Multifunctional dendrimer/combretastatin A4 inclusion complexes enable in vitro targeted cancer therapy

Mengen Zhang1 State Key Laboratory for Modification of Chemical Fibers and Polymer Materials, Donghua University, Shanghai, People’s Republic of China;2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China

Rui Guo2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China

Yin Wang2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China

Xueyan Cao2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China

Mingwu Shen2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China

Xiangyang Shi1 State Key Laboratory for Modification of Chemical Fibers and Polymer Materials, Donghua University, Shanghai, People’s Republic of China;2 College of Chemistry, Chemical Engineering and Biotechnology, Donghua University, Shanghai, People’s Republic of China;3 Centro de Química da Madeira, Universidade da Madeira, Campus da Penteada, Funchal, PortugalCorrespondence[email protected]

Abstract

Background

We report here a unique approach to using multifunctional dendrimer/combretastatin A4 (CA4) inclusion complexes for targeted cancer therapeutics.

Methods

Amine-terminated generation 5 polyamidoamine dendrimers were first partially acetylated to neutralize a significant portion of the terminal amines, and then the remaining dendrimer terminal amines were sequentially modified with fluorescein isothiocyanate as an imaging agent and folic acid as a targeting ligand. The multifunctional dendrimers formed (G5.NHAc-FI-FA) were utilized to encapsulate the anticancer drug, CA4, for targeted delivery into cancer cells overexpressing folic acid receptors.

Results

The inclusion complexes of G5.NHAc-FI-FA/CA4 formed were stable and are able to significantly improve the water solubility of CA4 from 11.8 to 240 μg/mL. In vitro release studies showed that the multifunctional dendrimers complexed with CA4 could be released in a sustained manner. Both 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide colorimetric assay and morphological cell observation showed that the inhibitory effect of the G5.NHAc-FI-FA/CA4 complexes was similar to that of free CA4 at the same selected drug concentration. More importantly, the complexes were able to target selectively and display specific therapeutic efficacy to cancer cells overexpressing high-affinity folic acid receptors.

Conclusion

Multifunctional dendrimers may serve as a valuable carrier to form stable inclusion complexes with various hydrophobic anticancer drugs with improved water solubility, for targeting chemotherapy to different types of cancer.

Keywords:

Supplementary figures

Figure S1 The photograph of G5.NHAc-FI-FA dendrimers (A) and G5.NHAc-FI-FA/CA4 complexes (B) dispersed in water at pH values of 5.0, 7.0, and 10.0.

Abbreviations: G5, generation 5; G5.NHAc-FI-FA, fluorescein isothiocyanate-modified and folic acid-modified G5 PAMAM dendrimers with acetyl terminal groups; CA4, combretastatin A4.

Figure S2 G5.NHAc-FI-FA/CA4 complexes dispersed in (A) water, (B) phosphate-buffered saline, and (D) cell culture medium. (C) Cell culture medium without complexes.

Abbreviations: G5, generation 5; G5.NHAc-FI-FA, fluorescein isothiocyanate-modified and folic acid-modified G5 PAMAM dendrimers with acetyl terminal groups; CA4, combretastatin A4.

Figure S3 Dose-dependent viability of KB cells treated with free CA4 and G5.NHAc-FI-FA/CA4 complexes.

Abbreviations: G5, generation 5; G5.NHAc-FI-FA, fluorescein isothiocyanatemodified and folic acid-modified G5 PAMAM dendrimers with acetyl terminal groups; CA4, combretastatin A4.

Figure S4 Confocal microscopic analysis of distribution of G5.NHAc-FI-FA/CA4 complexes into KB-HFAR cells. The cells were incubated with (A) phosphate-buffered saline for one hour, (B) G5.NHAc-FI-FA/CA4 complexes for one hour, and (C) G5.NHAc-FI-FA/CA4 complexes for 2 hours, respectively. In (B) and (C), cells were treated with Lyso Tracker Red (50 nM) for an additional one hour after treatment with G5.NHAc-FI-FA/CA4 complexes. The upper left of each panel shows the green fluorescence of fluorescein isothiocyanate, the upper right of each panel shows the red fluorescence of Lyso Tracker Red, the lower left of each panel shows the differential interference contrast images, and the lower right of each panel shows merged images with the above three modes.

Abbreviations: G5, generation 5; G5.NHAc-FI-FA, fluorescein isothiocyanate-modified and folic acid-modified G5 PAMAM dendrimers with acetyl terminal groups; CA4, combretastatin A4.

Acknowledgment

Rui Guo and Mengen Zhang equally contributed to this work. This research was funded by the National Natural Science Foundation of China (20974019 and 81101150), the Program for Professor of Special Appointment (Eastern Scholar) at Shanghai Institutions of Higher Learning, the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry, and the Fundamental Research Funds for the Central Universities (for RG, MS, and XS). RG thanks the Shanghai Natural Science Foundation (10ZR1400800). XS gratefully acknowledges the Fundação para a Ciência e a Tecnologia and the Santander Bank for supporting a Chair in Nanotechnology.

Disclosure

The authors report no conflicts of interest relevant to this research.