Enzyme-activated nanoconjugates for tunable release of doxorubicin in hepatic cancer cells.

Abstract

We report the synthesis of a series of aromatic azo-linkers (L1-L4), which are selectively recognized and cleaved by azoreductase enzymes present in the cytoplasm of hepatic cancer cells via a NADPH-dependent mechanism. We utilized L1-L4 azo-linkers to conjugate doxorubicin to generation 5 (G5) of poly(amidoamine) dendrimers to prepare G5-L(x)-DOX nanoconjugates. We incorporated electron-donating oxygen (O) or nitrogen (N) groups in the para and ortho positions of L1-L4 azo-linkers to control the electronegativity of G5-L(x)-DOX conjugates and investigated their cleavage by azoreductase enzymes and the associated release of loaded DOX molecules. Hammett σ values of G5-L(x)-DOX conjugates ranged from -0.44 to -1.27, which is below the reported σ threshold (-0.37) required for binding to azoreductase enzymes. Results show that incubation of G5-L1-DOX (σ = -0.44), G5-L2-DOX (σ = -0.71), G5-L3-DOX (σ = -1.00), and G5-L4-DOX (σ = -1.27) conjugates with human liver microsomal (HLM) enzymes and the S9 fraction isolated from HepG2 hepatic cancer cells results in release of 4%-8%, 17%, 60%, and 100% of the conjugated DOX molecules, respectively. These results show that increasing the electronegativity (i.e. lower σ value) of L1-L4 azo-linkers increases their susceptibility to cleavage by azoreductase enzymes. Intracellular cleavage of G5-L(x)-DOX nanoconjugates, release of conjugated DOX molecules, and cytotoxicity correlated with conjugate's electronegativity (σ value) was investigated, with G5-L4-DOX conjugate exhibiting the highest toxicity towards hepatic cancer cells with an IC50 of 13 nm ± 5 nm in HepG2 cells. Cleavage of G5-L(x)-DOX conjugates was specific to hepatic cancer cells as shown by low non-specific DOX release upon incubation with non-enzymatic insect proteins and the S9 fraction isolated from rat cardiomyocytes. These enzyme-activated G5-L(x)-DOX conjugates represent a drug delivery platform that can achieve tunable and cell-specific release of the loaded cargo in hepatic cancer cells.

• Authors: Scott H Medina, Maxim V Chevliakov, Gopinath Tiruchinapally, Yasemin Yuksel Durmaz, Sibu P Kuruvilla, Mohamed E H Elsayed
• Journal: Biomaterials (Biomaterials) Vol. 34 Issue 19 Pg. 4655-66 (Jun 2013) ISSN: 1878-5905 [Electronic] Netherlands
• PMID: 23523429 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2013 Elsevier Ltd. All rights reserved.

Chemical References

• Antibiotics, Antineoplastic
• Delayed-Action Preparations
• Dendrimers
• Nanoconjugates
• Poly(amidoamine)
• Polyamines
• Doxorubicin

Topics

• Animals
• Antibiotics, Antineoplastic (administration & dosage, pharmacology, toxicity)
• Cells, Cultured
• Delayed-Action Preparations (chemistry, metabolism, toxicity)
• Dendrimers (chemistry, metabolism, toxicity)
• Doxorubicin (administration & dosage, pharmacology, toxicity)
• Hep G2 Cells
• Humans
• Liver (drug effects, pathology)
• Liver Neoplasms (drug therapy, pathology)
• Myocytes, Cardiac (drug effects)
• Nanoconjugates (chemistry, toxicity)
• Polyamines (chemistry, metabolism, toxicity)
• Rats