cAmbly: A non-toxic cell-penetrating peptide derived from Amblyomin-X with targeted delivery to mitochondrial and cytoplasmic proteins.

The effective delivery of drugs remains a major challenge in the development of new therapeutic molecules. Several strategies have been employed to address this issue, with cell-penetrating peptides (CPPs) standing out due to their ability to traverse cell membranes with minimal cytotoxicity and their relatively straightforward synthesis when conjugated with other molecules. However, while CPPs can successfully enter the cytoplasm, they often lack specificity for particular organelles, leaving target engagement to the drug itself. In this study, we present cAmbly, a novel CPP derived from the antitumoral protein Amblyomin-X. Our findings demonstrate that cAmbly efficiently internalizes into T98G cells within 30 min of incubation and preferentially colocalizes with mitochondria, exhibiting a clear affinity for mitochondrial proteins. These results suggest that cAmbly could serve as a promising delivery vehicle for mitochondria-targeted drugs, such as BCL-2 or OXPHOS modulators, which are commonly employed in cancer treatment. Furthermore, we identified the C-terminal of cAmbly as the optimal site for conjugating molecules intended for intracellular delivery.