Abstract

E-selectin is a highly glycosylated protein overexpressed on the surface of endothelial cells within the tumor vasculature, especially at sites of active angiogenesis and metastasis. This localized overexpression raises the opportunity to target the tumor microenvironment by using nanocarriers capable of specific recognition by this protein. In this work, we report dual-peptide PAMAM dendrimer conjugates as novel nanocarriers with specific E-selectin-mediated uptake properties. The conjugates were obtained from fourth-generation PAMAM dendrimers that were partially acetylated and doubly conjugated with an E-selectin targeting peptide (CIELLQAR or CIELFQAR) and the cell-penetrating peptide pTAT. Acetylation degrees close to 50% were successfully achieved, with peptide substitution ratios corresponding to 3-4 pTAT units and 2-3 E-selectin targeting peptides per dendrimer, as determined from NMR analysis. The dual-peptide conjugated dendrimers showed excellent safety profiles, with negligible intrinsic cytotoxicity in HUVEC/TERT2 and T98G cells, as models for endothelial and tumor cells. Their E-selectin-mediated uptake was confirmed in endothelial cells overexpressing E-selectin and blocked in cells treated with an anti-E-selectin antibody, with the pTAT-CIELFQAR-conjugated dendrimer having a superior performance. The best dual-peptide dendrimer conjugate was also internalized by T98G cells, which was attributed to the pTAT cell internalization properties. In preliminary assays, this system proved capable of delivering doxorubicin to tumor cells, which highlights the potential of this functional dendrimer conjugate as a novel platform for targeted cancer therapy.