Synthesis of 4,5,6,7-tetrahydrothieno[3,2-c]pyridines and comparison with their isosteric 1,2,3,4-tetrahydroisoquinolines as inhibitors of phenylethanolamine N-methyltransferase.

A series of substituted 4,5,6,7-tetrahydrothieno[3,2-c]pyridines (THTPs) was synthesized and evaluated for their human phenylethanolamine N-methyltransferase (hPNMT) inhibitory potency and affinity for the alpha(2)-adrenoceptor. The THTP nucleus was suggested as an isosteric replacement for the 1,2,3,4-tetrahydroisoquinoline (THIQ) ring system on the basis that 3-thienylmethylamine (18) was more potent as an inhibitor of hPNMT and more selective toward the alpha(2)-adrenoceptor than benzylamine (15). Although the isosterism was confirmed, with similar influence of functional groups and chirality in both systems on hPNMT inhibitory potency and selectivity, the THTP compounds proved, in general, to be less potent as inhibitors of hPNMT than their THIQ counterparts, with the drop in potency being primarily attributed to the electronic properties of the thiophene ring. A hypothesis for the reduced hPNMT inhibitory potency of these compounds has been formed on the basis of molecular modeling and docking studies using the X-ray crystal structures of hPNMT co-crystallized with THIQ-type inhibitors and S-adenosyl-L-homocysteine as a template.

Exploring the active site of phenylethanolamine N-methyltransferase with 1,2,3,4-tetrahydrobenz[h]isoquinoline inhibitors.

1,2,3,4-Tetrahydrobenz[h]isoquinoline (THBQ, 11) is a potent, inhibitor of phenylethanolamine N-methyltransferase (PNMT). Docking studies indicated that the enhanced PNMT inhibitory potency of 11 (hPNMT K(i)=0.49microM) versus 1,2,3,4-tetrahydroisoquinoline (5, hPNMT K(i)=5.8microM) was likely due to hydrophobic interactions with Val53, Met258, Val272, and Val269 in the PNMT active site. These studies also suggested that the addition of substituents to the 7-position of 11 that are capable of forming hydrogen bonds to the enzyme could lead to compounds (14-18) having enhanced PNMT inhibitory potency. However, these compounds are in fact less potent at PNMT than 11. Furthermore, 7-bromo-THBQ (19, hPNMT K(i)=0.22mM), which has a lipophilic 7-substituent that cannot hydrogen bond to the enzyme, is twice as potent at PNMT than 11. This once again illustrates the limitations of docking studies for lead optimization.

Comparison of the binding of 3-fluoromethyl-7-sulfonyl-1,2,3,4-tetrahydroisoquinolines with their isosteric sulfonamides to the active site of phenylethanolamine N-methyltransferase.

3-Fluoromethyl-7-(N-substituted aminosulfonyl)-1,2,3,4-tetrahydroisoquinolines (14, 16, and 18-22) are highly potent and selective inhibitors of phenylethanolamine N-methyltransferase (PNMT). Molecular modeling studies with 3-fluoromethyl-7-(N-alkyl aminosulfonyl)-1,2,3,4-tetrahydroisoquinolines, such as 16, suggested that the sulfonamide -NH- could form a hydrogen bond with the side chain of Lys57. However, SAR studies and analysis of the crystal structure of human PNMT (hPNMT) in complex with 7 indicated that the sulfonamide oxygens, and not the sulfonamide -NH-, formed favorable interactions with the enzyme. Thus, we hypothesized that replacement of the sulfonamide -NH- with a methylene group could result in compounds that would retain potency at PNMT and that would have increased lipophilicity, thus increasing the likelihood they will cross the blood brain barrier. A series of 3-fluoromethyl-7-sulfonyl-1,2,3,4-tetrahydroisoquinolines (23-30) were synthesized and evaluated for their PNMT inhibitory potency and affinity for the alpha2-adrenoceptor. A comparison of these compounds with their isosteric sulfonamides (14, 16, and 18-22) showed that the sulfones were more lipophilic but less potent than their corresponding sulfonamides. Sulfone 24 (hPNMT Ki = 1.3 microM) is the most potent compound in this series and is quite selective for PNMT versus the alpha2-adrenoceptor, but 24 is less potent than the corresponding sulfonamide, 16 (hPNMT Ki = 0.13 microM). We also report the crystal structure of hPNMT in complex with sulfonamide 15, from which a potential hydrogen bond acceptor within the hPNMT active site has been identified, the main chain carbonyl oxygen of Asn39. The interaction of this residue with the sulfonamide -NH- is likely responsible for much of the enhanced inhibitory potency of the sulfonamides versus the sulfones.

Application of the Goldilocks effect to the design of potent and selective inhibitors of phenylethanolamine N-methyltransferase: balancing pKa and steric effects in the optimization of 3-methyl-1,2,3,4-tetrahydroisoquinoline inhibitors by beta-fluorination.

3-Methyl-1,2,3,4-tetrahydroisoquinolines (3-methyl-THIQs) are potent inhibitors of phenylethanolamine N-methyltransferase (PNMT), but are not selective due to significant affinity for the alpha(2)-adrenoceptor. Fluorination of the methyl group lowers the pK(a) of the THIQ amine from 9.53 (CH(3)) to 7.88 (CH(2)F), 6.42 (CHF(2)), and 4.88 (CF(3)). This decrease in pK(a) results in a reduction in affinity for the alpha(2)-adrenoceptor. However, increased fluorination also results in a reduction in PNMT inhibitory potency, apparently due to steric and electrostatic factors. Biochemical evaluation of a series of 3-fluoromethyl-THIQs and 3-trifluoromethyl-THIQs showed that the former were highly potent inhibitors of PNMT, but were often nonselective due to significant affinity for the alpha(2)-adrenoceptor, while the latter were devoid of alpha(2)-adrenoceptor affinity, but also lost potency at PNMT. 3-Difluoromethyl-7-substituted-THIQs have the proper balance of both steric and pK(a) properties and thus have enhanced selectivity versus the corresponding 3-fluoromethyl-7-substituted-THIQs and enhanced PNMT inhibitory potency versus the corresponding 3-trifluoromethyl-7-substituted-THIQs. Using the "Goldilocks Effect" analogy, the 3-fluoromethyl-THIQs are too potent (too hot) at the alpha(2)-adrenoceptor and the 3-trifluoromethyl-THIQs are not potent enough (too cold) at PNMT, but the 3-difluoromethyl-THIQs are just right. They are both potent inhibitors of PNMT and highly selective due to low affinity for the alpha(2)-adrenoceptor. This seems to be the first successful use of the beta-fluorination of aliphatic amines to impart selectivity to a pharmacological agent while maintaining potency at the site of interest.

Exploring the active site of phenylethanolamine N-methyltransferase with 3-hydroxyethyl- and 3-hydroxypropyl-7-substituted-1,2,3,4-tetrahydroisoquinolines.

3-Hydroxyethyl- and 3-hydroxypropyl-7-substituted-tetrahydroisoquinolines (9, 10, 16, and 17) were synthesized and evaluated for their phenylethanolamine N-methyltransferase (PNMT) inhibitory potency and affinity for the alpha(2)-adrenoceptor. Although alpha(2)-adrenoceptor affinity decreased for these compounds, selectivity was not gained over the parent 3-hydroxymethyl compounds (1, 2) due to a loss in PNMT inhibitory potency.