Shohei Shirakami

Discovery of 3,6-dihydroimidazo[4,5-d]pyrrolo[2,3-b]pyridin-2(1H)-one derivatives as novel JAK inhibitors.

Because Janus kinases (JAKs) play a crucial role in cytokine-mediated signal transduction, JAKs are an attractive target for the treatment of organ transplant rejection and autoimmune diseases such as rheumatoid arthritis (RA). To identify JAK inhibitors, we focused on the 1H-pyrrolo[2,3-b]pyridine derivative 3, which exhibited moderate JAK3 and JAK1 inhibitory activities. Optimization of 3 identified the tricyclic imidazo-pyrrolopyridinone derivative 19, which exhibited potent JAK3 and JAK1 inhibitory activities (IC50=1.1 nM, 1.5 nM, respectively) with favorable metabolic stability.

Synthesis and Evaluation of 1H-Pyrrolo[2,3-b]pyridine Derivatives as Novel Immunomodulators Targeting Janus Kinase 3

Janus kinases (JAKs) have been known to play crucial roles in modulating a number of inflammatory and immune mediators. Here, we describe a series of 1H-pyrrolo[2,3-b]pyridine derivatives as novel immunomodulators targeting JAK3 for use in treating immune diseases such as organ transplantation. In the chemical modification of compound 6, the introduction of a carbamoyl group to the C5-position and substitution of a cyclohexylamino group at the C4-position of the 1H-pyrrolo[2,3-b]pyridine ring led to a large increase in JAK3 inhibitory activity. Compound 14c was identified as a potent, moderately selective JAK3 inhibitor, and the immunomodulating effect of 14c on interleukin-2-stimulated T cell proliferation was shown. Docking calculations and WaterMap analysis of the 1H-pyrrolo[2,3-b]pyridine-5-carboxamide derivatives were conducted to confirm the substituent effects on JAK3 inhibitory activity.

AS2553627, a novel JAK inhibitor, prevents chronic rejection in rat cardiac allografts

ABSTRACT Janus family kinases (JAKs) are essential molecules for cytokine responses and attractive targets for the treatment of transplant rejection and autoimmune diseases. Several JAK inhibitors have shown demonstrable effects on acute rejection in experimental cardiac transplant models. However, little is known about the potential benefits of JAK inhibitors on chronic rejection outcomes such as vasculopathy and fibrosis. Here, we examined the pharmacological profile of a novel JAK inhibitor, AS2553627, and explored its therapeutic potential in chronic rejection as well as acute rejection in a rat cardiac transplant model. AS2553627 potently inhibited JAK kinases but showed no inhibition of other kinases, including TCR‐associated molecules. The compound also suppressed proliferation of IL‐2 stimulated human and rat T cells. In a rat cardiac transplant model, oral administration of AS2553627 alone or co‐administration with a sub‐therapeutic dose of tacrolimus effectively prolonged cardiac allograft survival, suggesting the efficacy in treating acute rejection. To evaluate the effect on chronic rejection, recipient rats were administered a therapeutic dose of tacrolimus for 90 days. In combination with tacrolimus, AS2553627 significantly reduced cardiac allograft vasculopathy and fibrosis that tacrolimus alone did not inhibit. AS2553627 at the effective dose in rat transplantation models did not significantly reduce reticulocyte counts in peripheral whole blood after in vivo erythropoietin administration, indicating a low risk for anemia. These results suggest that AS2553627 may be a therapeutic candidate for the prevention of not only acute but also chronic rejection in cardiac transplantation.

Synthesis and evaluation of novel 1H-pyrrolo[2,3-b]pyridine-5-carboxamide derivatives as potent and orally efficacious immunomodulators targeting JAK3.

