Antagonists of cannabinoid receptor 1 (CB1) have potential for the treatment of several diseases such as obesity, liver disease and diabetes. Recently, development of several CB1 antagonists was halted due to adverse central nervous system (CNS) related side effects observed with rimonabant, the first clinically approved CB1 inverse agonist. However, recent studies indicate that regulation of peripherally expressed CB1 with CNS-sparing compounds is a viable strategy to treat several important disorders. Our efforts aimed at rationally designing peripherally restricted CB1 antagonists have resulted in compounds that have limited blood-brain barrier (BBB) permeability and CNS exposure in preclinical in vitro and in vivo models. Typically, compounds with high topological polar surface areas (TPSAs) do not cross the BBB passively. Compounds with TPSAs higher than rimonabant (rimonabant TPSA = 50) and excellent functional activity with limited CNS penetration were identified. These compounds will serve as templates for further optimization.

In addition to its ability to rapidly alter metabolism, insulin is also able to regulate the expression of numerous genes via activation of the PI3-kinase (PI3-K), MAPK kinase (MEK)-ERK, or p38 pathways. Using differential screening of H4IIE cells, we have identified two members of the Egr zinc-finger transcription factor family of early response genes, Egr-1 and Krox20, whose transcription is induced by insulin treatment. Egr-1 may be involved in insulin's regulation of hepatic gene expression. Krox20 regulation and expression have been primarily studied in neural cells and tissues, but little has been previously reported on the presence of Krox20 in cells of hepatic origin or its regulation by insulin. In the present studies, insulin treatment rapidly increased transcription of both Egr-1 and Krox20. In cells pretreated with a PI3-K inhibitor, there was no reduction in the effect of insulin on Egr-1 and Krox20, but an increase in Egr-1 transcription. The rapid induction of ERK1/2 phosphorylation was completely blocked by pretreatment with a MEK1 inhibitor and was associated with a nearly complete inhibition of insulin-stimulated induction of both Egr-1and Krox20, indicating this pathway is necessary for insulin's effect on these genes. Finally, inhibition of the p38 pathway, followed by insulin addition, caused an additive induction of both Egr-1and Krox20. In conclusion, these genes are induced by insulin via coordinated regulation of the MEK-ERK and p38 pathways and, in the case of Egr-1, the PI3-K pathway.

Cannabinoid receptor 1 (CB1) antagonists are potentially useful for the treatment of several diseases. However, clinical development of several CB1 antagonists was halted due to central nervous system (CNS)-related side effects including depression and suicidal ideation in some users. Recently, studies have indicated that selective regulation of CB1 receptors in the periphery is a viable strategy for treating several important disorders. Past efforts to develop peripherally selective antagonists of CB1 have largely targeted rimonabant, an inverse agonist of CB1. Reported here are our efforts toward developing a peripherally selective CB1 antagonist based on the otenabant scaffold. Even though otenabant penetrates the CNS, it is unique among CB1 antagonists that have been clinically tested because it has properties that are normally associated with peripherally selective compounds. Our efforts have resulted in an orally absorbed compound that is a potent and selective CB1 antagonist with limited penetration into the CNS.

CB1 receptor antagonists that are peripherally restricted were targeted. Compounds with permanent charge as well as compounds that have increased polar surface area were made and tested against CB1 for binding and activity. Sulfonamide and sulfamide with high polar surface area and good activity at CB1 were rationally designed and pharmacologically tested. Further optimization of these compounds and testing could lead to the development of a new class of therapeutics to treat disorders where the CB1 receptor system has been implicated.

Insulin directly affects many aspects of cellular metabolism. An additional, poorly studied effect of insulin is the regulation of multiple genes whose products are important in many cellular functions. Using differential screening techniques, we identified insulin-regulated genes induced in insulin-treated rat H4IIE (H4) hepatoma cells. Two of the mRNAs identified were homologous to the previously characterized mouse pip 92 and rat CL-6 immediate-early genes. The pip 92 clone was initially isolated from mitogen-stimulated mouse Balb/c 3T3 fibroblasts, whereas the CL-6 clone was first isolated from regenerating rat liver. In this article, we demonstrated that in rat H4 cells, the transcription rates of both pip 92 and CL-6 are induced by insulin alone. Additionally, we showed that the transcription rates of two other genes, whose sequences are not homologous to any other sequences in gene bank ("novel genes"), were rapidly and transiently induced by insulin. These results demonstrate that insulin regulates the expression of several novel genes with a time-course similar to members of the immediate-early response gene family.

In the present work, insulin's regulation of expression of activating transcription factor 3 (ATF-3), the putative transcription factor proline-rich induced protein (Pip)92, and insulin-inducible gene-1 (Insig-1) (an ER resident protein involved in regulation of sterol-responsive element-binding protein 1 activation) have been examined in a liver-derived cell line (rat H4IIE hepatoma cells). We report that: 1) insulin-induced transcription of ATF-3, Pip92, and Insig-1 required MEK-ERK activation; 2) insulin-induced transcription of ATF-3 and Pip92 reached maximum levels within 15 min and was blocked by wortmannin but not LY294002; 3) in contrast, the maximum level of insulin-induced transcription of Insig-1 was delayed and was not blocked by either wortmannin or LY294002; 4) insulin activated ERK1/2 in two distinct phases, a rapid peak and a later plateau; 5) the delayed plateau phase of insulin-induced ERK1/2 activation was partially phosphatidylinositol 3-OH-kinase dependent; and 6) however, the rapid, insulin-induced peak of ERK1/2 activation was blocked by wortmannin but not LY294002.

Reduced circulating apelin peptides may be associated with preeclampsia. The apelinergic system should be further investigated to elucidate its role in preclampsia and other hypertensive maternal disorders.

Antagonists of the CB1 receptor can be useful in the treatment of several diseases including obesity, diabetes, and liver disease. However, to date, the only clinically approved CB1 receptor antagonist, rimonabant, was withdrawn due to adverse CNS related side effects such as depression and suicidal ideation. Since rimonabant’s withdrawal, several groups have begun pursuing peripherally selective CB1 antagonists. These compounds are expected to be devoid of undesirable CNS related effects but maintain efficacy through antagonism of peripherally expressed CB1 receptors within target tissues. Reported here are our latest results toward development of a peripherally selective analog of the diphenyl purine CB1 antagonist otenabant 1. Compound 9 (N-{1-[8-(2-Chlorophenyl)-9-(4-chlorophenyl)-9H-purin-6-yl]piperidin-4-yl}pentanamide) is a potent, orally absorbed antagonist of the CB1 receptor that is >50-fold selective for CB1 over CB2, highly selective for the periphery in a rodent model, and without efficacy in a series of in vivo assays designed to evaluate its ability to mitigate the central effects of Δ9-THC through the CB1 receptor.