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Transferrin Receptor Targeted Cellular Delivery of Doxorubicin Via a Reduction-Responsive Peptide-Drug Conjugate

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Abstract

Purpose

Transferrin receptors (TfRs) are overexpressed in tumor cells but are scarce in normal tissues, which makes TfR an attractive target for drug treatment of cancer. The objective of this study was to evaluate the potential of BP9a (CAHLHNRS) as a peptide vector for constructing TfR targeted peptide-drug conjugates and selective drug delivery.

Methods

Doxorubicin (DOX) was connected to BP9a via a disulfide-intercalating linker to afford a reduction-responsive BP9a-SS-DOX conjugate. By using HepG2 human liver cancer cells and L-O2 normal hepatic cells as TfR over-expressing and low-expressing in vitro models, respectively, TfR mediated cellular uptake of this conjugate was studied by using flow cytometry and confocal laser scanning microscopy. The in vitro cytotoxicities of the conjugate against HepG2 and L-O2 cells were examined by cell counting kit-8 (CCK-8) assay to evaluate its tumorous specificity.

Results

Cellular uptake and TfR blockage test results showed that the BP9a-SS-DOX conjugate gained entry into HepG2 cells via endocytosis mediated by TfR and mainly accumulated in cytoplasm. The in vitro antiproliferative activity of this conjugate against HepG2 cells (IC50 6.21 ± 1.12 μM) was approximately one-sixth of that of free DOX (IC50 1.03 ± 0.13 μM). However, its cytotoxic effect on L-O2 cells was obviously reduced compared with that of free DOX.

Conclusions

The BP9a-SS-DOX conjugate showed specific antiproliferative activity against HepG2 liver cancer cells. Our study suggests that BP9a has the potential to target chemotherapeutic agents to tumor cells over-expressing TfR and facilitate selective drug delivery.

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Abbreviations

CCK-8:

Cell counting kit-8

CTC:

Chlorotrityl chloride

DAPI:

4′, 6-diamidino-2-phenylindole

DIC:

N, N′-diisopropylcarbodiimide

DIPEA:

N, N-diisopropylethylamine

DMF:

N, N-dimethylformamide

DMSO:

Dimethyl sulfoxide

DOX:

Doxorubicin

EDT:

1, 2-ethanedithiol

EGFP:

Enhanced green fluorescence protein

ESI MS:

Electrospray ionization mass spectrometry

Fmoc:

9-fluorenylmethoxycarbonyl

HOBt:

1-hydroxybenzotriazole

MFI:

Mean fluorescence intensity

RP-HPLC:

Reversed-phase high-performance liquid chromatography

SPDP:

3-(2-pyridyldithio) propionic acid N-hydroxysuccinimide ester

TEA:

Triethylamine

TFA:

Trifluoroacetic acid

TfR:

Transferrin receptor

TIS:

Triisopropylsilane

TLC:

Thin-layer chromatography

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Authors and Affiliations

  1. Hebei Province Key Laboratory of Research and Development of Traditional Chinese Medicine, Institute of Chinese Mateia Medica, Chengde Medical University, Chengde, 067000, China

    Songtao Li, Hongling Zhao, Xiaoxia Mao, Ruxing Wang, Qi Liu & Guiqin Zhao

  2. Institute of Basic Medicine, Chengde Medical University, Chengde, 067000, China

    Yanfang Fan & Xiujun Liang

  3. Department of Immunology, Chengde Medical University, Chengde, 067000, China

    Lijun Xiao & Jianping Wang

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  1. Songtao Li
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  2. Hongling Zhao
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  3. Xiaoxia Mao
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  4. Yanfang Fan
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Li, S., Zhao, H., Mao, X. et al. Transferrin Receptor Targeted Cellular Delivery of Doxorubicin Via a Reduction-Responsive Peptide-Drug Conjugate. Pharm Res 36, 168 (2019). https://doi.org/10.1007/s11095-019-2688-2

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  • DOI: https://doi.org/10.1007/s11095-019-2688-2

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