Janus kinases (JAKs) regulate various inflammatory and immune responses and are targets for the treatment of inflammatory and immune diseases. As a novel class of immunomodulators targeting JAK3, 1H-pyrrolo[2,3-b]pyridine-5-carboxamide derivatives are promising candidates for treating such diseases. In chemical modification of lead compound 2, the substitution of a cycloalkyl ring for an N-cyanopyridylpiperidine in C4-position was effective for increasing JAK3 inhibitory activity. In addition, modulation of physical properties such as molecular lipophilicity and basicity was important for reducing human ether-a-go-go-related gene (hERG) inhibitory activity. Our optimization study gave compound 31, which exhibited potent JAK3 inhibitory activity as well as weak hERG inhibitory activity. In cellular assay, 31 exhibited potent immunomodulating effect on IL-2-stimulated T cell proliferation. In a pharmacokinetic study, good metabolic stability and oral bioavailability of 31 were achieved in rats, dogs, and monkeys. Further, 31 prolonged graft survival in an in vivo rat heterotopic cardiac transplant model.

THU0101 ASP015K: A Novel Jak Inhibitor Demonstrated Potent Efficacy in Adjuvant-Induced Arthritis Model in Rats

Background The Janus kinase (JAK) family of enzymes plays a key role in cytokine signaling, which is involved in the pathogenic events of immune-mediated disorders such as rheumatoid arthritis (RA). Objectives The aim of this study was to identify pharmacological profiles of a newly synthesized JAK inhibitor, ASP015K, and to estimate its therapeutic potential in the treatment of RA patients using an experimental animal model. Methods In vitro enzyme inhibition assays were conducted against JAK1, JAK2, JAK3 and tyrosine kinase 2 (TYK2) enzymes. Cell-based assays were also conducted to assess the selectivity of ASP015K for signaling via JAK1/JAK3 over JAK2/JAK2. JAK1/3 activation was evaluated by interleukin (IL)-2-stimulated T cell proliferation; JAK2/2 action was evaluated by erythropoietin (EPO)-stimulated erythroleukemia cell proliferation. Phosphorylation of signal transducer and activator of transcription5 (STAT5) was quantified by flow cytometry as a biomarker of JAK activity in vitro in human whole blood and ex vivo in rat whole blood. In order to evaluate the potential efficacy of ASP015K to reduce clinical signs and symptoms of RA as well as disease progression, the reduction of paw swelling and ankle bone destruction in adjuvant-induced arthritic (AIA) rats were assessed after both prophylactic and therapeutic dosing regimens of ASP015K. Results ASP015K was shown to inhibit JAK enzymes, with moderate selectivity for JAK3. ASP015K suppressed the IL-2-induced proliferation of human T cells with greater potency than EPO-induced proliferation of human erythroleukemia cells. ASP015K inhibited STAT5 phosphorylation (pSTAT5) in human whole blood in a concentration related manner. Additionally, after oral administration, ASP015K also suppressed pSTAT5 in rat whole blood. In the rat AIA model, the hind paw volume gradually increased after adjuvant injection and ankle bone destruction was established. In a prophylactic dosing regimen, the increase in paw volume was significantly decreased by oral administration of ASP015K in a dose-dependent manner. Similar findings in ankle bone destruction score were observed. In a therapeutic dosing regimen, paw swelling and ankle bone destruction score were also suppressed in a dose-dependent manner. Conclusions Data from the current study demonstrate that ASP015K inhibits human JAK enzymes with moderate selectivity against JAK1/3 over JAK2/2, which may translate to less hematological side effects observed in the clinic. ASP015K improved paw swelling and ankle bone destruction after both prophylactic and therapeutic dosing regimens in the rat AIA model. These data together with the finding that ASP015K suppressed pSTAT5 in whole blood after oral administration in rats suggest that the effects of ASP015K on AIA model are due to JAK inhibition. These data also suggest that ASP015K has the potential to reduce clinical signs and symptoms as well as prevent disease progression in RA patients and warrants further clinical investigation. Disclosure of Interest S. Yamazaki Employee of: Astellas Pharma Inc., H. Morio Employee of: Astellas Pharma Inc., M. Inami Employee of: Astellas Pharma Inc., M. Ito Employee of: Astellas Pharma Inc., Y. Fujii Employee of: Astellas Pharma Inc., K. Hanaoka Employee of: Astellas Pharma Inc., K. Yamagami Employee of: Astellas Pharma Inc., K. Okuma Employee of: Astellas Pharma Inc., Y. Morita Employee of: Astellas Pharma Inc., S. Shirakami Employee of: Astellas Pharma Inc., T. Inoue Employee of: Astellas Pharma Inc., S. Miyata Employee of: Astellas Pharma Inc., Y. Higashi Employee of: Astellas Pharma Inc., N. Seki Employee of: Astellas Pharma Inc.

