Interleukin-1 receptor-associated kinase 3 (IRAK3) is a pseudokinase mediator in the human inflammatory pathway, and ablation of its function is associated with enhanced antitumor immunity. Traditionally, pseudokinases have eluded "druggability" and have not been considered tractable targets in the pharmaceutical industry. Herein we disclose a CRISPR/Cas9-mediated knockout of IRAK3 in monocyte-derived dendritic cells that results in an increase in IL-12 production upon lipopolysaccharide (LPS) stimulation. Furthermore, we disclose and characterize Degradomer D-1, which displays selective proteasomal degradation of IRAK3 and reproduces the 1L-12p40 increases observed in the CRISPR/Cas9 knockout.

We performed a high-throughput phenotypic whole cell screen of Mycobacterium tuberculosis against a diverse chemical library of approximately 100,000 compounds from the AbbVie corporate collection and identified 24 chemotypes with anti-tubercular activity. We selected two series for further exploration and conducted structure–activity relationship studies with new analogs for the 4-phenyl piperidines (4PP) and phenylcyclobutane carboxamides (PCB). Strains with mutations in MmpL3 demonstrated resistance to both compound series. We isolated resistant mutants for the two series and found mutations in MmpL3. These data suggest that MmpL3 is the target, or mechanism of resistance for both series.

There is a significant need for improved treatments for onchocerciasis and lymphatic filariasis, diseases caused by filarial worm infection. In particular, an agent able to selectively kill adult worms (macrofilaricide) would be expected to substantially augment the benefits of mass drug administration (MDA) with current microfilaricides, and to provide a solution to treatment of onchocerciasis / loiasis co-infection, where MDA is restricted. We have identified a novel macrofilaricidal agent, Tylosin A (TylA), which acts by targeting the worm-symbiont Wolbachia bacterium. Chemical modification of TylA leads to improvements in anti-Wolbachia activity and oral pharmacokinetic properties; an optimized analog (ABBV-4083) has been selected for clinical evaluation.

Hepatitis C virus (HCV) nucleoside inhibitors have been a key focus of nearly 2 decades of HCV drug research due to a high barrier to drug resistance and pan-genotypic activity profile provided by molecules in this drug class. Our investigations focused on several potent 2'-halogenated uridine-based HCV polymerase inhibitors, resulting in the discovery of novel 2'-deoxy-2'-dihalo-uridine analogs that are potent inhibitors in replicon assays for all genotypes. Further studies to improve in vivo performance of these nucleoside inhibitors identified aminoisobutyric acid ethyl ester (AIBEE) phosphoramidate prodrugs 18a and 18c, which provide high levels of the active triphosphate in dog liver. AIBEE prodrug 18c was compared with sofosbuvir (1) by co-dosing both compounds by oral administration in dog (5 mg/kg each) and measuring liver concentrations of the active triphosphate metabolite at both 4 and 24 h post dosing. In this study, 18c provided liver triphosphate concentrations that were 6-fold higher than sofosbuvir (1) at both biopsy time points, suggesting that 18c could be a highly effective agent for treating HCV infected patients in the clinic.

ABBV-168 is a dihalogenated nucleotide under investigation for the treatment of hepatitis C virus. Three synthetic routes aimed at achieving the stereoselective installation of the C2' gem-Br,F substitution and subsequent Vorbruggen glycosylation were explored to prepare the penultimate nucleoside intermediate. Development culminated in a route to ABBV-168 featuring a de novo chromatography-free furanose synthesis, protecting group-directed Vorbruggen glycosylation, and highly selective phosphoramidation to furnish the API.

A HTS campaign identified compound 1, an excellent hit-like molecule to initiate medicinal chemistry efforts to optimize a dual ROCK1 and ROCK2 inhibitor. Substitution (2-Cl, 2-NH2, 2-F, 3-F) of the pyridine hinge binding motif or replacement with pyrimidine afforded compounds with a clean CYP inhibition profile. Cocrystal structures of an early lead compound were obtained in PKA, ROCK1, and ROCK2. This provided critical structural information for medicinal chemistry to drive compound design. The structural data indicated the preferred configuration at the central benzylic carbon would be (R), and application of this information to compound design resulted in compound 16. This compound was shown to be a potent and selective dual ROCK inhibitor in both enzyme and cell assays and efficacious in the retinal nerve fiber layer model after oral dosing. This tool compound has been made available through the AbbVie Compound Toolbox. Finally, the cocrystal structures also identified that aspartic acid residues 176 and 218 in ROCK2, which are glutamic acids in PKA, could be targeted as residues to drive both potency and kinome selectivity. Introduction of a piperidin-3-ylmethanamine group to the compound series resulted in compound 58, a potent and selective dual ROCK inhibitor with excellent predicted drug-like properties.

