| Abstract |
The important role of self-assembly in biological systems has inspired the scientific community to create artificial materials with defined structures and customized properties. Thus, both biological and synthetic building blocks are the subject of extensive research in the field of self-assembly. Τhe value of peptides as selfassembled building blocks has been largely recognized in recent years. In particular, the covalent coupling of diphenylalanine, known for its self-assembling properties, has been extensively studied in the past, leading to the creation of well-organized structures. Motivated by previous studies, we have developed a hybrid triad of diphenylalanineporphyrin-lysine, FF-DMP-PCP-Lys-(COOMe)3. The coupling of both diphenylalanine and lysine to the porphyrin derivative took place via an amide bond. The complete characterization of the final product was performed through mass spectrometry (MALDI-TOF), 1H and 13C NMR spectroscopy as well as UV-Vis absorption spectroscopy. The tendency of the FF-DMP-PCP-Lys-(COOMe)3 compound to selfassemble was examined using Electronic Scanning Microscopy, which showed the formation of spherical nanostructures. At the same time, bio-imaging often requires specific labeling of biomolecules, making it necessary to develop new, non-invasive methods for their detection. A very promising method that meets the above criteria is fluorescent labeling using fluorescent probes or tags. In order to detect biomolecules such as proteins and peptides, fluorescent proteins or fluorescent chromophores are often used. One of the most well-known labeling methods is the metallochelate coupling of fluorophores with the amino acid residues (eg His) of the peptide/protein to be analyzed. However, there are few references in the literature that include porphyrins as fluorescents in cases of metallochelate coupling. Thus, we propose the synthesis of a new water-soluble porphyrin dyad with a chellating ligand for peptide labeling (RGD-SGAITIG-H) bearing histidine (His). Peptide labeling is achieved by metallochelate coupling of the histidine imidazole and the NTA-containing porphyrin dyad via Ni2+ . The synthesis of the dyad, TriPyPLys(COOH) )3, takes place by the formation of an amide bond between porphyrin and the chelating ligand (Lys-NTA). The complete characterization of the TriPyP- xvi Lys(COOH) )3 was performed via 1H and 13C NMR spectroscopy, UV-Vis absorption and fluorescence spectroscopy, and fluorescence life-time. The final complex was characterized by UV-Vis, CD and IR spectroscopies. At the same time, DLS and zeta potential measurements were performed to determine the size and charge of the formed nanostructures. The ability of intracellular transport and the localization of the complex in cell lines of human breast adenocarcinoma MCF-7 were studied using confocal microscopy. CD44 protein receptor has been found to be overexpressed by many tumors and is recognized as one of the most common Cancer Stem Cells surface markers, making it a promising receptor for therapeutic targeting. At the same time, aptamers are synthetic single-stranded oligonucleotides or peptides that can act against almost any target including ions, molecules, peptides, proteins and even entire living cells such as cancer cells. Thus, aptamers constitute promising molecules with high potential in biomedical applications. Inspired by the forenamed, we recommend the synthesis of a dyad of porphyrinpolyethylene glycol, Zn-TPP-PEG-MAL that carries free maleimide. The dyad-target is conjugated by a thiol-maleimide "click" reaction to an RNA aptamer (Apt1) containing 2′-F-pyrimidine and has been studied to act against the CD44 biomarker. Thus, we aim to bind the porphyrin-aptamer system to cancer cells and to study it as a specific drug delivery system leading to cell apoptosis through photodynamic therapy.
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