Elsevier

Composites Part B: Engineering

Volume 221, 15 September 2021, 109031
Composites Part B: Engineering

Injectable halloysite-g-chitosan hydrogels as drug carriers to inhibit breast cancer recurrence

https://doi.org/10.1016/j.compositesb.2021.109031Get rights and content

Highlights

  • The gels have thermosensitivity, injectability and good mechanical properties as slow and controlled release drug carriers.

  • The gels have good biocompatibility, and exert excellent tumoricidal effect in vitro and in vivo after loading doxorubicin.

  • The hydrogels can prevent tumor recurrence efficiently while promoting new tissue generation after tumor resection.

Abstract

Breast cancer is one of the frequently occurring malignant tumors in the world. Breast resection, high tumor recurrence rate and high dose chemotherapy cause great pain to patients. Is there a way to prevent tumor recurrence efficiently while promoting new tissue generation after tumor resection? Based on this, an injectable biocompatible thiolated chitosan (CSSH) hydrogel with pH-response was designed to be used for accurate treatment of tumor cells in the initial stage of tissue resection and for tissue reconstruction later. To improve the mechanical strength of hydrogels, reduce the toxic and side effects, and delay the release of doxorubicin (DOX), halloysite nanotubes (HNTs) were modified to obtain thiolated HNTs (HNTs-SH), then loaded DOX (DOX@HNTs-SH), and further cross-linked CSSH to form DOX@CSSH/HNTs-SH Gel. The results showed that HNTs-SH can be evenly dispersed in the gel matrix, which improve the compressive strength of the hydrogels. In vitro DOX release experiments showed that the hydrogels were pH sensitive, DOX released slowly at normal physiological pH, but released quickly in the acidic microenvironment of tumors. Cell experiments showed that DOX@CSSH/HNTs-SH Gel released DOX can be taken up by MCF-7, thereby effectively inhibited its growth. In vivo recurrence experiments showed that the carriers can reduce the toxic and side effects of DOX, and effectively inhibited the recurrence and repair the defect tissue after tumor resection. Therefore, the drug-loaded gel is expected to be used as a carrier for local accurate and effective drug release, especially for inhibiting recurrence and repairing defective tissues after tumor resection.

Introduction

Cancer is one of the diseases that seriously threatens human life [1]. Surgical treatment is easy to produce residual tumor cells, cause trauma, and relapse after surgery. Chemotherapy will be affected by the toxic and side effects, drug resistance, poor targeting and poor water solubility of anticancer drugs [2]. Therefore, it is of great significance to design a drug carrier that can effectively load drugs, inhibit postoperative tumor recurrence and treat surgical wounds.

The method of drug administration is very important. For example, intravenous injection of the drug solution or oral administration is easy to be metabolized, cannot effectively reach the lesion site, has low drug availability, requires repeated administration, and has strong toxic and side effects. Therefore, it is very important to continuously release the drugs at the lesion site. The injectable thermosensitive hydrogel can form hydrogel at physiological temperature, fill the defect site and slowly release the drugs locally, thereby improving the availability of the drugs [3]. Therefore, injectable thermosensitive hydrogels have good application prospects in sustained-release drug carriers.

Chitosan (CS), as a naturally positive alkaline polysaccharide, has the advantages of degradability, antibacterial properties, and good biocompatibility, but it is insoluble in water, and the mechanical strength of chitosan-based hydrogel is poor, which limits its applications [4]. The in-situ injectable CS hydrogel is fluid at room temperature, can be injected locally in the body for rapid gelation under the physiological temperature conditions, filling defects in different shapes, and mixing various therapeutic drugs [5], thereby reducing surgical trauma and reducing costs, and is widely used in drug delivery [6]. The preparation methods of injectable CS hydrogels are mainly physical cross-linking, UV-induced cross-linking or the introduction of chemical cross-linking agents, but the pure physical cross-linking hydrogels have poor mechanical strength and long gel time, such as chitosan/β-Sodium glycerophosphate gel [7], chemically cross-linked hydrogels have good strength, while photoinitiators or chemical cross-linking agents have certain potential toxic effects on cells, and chemically cross-linked gels can not degrade [8,9]. The poor compressibility and elasticity of hydrogels are not conducive to its in-situ injectable formation and postoperative suture. Therefore, it is of great significance to prepare a chitosan hydrogel that can enhance mechanical properties and avoiding cytotoxicity.

Halloysite nanotubes (HNTs) is one-dimensional hollow tubular nanomaterials, its molecular formula is Al2Si2O5(OH)4.nH2O, its length is between 200 nm and 2 μm, and its outer diameter and inner diameter are 50–70 nm and 10–20 nm, respectively. The inner surface is aluminum hydroxyl group, and the outer surface is composed of silicon hydroxyl group [10]. It has many advantages in the application of biomaterials, including the hollow structure with high length to diameter ratio, different chemical groups on the inner and outer surfaces, high water dispersion stability, high adsorption, good biocompatibility, environmental friendliness and high biosecurity [11]. Because HNTs has high length to diameter ratio, high adsorption and the silicon hydroxyl groups, it can adsorb doxorubicin (DOX) [12,13], decorate with CS as effective reinforcement in the system [14,15], enhance gel strength [16], modify the surface of HNTs with polyethylene glycol (PEG) and folic acid (FA) [17]. In summary, it can be seen that HNTs with good biocompatibility can be used as drug carriers, load and slow-release drugs, prolong the action time of drugs, improve the efficiency of drugs entering tumor cells, and increase the strength of hydrogels. Therefore, it has potential application value in the fields of drug carriers and tissue engineering scaffolds. In this study, HNTs can be modified through the surface silicon hydroxyl groups to produce reactive functional groups, load and slow-release drugs, and improve the strength of the hydrogels.

