Boronic acid-rich dendrimer for efficient intracellular peptide delivery

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  • ReceivedSep 25, 2019
  • AcceptedOct 24, 2019
  • PublishedNov 20, 2019


Interests in intracellular peptide delivery have continued to grow, significantly fueled by the importance of peptides and their mimetics in modern cell biology and pharmaceutical industry. However, efficient intracellular delivery of membrane-impermeable peptides of different polarities remains a challenging task. In this study, we develop a general and robust strategy for intracellular peptide delivery by using a boronic acid-rich dendrimer. The designed material is capable of transporting peptides with different polarities and charge properties into the cytosol of various cell lines without inducing additional cytotoxicity. The transduction efficacy and proteolytic stability of cargo peptides delivered by the boronic acid-rich dendrimer are much superior to peptides conjugated with cell penetrant peptides such as octaarginine. In addition, the bioactivities of pro-apoptotic peptides are maintained after intracellular delivery. This study provides a versatile and robust platform for the intracellular delivery of membrane-impermeable peptides.

Funded by

the National Natural Science Foundation of China(21725402)

Science and Technology Commission of Shanghai Municipality(17XD1401600)

and Guangdong Innovative and Entrepreneurial Research Team Program(2016ZT06C322)


This work was supported by the National Natural Science Foundation of China (21725402), the Science and Technology Commission of Shanghai Municipality (17XD1401600), and Guangdong Innovative and Entrepreneurial Research Team Program (2016ZT06C322). We thank the supports from the Flow Cytometry Core Facility and the Confocal Microscopy Facility at ECNU.

Interest statement

The authors declare that they have no conflict of interest.

Contributions statement

Lv J synthesized and tested the peptide delivery efficacy of the polymer; Liu C performed part of peptide delivery experiments; Lv K contributed to the characterization of BDP/peptide complexes; Cheng Y and Wang H contributed to the theoretical analysis; Cheng Y and Lv J wrote the paper; all authors contributed to the general discussion.

Author information

Jia Lv received her PhD degree from the East China Normal University in 2018. She is currently working as a postdoctoral fellow at South China University of Technology. Her research interests mainly focus on the intracellular delivery of biomacromolecules such as proteins and peptides.

Yiyun Cheng is a full professor of biomedical engineering at the School of Life Sciences, East China Normal University. He received his PhD degree from the University of Science and Technology of China and was a postdoctoral fellow at Washington University in St. Louis, MO. His research interests focus on the rational design of polymers for the delivery of biomacromolecules such as DNA, RNA, proteins and peptides.


Supplementary information

Supporting data are available in the online version of the paper.


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  • Figure 1

    Synthesis of BDP and its efficacies in cytosolic delivery of peptides with various properties. (a) Synthesis of BDP and the preparation of BDP/peptide complexes. (b) Properties of cargo peptides P1–P6, the hydrophobicity was calculated according to Kyte-Doolittle hydrophobicity value (Kd) [48]. Intracellular peptide delivery into HeLa cells by BDP for 4 h was measured by flow cytometry (c) and confocal microscopy (d). Peptides without addition of BDP (peptide only) were used as negative controls. The concentrations of peptides and BDP were 13.3 and 20 μg mL−1, respectively.

  • Figure 2

    The comparison of BDP with CPP. (a) Confocal images of HeLa cells incubated with BDP/peptide, TAT- and R8-conjugated peptides for 4 h. (b) Flow cytometry of cells incubated by BDP/P4, R8-P4, TAT-P4, respectively, for 4 h. The samples were pre-treated without or with 4 μg mL−1 trypsin (Trp) for 2 h before incubation with cells. (c) Relative fluorescence intensities of HeLa cells treated with BDP/peptide, R8-peptide, or TAT-peptide, respectively, for 4 h. The samples were pre-treated without or with 4 μg mL−1 trypsin for 2 h before incubation with cells. For the samples without trypsin treatment, the relative fluorescence intensity was set as 100%. The concentrations of peptides (P2, P4 and P5) and BDP were 13.3 and 20 μg mL−1, respectively. The molar concentrations of R8- and TAT-conjugated peptides were equal to those of P2, P4 and P5 in the BDP complexes, respectively.

  • Figure 3

    Internalization pathways of BDP/P2 complex by HeLa cells and peptide delivery efficacy of BDP on other cell lines. Cells were pre-incubated with inhibitors including EIPA (inhibitor of macropinocytosis), genistein (inhibitor of caveolin-involved pathway), CPZ (inhibitor of clathrin-involved pathway) and MβCD (inhibitor of lipid raft) for 1 h before intracellular peptide delivery. The cells treated with BDP/P2 complex for 4 h were observed by confocal microscope (a) or quantitatively measured with flow cytometry (b). (c) Confocal images of HEK 293, MDA-MB-231, MCF-7, BAT, and iWAT treated with BDP/P2 complexes for 4 h. The concentrations of P2 and BDP were 13.3 and 20 μg mL−1, respectively. The scale bar is 30 μm.

  • Figure 4

    Intracellular delivery of AG5 by BDP. Fluorescence images (a) and intensity (b) of HeLa cells treated with BDP/AG5-FITC, R8-AG5-FITC, and AG5-FITC, respectively, for 4 h. (c) LDH release from cells treated with AG5, BDP, BDP/AG5 complex and R8-AG5 for 24 h. (d) Confocal images of cells with calcein treatment after incubation with AG5, BDP, BDP/AG5 complex and R8-AG5 for 24 h, respectively. (e) Apoptosis of cells treated with AG5, BDP, BDP/AG5 complex, and R8-AG5 for 24 h. The concentrations of BDP, AG5-FITC and AG5 were 20, 13.3, and 83.3 μg mL−1, respectively. The molar concentration of R8-AG5 was equal to that of AG5 used in BDP/AG5 complex.

  • Figure 5

    Intracellular delivery of CC11 by BDP. Fluorescence images (a) and mean fluorescence intensity (b) of cells incubated with BDP/CC11 complex and CC11, respectively, for 4 h. The peptide was labeled with FITC to monitor its intracellular delivery. (c) LDH release from HeLa cells treated with BDP/CC11, BDP, and CC11 for 24 h. (d) Apoptosis of HeLa cells incubated with CC11, BDP, and BDP/CC11 complex for 24 h. The concentrations of BDP, CC11-FITC, and CC11 were 20, 13.3, and 50 μg mL−1, respectively.

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