Identification and Validation of Carbonic Anhydrase II as the First Target of the Anti-Inflammatory Drug Actarit.

Ghita Ghislat,Pedro J Ballester,Taufiq Rahman

doi:10.3390/biom10111570

Abstract

Background and purpose: Identifying the macromolecular targets of drug molecules is a fundamental aspect of drug discovery and pharmacology. Several drugs remain without known targets (orphan) despite large-scale in silico and in vitro target prediction efforts. Ligand-centric chemical-similarity-based methods for in silico target prediction have been found to be particularly powerful, but the question remains of whether they are able to discover targets for target-orphan drugs. Experimental Approach: We used one of these in silico methods to carry out a target prediction analysis for two orphan drugs: actarit and malotilate. The top target predicted for each drug was carbonic anhydrase II (CAII). Each drug was therefore quantitatively evaluated for CAII inhibition to validate these two prospective predictions. Key Results: Actarit showed in vitro concentration-dependent inhibition of CAII activity with submicromolar potency (IC50 = 422 nM) whilst no consistent inhibition was observed for malotilate. Among the other 25 targets predicted for actarit, RORγ (RAR-related orphan receptor-gamma) is promising in that it is strongly related to actarit’s indication, rheumatoid arthritis (RA). Conclusion and Implications: This study is a proof-of-concept of the utility of MolTarPred for the fast and cost-effective identification of targets of orphan drugs. Furthermore, the mechanism of action of actarit as an anti-RA agent can now be re-examined from a CAII-inhibitor perspective, given existing relationships between this target and RA. Moreover, the confirmed CAII-actarit association supports investigating the repositioning of actarit on other CAII-linked indications (e.g., hypertension, epilepsy, migraine, anemia and bone, eye and cardiac disorders).

Highlights

Discovering the molecular targets of a molecule is important for its potential use as a drug [1,2]
An early example of these methods is the Similarity Ensemble Approach (SEA), which constructs a statistical model for each considered target and uses the ensemble of models to predict which targets interact with the investigated molecule
The first version (v1) of this tool exploited a knowledgebase with 184,912 molecules retrieved from release 20 of the ChEMBL database [11] and were annotated with 3046 single-protein targets

Summary

Introduction

Discovering the molecular targets of a molecule is important for its potential use as a drug [1,2]. SEA has been prospectively applied to de-orphanize drugs without known protein targets [7] These authors analysed a set of 1431 world-wide approved drugs. We developed MolTarPred [4], a -similarity-based method able to predict more than 4500 protein targets, and made it freely available as a webserver [10]. MolTarPred revealed that one of these target-orphan drugs (actarit) has potent activity against human carbonic anhydrase II (CAII) This discovery may be helpful to understand the mechanism of action of actarit in rheumatoid arthritis (RA) and opens the door to repositioning this anti-inflammatory drug to other CAII-linked indications

Materials and Methods

CAII Activity Assay

Materials

Results