Integrative reverse-screening approaches for target discovery: the case of hydroxytyrosyl punicate

Reverse-screening methodologies have emerged as powerful tools for identifying molecular targets of bioactive compounds, complementing experimental approaches and accelerating drug discovery. Recent developments in integrative strategies combining multiple databases of protein-ligand interactions, gene expression profiles, and structural information, offer improved accuracy and broader coverage in mapping compound-target networks. In this review, we highlight the principles, strengths, and limitations of these integrative reverse-screening approaches, with particular attention to their application in natural product research. As an illustrative case study, we discuss hydroxytyrosyl punicate (HT-PA), a synthetic phenolipid derived from hydroxytyrosol and punicic acid, which exhibits antiproliferative and antiparasitic effects. The application of multi-database reverse screening to HT-PA identified potential targets, including arachidonate 5-lipoxygenase (ALOX5), transient receptor potential channels (TRPs), and peroxisome proliferator-activated receptors (PPARs), which are central to inflammation, metabolism, and pain regulation. This case exemplifies how integrative computational frameworks can provide mechanistic insights, prioritize targets for experimental validation, and guide the therapeutic development of natural product derivatives. More broadly, we argue that multi-database reverse screening represents a versatile platform for uncovering the molecular basis of bioactivity and advancing rational drug discovery from complex natural compounds.