TRPV1 (transient receptor potential vanilloid family type 1) is a nonselective cation channel that is activated and/or sensitized by noxious heat, protons, and other endogenous molecules released following tissue injury. In addition, a role for TRPV1 in mechanotransmission is emerging. We have recently reported that a selective TRPV1 receptor antagonist, A‐889425, reduces mechanical allodynia and spinal neuron responses to mechanical stimulation of complete Freund's adjuvant (CFA)‐inflamed rat hind paws. The population of peripheral nerve fibers through which TRPV1 antagonists mediate their effect on mechanotransmission have not yet been described. The objective of this study was to characterize TRPV1‐mediated modulation of mechanically evoked activity in sensory axons innervating rat hind paws. We used an in vitro skin‐nerve preparation to record neural activity from single axons isolated from rat tibial nerve. Single fibers were classified by conduction velocity, mechanical threshold, and stimulus–response relationships. We used A‐889425 to investigate uninjured and inflamed skin afferent neuron populations to evoked mechanical stimulation. Application of A‐889425 had no effect on the mechanical responsiveness of Aδ and C‐fiber units innervating uninjured skin. In contrast, A‐889425 inhibited responses of slowly conducting Aδ fiber units to noxious mechanical stimulation in a population of axons innervating CFA‐inflamed hind paws. These data support a role for TRPV1 in mechanotransmission following peripheral inflammation, and highlight the importance of a distinct subclass of primary afferent neurons in mediating this effect. Synapse, 2012. © 2011 Wiley Periodicals, Inc.

The synthesis and SAR of a series of indazole TRPV1 antagonists leading to the discovery of 21 (ABT-116) is described. Biological studies demonstrated potent in vitro and in vivo activity for 21, as well as suitable physicochemical and pharmacokinetic properties for advancement to clinical development for pain management.

Inhibition of FBPase is considered a promising way to reduce hepatic gluconeogenesis and therefore could be a potential approach to treat type 2 diabetes. Herein we report the discovery of a series of purine phosphonic acids as AMP mimics targeting the AMP site of FBPase, which was achieved using a structure-guided drug design approach. These non-nucleotide purine analogues inhibit FBPase in a similar manner and with similar potency as AMP. More importantly, several purine analogues exhibited potent cellular and in vivo glucose-lowering activities, thus achieving proof-of-concept for inhibiting FBPase as a drug discovery target. For example, compounds 4.11 and 4.13 are as equipotent as AMP with regard to FBPase inhibition. Furthermore, compound 4.11 inhibited glucose production in primary rat hepatocytes and significantly lowered blood glucose levels in fasted rats.

In order to enhance understanding of TRPV1 contributions to thermoregulation, we measured the effects of a TRPV1 receptor antagonist, A-889425, on thermoregulatory neurons in the medial preoptic area of the hypothalamus (mPOA) of rats while simultaneously monitoring rectal temperature (Tr). Administration of A-889425 (4 μmol/kg, i.v.) significantly increased Tr by 0.42±0.02 °C in anesthetized rats. Warm-sensitive (WS) neurons in the mPOA increase firing in response to body warming, and when active stimulate heat loss and inhibit heat production. WS neurons were initially inhibited by A-889425. Subsequently, WS neuronal activity diverged, differentiating WS neurons into two subgroups. One group of WS neurons continued to be inhibited during the recording period while another group of “biphasic” WS neurons increased firing as Tr increased. Cold-sensitive (CS) neurons fire at a higher rate during cooling of the body, and when active, may contribute to heat production. Injection of A-889425 affected CS neurons in a manner opposite to the biphasic WS neurons; activity was initially increased followed by a later decrease. Direct administration of A-889425 into the mPOA (10 and 30 nmol) or spinal cord (30 nmol) did not affect Tr. Disruption of abdominal TRPV1 receptor function by injection of the TRPV1 receptor agonist, resiniferatoxin (20 μg/kg, i.p.), 9–15 days prior to experiments, blocked the effects of systemically injected A-889425 on Tr and mPOA neuronal activity. These data demonstrate that antagonist block of abdominal TRPV1 receptors indirectly modulates activity of thermoregulatory neurons in the mPOA in a manner that is consistent with producing an acute rise in body temperature.

TRPV1 receptors are activated and/or modulated by noxious heat, capsaicin, protons and other endogenous agents released following tissue injury. There is a growing appreciation that this molecular integrator may also have a role in mechanosensation. To further understand this role, we investigated the systemic and site-specific effects of a selective TRPV1 receptor antagonist, A-889425, on low-intensity mechanical stimulation in inflamed rats. Systemic administration of A-889425 (30 and 100 micromol/kg po) reduced mechanical allodynia in complete Freund's adjuvant (CFA)-inflamed rats. Systemic A-889425 (3 and 10 micromol/kg iv) also decreased the responses of spinal wide dynamic range (WDR) neurons to low-intensity mechanical stimulation in CFA-inflamed but not uninjured rats. This effect of A-889425 was likely mediated via multiple sites since local injection of A-889425 into the spinal cord (1-3 nmol), ipsilateral hindpaw (200 nmol), and cerebral ventricles (30-300 nmol) all attenuated WDR responses to low-intensity mechanical stimulation. In addition to an effect on mechanotransmission, systemic administration of A-889425 reduced the spontaneous firing of WDR neurons in inflamed but not uninjured rats. Spontaneous firing is elevated after injury and may reflect ongoing pain in the animal. Local injection experiments indicated that this effect of A-889425 on spontaneous firing was mainly mediated via TRPV1 receptors in the spinal cord. Thus the current data demonstrate that TRPV1 receptors have an enhanced role after an inflammatory injury, impacting both low-intensity mechanotransmission and possibly spontaneous pain. Furthermore this study delineates the differential contribution of central and peripheral TRPV1 receptors to affect spontaneous or mechanically evoked firing of WDR neurons.