Baricitinib and tofacitinib off-target profile, with a focus on Alzheimer’s disease

Introduction: Janus Kinase (JAK) inhibitors were recently identified as promising drug candidates for repurposing in Alzheimer’s disease (AD) due to their capacity to suppress inflammation via modulation of JAK/STAT signalling pathways. Besides interaction with primary therapeutic targets, JAK inhibitor drugs frequently interact with unintended, often unknown, biological off-targets, leading to associated effects. Nevertheless, the relevance of JAK inhibitors off-target interactions in the context of AD remains unclear. Methods: Putative off-targets of baricitinib and tofacitinib were predicted using a machine learning (ML) approach. After screening scientific literature, off-targets were filtered based on their relevance to AD. Targets that had not been previously identified as off-targets of baricitinib or tofacitinib were subsequently tested using biochemical or cell-based assays. From those, active concentrations were compared to bioavailable concentrations in the brain predicted by physiologically based pharmacokinetic (PBPK) modelling. Results: With the aid of ML and in vitro activity assays, we identified two enzymes previously unknown to be inhibited by baricitinib, namely casein kinase 2 subunit alpha 2 (CK2-alpha-2) and dual leucine zipper kinase (MAP3K12), both with Kd values of 5.8 uM. Predicted maximum concentrations of baricitinib in brain tissue using PBPK modelling range from 1.3 to 23 nM, which is two to three orders of magnitude below the corresponding binding constant. Conclusion: In this study, we extended the list of baricitinib off-targets that are potentially relevant for AD progression and predicted drug distribution in the brain. The results suggest a low likelihood of successful repurposing in AD due to low brain permeability, even at the maximum recommended daily dose. While additional research is needed to evaluate the potential impact of the off-target interaction on AD, the combined approach of ML target prediction, in vitro confirmation, and PBPK modelling may help prioritise drugs with a high likelihood of being effectively repurposed for AD.