A general synthetic pathway to these tricyclic structures is

A general synthetic pathway to these tricyclic structures is described in (detailed procedures can be accessed in the patent application WO 2011/014520 A2). Benzylation of the appropriate ketoester using KHMDS, followed by treatment with DMF-dimethyl acetal yielded the racemic (dimethylamino)-methylene ketone. The optimal synthetic pathway for enantiomerically pure analogs proved to be analytical resolution (preparative chiral chromatography) at this stage. The final products were afforded by acid-catalyzed condensation with the appropriate 4-aryl aminopyrazole, which was derived from the corresponding aryl acetonitrile via condensation with ethyl formate and subsequent cyclization with hydrazine. In line with previous observations, the () enantiomer was significantly more active than the () enantiomer . The ethyl and methyl esters had similar activities and properties for a variety of analogs (e.g. vs ). The 3-fluoro substitution on the benzyl group (R=F in ) did not bring about any major changes (entries vs ). Expansion of the carbocyclic ring beyond the 5-membered carbacycle in led to reduced GPR119 activity (entries –). As described in the preceding report, SAR around the position 3 and 7 of the core heterocycle was rather steep, the more polar substituents being particularly not tolerated. Indeed, small changes in the -substituent of the 3-aryl group led to a marked drop in Caspase-8, human recombinant protein australia agonist potency and efficacy (entries –). In an effort to decrease hydrophobicity and increase solubility in the series, we thus focused our attention to the newly formed carbacycle in the tricyclic core. To our surprise, either inserting a heteroatom position 7 of the core heterocycle (entries and ) or next to the quarternary carbon (entries and ) did not result in a complete loss of GPR119 agonist activity. Even the introduction of a physiological-conditions ionizable secondary amine was well tolerated, if not beneficial to GPR119 activity. Please note that due to the heteroatom next to the quarternary chiral center, the absolute stereochemical assignment changes from () to (). Interestingly, methylation to a tertiary amine (entry ) was less tolerated. Compound was labile in the presence of human microsomes (ER=0.95), and to a lesser extent in mouse microsomes (ER=0.61). A metabolic ID study revealed that a major metabolite was formed via oxidation of the CN bond in the 5-membered ring. Hence oxidized analogs and were synthesized and tested for their GPR119 agonist activity. Both compounds had significantly lower activity compared to . Calculated log values indicated a significant 2-log drop going from the carbacycle to the secondary amine . Although there were no precipitation issues observed for these analogs, PK studies in mice showed similarly low oral exposure for amine compared to the carbon analog , indicating that the changes in the 5-membered ring alone were insufficient to positively impact exposures. Next, we focused our attention on the ester functionality (). The ester group was seen as a potential liability, although in vitro plasma stability studies did not indicate an inherent instability of the ester group, probably due to steric crowding. Previous investigations deemed the ester essential for GPR119 activity, although a few amide replacements were identified. Transferring these findings to the tricyclic series such as in analog maintained good GPR119 activity and high plasma stability, but solubilities remained low (<5μM) and extraction ratios remained high (ER=0.9; ER=0.84). Hence, oral exposure in mice did not improve, in fact it was lower (AUC=0.07μM*h) than for the corresponding ester (AUC=0.31μM*h). In addition, we found that the tricyclic series also tolerated small alkyl groups in place of the ester functionality (entries and ). Specifically, the cyclopropyl analog led to an only 10-fold drop in potency (similar on the rodent receptor), but markedly increased metabolic stability in all species (ER∼0.3–0.6).