A selective prostaglandin E2 receptor subtype 2 (EP2) antagonist increases the macrophage-mediated clearance of amyloid-beta plaques.

A high-throughput screen resulted in the discovery of benzoxazepine 1, an EP2 antagonist possessing low microsomal stability and potent CYP3A4 inhibition. Modular optimization of lead compound 1 resulted in the discovery of benzoxazepine 52, a molecule with single-digit nM binding affinity for the EP2 receptor and significantly improved microsomal stability. It was devoid of CYP inhibition and was ∼4000-fold selective against the other EP receptors. Compound 52 was shown to have good PK properties in CD-1 mice and high CNS permeability in C57Bl/6s mice and Sprague-Dawley rats. In an ex vivo assay, it demonstrated the ability to increase the macrophage-mediated clearance of amyloid-beta plaques from brain slices in a dose-dependent manner.

Discovery of AM-7209, a potent and selective 4-amidobenzoic acid inhibitor of the MDM2-p53 interaction.

Structure-based rational design and extensive structure-activity relationship studies led to the discovery of AMG 232 (1), a potent piperidinone inhibitor of the MDM2-p53 association, which is currently being evaluated in human clinical trials for the treatment of cancer. Further modifications of 1, including replacing the carboxylic acid with a 4-amidobenzoic acid, afforded AM-7209 (25), featuring improved potency (KD from ITC competition was 38 pM, SJSA-1 EdU IC50 = 1.6 nM), remarkable pharmacokinetic properties, and in vivo antitumor activity in both the SJSA-1 osteosarcoma xenograft model (ED50 = 2.6 mg/kg QD) and the HCT-116 colorectal carcinoma xenograft model (ED50 = 10 mg/kg QD). In addition, 25 possesses distinct mechanisms of elimination compared to 1.

Novel inhibitors of the MDM2-p53 interaction featuring hydrogen bond acceptors as carboxylic acid isosteres.

We previously reported the discovery of potent and selective morpholinone and piperidinone inhibitors of the MDM2-p53 interaction. These inhibitors have in common a carboxylic acid moiety that engages in an electrostatic interaction with MDM2-His96. Our continued search for potent and diverse inhibitors led to the discovery of novel replacements for these acids uncovering new interactions with the MDM2 protein. In particular, using pyridine or thiazole as isosteres of the carboxylic acid moiety resulted in very potent analogues. From these, AM-6761 (4) emerged as a potent inhibitor with remarkable biochemical (HTRF IC50 = 0.1 nM) and cellular potency (SJSA-1 EdU IC50 = 16 nM), as well as favorable pharmacokinetic properties. Compound 4 also shows excellent antitumor activity in the SJSA-1 osteosarcoma xenograft model with an ED50 of 11 mg/kg. Optimization efforts toward the discovery of these inhibitors as well as the new interactions observed with the MDM2 protein are described herein.

Selective and potent morpholinone inhibitors of the MDM2-p53 protein-protein interaction.

We previously reported the discovery of AMG 232, a highly potent and selective piperidinone inhibitor of the MDM2-p53 interaction. Our continued search for potent and diverse analogues led to the discovery of novel morpholinone MDM2 inhibitors. This change to a morpholinone core has a significant impact on both potency and metabolic stability compared to the piperidinone series. Within this morpholinone series, AM-8735 emerged as an inhibitor with remarkable biochemical potency (HTRF IC50 = 0.4 nM) and cellular potency (SJSA-1 EdU IC50 = 25 nM), as well as pharmacokinetic properties. Compound 4 also shows excellent antitumor activity in the SJSA-1 osteosarcoma xenograft model with an ED50 of 41 mg/kg. Lead optimization toward the discovery of this inhibitor as well as key differences between the morpholinone and the piperidinone series will be described herein.

Discovery of AMG 232, a potent, selective, and orally bioavailable MDM2-p53 inhibitor in clinical development.

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).

Structure-based design of novel inhibitors of the MDM2-p53 interaction.

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.

Novel pyrrolidine melanin-concentrating hormone receptor 1 antagonists with reduced hERG inhibition.