Design, synthesis, and evaluation of 4,6-diaminonicotinamide derivatives as novel and potent immunomodulators targeting JAK3

In organ transplantation, T cell-mediated immune responses play a key role in the rejection of allografts. Janus kinase 3 (JAK3) is specifically expressed in hematopoietic cells and associated with regulation of T cell development via interleukin-2 signaling pathway. Here, we designed novel 4,6-diaminonicotinamide derivatives as immunomodulators targeting JAK3 for prevention of transplant rejection. Our optimization of C4- and C6-substituents and docking calculations to JAK3 protein confirmed that the 4,6-diaminonicotinamide scaffold resulted in potent inhibition of JAK3. We also investigated avoidance of human ether-a-go-go related gene (hERG) inhibitory activity. Selected compound 28 in combination with tacrolimus prevented allograft rejection in a rat heterotopic cardiac transplantation model.

Discovery and structural characterization of peficitinib (ASP015K) as a novel and potent JAK inhibitor

Janus kinases (JAKs) are considered promising targets for the treatment of autoimmune diseases including rheumatoid arthritis (RA) due to their important role in multiple cytokine receptor signaling pathways. Recently, several JAK inhibitors have been developed for the treatment of RA. Here, we describe the identification of the novel orally bioavailable JAK inhibitor 18, peficitinib (also known as ASP015K), which showed moderate selectivity for JAK3 over JAK1, JAK2, and TYK2 in enzyme assays. Chemical modification at the C4-position of lead compound 5 led to a large increase in JAK inhibitory activity and metabolic stability in liver microsomes. Furthermore, we determined the crystal structures of JAK1, JAK2, JAK3, and TYK2 in a complex with peficitinib, and revealed that the 1H-pyrrolo[2,3-b]pyridine-5-carboxamide scaffold of peficitinib forms triple hydrogen bonds with the hinge region. Interestingly, the binding modes of peficitinib in the ATP-binding pockets differed among JAK1, JAK2, JAK3, and TYK2. WaterMap analysis of the crystal structures suggests that unfavorable water molecules are the likely reason for the difference in orientation of the 1H-pyrrolo[2,3-b]pyridine-5-carboxamide scaffold to the hinge region among JAKs.

Design, synthesis, and biological evaluation of novel biphenyl-4-carboxamide derivatives as orally available TRPV1 antagonists

A new series of transient receptor potential vanilloid type 1 (TRPV1) antagonists were designed and synthesized from N-(3-hydroxyphenyl)-2-(piperidin-1-ylmethyl)biphenyl-4-carboxamide hydrochloride (8). SAR studies identified (R)-N-(1-methyl-2-oxo-1,2,3,4-tetrahydro-7-quinolyl)-2-[(2-methylpyrrolidin-1-yl)methyl]biphenyl-4-carboxamide hydrochloride (ASP8370, 7), as a compound with high aqueous solubility, satisfactory stability in human liver microsomes, and reduced CYP3A4 inhibition. ASP8370 was selected as a clinical development candidate with significant ameliorative effects on neuropathic pain. SAR studies also revealed the structural mechanisms underlying the switching between TRPV1 antagonism and agonism.

Discovery of tricyclic dipyrrolopyridine derivatives as novel JAK inhibitors

Janus kinases (JAKs) play a crucial role in cytokine mediated signal transduction. JAK inhibitors have emerged as effective immunomodulative agents for the prevention of transplant rejection. We previously reported that the tricyclic imidazo-pyrrolopyridinone 2 is a potent JAK inhibitor; however, it had poor oral absorption due to low membrane permeability. Here, we report the structural modification of compound 2 into the tricyclic dipyrrolopyridine 18a focusing on reduction of polar surface area (PSA), which exhibits potent in vitro activity, improved membrane permeability and good oral bioavailability. Compound 18a showed efficacy in rat heterotopic cardiac transplants model.