Unveiling 2-isopropyl-5-methylphenol's immunomodulatory potential in breast cancer: A synergistic computational and laboratory investigation.

Tumor cells evade immune surveillance and promote their growth by modulating immune checkpoint molecules (ICMs). Various breast cancer subtypes have demonstrated the involvement of these molecules, promoting the investigation of natural compounds for immunomodulatory potential. 2-Isopropyl-5-methylphenol, a bioactive monoterpenoid found in several plant species, was evaluated for its impact on breast cancer immunomodulation. Computational and in-vitro studies were conducted to evaluate 2-Isopropyl-5-methylphenol's immunomodulatory potential. Molecular docking of ICMs (CTLA-4, PD-1, and PD-L1) with 2-Isopropyl-5-methylphenol was performed using AutoDock Vina. Additional tools utilized were BIOVIA Discovery Studio 2021, PyMOL, and LigPlot. The mRNA expression of ICMs upon treatment of breast cancer cells (MDA-MB-231) with varying concentrations (50, 100, and 200 µg) of 2-Isopropyl-5-methylphenol was measured by qPCR. Docking complexes of CTLA-4, PD-1, and PD-L1 with 2-Isopropyl-5-methylphenol showed binding free energies of -4.3 kcal/mol, -5.2 kcal/mol and -4.4 kcal/mol, respectively. Involvement of different key amino acid residues strengthened 2-Isopropyl-5-methylphenol's binding with ICMs. Expression analysis revealed 2-Isopropyl-5-methylphenol downregulated CTLA-4 expression while upregulated PD-1 and PD-L1 expression. This study highlights the immunomodulatory potential of the bioactive monoterpene 2-Isopropyl-5-methylphenol. Supported by computational and in-vitro findings, 2-Isopropyl-5-methylphenol could be considered as a potential target for anti-tumor drug development.