We recently reported the discovery of AM-8553 (1), a potent and selective piperidinone inhibitor of the MDM2-p53 interaction. Continued research investigation of the N-alkyl substituent of this series, focused in particular on a previously underutilized interaction in a shallow cleft on the MDM2 surface, led to the discovery of a one-carbon tethered sulfone which gave rise to substantial improvements in biochemical and cellular potency. Further investigation produced AMG 232 (2), which is currently being evaluated in human clinical trials for the treatment of cancer. Compound 2 is an extremely potent MDM2 inhibitor (SPR KD = 0.045 nM, SJSA-1 EdU IC50 = 9.1 nM), with remarkable pharmacokinetic properties and in vivo antitumor activity in the SJSA-1 osteosarcoma xenograft model (ED50 = 9.1 mg/kg).

FFA2 (GPR43) has been identified as a receptor for short-chain fatty acids (SCFAs) that include acetate and propionate. FFA2 is highly expressed in islets, a subset of immune cells, and adipocytes. Although the potential roles of FFA2 activation in these tissues have previously been described, the physiological functions are still unclear. The potency for SCFAs on FFA2 is low, in the high micromolar to millimolar concentrations. To identify better pharmacological tools to study receptor function, we used high-throughput screening (HTS) to discover a series of small molecule phenylacetamides as novel and more potent FFA2 agonists. This series is specific for FFA2 over FFA1 (GPR40) and FFA3 (GPR41), and it is able to activate both the G␣ q and G␣ i pathways in vitro on Chinese hamster ovary cells stably expressing FFA2. Treatment of adipocytes with these compounds also resulted in G␣ i -dependent inhibition of lipolysis similar to that of endogenous ligands (SCFAs). It is noteworthy that these compounds not only acted as FFA2 agonists but also exhibited positive cooperativity with acetate or propionate. The observed allosteric modulation was consistent in all the functional assays that we have explored, including cAMP, calcium mobilization, guanosine 5Ј-[␥-thio]triphosphate binding, and lipolysis. Molecular modeling analysis of FFA2 based on human ␤ 2 -adrenergic receptor structure revealed potential nonoverlapping binding sites for the endogenous and synthetic ligands, further providing insight into the binding pocket for the allosteric interactions. This is the first report describing the identification of novel allosteric modulators with agonist activity for FFA2, and these compounds may serve as tools for further unraveling the physiological functions of the receptor and its involvement in various diseases.

Structure-based rational design led to the discovery of novel inhibitors of the MDM2-p53 protein-protein interaction. The affinity of these compounds for MDM2 was improved through conformational control of both the piperidinone ring and the appended N-alkyl substituent. Optimization afforded 29 (AM-8553), a potent and selective MDM2 inhibitor with excellent pharmacokinetic properties and in vivo efficacy.

A novel nortriterpene, termed correolide, purified from the tree Spachea correae, inhibits Kv1.3, a Shaker-type delayed rectifier potassium channel present in human T lymphocytes. Correolide inhibits 86Rb+ efflux through Kv1.3 channels expressed in CHO cells (IC50 86 nM; Hill coefficient 1) and displays a defined structure-activity relationship. Potency in this assay increases with preincubation time and with time after channel opening. Correolide displays marked selectivity against numerous receptors and voltage- and ligand-gated ion channels. Although correolide is most potent as a Kv1.3 inhibitor, it blocks all other members of the Kv1 family with 4-14-fold lower potency. C20-29-[3H]dihydrocorreolide (diTC) was prepared and shown to bind in a specific, saturable, and reversible fashion (Kd = 11 nM) to a single class of sites in membranes prepared from CHO/Kv1.3 cells. The molecular pharmacology and stoichiometry of this binding reaction suggest that one diTC site is present per Kv1.3 channel tetramer. This site is allosterically coupled to peptide and potassium binding sites in the pore of the channel. DiTC binding to human brain synaptic membranes identifies channels composed of other Kv1 family members. Correolide depolarizes human T cells to the same extent as peptidyl inhibitors of Kv1.3, suggesting that it is a candidate for development as an immunosuppressant. Correolide is the first potent, small molecule inhibitor of Kv1 series channels to be identified from a natural product source and will be useful as a probe for studying potassium channel structure and the physiological role of such channels in target tissues of interest.

The eukaryotic initiation factor 4E (eIF4E) plays a central role in the initiation of gene translation and subsequent protein synthesis by binding the 5' terminal mRNA cap structure. We designed and synthesized a series of novel compounds that display potent binding affinity against eIF4E despite their lack of a ribose moiety, phosphate, and positive charge as present in m7-GMP. The biochemical activity of compound 33 is 95 nM for eIF4E in an SPA binding assay. More importantly, the compound has an IC(50) of 2.5 μM for inhibiting cap-dependent mRNA translation in a rabbit reticular cell extract assay (RRL-IVT). This series of potent, truncated analogues could serve as a promising new starting point toward the design of neutral eIF4E inhibitors with physicochemical properties suitable for cellular activity assessment.