Abstract A179: NMS-P113, a novel orally available JAK2 selective inhibitor
Authors: Gnocchi Paola , Brasca Maria Gabriella , Amboldi Nadia , Avanzi Nilla , Bindi Simona , Canevari Giulia , Casero Daniele , Ceruti Roberta , Ciomei Marina , Cribioli Sabrina , Cristiani Cinzia , Nesi Marcella , Pastori Wilma , Patton Veronica , Pellizzoni Cinzia , Texido Gemma , Ardini Elena , Felder Eduard R. , Isacchi Antonella , Donati Daniele , Galvani Arturo 
Published: May 2016

Abstract
The Janus Kinases (JAK1, JAK2, JAK3, TYK2) are non-receptor tyrosine kinases that play important roles in hematopoiesis and immune response. In particular, gene ablation of JAK1 or JAK2 in the mouse is incompatible with life, due to neurological defects/immunodeficency and lack of erythropoiesis, respectively, whereas that of JAK3 or TYK2 is associated with severe immunodeficiency.
Activating mutations of JAKs are found in association with malignant transformation. The best characterized gain-of-function mutation, JAK2-V617F in the pseudo-kinase domain of JAK2, is present in hematopoietic cells of patients with myeloproliferative disorders (MPD). In particular, the JAK2-V617F mutation is found in &gt;95% of patients with polycythemia vera (PV), circa 50% of patients with essential thrombocythemia (ET), and circa 50% of myelofibrosis (MF) patients. Recently, a central role of JAK2 has been described in upregulation of the immune checkpoint component PD-L1 mediated by IFN-γ or by chromosome 9p24.1 amplification, suggesting that its inhibition might provide a new strategy to increase immune-mediated tumor inhibition in specific contexts.
Although JAK inhibitors have been approved in oncological and autoimmune settings (e.g. the JAK1/JAK2 inhibitor ruxolitinib in MF and the pan-JAK inhibitor tofacitinib in rheumatoid arthritis) and multiple agents are in clinical testing, JAK2 selective compounds might be provide an advantage for long-term MPD therapy or in association with immunotherapy, given that inhibition of other JAK family members leads to immunosuppressive effects.
Due to high homology amongst JAK family kinases within the ATP binding pocket, discovery of selective JAK2 inhibitors has represented a significant challenge. Here, we report the characterization of NMS-P113, a potent and selective JAK2 inhibitor. Screening of the NMS compound collection led to the identification of a pyrrole series with promising activity against JAK2. An optimization program led to identification of the potent and orally bioavailable JAK2 inhibitor NMS-P113. In biochemical assay this compound possesses low nanomolar potency against JAK2 (IC50 3 nM), with preferential activity over other JAK family members and high selectivity against a panel of 60 further kinases. In cellular assay, NMS-P113 potently inhibits proliferation of the JAK2 dependent SET-2 human megakaryoblastic leukemia line, derived from an ET patient and which harbors the JAK2-V617F mutation, as well as of Ba/F3 cells engineered to express constitutively activated JAK2 (IC50s circa 200 nM). NMS-P113 is 10-fold less active against the DERL-7 T-cell lymphoma cell line (which is dependent upon JAK1/JAK3) and has poor antiproliferative activity in JAK independent lines. Oral administration of NMS-P113 results in dose-related normalization of spleen weight and erythrocyte precursors count in an erythropoietin-induced model of PV in the mouse. Mechanism of action of NMS-P113, as assessed by inhibition of P-STAT5, was confirmed in vitro and in vivo.
A favorable ADME profile with high oral bioavailability, together with permissive therapeutic safety margins in test species, indicate that NMS-P113 is suitable for further development, affording the possibility of reduced undesirable immunosuppressive activities compared to inhibitors that target multiple members of the JAK family.
Citation Format: Paola Gnocchi, Maria Gabriella Brasca, Nadia Amboldi, Nilla Avanzi, Simona Bindi, Giulia Canevari, Daniele Casero, Roberta Ceruti, Marina Ciomei, Sabrina Cribioli, Cinzia Cristiani, Marcella Nesi, Wilma Pastori, Veronica Patton, Cinzia Pellizzoni, Gemma Texido, Elena Ardini, Eduard R. Felder, Antonella Isacchi, Daniele Donati, Arturo Galvani. NMS-P113, a novel orally available JAK2 selective inhibitor. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A179.