Selective Modulation of PAR-2-Driven Inflammatory Pathways by Oleocanthal: Attenuation of TNF-α and Calcium Dysregulation in Colorectal Cancer Models

Abstract

Colorectal cancer (CRC) remains a principal contributor to oncological mortality worldwide, with chronic inflammation serving as a fundamental driver of its pathogenesis. Protease-activated receptor-2 (PAR-2), a G-protein-coupled receptor, orchestrates inflammation-driven tumorigenesis by potentiating NF-κB and Wnt/β-catenin signaling, thereby fostering epithelial–mesenchymal transition (EMT), immune evasion, and therapeutic resistance. Despite its pathological significance, targeted modulation of PAR-2 remains an underexplored avenue in CRC therapeutics. Oleocanthal (OC), a phenolic constituent of extra virgin olive oil, is recognized for its potent anti-inflammatory and anti-cancer properties; however, its regulatory influence on PAR-2 signaling in CRC is yet to be elucidated. This study interrogates the impact of OC on PAR-2-mediated inflammatory cascades using HT-29 and Caco-2 CRC cell lines subjected to lipopolysaccharide (LPS)-induced activation of PAR-2. Expression levels of PAR-2 and TNF-α were quantified through Western blotting and RT-PCR, while ELISA assessed TNF-α secretion. Intracellular calcium flux, a pivotal modulator of PAR-2-driven oncogenic inflammation, was evaluated AcademicEditor: CarmineStolfi Received: 7February2025 Revised: 15March2025 Accepted: 19March2025 Published: 24 March2025 Citation: Patnaik, R.; Varghese, R.L.; Banerjee, Y. Selective Modulation of PAR-2-DrivenInflammatory PathwaysbyOleocanthal: Attenuation of TNF-αandCalciumDysregulation in Colorectal Cancer Models. Int. J. Mol. Sci. 2025, 26, 2934. https:// doi.org/10.3390/ijms26072934 Copyright: ©2025bytheauthors. Licensee MDPI,Basel,Switzerland. This article is an open access article distributed under the termsand conditions of the Creative Commons Attribution (CC BY)license (https://creativecommons.org/ licenses/by/4.0/). via Fluo-4 calcium assays. LPS markedly elevated PAR-2 expression at both mRNA and protein levels in CRC cells (p < 0.01, one-way ANOVA). OC administration (20–150 µg/mL) elicited a dose-dependent suppression of PAR-2, with maximal inhibition at 100–150 µg/mL (p < 0.001, Tukey’s post hoc test). Concomitant reductions in TNF-α transcription (p < 0.01) and secretion (p < 0.001) were observed, corroborating the anti-inflammatory efficacy of OC. Additionally, OC ameliorated LPS-induced calcium dysregulation, restoring intracellular calcium homeostasis in a concentration-dependent manner (p < 0.01). Crucially, OC exhibited selectivity for PAR-2, leaving PAR-1 expression unaltered (p > 0.05), underscoring its precision as a therapeutic agent. These findings position OC as a selective modulator of PAR-2-driven inflammation in CRC, disrupting the pro-tumorigenic microenvironment through attenuation of TNF-α secretion, calcium dysregulation, and oncogenic signaling pathways. This study furnishes mechanistic insights into OC’s potential as a nutraceutical intervention in inflammation-associated CRC. Given the variability in OC bioavailability and content in commercial olive oil, future investigations should delineate optimal dosing strategies and in vivo efficacy to advance its translational potential in CRC therapy.