Synthesis and Biological Studies on (KLAKLAK)2-NH2 Analog Containing Unnatural Amino Acid β-Ala and Conjugates with Second Pharmacophore.

Sirine Jaber,Tsvetelina Foteva,Veronika Karadjova,Nelly Georgieva,Dancho Danalev,Ivan Iliev,Ivan Givechev,Emilia Naydenova,Elena Ivanova,Veronica Nemska

doi:10.3390/molecules26237321

Abstract

(1) Background: Peptides are good candidates for anticancer drugs due to their natural existence in the body and lack of secondary effects. (KLAKLAK)2 is an antimicrobial peptide that also shows good anticancer properties. (2) Methods: The Solid Phase Peptide Synthesis (Fmoc-strategy) was used for the synthesis of target molecules, analogs of (KLAKLAK)2-NH2. The purity of all compounds was monitored by HPLC, and their structures were proven using mass spectrometry. Cytotoxicity and antiproliferative effects were studied using 3T3 NRU and MTT tests, respectively. For determination of antimicrobial activity, the disc-diffusion method was used. Hydrolytic stability at three pH values, which mimic the physiological pH in the body, was investigated by means of the HPLC technique. (3) Results: A good selective index against MCF-7 tumor cell lines, combined with good cytotoxicity and antiproliferative properties, was revealed for conjugates NphtG-(KLAKLAK)2-NH2 and Caf-(KLAKLAK)2-NH2. The same compounds showed very good antifungal properties and complete hydrolytic stability for 72 h. The compound Caf-(KLβ-AKLβ-AK)2-NH2 containing β-Ala in its structures exhibited good antimicrobial activity against Escherichia coli K12 407 and Bacillus subtilis 3562, in combination with very good antiproliferative and cytotoxic properties, as well as hydrolytic stability. (4) Conclusions: The obtained results reveal that all synthesized conjugates could be useful for medical practice as anticancer or antimicrobial agents.

Highlights

Millions of people worldwide suffer from some kind of cancer
Taking into account that the introduction of unnatural amino acids in the primary structure of peptides could lead to higher activity and stability, in our previous work, we reported the synthesis and some investigations of the anticancer ability of the monomer KLAKLAK-NH2 peptide containing amino acids β-Ala and nor-Leu (Nle) in its structure [25]
A series of analogs of (KLAKLAK)2 were synthesized as C-terminal amides with the general structure (LysLeuXLysLeuXLys)2-NH2, where X is Ala or β-Ala

Summary

Introduction

Millions of people worldwide suffer from some kind of cancer. According to the data of the GLOBOCAN database, cancer diseases increased in 2020 to 19.3 million new cases, and by 2040, the database predicts more than 30 million [1]. Possible alternatives to conventional chemotherapeutics are different natural products with established anticancer properties [2,3] and molecules that can be associated with the normal function of the body as part of its metabolic processes such as peptides, carbohydrates, etc. The latter possess different biological activities such as antiviral [4,5], anticancer [6,7], analgesic [8,9], etc. They have the advantage that, as part of the structure and metabolic processes of the body, they do not show any secondary effects, or any undesired effects are relatively weak

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Conclusion