Abstract
Crotamine is a highly cationic; cysteine rich, cross-linked, low molecular mass cell penetrating peptide (CPP) from the venom of the South American rattlesnake. Potential application of crotamine in biomedicine may require its large-scale purification. To overcome difficulties related with the purification of natural crotamine (nCrot) we aimed in the present study to synthesize and characterize a crotamine analog (sCrot) as well investigate its CPP activity. Mass spectrometry analysis demonstrates that sCrot and nCrot have equal molecular mass and biological function—the capacity to induce spastic paralysis in the hind limbs in mice. sCrot CPP activity was evaluated in a wide range of tumor and non-tumor cell tests performed at different time points. We demonstrate that sCrot-Cy3 showed distinct co-localization patterns with intracellular membranes inside the tumor and non-tumor cells. Time-lapse microscopy and quantification of sCrot-Cy3 fluorescence signalss in living tumor versus non-tumor cells revealed a significant statistical difference in the fluorescence intensity observed in tumor cells. These data suggest a possible use of sCrot as a molecular probe for tumor cells, as well as, for the selective delivery of anticancer molecules into these tumors.
Highlights
Crotamine, composed of 42 amino acids, is a myo-neurotoxin derived from Crotalus durissus terrificus venom that belongs to the reptilian β-defensins—a group of small cationic antimicrobial peptides—that present high sequence variability preservation and the same three-dimensional structure
Successful achievements over the past years with the use of cell penetrating peptide (CPP) in various preclinical models have revealed their remarkable potential for clinical application [14]
The CPP activity of synthesize and characterize a crotamine analog (sCrot) reported here was extensively investigated in different cell types, whether tumor or not, at different time points, concentrations and in two and three dimension models
Summary
Crotamine, composed of 42 amino acids, is a myo-neurotoxin derived from Crotalus durissus terrificus venom that belongs to the reptilian β-defensins—a group of small cationic antimicrobial peptides—that present high sequence variability preservation and the same three-dimensional structure. Cationic CPPs are short arginine and lysine rich positively charged sequences [5,6] They can penetrate usually impermeable cell membranes and may trigger actions in the cytoplasm or the nucleus of cells, or both [7,8,9,10,11,12,13]. Despite the great potential of CPPs as a new therapeutic strategy, a limitation is emergent, due to the lack of selectivity of CPPs for specific cell types or cell organelles. This is a major obstacle to the clinical application of CPPs as, Molecules 2018, 23, 968; doi:10.3390/molecules23040968 www.mdpi.com/journal/molecules
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