Glucose oxidase and L-arginine functionalized black phosphorus nanosheets for multimodal targeted therapy of glioblastoma

https://doi.org/10.1016/j.cej.2021.132898Get rights and content

Highlights

  • Development of highly biocompatible and stable black phosphorus nanosheets.

  • Combination of photothermal, starvation and nitric oxide for glioblastoma therapy.

  • Superior capability in blood–brain barrier penetration and glioblastoma targeting.

  • Excellent in vivo antitumor performance on orthotopic glioblastoma model in mice.

Abstract

Glioblastoma (GBM) is the most common type of primary malignant brain tumor with few innovative therapies. Developing an efficient and “green” synergistic anticancer strategy for GBM treatment remains a pressing need. Herein, a novel strategy that combines photothermal therapy (PTT), tumor starvation and nitric oxide (NO) therapy based on functionalized black phosphorus nanosheets (BP) is developed. NO-functionalized BP (BPA) is prepared by esterification reaction between the carboxyl group of L-arginine (Arg) and the hydroxyl group (P-OH) formed from the preliminary oxidation on the surface of BP. Then glucose oxidase (GOx) is further introduced to Arg by amidation to form a multimodal nanodrug (BPAG). The mild chemical modification empowers BP with superior stability under physiological condition and induce release of H2O2 and NO by the cascaded oxidation of glucose and Arg. This process can be significantly accelerated by PTT. To facilitate BPAG with tumor-targeting ability, the macrophage membrane is used to coat the nanoparticles under ultrasonic condition. The membrane-coated BPAG (M@BPAG) improves penetration through blood–brain barrier for GBM targeting. Taken together, M@BPAG combines GBM targeting, H2O2-NO release, and PTT effect, leading to reprogramming the tumor immune microenvironment and a significant synergistic antitumor performance without systemic toxicity.

Introduction

In the past decades, traditional strategies including surgical resection, chemotherapy and radiation therapy have been applied for the treatment of glioblastoma (GBM), which is one of the most common primary brain cancers with devastating prognosis [1]. However, innovative treatments are urgently needed because recurrence and metastasis of GBM frequently occur owing to the unsatisfactory effect of traditional mono-modal therapies [2], [3], [4], [5]. Numerous evidences have suggested that combination of different approaches for cancer treatment improves overall survival compared to mono-modal therapy in GBM [6], [7], [8], [9], [10]. However, the combination of routine treatments such as chemotherapy, radiotherapy or immune check point inhibitors often cause drug resistance and systematic toxicity [2], [3], [7]. Therefore, exploring a combination of innovative treatment paradigm is highly desired for GBM management.

Recently, the photothermal, tumor cell starvation and gas therapies have attracted enormous attentions in preclinical research [11]. These novel treatment modalities have been suggested to be promising candidates for treatment of multiple tumors due to their minimal invasiveness toward surrounding normal tissues and less side effect [12], [13], [14], [15], [16]. Photothermal therapy (PTT), which depends on the efficient conversion of near-infrared (NIR) light energy into heat by photo-absorbing agents and irradiation at target sites, could destruct tumor cells without damaging normal tissues [17], [18], [19], [20], [21]. However, employing PTT alone is hard to eliminate cancer cells completely because the heat distribution within tumor tissue is heterogeneous and hyperthermia might kill immune cells that have antitumor activity [12], [22]. Therefore, it is necessary to seek highly efficient antitumor strategies to synergize with PTT. Cancer cell starvation has been demonstrated as a novel “green” therapy that could directly cut off the nutrients supplied to tumor cells resulting in cell death [14]. The starving-like agent glucose oxidase (GOx) exerts more effective therapeutic efficacy than conventional starvation treatment through assisting the conversion of glucose into gluconic acid and H2O2, which subsequently oxidize L-arginine (Arg) into nitric oxide (NO) for gas therapy [23]. NO has been reported to have antitumor activity with various mechanisms including directly inducing cell apoptosis at high concentrations, reversing multidrug resistance by inhibiting P-Glycoprotein expression [24], reprogramming tumor immune microenvironments and more [25], [26]. Therefore, developing an integrated platform to realize the combination of PTT, GOx and NO therapy could exert a synergistic antitumor effect in GBM.

In this study, we utilized black phosphorus nanosheets (BP) as a photothermal agent to explore the combined antitumor activity of PTT, GOx and NO. BP has been demonstrated to have a high surface to volume ratio for facilitating drug-loading [27], [28]. BP exhibits excellent biocompatibility and biodegradability, low biotoxicity and high photothermal conversion efficiency under NIR irradiation [29], [30], [31], [32], [33]. However, BP lacks stability when exposes to air and moisture, that limits its clinical application in PTT and drug delivery [34], [35], [36], [37]. To address this issue, we introduced a functional molecule Arg to the surface of BP (BPA) to serve as a NO donor via esterification reaction between the carboxyl group of Arg and the hydroxyl group (P-OH) derived from the preliminary oxidation of BP, which could also prevent BP from further oxidative degradation. To stimulate the release of NO, GOx was linked to Arg by amidation to construct the multimodal nanodrug (BPAG). BPAG was able to release H2O2 via oxidizing glucose using the catalysis of GOx followed by NO generation from the oxidization of Arg. Moreover, the release of H2O2 and NO could be significantly accelerated by 808 nm NIR irradiation. Furthermore, we coated BPAG with the macrophage membrane to facilitate the penetration through blood–brain barrier (BBB) and improve GBM-targeting ability [38], [39]. The membrane coated BPAG (M@BPAG) was subsequently administered by intravenous injection and actively targeted to GBM tissues exhibiting strong antitumor activity via the combinational effect of PTT, GOx and NO without systematic toxicity. In short, an integrated nanodrug with potential clinical application for GBM therapy is engineered and characterized by BBB permeability, GBM targeting, rapidly cascaded H2O2-NO release, and PTT effect, which exhibits excellent antitumor performance in vivo.

