4-carboxylic Acid (25). Deprotonation Method A. To a suspension of 25 (1 equiv.) in MeOH (3 ml), MeOLi (2 equiv.) was added, and the now clear soln. was stirred 20 min at 208. Volatiles were removed in vacuo, and the resulting white foam was dried overnight under high vacuum (ca. 10 À6 Torr). The residue was redissolved in dry THF (10 ml) and cooled to À 788.
Deprotonation Method B. To a soln. of 25 (1 equiv.) in dry THF (8 ml), LiH (2 equiv.) was added. The mixture was stirred 15 min at 208, and then cooled to À 788.
Br/Li Exchange and Quenching with Electrophile. t-BuLi (2.5 equiv.) was added dropwise to the soln. via a syringe, and the resulting dark yellow mixture was stirred 30 min at À 788. The desired electrophile was added, and stirring was continued for 30 min at À 788. The cooling bath was removed, and the mixture was stirred another 2 h at 208, followed by acidification with 10% aq. AcOH soln. and extraction with AcOEt (2 30 ml). The org. fractions were combined, dried (MgSO 4 ), and evaporated in vacuo.
The residue was purified using CC (SiO 2 ; hexane/AcOEt/AcOH) to yield the desired compound as a yellowish to colorless solid.
General Procedure 5 (GP 5) for the Synthesis of (Diarylmethyldioxy)-catechol-carboxylates via Suzuki Reaction. Method A. To a soln. of 39 (1 equiv.) and [Pd(PPh 3 ) 4 ] (0.05 equiv.) in toluene (10 ml), a soln. of the desired arylboronic acid (4 equiv.) in EtOH (1.5 ml), and a soln. of K 2 CO 3 (6 equiv.) in H 2 O (1 ml) were added. This mixture was heated to reflux for 4 h. After cooling to 208, the mixture was partitioned between AcOEt and H 2 O. The org. phase was washed twice with sat. aq. NaCl soln., dried (MgSO 4 ), and evaporated in vacuo. The residue was purified by CC (SiO 2 ; hexane/AcOEt 20 : 1 ! 5 : 1) to afford the desired compound as a colorless solid.
Method B. To a soln. of 39 (1 equiv.) in dry toluene (20 ml), [Pd(PPh 3 ) 4 ] (0.05 equiv.), bis(pinacolato)diboron (1.3 equiv.), and AcOK (1.5 equiv.) were added, and the mixture was heated to reflux for 4 h. After cooling to 208, the mixture was filtered over Celite. [Pd(PPh 3 ) 4 ] (0.05 equiv.), the desired aryl or benzyl bromide (1.2 equiv.), and a soln. of K 2 CO 3 (5 equiv.) in H 2 O (3 ml) were added to the yellowish soln., and the mixture was heated to reflux for 16 h. After cooling to 208, the mixture was partitioned between H 2 O and AcOEt, and the org. phase was washed twice with sat. aq. NaCl soln. (20 ml), dried (MgSO 4 ), and evaporated under reduced pressure. The residue was purified by CC (SiO 2 ; hexane/ AcOEt 20 : 1 ! 9 : 1) to yield the desired compound as a colorless solid.
General Procedure 6 (GP 6) for the Amide Coupling of Catechol-carboxylic Acid Building Blocks with Primary Amine 20. To a soln. of the corresponding carboxylic acid building block (1 equiv.) in CH 2 Cl 2 (5 ml), EDC•HCl (1.5 equiv.) and N-hydroxysuccinimide (1.3 equiv.) were added, and the mixture was stirred 1 h at 208. After the addition of amine 20 (0.7 -1 equiv.) and Et 3 A C H T U N G T R E N N U N G N (0.1 ml), stirring was continued for 16 h at 208. The mixture was partitioned between CH 2 Cl 2 and H 2 O, and the org. phase was washed with sat. aq. NaCl soln., dried (MgSO 4 ), and evaporated in vacuo. The residue was purified by CC (SiO 2 ; CH 2 Cl 2 /MeOH 20 : 1) to yield the desired compound as a colorless foam.
General Procedure 7 (GP 7) for the Acid-Catalyzed Deprotection of Acetonide and Diarylmethylketal Protecting Groups. The protected precursor was treated with a mixture of TFA/H 2 O 1 : 1 (3 ml) at 08 for 20 -60 min. Volatiles were removed under high vacuum at ambient temp. The residue was redissolved in Me 2 A C H T U N G T R E N N U N G SO (2 -3 ml) and purified by prep. RP-HPLC. The combined product fractions were evaporated to dryness by lyophilization.
Methyl 6-Bromo-2,2-diphenyl-1,3-benzodioxole-4-carboxylate (39). Compound 41 (3 g, 12.87 mmol) [20] was reacted with dichloro(diphenyl)methane (3.7 ml, 4.58 g, 19.31 mmol) according to GP 2.1, Workup Method A, to afford 39 (4 g, 76%). Colorless solid. M.p. 146 -1488. IR (KBr): 3079w, 2950w, 1718s, 1467s, 1355s, 1238s, 1204s, 1043s, 1013s, 944m, 867m, 780s. 1 H-NMR (300 MHz, CDCl 3 ): 3.94 (s, 3 H); 7.14 (d, J = 1.9, 1 H); 7. 38 -7.41 (m, 6 H); 7.55 -7.59 (m, 5 H). 13 C-NMR (75 MHz, CDCl 3 ): 52.4; 112.6; 113.7; 115.6; 119.1; 125.0; 126.2; 128.3; 129.4; 139.0; 147.5; 149.1;