Background and Aims Oral systemic pan-Janus kinase [JAK] inhibition is effective for ulcerative colitis [UC] but is limited by toxicities. We describe preclinical to clinical translation of TD-1473—an oral gut-selective pan-JAK inhibitor—from in vitro characterization through a Phase 1b study in patients with UC. Methods TD-1473 JAK inhibition potency was evaluated in vitro; plasma pharmacokinetics, safety and efficacy were assessed in mice. In a first-time-in-human study, plasma pharmacokinetics and safety were assessed after single and multiple [14 days] ascending doses administered orally to healthy subjects. The Phase 1b study randomized patients with moderately to severely active UC to receive once-daily oral TD-1473 20, 80 or 270 mg, or placebo for 28 days. Plasma and colonic tissue concentrations were measured; safety was assessed; and efficacy was evaluated by UC clinical parameters, disease-surrogate biomarkers, endoscopy, histology and colonic tissue JAK signalling. Results TD-1473 exhibited potent pan-JAK inhibitory activity in vitro. Oral TD-1473 administration to mice achieved high, biologically active colonic tissue concentrations with low plasma exposure and decreased oxazolone-induced colitis activity without reducing blood cell counts vs placebo. TD-1473 administration in healthy human subjects and patients with UC yielded low plasma exposure and was generally well tolerated; treatment in patients with UC resulted in biologically active colonic tissue concentrations and descriptive trends toward reduced clinical, endoscopic and histological disease activity vs placebo. Conclusion Gut-selective pan-JAK inhibition with TD-1473 administration resulted in high intestinal vs plasma drug exposure, local target engagement, and trends toward reduced UC disease activity. [Clinicaltrials.gov NCT02657122, NCT02818686]

1 5-Hydroxytryptamine (5-HT) receptors mediating contraction and relaxation are present in Cynomolgus monkey isolated jugular vein denuded of endothelium. 2 In the absence of spasmogen, a-methyl-5-HT and sumatriptan contracted the tissues with potency values (pEC50) of 6.8 (n = 2) and 6.4 + 0.1 (mean + s.e. mean, n = 3), respectively. In contrast, 5-HT caused an initial contraction (10 nM -1 pM), followed by relaxation (1 pMM-32 yM). The contractile effect of x-methyl-5-HT was antagonized by ketanserin with a pKB value of 8.1 (n = 2). 5-Carboxamidotryptamine (5-CT), 5-methoxytryptamine (5-MeOT) and 8-OH-DPAT did not contract or relax the tissues in the absence of spasmogen.3 In tissues precontracted with U46619 (10 nM) and in the presence of 5-HTIA, 5-HTIB, 5-HT2A, 5-HT3, and 5-HT4 receptor blockade, 5-CT and 5-MeOT caused endothelium-independent relaxation with potency values of 7.5+0.1 (n=21) and 5.7+0.1 (n=4), respectively. The potency of 5-HT was 7.2 (n=2) while oa-methyl-5-HT did not start to relax the tissues below a concentration of 10 MM.4 Relaxations elicited by 5-CT were antagonized by the following compounds (with pKB values in parentheses): methiothepin (9.7), mesulergine (8.1), metergoline (8.0), clozapine (7.8), mianserin (7.7), spiperone (7.3), ritanserin (7.1), methysergide (7.0) and ketanserin (5.7). 5 It is concluded that the 5-HT receptor mediating endothelium-independent relaxation may be a functional correlate of the putative 5-ht7 receptor.