We discovered novel pyrrolidine MCHR1 antagonist 1 possessing moderate potency. Profiling of pyrrolidine 1 demonstrated that it was an inhibitor of the hERG channel. Investigation of the structure-activity relationship of this class of pyrrolidines allowed us to optimize the MCHR1 potency and decrease the hERG inhibition. Increasing the acidity of the amide proton by converting the benzamide in lead 1 to an anilide provided single digit nanomolar MCHR1 antagonists while replacing the dimethoxyphenyl ring of 1 with alkyl groups possessing increased polarity dramatically reduced the hERG inhibition.

Evaluation of indenoisoquinoline topoisomerase I inhibitors using a hollow fiber assay.

The indenoisoquinolines are a novel class of non-camptothecin topoisomerase I (Top1) inhibitors whose mechanism of action involves trapping the covalent complex formed between DNA and Top1 during cellular processes. As an ongoing evaluation of the indenoisoquinolines for Top1 inhibition and anticancer activity, indenoisoquinoline analogs have been screened in the National Cancer Institute's hollow fiber assay (HFA). Some of the derivatives demonstrated significant activity at intraperitoneal and subcutaneous fiber placement sites, along with net cancer cell kill in one or more cell lines.

Design, synthesis, and biological evaluation of cytotoxic 11-alkenylindenoisoquinoline topoisomerase I inhibitors and indenoisoquinoline-camptothecin hybrids.

The indenoisoquinolines are a novel class of topoisomerase I (top1) inhibitors that are cytotoxic in cancer cell cultures and are therefore under development as potential anticancer agents. As inhibitors of the DNA religation reaction occurring after DNA cleavage by the enzyme, they are classified as top1 poisons, similar to the camptothecins. Two strategies were employed in order to further develop the structure-activity relationships of the indenoisoquinolines and enhance their therapeutic potential. The first strategy involved the synthesis of indenoisoquinoline-camptothecin hybrid molecules to take advantage of a proposed structural analogy between the indenoisoquinolines and camptothecin. The desired hybrids were synthesized by reaction of halogenated phthalides with a dihydropyrroloquinoline. The second strategy involved the attachment of various alkenyl substituents to the C-11 position of the indenoisoquinolines, which were assumed to project into the DNA minor groove. The required C-11-substituted indenoisoquinolines were synthesized by McMurry reactions of 11-ketoindenoisoquinolines with aldehydes, and the geometries of the resulting alkenes were established by nuclear Overhauser effect difference NMR spectroscopy. All of the new indenoisoquinolines were examined for cytotoxicity in human cancer cell cultures as well as for activity vs top1. Although the indenoisoquinoline-camptothecin hybrid molecules proved to be less cytotoxic and displayed less activity against top1, an analogue incorporating a 3'-aminoalkenyl substituent at the C-11 position of the indenoisoquinoline system was significantly more potent than the prototype indenoisoquinoline in both assays. These results indicate that C-11 aminoalkyl substituents that are assumed to project into the minor groove enhance the cytotoxicity and top1 inhibitory activity of the parent indenoisoquinoline system.

Synthesis of new dihydroindeno[1,2-c]isoquinoline and indenoisoquinolinium chloride topoisomerase I inhibitors having high in vivo anticancer activity in the hollow fiber animal model.

A number of novel dihydroindenoisoquinolines and indenoisoquinolinium salts were synthesized and examined for cytotoxicity in cancer cell cultures and for inhibition of topoisomerase I (top1). The top1-mediated DNA cleavage patterns produced in the presence of several of the new analogues were also investigated, and a few of the more potent compounds were examined for activity in hollow fiber animal models. Very cytotoxic dihydroindenoisoquinoline and isoquinolinium salts were obtained with mean graph midpoints (MGMs) for growth inhibition in the low submicromolar range. Two of the new dihydroindenoisoquinolines were found to be weaker top1 inhibitors than the lead compound 1, while two of the indenoisoquinolinium salts were more potent. The top1-mediated DNA cleavage patterns of the indenoisoquinolines examined were found to be similar to each other but different from that of camptothecin. Several of the more potent indenoisoquinolines displayed promising anticancer activities in hollow fiber animal models.