Pyridine N-oxides were converted to 2-aminopyridines in a one-pot fashion using Ts2O-t-BuNH2 followed by in situ deprotection with TFA. The amination proceeded in high yields, excellent 2-/4-selectivity, and with good functional group compatibility. 2-Amino (iso)quinolines were also obtained in the same manner. Combined with the simple oxidation of pyridines to pyridine N-oxides, this method provides a general and efficient way for amination of 2-unsubstituted pyridines.

No abstract

Lipophilicity has a dominant effect on many parameters that determine unbound drug exposure as well as drug potency. Despite this, analysis of a large body of drug data indicates lipophilicity has no consistent directional impact on dose. This can be rationalized based on the interplay of the effects of lipophilicity on individual parameter values in pharmacokinetic equations. We believe this undermines the effectiveness of strategies that target specific ranges for drug parameters for which lipophilicity plays such a dominant role. As a result, our research organization no longer leverages the common approach of screening for low intrinsic clearance in vitro to target high unbound exposure in vivo. Instead, we advocate for approaches less biased to lipophilicity through optimization of key parameter ratios controlling dose. We believe this improves efficiency in drug discovery by enabling exploration of broad physicochemical space.

Integrins alpha9beta1 and alpha4beta1 form a distinct structural class, but while alpha4beta1 has been subjected to extensive study, alpha9beta1 remains poorly characterized. We have used the small molecule N-(benzenesulfonyl)-(L)-prolyl-(L)-O-(1-pyrrolidinylcarbonyl)tyrosine (3) to investigate the biochemical properties of alpha9beta1 and directly compare these properties with those of alpha4beta1. Compound 3 has a high affinity for both integrins with K(D) values of < or =3 and 180 pM for alpha9beta1 in 1 mM Mn2+ (activating) and 1 mM Ca2+ and 1 mM Mg2+ (nonactivating) conditions and < or =5 and 730 pM for alpha4beta1 under the corresponding conditions. Ca2+ treatment promoted the binding of 3 to both integrins (EC50 = 30 microM Ca2+ in both cases). Compound 3 binding to both integrins was also stimulated by the addition of the activating monoclonal antibody TS2/16. These findings indicate that the mechanisms by which metal ions and TS2/16 regulate ligand binding to alpha9beta1 and alpha4beta1 are similar. The binding of 3 to both integrins induced the mAb 9EG7 LIBS epitope, a property consistent with occupancy of the receptor's ligand binding site by 3. But whereas EGTA treatment inhibited the binding of 9EG7 to alpha4beta1, it stimulated the binding of 9EG7 to alpha9beta1. The 9EG7 and TS2/16 effects point to contributions of the beta1-chains on binding. Cross-linking data revealed that the integrin alpha-chains are also involved in binding the small molecule, as stable linkages were observed on both the alpha9 chain of alpha9beta1 and the alpha4 chain of alpha4beta1. Extensive structure-activity analyses with natural and synthetic ligands indicate distinct features of the ligand binding pockets. Most notable was the estimated >1000-fold difference in the affinity of the integrins for VCAM-1, which binds alpha4beta1with an apparent K(D) of 10 nM and alpha9beta1 with an apparent K(D) of >10 microM. Differences were also seen in the binding of alpha9beta1 and alpha4beta1 to osteopontin. Compound 3 competed effectively for the binding of VCAM-1 and osteopontin to both integrins. While these studies show many similarities in the biochemical properties of alpha9beta1 and alpha4beta1, they identify important differences in their structure and function that can be exploited in the design of selective alpha9beta1 and alpha4beta1 inhibitors.

This article is available online at http://dmd.aspetjournals.org ABSTRACT:Zileuton, a 5-lipoxygenase inhibitor, was evaluated as an inhibitor of cytochrome P450 activity in human liver microsomes. In the absence of preincubation, the racemate was found to be a weak inhibitor (IC 50 > 100 M) of phenacetin O-deethylation (POD) (CYP1A2), paclitaxel 6␣-hydroxylation (CYP2C8), diclofenac 4-hydroxylation (CYP2C9), (S)-mephenytoin 4-hydroxylation (CYP2C19), bufuralol 1-hydroxylation (CYP2D6), testosterone 6␤-hydroxylation (CYP3A4), chlorzoxazone 6-hydroxylation (CYP2E1), and bupropion hydroxylation (CYP2B6). When preincubated with NADPH-fortified human liver microsomes in the absence of substrate, zileuton (racemate) was shown to inhibit POD. The effect was NADPH-, time-, and concentration-dependent, and was characterized by a k inact (maximal rate of enzyme inactivation) and apparent K I (inhibitor concentration that supports half the maximal rate of inactivation) of 0.035 min ]. In addition, the inhibition of CYP1A2 was not reversed in the presence of reduced glutathione, catalase, and superoxide dismutase and was refractory to dialysis. Therefore, zileuton was characterized as a mechanism-based inhibitor of human liver microsomal CYP1A2. Mechanism-based inhibition of CYP1A2 may explain why zileuton decreases the oral clearance of antipyrine, propranolol, (R)-warfarin, and theophylline, at doses that have a minimal effect on the pharmacokinetics of (S)-warfarin, phenytoin, and terfenadine. Zileuton (Zyflo;thien-2-ylethyl)-N-hydroxyurea), a substituted hydroxamic acid, is a potent and selective 5-lipoxygenase inhibitor that has been approved for the prevention and chronic treatment of asthma (Isreal et al., 1990;Carter et al., 1991;Bell et al., 1992;Wenzel and Kamada, 1996;Dube et al., 1999). The drug contains a single chiral center (Fig. 1), is administered as a racemic mixture of (R)-(ϩ)-and (S)-(Ϫ)-enantiomers, and is characterized by a plasma half-life of about 4 h (Wong et al., 1995). As a result of the short half-life, zileuton has to be dosed four times a day (q.i.d.), and effective inhibition (70 -80%) of leukotriene B 4 biosynthesis can be attained using a 600-mg q.i.d. dosing regimen (Awni et al., 1995a).Zileuton is well absorbed, and the majority of the dose (ϳ75%) is recovered in the urine of healthy volunteers as the N-hydroxy glucuronides of both the (R)-(ϩ)-and (S)-(Ϫ)-enantiomers (Wong et al., 1995). Although N-hydroxy glucuronidation has been confirmed with human liver microsomes, zileuton is metabolized in the presence of NADPH (Machinist et al., 1995b;Sweeny and Nellans, 1995). In fact, CYP1A2 and CYP2C9 catalyze ring hydroxylation, whereas sulfoxidation is catalyzed largely by CYP3A4. Both reactions conform to Michaelis-Menten kinetics in human liver microsomes and are described by relatively high K m values (Ն 0.2 mM).Zileuton also has been shown to be a weak inhibitor of human liver microsomal CYP3A4, CYP2C9, and CYP2D6 activity in vitro (Machinist et al., 1995b). As anticipated, zileuton has a m...