Elsevier

Acta Biomaterialia

Volume 68, 1 March 2018, Pages 308-319
Acta Biomaterialia

Full length article
pH-responsive gold nanoclusters-based nanoprobes for lung cancer targeted near-infrared fluorescence imaging and chemo-photodynamic therapy

https://doi.org/10.1016/j.actbio.2017.12.034Get rights and content

Abstract

Nanoparticle-based drug delivery systems have drawn a great deal of attention for their opportunities to improve cancer treatments over intrinsic limits of conventional cancer therapies. Herein, we developed the polypeptide-modified gold nanoclusters (GNCs)-based nanoprobes for tumor-targeted near-infrared fluorescence imaging and chemo-photodynamic therapy. The nanoprobes comprise of tetra-functional components: i) polyethylene glycol (PEG) shell for long blood circulation and better biocompatibility; ii) MMP2 polypeptide (CPLGVRGRGDS) for tumor targeting; iii) cis-aconitic anhydride-modified doxorubicin (CAD) for pH-sensitive drug release; iv) photosensitizer chlorin e6 (Ce6) for photodynamic therapy and fluorescence imaging. The in vitro results demonstrated that the as-synthesized nanoprobes could be efficiently internalized into A549 cells and then significantly enhance the mortality of cancer cells compared with free Ce6 and doxorubicin. For in vivo tests, the nanoprobes showed excellent tumor targeting ability, long blood circulation time, and could remarkably inhibit the growth of tumor. Our results will help to advance the design of combination strategies to enhance the efficacy of imaging-guided cancer therapy.

Statement of Significance

The as-prepared CDGM NPs could accumulate into the tumor tissue with the enhanced permeability and retention (EPR) effect as well as the active tumor targeting ability from the MMP2 polypeptides.

With the acid-sensitive linker, the doxorubicin (DOX) would be released from the synthesized nanoparticles after exposing to the acid tumor microenvironment.

The CDGM NPs exhibit excellent tumor targeting ability and could remarkably suppress the growth of tumor compared with free Ce6 and DOX.

Introduction

Lung carcinoma, the highest prevalence and mortality of cancer, continues to be an unabating disease until now [1]. Chemotherapy plays an important role in current cancer treatments. However, the chemotherapy drugs with poor pharmacokinetics are easily diffused throughout the normal tissue of patients, resulted in severe systemic side effects and unsatisfactory curative effects [2], [3], [4]. Combining multiple cancer treatment modalities are regarded as a promising way to enhance anticancer efficiency and minimize the side effects [5], [6]. Recently, photosensitizers combined with nanomaterials as multifunctional photodynamic therapy (PDT) agents have become the popular choice for cancer therapy research [7]. PDT is a clinically curative technique used for tumor treatment by inducing apoptosis or necrosis of cancer cells via the activation of certain chemical reaction [8], [9]. Most importantly, PDT can significantly improve the quality of patients’ life by irradiation at an appropriate wavelength and can be used simultaneously with chemotherapy without compromising the therapeutic efficiency [9]. Moreover, photosensitizer, an essential element of PDT, can also be employed in cancer imaging diagnosis as it can emit the near-infrared (NIR) light [8], [10]. Therefore, the NIR photosensitizer like chlorin e6 (Ce6) is widely used for the research of both cancer PDT treatment and fluorescence imaging [8], [11], [12], [13]. Unfortunately, most of the photosensitizers are hydrophobic which makes them insoluble under physiological conditions and hampered their clinical application.

Developing rapid and efficient delivery systems for improving the water solubility and tumor accumulation effect of photosensitizer may be a promising strategy for their application in cancer imaging and therapy. Glutathione-protected gold nanoclusters (GNCs) have attracted increasing attention in the field of bioimaging, sensing, and drug delivery [14]. In contrast to the spherical gold nanoparticles, the GNCs exhibit fluorescence in the visible to near-infrared (NIR) region while have no surface plasmon resonance in the visible region [14]. Additionally, GNCs possess sizes about 2 nm and they draw great attention due to the excellent biocompatibility and less renal toxicity [15], [16], [17], [18], [19]. Furthermore, chemical composition on the surface of GNCs makes it possible to be covalently conjugated with therapeutic or diagnostic agent for cancer diagnosis and therapy [19], [20]. For example, Li et al. [21] reported that the cisplatin (Pt) prodrug and folic acid (FA) modified GNCs (FA-GNC-Pt) could be used for bioimaging and targeting treatment on breast cancers. GNCs-based nanoprobes which cross-linking FA (folic acid), PEG (polyethylene glycol) with GNCs and loading with photosensitizer (Ce6), were also applied for imaging and targeted therapy of tumors [22]. That is, the GNCs would provide a promising way for drug and photosensitizer delivery and subsequently for cancer chemo-photodynamic combination therapy. However, the clinic application of GNCs is limited without the tumor targeting properties, a very necessary ability for tumor diagnosis and therapy. Deducibly, targeting ligands modification on the surface would improve the clinic cancer diagnosis and therapy ability of the GNCs. Matrix metalloproteinases (MMPs) belong to the zinc and calcium-dependent family and are usually associated with cancer proliferation, metastasis, and angiogenesis [23], [24]. MMP2 are one kind of MMPs, which are elevated in various human cancers, have been widely developed as linker or target moieties for tumor-imaging diagnosis and therapy [12], [25], [26].

