Janus kinase 2 (JAK2), is a non-receptor tyrosine kinase that regulates hematopoiesis through the JAK-STAT signaling pathway. Under physiological conditions, the ATP-binding pseudokinase (JH2) domain of JAK2 controls the activity of the canonical kinase (JH1) domain. The V617F mutation in JAK2 JH2 domain results in aberrant activation of the JH1 domain, and is the underlying cause of various myeloproliferative disorders (MPDs), including polycythemia vera, myelofibrosis, and essential thrombocythemia. Drug discovery efforts for MPDs have been focused on targeting the JAK2 JH1 domain, yielding agents that are non-selective, multi-kinase binders. An alternative strategy, with the potential to provide more selective therapies, would be to reduce the hyperactivation of the pathogenic JAK2 JH2V617F through targeting the JAK2 JH2 domain. The value of targeting the ATP site of the JH2 domain has been suggested by a mutagenesis study, but has not been proven using small-molecule modulators. To provide insights on the therapeutic potential of targeting JH2 with small molecules, we previously identified potent and selective JAK2 JH2 ligands with a diaminotriazole core. In this study we sought to optimize the leads of this series to enable testing in cells. We first aimed to improve their binding affinity using de novo design, exploration of heterocyclic substituents, and leveraging knowledge of new pharmacophore elements. Through this strategy, novel lead compounds were identified, exhibiting up to 10-fold improved binding and more than 100-fold selectivity. PK optimization of the leads was also necessary to enable cell testing. Integrating computer-aided design, synthesis, and efficient assays for in-vitro PK assessment led to cell-permeable JAK2 JH2 ligands that were used as probes to shed light on the JH2 targeting hypothesis.

Lead optimization strategies for JAK2 pseudokinase (JH2) binders: A remodeling journey to improve affinity, selectivity, and cell permeability of a diaminotriazole series

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.