Rational design and structural Bioinformatics-Driven discovery of tetrapeptide inhibitors for LIMK-Targeted cancer therapy.

LIM kinases (LIMK1 and LIMK2) serve as critical regulators of microtubule dynamics and mitotic progression, positioning them as promising targets for cancer therapy. However, most existing LIMK inhibitors lack specificity and cause toxic side effects due to the high sequence conservation between LIMK isoforms. Notably, no peptide-based inhibitors targeting the conserved ATP-binding pocket of LIMKs have been systematically investigated to date. To address this, an integrative structural bioinformatics approach, such as virtual screening of oligopeptide libraries, molecular dynamics simulations (MDS), and binding free energy analyses, was implemented to identify selective and less toxic peptide inhibitors. Among the top candidates, peptides with known anti-cancer, anti-inflammatory, and anti-angiogenic properties were prioritized. Interaction profiling revealed four and five tetrapeptides that closely mimicked interactions of LIMKi3 (BMS-5), a known Type-I inhibitor, with LIMK1 and LIMK2, respectively were shortlisted. Notably, MD and binding free energy analyses identified WRHF as a potent LIMK1 inhibitor with greater affinity than LIMKi3, while YYRW and WWHW exhibited superior stability and binding strength toward LIMK2. Overall, WRHF (LIMK1), YYRW, and WWHW (LIMK2) emerged as stable and high-affinity tetrapeptide inhibitors, demonstrating strong potential as selective, low-toxicity therapeutics for LIMK-targeted cancer treatment. However, in vitro and in vivo validation are essential to confirm their inhibitory efficacy and therapeutic potential, paving the way for the development of next-generation anti-cancer agents with reduced off-target effects.