In recent decades, many nanoscale drug delivery systems have been designed to improve the therapeutic efficacy [27], [28]. As an appealing quest for therapeutic purposes, development of stimuli-responsive nanoplatforms are one of the most effective strategies for cancer therapy since they can realize accelerated drug release at the target site [29]. pH controlled delivery has been widely used to design responsive nanoparticles because the tumor extracellular environment is more acidic (pH ∼ 6.5) compared with normal tissues and blood (∼7.4) [30], [31], [32], [33]. In addition, up-regulation of certain enzymes such as MMP2 in pathological conditions (cancer), can be developed to delivery drugs to tumor regions [34], [35]. To the best of our knowledge, pH- responsive drug delivery systems and MMP2 polypeptides as target moiety based on GNCs have not yet been reported.

Herein, we designed a novel type of gold nanoclusters-based tumor targeted Ce6-DOX(doxorubicin)-GNCs-MMP2 polypeptide nanoparticles (CDGM NPs) for tumor targeted imaging and combination therapy (Scheme 1). The CDGM NPs were prepared by chemical conjugation of Ce6, pH-sensitive cis-aconitic anhydride-modified doxorubicin (CAD), and polyethylene glycol (PEG) modified polypeptide (CPLGVRGRGDS) to the GNCs. These multifunctional CDGM NPs would accumulate into the tumor tissue with the enhanced permeability and retention (EPR) effect as well as the active tumor targeting ability from the polypeptides. With the PEG on the surface, the blood circulation time and biocompatibility of the GNCs could be improved significantly [36]. And accelerated release of DOX could also be realized due to the acid tumor microenvironment (pH = 6.5), where the acid-sensitive cis-aconityl linkage between the DOX and the cis-aconitic anhydride (CA) would be hydrolyzed. Our elaborate design of CDGM NPs provides a great potential in simultaneous cancer chemo-photodynamic therapy and fluorescence imaging application.

Section snippets

Materials

Doxorubicin hydrochloride (DOX·HCl) was purchased from Dalian Meilun Biotech Co., Ltd (Dalian, China). MMP2-cleavable polypeptide (CPLGVRGRGDS) with a thiol group was supplied from ChinaPeptides Co., Ltd (Shanghai, China). Cis-Aconitic anhydride (CA) was obtained from Meryer Chemical Technology (Shanghai, China). Chlorin e6 (Ce6) was obtained from Frontier Scientific (Utah, USA). Gold chloride trihydrate (HAuCl4·3H2O, 99%) and GSH (98%) were purchased from Sinopharm Chemical Reagent Co., Ltd.

Synthesis and characterization of CDGM NPs

GSH-capped GNCs (Fig. S2A) with average size about 2 nm (Fig. 1A) were synthesized according to a modified method [22], [38]. The CDGM NPs were synthesized as follows. Briefly, cis-aconitic anhydride modified doxorubicin (DOX) (Fig. S2B) and Ce6 were simply conjugated to the amine groups of GNCs to obtain Ce6-DOX-GNCs nanoparticles (CDG NPs) (Fig. S3A). Then, CDGM NPs were generated by chemically linking mPEG-modified MMP2 polypeptides with CDG NPs (Fig. S3B). The as-prepared CDGM NPs show

Conclusion

GNCs-based nanoprobes for lung cancer NIR fluorescence imaging and chemo-photodynamic therapy were successfully prepared and then employed for drug delivery and biomedical imaging applications. The CDGM NPs showed good monodispersity and high stability in aqueous solution. They possessed the ability to promote the cellular uptake of Ce6 and DOX and exhibited significantly higher cancer cell growth inhibition than Mix (free Ce6&DOX) and Ce6-DOX-GNCs nanoparticles (CDG NPs) at equivalent

Acknowledgments

This work is supported by the National Basic Research Program (973 Project) (Project No. 2015CB931802), the National Key Research and Development Program of China (Grant No. 2017FYA0205301), the National Natural Scientific Foundation of China (Grant Nos. 81401458, 81225010, 81028009, and 31170961), the 863 projects of China (Project No. 2014AA020701), China Postdoctoral Science Foundation (Grant No. 2017M621486) and the Shanghai Science and Technology Fund (Grant No. 14ZR1432400 and 13NM1401500

Competing interests

The authors declare no conflicts of interest.

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