BackgroundAn unmet need remains for safe and effective treatments to induce and maintain remission in inflammatory bowel disease (IBD) patients. The Janus kinase (JAK) inhibitor, tofacitinib, has demonstrated robust efficacy in ulcerative colitis patients although, like other systemic immunosuppressants, there may be safety concerns associated with its use. This preclinical study evaluated whether modulating intestinal inflammation via local JAK inhibition can provide efficacy without systemic immunosuppression.MethodsThe influence of tofacitinib, dosed orally or intracecally, on oxazolone-induced colitis, oxazolone or interferon-γ (IFNγ)-induced elevation of colonic phosphorylated signal transducer and activator of transcription1 (pSTAT1) levels, and basal splenic natural killer (NK) cell counts was investigated in mice.ResultsTofacitinib, dosed orally or intracecally, inhibited, with similar efficacy, oxazolone-induced colitis, represented by improvements in the disease activity index and its sub-scores (body weight, stool consistency and blood content). Intracecal dosing of tofacitinib resulted in a higher colon:plasma drug exposure ratio compared to oral dosing. At equieffective oral and intracecal doses, colonic levels of tofacitinib were similar, while the plasma levels for the latter were markedly lower, consistent with a lack of effect on splenic NK cell counts. Tofacitinib, dosed orally, intracecally, or applied to the colonic lumen in vitro, produced dose-dependent, and maximal inhibition of oxazolone or IFNγ-induced STAT1 phosphorylation in the colon.ConclusionsLocalized colonic JAK inhibition, by intracecal delivery of tofacitinib, provides colonic target engagement and efficacy in a mouse colitis model at doses which do not impact splenic NK cell counts. Intestinal targeting of JAK may permit separation of local anti-inflammatory activity from systemic immunosuppression, and thus provide a larger therapeutic index compared to systemic JAK inhibitors.

Through application of our multivalent approach to drug discovery we previously reported the first discovery of dual pharmacology MABA bronchodilators, exemplified by 1. Herein we describe the subsequent lead optimization of both muscarinic antagonist and β2 agonist activities, through modification of the linker motif, to achieve 24 h duration of action in a guinea pig bronchoprotection model. Concomitantly we targeted high lung selectivities, low systemic exposures and identified crystalline forms suitable for inhalation devices. This article culminates with the discovery of our first clinical candidate 12f (TD-5959, GSK961081, batefenterol). In a phase 2b trial, batefenterol produced statistical and clinically significant differences compared to placebo and numerically greater improvements in the primary end point of trough FEV1 compared to salmeterol after 4 weeks of dosing in patients with moderate to severe chronic obstructive pulmonary disease (COPD).

The objective of the present studies was to characterize the pharmacologic properties of GSK-961081 [TD-5959; (R)-1-(3-((2-chloro-4-(((2-hydroxy-2-(8-hydroxy-2-oxo-1,2-dihydroquinolin-5-yl) ethyl)amino)methyl)-5-methoxyphenyl)amino)-3-oxopropyl) piperidin-4-yl [1,19-biphenyl]-2-ylcarbamate], a novel first-in-class inhaled bifunctional compound possessing both muscarinic antagonist (MA) and b 2 -adrenoceptor agonist (BA) properties (MABA). In competition radioligand binding studies at human recombinant receptors, GSK-961081 displayed high affinity for hM 2 (K i 5 1.4 nM), hM 3 muscarinic receptors (K i 5 1.3 nM) and hb 2 -adrenoceptors (K i 5 3.7 nM). GSK-961081 behaved as a potent hb 2 -adrenoceptor agonist (EC 50 5 0.29 nM for stimulation of cAMP levels) with 440-and 320-fold functional selectivity over hb 1 -and hb 3 -adrenoceptors, respectively. In guinea pig isolated tracheal tissues, GSK-961081 produced smooth muscle relaxation through MA (EC 50 5 50.2 nM), BA (EC 50 524.6 nM), and MABA (EC 50 5 11 nM) mechanisms. In the guinea pig bronchoprotection assay, inhaled GSK-961081 produced potent, dose-dependent inhibition of bronchoconstrictor responses via MA (ED 50 5 33.9 mg/ml), BA (ED 50 5 14.1 mg/ml), and MABA (ED 50 5 6.4 mg/ml) mechanisms. Significant bronchoprotective effects of GSK-961081 were evident in guinea pigs via MA, BA, and MABA mechanisms for up to 7 days after dosing. The lung selectivity index of GSK-961081 in guinea pigs was 55-to 110-fold greater than that of tiotropium with respect to systemic antimuscarinic antisialagogue effects and was 10-fold greater than that of salmeterol with respect to systemic b 2 -adrenoceptor hypotensive effects. These preclinical findings studies suggest that GSK-961081 has the potential to be a promising next-generation inhaled lung-selective bronchodilator for the treatment of airway diseases, including chronic obstructive pulmonary disease.