Tumor-targeted intracellular delivery of anticancer drugs through the mannose-6-phosphate/insulin-like growth factor II receptor.

Tumor-targeting of anticancer drugs is an interesting approach for the treatment of cancer since chemotherapies possess several adverse effects. In the present study, we propose a novel strategy to deliver anticancer drugs to the tumor cells through the mannose-6-phosphate/insulin-like growth factor receptor (M6P/IGF-IIR) which are abundantly expressed in several human tumors. We developed a drug carrier against M6P/IGF-II receptor by modifying human serum albumin (HSA) with M6P moieties. M6P-HSA specifically bound and internalized into M6P/IGF-IIR-expressing B16 melanoma cells as demonstrated with radioactive studies and anti-HSA immunostaining. In vivo, M6P-HSA rapidly accumulated in subcutaneous tumors in tumor and stromal components after an intravenous injection. To demonstrate the application of M6P-HSA as a drug carrier, we coupled doxorubicin to it. Dox-HSA-M6P conjugate could release doxorubicin at lysosomal pH and showed M6P-specific binding and uptake in tumor cells. In vitro, a short exposure with Dox-HSA-M6P induced killing of tumor cells, which could be blocked by excess M6P-HSA. In vivo, Dox-HSA-M6P distributed to tumors and some other organs while free doxorubicin distributed to all organs but slightly to tumors. In B16 tumor-bearing mice, Dox-HSA-M6P significantly inhibited the tumor growth whereas an equimolar dose of free doxorubicin did not show any anti-tumor effect. In addition, targeted doxorubicin did not show any side-effects on liver and kidney function tests, body weight and blood cell counts. In conclusion, M6P-HSA is a suitable carrier for delivery of anticancer drugs to tumors through M6P/IGF-IIR. Improved antitumor effects of the targeted doxorubicin by M6P-HSA suggest that this novel approach may be applied to improve the therapeutic efficacy of anticancer drugs.

The antiproliferative drug doxorubicin inhibits liver fibrosis in bile duct-ligated rats and can be selectively delivered to hepatic stellate cells in vivo.

Hepatic stellate cell (HSC) proliferation is a key event in liver fibrosis; therefore, pharmacological intervention with antiproliferative drugs may result in antifibrotic effects. In this article, the antiproliferative effect of three cytostatic drugs was tested in cultured rat HSC. Subsequently, the antifibrotic potential of the most potent drug was evaluated in vivo. As a strategy to overcome drug-related toxicity, we additionally studied how to deliver this drug specifically to HSC by conjugating it to the HSC-selective drug carrier mannose-6-phosphate-modified human serum albumin (M6PHSA). We investigated the effect of cisplatin, chlorambucil, and doxorubicin (DOX) on 5-bromo-2'-deoxyuridine incorporation in cultured HSC and found DOX to be the most potent drug. Treatment of bile duct-ligated (BDL) rats with daily i.v. injections of 0.35 mg/kg DOX from day 3 to 10 after BDL reduced alpha-smooth muscle actin-stained area in liver sections from 8.5 +/- 0.8 to 5.1 +/- 0.9% (P < 0.01) and collagen-stained area from 13.1 +/- 1.3 to 8.9 +/- 1.5% (P < 0.05). DOX was coupled to M6PHSA, and the organ distribution of this construct (M6PHSA-DOX) was investigated. Twenty minutes after i.v. administration, 50 +/- 6% of the dose was present in the livers, and colocalization of M6PHSA-DOX with HSC markers was observed. In addition, in vitro studies showed selective binding of M6PHSA-DOX to activated HSC. Moreover, M6PHSA-DOX strongly attenuated HSC proliferation in vitro, indicating that active drug is released after uptake of the conjugate. DOX inhibits liver fibrosis in BDL rats, and HSC-selective targeting of this drug is possible. This may offer perspectives for the application of antiproliferative drugs for antifibrotic purposes.

Membrane-permeant derivatives of mannose-1-phosphate.

For treatment of congenital disorder of glycosylation type Ia (CDG-Ia) membrane-permeant derivatives of mannose-1-phosphate are required. Employing biologically cleavable phosphate protecting groups advantageous precursor derivatives could be synthesized following a facile approach. Their enzymatic cleavages using esterase from porcine liver (E.C. 3.1.1.1) were investigated.

Characteristics of the hepatic stellate cell-selective carrier mannose 6-phosphate modified albumin (M6P(28)-HSA).

BACKGROUND/AIMS: Drug targeting to hepatic stellate cells (HSC) may improve the pharmacological effects of antifibrotic drugs. Recently, albumin substituted with 28 mannose 6-phosphate moieties (M6P(28)-HSA) was found to distribute selectively to HSC in fibrotic rat livers. To assess whether this albumin can be used as a carrier for intracellular drug delivery, we explored the cellular handling of M6P(28)-HSA in HSC. METHODS/RESULTS: Application of competitive substrates for the M6P/IGFII receptor or other receptors showed that the binding of M6P-HSA to the M6P/IGFII receptor is specific. Binding was strong to activated HSC, but not to quiescent HSC. Furthermore, M6P(28)-HSA was extensively internalized by these cells. Using monensin, a specific inhibitor of the lysosomal pathway, proof was obtained that M6P-HSA is endocytosed via this route. The experiments performed with tissue slices, prepared from rat and human livers, revealed a specific binding and uptake of M6P(28)-HSA in both normal and cirrhotic livers. In livers from cirrhotic patients, HSC contributed predominantly to the uptake of this neoglycoprotein. CONCLUSIONS: Based on our in vivo data demonstrating the HSC-selectivity and on our in vitro data demonstrating binding and rapid internalization in activated HSC, we conclude that M6P(28)-HSA is applicable as a stellate cell-selective carrier for antifibrotic drugs that act intracellularly. This may have implications for the design of new strategies for the treatment of liver fibrosis.