Synthesis of Temporin-SHa Retro Analogs with Lysine Addition/Substitution and Antibiotic Conjugation to Enhance Antibacterial, Antifungal, and Anticancer Activities

In the face of rising the threat of resistant pathogens, antimicrobial peptides (AMPs) offer a viable alternative to the current challenge due to their broad-spectrum activity. This study focuses on enhancing the efficacy of temporin-SHa derived NST-2 peptide (<b>1</b>), which is known for its antimicrobial and anticancer activities. We synthesized new analogs of <b>1</b> using three strategies, i.e., retro analog preparation, lysine addition/substitution, and levofloxacin conjugation. Analogs were tested in terms of their antibacterial, antifungal, and anticancer activities. Analog <b>2,</b> corresponding to retro analog of NST-2, was found to be more active but also more hemolytic, reducing its selectivity index and therapeutic potential. The addition of lysine (in analog <b>3</b>) and lysine substitution (in analog <b>7</b>) reduced the hemolytic effect resulting in safer peptides. Conjugation with levofloxacin on the lysine side chain (in analogs <b>4</b> and <b>5</b>) decreased the hemolytic effect but unfortunately also the antimicrobial and anticancer activities of the analogs. Oppositely, conjugation with levofloxacin at the N-terminus of the peptide via the β-alanine linker (in analogs <b>6</b> and <b>8</b>) increased their antimicrobial and anticancer activity but also their hemolytic effect, resulting in less safe/selective analogs. In conclusion, lysine addition/substitution and levofloxacin conjugation, at least at the N-terminal position through the β-alanine linker, were found to enhance the therapeutic potential of retro analogs of NST-2 whereas other modifications decreased the activity or increased the toxicity of the peptides.