In recent years, cancer patients and death cases have increased every year, and the high incidence of breast cancer is threatening human life [18]. Dox as an anticancer drug, the direct injection of DOX will be released prematurely outside the cell, affecting normal cell growth and causing strong side effects. It has been reported that pH and redox-sensitive mesoporous silicone hybrid nanoparticles encapsulated Dox for intracellular delivery [19]. HNTs is a kind of tubular inorganic material with good biocompatibility, negatively charged outer surface with silanols, can be loaded with curcumin-Au and coated CS with pH and near-infrared responsive for cancer drug delivery [20], and can also be loaded with DOX to reduce its side effects [21]. The hydrogel composed of sodium carboxymethyl cellulose and CS can enhance its compressive mechanical properties by adding HNTs and graphene oxide [22]. In summary, in order to improve the mechanical strength of natural polysaccharide-based chitosan hydrogel and design an anti-cancer drug carrier that can reduce the strong cytotoxicity of DOX, this study modified halloysite and chitosan by sulfhydrylization and added β-Glycerophosphate sodium to adjust the pH of the system and the temperature sensitivity of the hydrogel, to form DOX-loaded thiolated halloysite cross-linked thiolated chitosan hydrogels through electrostatic interaction and cross-linking of disulfide bonds. It is expected that it can improve the mechanical properties of the hydrogels, slow and controlled release and reduce the cytotoxicity of DOX.

In order to attach reactive functional groups, reduce the release rate and toxic side effects of DOX, halloysite nanotubes (HNTs) are modified by 3-mercaptopropyl trimethoxysilane (KH590) to obtain thiolated halloysite nanotubes (HNTs-SH) and load DOX (DOX@HNTs-SH) (Scheme 1 A). Scheme 1 B shows the formation process of DOX-loaded thiolated halloysite nanotubes (DOX@HNTs-SH) cross-linked thiolated chitosan hydrogel (DOX@CSSH/HNTs-SH Gel) through the interaction mechanism of disulfide bonds and electrostatic interaction. Scheme 1C is schematic diagram of the drug-loaded gel precursor applied to local accurate and sustained delivery of the DOX after tumor resection by in-situ injection molding, thereby inhibiting tumor recurrence. The DOX@CSSH/HNTs-SH Gel are expected to be designed as a promising new drug delivery vehicle for local, accurate and continuous delivery of DOX, and to inhibit tumor recurrence and promote tissue regeneration after breast cancer resection.

Section snippets

Materials

Chitosan (CS, degree of deacetylation = 75–85%, medium molecular weight = 190–310 kg/mol, viscosity = 200–800 cps, product of Iceland) was purchased from Sigma-Aldrich. Beta-glycerophosphate acid disodium salt pentahydrate (β-GP, 98%, Acros) was purchased from Qi Yun Biotechnology Co., Ltd. (China). Halloysite nanotubes (HNTs) were purchased from Guangzhou Runwo Materials Technology Co., Ltd. (China). 3-mercaptopropyl trimethoxysilane (KH590) was purchased from Energy Chemical Saen Chemical

Characterization of the HNTs-SH

It can be seen from Table 1 that the HNTs are negatively charged at pH = 6.5 and pH = 8.5, which are - 20.03 ± 0.15 mV and - 45.27 ± 2.37 mV, respectively, indicating that the change of pH does not change their negative charge. The negatively charged surface of halloysite nanotubes can form electrostatic interactions with positively charged CSSH to form coating layers. The halloysite nanotubes are nano-sized, the average size is 236.97 ± 1.72 nm (pH = 6.5) and 240.43 ± 0.29 nm (pH = 8.5), PDI

Conclusions

To improve the mechanical properties of CSSH Gel, load and slow-release water-soluble DOX to reduce its cytotoxicity, HNTs were modified by thiol groups, then loaded with DOX, and added to the gel precursor to form DOX@CSSH/HNTs-SH Gel. The results showed that the HNTs-SH could be uniformly dispersed in the gel matrix to improve the compressive strength of the gel, and the gel was pH sensitive to release DOX quickly in the tumor acidic microenvironment to be taken up by MCF-7 cells, and

Author statement

Riwang Li: Investigation, Methodology, Data curation, Writing - original draft., Yuhui Zhang: Investigation, Methodology, Writing - review & editing. Zhen Lin: Conceptualization, Resources., Qiqi Lei: Investigation, Methodology, Data curation. Yi Liu: Writing - review & editing. Xinyang Li: Investigation, Methodology, Data curation, Mingxian Liu: Data curation, Conceptualization, Supervision. Gang Wu: Data curation, Conceptualization, Supervision. Simin Luo: Data curation, Conceptualization,

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

This work was supported by the National Key Research and Development Project (2019YFD0901905), the National Natural Science Foundation of China (81472089), Guangdong Provincial Medical Scientific Research Foundation (B2019038), the Fundamental Research Funds for the Central Universities (21620452,21619408), Natural Science Foundation of Guangdong Province (2019A1515011553).

References (35)

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