Design, Synthesis, and Structure–Activity Relationship of Novel Human TLR1/2 Agonists for Potential Immunotherapy

Abstract Capitalizing on the advantages of Toll-like receptor 2 (TLR2) small-molecule agonists combining high potency and low toxicity in cancer therapy, we designed and synthesized a novel series of thiourea-based TLR2 small-molecule agonists and identified a low-toxicity TLR1/2-specific small molecule agonist SMU-C409 (EC50 = 65 ± 3 nM). SMU-C409 incorporates key optimizations: a carbonyl conservation strategy eliminates carboxylesterase-mediated degradation, and a quinoline (N-heterocycle) enables salt formation that boosts solubility by 3.4-fold (free base) and 5.5-fold (hydrochloride) relative to the lead compound SMU-C80 (EC50 = 43 ± 5 nM), while retaining potent agonistic activity and exhibiting superior plasma stability, which is critical for favorable pharmacokinetics and efficacy. Mechanistically, SMU-C409 activates TLR1/2, recruits MyD88, induces NF-κB phosphorylation, and stimulates TNF-α/IL-1β secretion; in vitro studies confirm robust immune cell activation and antitumor immunomodulation. SMU-C409 overcomes core limitations of TLR2 agonists while maintaining high potency, positioning it as a promising candidate for advancing cancer immunotherapy.