Section snippets

Materials and reagents

Black phosphorus nanosheets (BP) dispersion was purchased from Kunming Black Phosphorus Technology Co., Ltd. (Kunming, China). L-arginine (Arg) was purchased from Yuanye Biological Technology Co., Ltd. (Shanghai, China). Glucose oxidase (GOx) was purchased from Jiuding Chemical Technology Co., Ltd. (Chengdu, China). 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydro (EDC), N-Hydroxy succinimide (NHS) and 4-Dimethylaminopyridine (DMAP) were purchased from Energy Chemical Co., Ltd. (Shanghai,

Preparation and characterization of M@BPAG

The Arg functionalized black phosphorus nanosheets (BPA) was prepared by the esterification reaction between the carboxyl (–COOH) group of Arg and the hydroxyl group (P-OH) formed from the preliminary oxidation on the surface of BP [43], [44]. The carboxyl group on GOx was utilized to conjugate BPA under the catalysis of EDC and NHS to form multifunctional BPAG (Fig. 1A). Both the XPS and Raman spectra were performed to demonstrate the reaction on the surface of BP. XPS and Raman results

Conclusion

In summary, we have successfully developed an integrated nanodrug (M@BPAG) that combined PTT, GOx with NO for GBM targeted therapy based on GOx and Arg functionalized BP. The mild chemical modification empowered BP with superior stability and cascaded release of H2O2 and NO in the presence of glucose, which could be accelerated by NIR irradiation. The M@BPAG with NIR irradiation converted cold tumors to hot ones by elevating the infiltration of CD8+ T cells and M1 macrophages while decreasing

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 Fundamental Research Funds for the Central Universities (2632020ZD13), the National Natural Science Foundation of China (NSFC 31800154, 51903088), the China Postdoctoral Science Foundation (2021M693953), the Jiangsu Province Postdoctoral Science Foundation (2021K385C), and the Natural Science Foundation of Jiangsu Province (BK20211221).

References (50)

  • S. Shen et al.

    Engineered nanoparticles disguised as macrophages for trapping lipopolysaccharide and preventing endotoxemia

    Biomaterials

    (2019)
  • C. Zhang et al.

    Biomimetic carbon monoxide nanogenerator ameliorates streptozotocin induced type 1 diabetes in mice

    Biomaterials

    (2020)
  • A.C. Tan et al.

    Management of glioblastoma: State of the art and future directions

    CA. Cancer J. Clin.

    (2020)
  • M. Lim et al.

    Current state of immunotherapy for glioblastoma

    Nat. Rev. Clin. Oncol.

    (2018)
  • B. Campos et al.

    A comprehensive profile of recurrent glioblastoma

    Oncogene

    (2016)
  • M.E. Davis

    Glioblastoma: overview of disease and treatment

    Clin. J. Oncol. Nurs.

    (2016)
  • A. Shergalis et al.

    Current challenges and opportunities in treating glioblastoma

    Pharmacol. Rev.

    (2018)
  • D.S. Pellosi et al.

    Targeted and synergic glioblastoma treatment: multifunctional nanoparticles delivering verteporfin as adjuvant therapy for temozolomide chemotherapy

    Mol. Pharm.

    (2019)
  • F.C. Lam et al.

    Enhanced efficacy of combined temozolomide and bromodomain inhibitor therapy for gliomas using targeted nanoparticles

    Nat. Commun.

    (2018)
  • Z. Xie et al.

    Emerging combination strategies with phototherapy in cancer nanomedicine

    Chem. Soc. Rev.

    (2020)
  • X. Li et al.

    Clinical development and potential of photothermal and photodynamic therapies for cancer

    Nat. Rev. Clin. Oncol.

    (2020)
  • S. Yu et al.

    Advances in nanomedicine for cancer starvation therapy

    Theranostics

    (2019)
  • Y.H. Song et al.

    Ferrimagnetic mPEG-b-PHEP copolymer micelles loaded with iron oxide nanocubes and emodin for enhanced magnetic hyperthermia-chemotherapy

    Natl. Sci. Rev.

    (2020)
  • L. Yu et al.

    Gas-generating nanoplatforms: material chemistry, multifunctionality, and gas therapy

    Adv. Mater.

    (2018)
  • W. Li et al.

    Targeting photodynamic and photothermal therapy to the endoplasmic reticulum enhances immunogenic cancer cell death

    Nat. Commun.

    (2019)
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