Characterization of Selective and Potent JAK1 Inhibitors Intended for the Inhaled Treatment of Asthma

Purpose: Janus kinase 1 (JAK1) is implicated in multiple inflammatory pathways that are critical for the pathogenesis of asthma, including the interleukin (IL)-4, IL-5, IL-13, and thymic stromal lymphopoietin cytokine signaling pathways, which have previously been targeted to treat allergic asthma. Here, we describe the development of AZD0449 and AZD4604, two novel and highly selective JAK1 inhibitors with promising properties for inhalation. Methods: The effects of AZD0449 and AZD4604 in JAK1 signaling pathways were assessed by measuring phosphorylation of signal transducer and activator of transcription (STAT) proteins and chemokine release using immunoassays of whole blood from healthy human volunteers and rats. Pharmacokinetic studies performed on rats evaluated AZD0449 at a lung deposited dose of 52 mu g/kg and AZD4604 at 30 mu g/kg. The efficacy of AZD0449 and AZD4604 was assessed by evaluating lung inflammation (cell count and cytokine levels) and the late asthmatic response (average enhanced pause [Penh]). Results: Both compounds inhibited JAK1-dependent cytokine signaling pathways in a dose-dependent manner in human and rat leukocytes. After intratracheal administration in rats, both compounds exhibited low systemic exposures and medium-to-long terminal lung half-lives (AZD0449, 34 hours; AZD4604, 5 hours). Both compounds inhibited STAT3 and STAT5 phosphorylation in lung tissue from ovalbumin (OVA)-challenged rats. AZD0449 and AZD4604 also inhibited eosinophilia in the lung and reduced the late asthmatic response, measured as Penh in the OVA rat model. Conclusion: AZD0449 and AZD4604 show potential as inhibitors of signaling pathways involved in asthmatic immune responses, with target engagement demonstrated locally in the lung. These findings support the clinical development of AZD0449 and AZD4604 for the treatment of patients with asthma

Effects of (2R)-(3-amino-2-fluoropropyl)sulphinic acid (AFPSiA) on transient lower oesophageal sphincter relaxation in dogs and mechanism of hypothermic effects in mice

1. The effects of the novel GABA analogue (2R)-(3-amino-2-fluoropropyl)sulphinic acid (AFPSiA) on transient lower oesophageal sphincter relaxations (TLOSRs) were studied in the dog. In addition, the GABA(A)/GABA(B) selectivity was determined in vitro and in vivo, and the pharmacokinetics and the metabolism of the compound were studied in the dog and rat. 2. TLOSRs were reduced by 55±8% after intragastric administration of AFPSiA at 14 μmol kg(−1) and did not decrease further at higher doses. When evaluated 2 and 4 h after administration, the effect declined to 37±6 and 16±9%, respectively. Spontaneous swallowing was only significantly inhibited at 100 μmol kg(−1). 3. The oral availability of AFPSiA was 52±17 and 71±4% in the dog and rat, respectively. A fraction of AFPSiA was oxidised to the corresponding sulphonate, (2R)-(3-amino-2-fluoropropyl)sulphonic acid (AFPSoA) after oral administration to the rat and dog. 4. In rat brain membranes, AFPSiA was found to have ten times higher affinity for rat brain GABA(B) (K(i)=47±4.4 nM) compared to GABA(A) (K(i)=430±46 nM) binding sites. The compound was a full agonist at human recombinant GABA(B(1a,2)) receptors (EC(50)=130±10 nM). In contrast, the metabolite AFPSoA was considerably more selective for binding to rat brain GABA(A) (K(i)=37±3.1 nM) vs GABA(B) (K(i)=6800±280 nM) receptors. 5. In the mouse, high doses (1–8 mmol kg(−1)) of AFPSiA induced a rapid and mild hypothermia followed by a profound and sustained hypothermia at the higher doses tested (6 and 8 mmol kg(−1)). This effect was unaffected by the selective GABA(B) receptor antagonist CGP62349. AFPSoA (1 and 2 mmol kg(−1)) produced transient and moderate hypothermia while the hypothermic response was considerably larger at 4 mmol kg(−1). 6. It is concluded that AFPSiA inhibits but does not abolish TLOSRs in the dog. High doses of the compound induce hypothermia in the mouse, which probably is attributable to activation of the GABA(A) receptor. The latter effect may be caused both by AFPSiA and its oxidised sulphonic acid metabolite AFPSoA