(W0930-07-37) Development of MMP2-Sensitive Carbon Nanotubes for Tumor-Targeted Drug Delivery

Purpose: Discrete multiwalled carbon nanotubes (dMWCNTs) are carbon nanotubes with high surface-to-volume ratio and tunable surface functionalization. Its ability to deliver cargos such as small molecules, peptides, nucleic acids, etc. have been demonstrated in previous studies. The purpose of this study is to engineer the dMWCNTs and develop a novel MMP2-sensitive CNT-based nanocarrier for tumor-targeted drug delivery.

Methods: We synthesized the MMP2-sensitive copolymer, poly(ethylene glycol) 2k-peptide (pp)-phosphoethanolamine (PEG-pp-PE), which showed tumor targetability. In this study, the PEG-pp-PE was used to modify the dMWCNTs to impart the MMP2 sensitivity into the nanocarriers (dMWCNT/PEG-pp-PE). The dMWCNT/PEG-pp-PE were characterized for their particle size, zeta potential, morphology, stability, and MMP2 sensitivity. The dMWCNT/PEG-pp-PE was further labeled with Rhodamine-PE (Rh-PE) for studying the cellular uptake and tissue penetration in various cell monolayers (4T1, Hela, etc.) and the 3D tumor cell spheroid model. To study the drug loading and release, doxorubicin (DOX) was loaded to the dMWCNT/PEG-pp-PE. The dMWCNTs’ drug release profile was investigated in the presence or absence of MMP2 by a dialysis method. The cytotoxicity of the DOX-loaded MMP2-sensitive dMWCNTs were evaluated in various cancer cells. The in vivo biodistribution and anticancer activity of the dMWCNT/PEG-pp-PE are under investigation using the 4T1-bearing mice as the model.

Results: Modification of the dMWCNTs (length: 450-850 nm, diameter: 12-14 nm) by PEG-pp-PE significantly decreased dMWCNTs’ particle size, probably due to the improved self-assembly (folding) and nanoparticle stability. Among the formulations, the DOX-loaded dMWCNT/PEG-pp-PE had a mean particle size of around 125±30 nm. The 4-h MMP2 incubation increased the dMWCNTs’ particle size to around 188±25 nm. This was because that the MMP2 cleaved the peptide linker (pp) and de-shielded the PEG layer from the dMWCNTs, resulting in the dMWCNTs’ “unfolding or swelling”. The DOX encapsulation efficiency in dMWCNT/PEG-pp-PE was around 75%. The loaded DOX showed sustained release under the simulated “sink” condition. In the cancer cells, the MMP2 pre-incubation almost doubled the cellular uptake of the dMWCNT/PEG-pp-PE in the absence of MMP2. The MMP2-dependent cell uptake/penetration of the dMWCNT/PEG-pp-PE was also observed in the 3D tumor cell model. As a result, the dMWCNT/PEG-pp-PE improved the DOX’s anticancer activity while the empty nanoparticles didn’t show significant toxicity in the cells.

Conclusion: The excellent physicochemical properties, stability, MMP2 sensitivity, drug loading and release, and in vitro data suggest that the MMP2-sensitive dMWCNTs might be a promising nanocarrier for tumor-targeted drug delivery.