Narcolepsy type 1 (NT1) is a chronic neurological disorder that impairs the brain’s ability to control sleep-wake cycles. Current therapies are limited to the management of symptoms with modest effectiveness and substantial adverse effects. Agonists of the orexin receptor 2 (OX2R) have shown promise as novel therapeutics that directly target the pathophysiology of the disease. However, identification of drug-like OX2R agonists has proven difficult. Here we report cryo-electron microscopy structures of active-state OX2R bound to an endogenous peptide agonist and a small-molecule agonist. The extended carboxy-terminal segment of the peptide reaches into the core of OX2R to stabilize an active conformation, while the small-molecule agonist binds deep inside the orthosteric pocket, making similar key interactions. Comparison with antagonist-bound OX2R suggests a molecular mechanism that rationalizes both receptor activation and inhibition. Our results enable structure-based discovery of therapeutic orexin agonists for the treatment of NT1 and other hypersomnia disorders.

As part of an effort to examine the utility of antibody-drug conjugates (ADCs) beyond oncology indications, a novel pyrophosphate ester linker was discovered to enable the targeted delivery of glucocorticoids. As small molecules, these highly soluble phosphate ester drug linkers were found to have ideal orthogonal properties: robust plasma stability coupled with rapid release of payload in a lysosomal environment. Building upon these findings, site-specific ADCs were made between this drug linker combination and an antibody against human CD70, a receptor specifically expressed in immune cells but also found aberrantly expressed in multiple human carcinomas. Full characterization of these ADCs enabled procession to in vitro proof of concept, wherein ADCs 1-22 and 1-37 were demonstrated to afford potent, targeted delivery of glucocorticoids to a representative cell line, as measured by changes in glucocorticoid receptor-mediated gene mRNA levels. These activities were found to be antibody-, linker-, and payload-dependent. Preliminary mechanistic studies support the notion that lysosomal trafficking and enzymatic linker cleavage are required for activity and that the utility for the pyrophosphate linker may be general for internalizing ADCs as well as other targeted delivery platforms.

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We have continued to explore the 3,3-dialkyl-5-aryloxindole series of progesterone receptor (PR) modulators looking for new agents to be used in female healthcare: contraception, fibroids, endometriosis, and certain breast cancers. Previously we reported that subtle structural changes with this and related templates produced functional switches between agonist and antagonist properties ( Fensome et al. Biorg. Med. Chem. Lett. 2002, 12, 3487; 2003, 13, 1317 ). We herein report a new functional switch within the 5-(2-oxoindolin-5-yl)-1 H-pyrrole-2-carbonitrile class of compounds. We found that the size of the 3,3-dialkyl substituent is important for controlling the functional response; thus small groups (dimethyl) afford potent PR antagonists, whereas larger groups (spirocyclohexyl) are PR agonists. The product from our optimization activities in cell-based systems and also for kinetic properties in rodents and nonhuman primates was 5-(7-fluoro-3,3-dimethyl-2-oxo-2,3-dihydro-1 H-indol-5-yl)-1-methyl-1 H-pyrrole-2-carbonitrile 27 (WAY-255348), which demonstrated potent and robust activity on PR antagonist and contraceptive end points in the rat and also in cynomolgus and rhesus monkeys including ovulation inhibition, menses induction, and reproductive tract morphology.

In an effort to examine the utility of antibody-drug conjugates (ADCs) beyond oncology indications, a novel phosphate bridged Cathepsin B sensitive linker was developed to enable the targeted delivery of glucocorticoids. Phosphate bridging of the Cathepsin B sensitive linkers allows for payload attachment at an aliphatic alcohol. As small molecule drug-linkers, these aqueous soluble phosphate containing drug-linkers were found to have robust plasma stability coupled with rapid release of payload in a lysosomal environment. Site-specific ADCs were successfully made between these drug-linkers and an antibody against human CD70, a receptor specifically expressed in immune cells but also found aberrantly expressed in multiple human carcinomas. These ADCs demonstrated in vitro targeted delivery of glucocorticoids to a representative cell line as measured by changes in glucocorticoid receptor (GR) mediated gene mRNA levels. This novel linker expands the scope of potential ADC payloads by allowing an aliphatic alcohol to be a stable, yet cleavable attachment site. This phosphate linker may have broad utility for internalizing ADCs as well as other targeted delivery platforms.

Glucocorticoids (GCs) are excellent anti-inflammatory drugs but are dose-limited by on-target toxicity. We sought to solve this problem by delivering GCs to immune cells with antibody-drug conjugates (ADCs) using antibodies containing site-specific incorporation of a non-natural amino acid, novel linker chemistry for in vitro and in vivo stability, and existing and novel glucocorticoid receptor (GR) agonists as payloads. We directed fluticasone propionate to human antigen-presenting immune cells to afford GR activation that was dependent on the targeted antigen. However, mechanism of action studies pointed to accumulation of free payload in the tissue culture supernatant as the dominant driver of activity and indeed administration of the ADC to human CD74 transgenic mice failed to activate GR target genes in splenic B cells. Suspecting dissipation of released payload, we designed an ADC bearing a novel GR agonist payload with reduced permeability which afforded cell-intrinsic activity in human B cells. Our work shows that antibody-targeting offers significant potential for rescuing existing and new dose-limited drugs outside the field of oncology.

The diphenylsulfonyl sulfonamide scaffold represented by 1 (WAY-316606) are small molecule inhibitors of the secreted protein sFRP-1, an endogenous antagonist of the secreted glycoprotein Wnt. Modulators of the Wnt pathway have been proposed as anabolic agents for the treatment of osteoporosis or other bone-related disorders. Details of the structure-activity relationships and biological activity from the first structural class of this scaffold will be discussed.

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