Abstract
Considering our interest in the use of peptides as potential target-specific drugs or as delivery vectors of metallodrugs for various biomedical applications, it is crucial to explore improved synthetic methodologies to accomplish the highest peptide crude purity in the shortest time possible. Therefore, we compared “classical” fluorenylmethoxycarbonyl (Fmoc)-solid phase peptide synthesis (SPPS) with ultrasound(US)-assisted SPPS based on the preparation of three peptides, namely the fibroblast growth factor receptor 3(FGFR3)-specific peptide Pep1 (VSPPLTLGQLLS-NH2) and the novel peptides Pep2 (RQMATADEA-NH2) and Pep3 (AAVALLPAVLLALLAPRQMATADEA-NH2), which are being developed aimed at interfering with the intracellular protein-protein interaction(PPI) RANK-TRAF6. Our results demonstrated that US-assisted SPPS led to a 14-fold (Pep1) and 4-fold time reduction (Pep2) in peptide assembly compared to the “classical” method. Interestingly, US-assisted SPPS yielded Pep1 in higher purity (82%) than the “classical” SPPS (73%). The significant time reduction combined with high crude peptide purity attained prompted use to apply US-assisted SPPS to the large peptide Pep3, which displays a high number of hydrophobic amino acids and homooligo-sequences. Remarkably, the synthesis of this 25-mer peptide was attained during a “working day” (347 min) in moderate purity (approx. 49%). In conclusion, we have reinforced the importance of using US-SPPS towards facilitating the production of peptides in shorter time with increased efficacy in moderate to high crude purity. This is of special importance for long peptides such as the case of Pep3.
Highlights
Owing to our interest in the study of target-specific molecules that can interfere with relevant protein-protein interactions (PPI) or that can act as delivery vectors of metallodrugs for biomedical applications, namely cancer theranostics, we have been actively involved in the design and biological evaluation of relevant targeting peptides [1,2,3,4,5,6]
Targeting fibroblast growth factor receptor (FGFR) has been considered a promising approach for the therapy of various cancers, with many drug candidates reaching clinical trials, namely highly specific molecules such as peptides [8]
Is significantly higher than that observed in “classical” solid phase peptide synthesis (SPPS) (42%) (Table 2). This successful accomplishment prompted us to apply the same methodology to the synthesis of the 9-mer peptide RQMATADEA-NH2 (Pep2), a potential inhibitor of the improvement is highly relevant as it reduces the production costs and facilitates the purification of final peptide Pep1
Summary
Owing to our interest in the study of target-specific molecules that can interfere with relevant protein-protein interactions (PPI) or that can act as delivery vectors of metallodrugs for biomedical applications, namely cancer theranostics, we have been actively involved in the design and biological evaluation of relevant targeting peptides [1,2,3,4,5,6]. The ruthenium peptide conjugate TM34-Pep obtained after conjugation was more active against the FGFR-overexpressing breast cancer cells (SKBR3, FGFR+) than against those that did not overexpress that receptor (MDAMB231, FGFR–) These findings highlighted the importance of using FGFR-targeting peptides for selective drug delivery and prompted us to develop further improved cytotoxic conjugates with higher selectivity. Sonication does not cause the racemization of sensible residues as clearly demonstrated by Merlino et al and Wołczanski et al and can be applied to assemble peptides bearing amino acid residues prone to racemization [19,20] Inspired by these achievements, we decided to prepare Pep using this approach and compare it with the “classical” standard synthetic methodology in the absence of ultrasound irradiation. The latter is recognized as a potential therapeutic target for bone-related diseases, including bone cancer metastasis [22,23,24]
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