New Temporin A Analogues Modified in Positions 1 and 10-Synthesis and Biological Studies

Background/Objectives: With growing antimicrobial resistance, the overuse of antibiotics, and stagnation in the discovery of new antibiotics, a novel alternative is required to overcome hard-to-treat infections. Antimicrobial peptides (AMPs) show great potential as a possible alternative to standard chemotherapeutics. Temporins are a group of AMPs that have been under the spotlight in numerous studies. Herein, we report the design and synthesis of Temporin A modified in position 1, where the proteinogenic amino acid Phe is replaced by Tyr or fluorinated Phe. In addition, in other analogues, in position 10, the Ser residue is replaced by Tyr or Thr. The aim of all modifications in the primary structure of the native Temporin A is to study the influence of the changes made on the antibacterial properties, antiproliferative activity, and hydrolytic stability of the newly synthesized molecules. Methods: The Fmoc/OBu^t SPPS strategy was employed for the synthesis of the novel-designed analogues. The antibacterial activity was evaluated with both disk diffusion and broth microdilution methods. The BALB 3T3 NRU test and MTT dye reduction assay were used to determine safety and antiproliferative activity. Results: The investigated analogues have low toxicity and are photosafe. The greatest selectivity was shown by DTTyr10 towards MCF-7 cells. DT4F, containing fluorinated Phe in position 1, was the most effective antibacterial agent among the new compounds. The incorporation of Thr in position 10, in comparison with the natural Ser residue, led to an increase in the antiproliferative effect of the new peptide. Conclusions: The obtained structure-activity relationship data show that the most promising compound in the tested series is FLPLIGRVL-Y-GILNH₂, where the Ser residue in position 10 is replaced by a more hydrophobic OH-containing Tyr residue. The analogue containing fluorinated Phe in position 1, DT4F, has the highest antiproliferative effect against both tested tumor cell lines, combined with good antibacterial properties at the lowest MIC (80 µg/mL), but it is more cyto- and phototoxic than the parent DTA molecule and is not stable at pH 9 for a 24 h period.