Nitrogen and boron dual-doped graphene quantum dots for near-infrared second window imaging and photothermal therapy

Highlights

• A bifunctional theranostic nanoparticle (BFNP) is developed.

• BFNP is a contrast agent for both magnetic resonance imaging and fluorescence imaging.

• BFNP converts NIR light into heat enabling photothermal therapy.

• BFNP is benign to organs and healthy tissue.

• Imaging and therapeutic efficacy of BFNP is demonstrated in a mouse model.

Abstract

Fluorescence imaging of biological systems in the second near-infrared window (NIR-II) has recently drawn much attention because of its negligible background noise of autofluorescence and low tissue scattering. Here we present a new NIR-II fluorescent agent, graphene quantum dots dual-doped with both nitrogen and boron (N-B-GQDs). N-B-GQDs have an ultra-small size (∼5 nm), are highly stable in serum, and demonstrate a peak fluorescent emission at 1000 nm and high photostability. In addition to the NIR-II imaging capability, N-B-GQDs efficiently absorb and convert NIR light into heat when irradiated by an external NIR source, demonstrating a photothermal therapeutic effect that kills cancer cells in vitro and completely suppresses tumor growth in a glioma xenograft mouse model. N-B-GQDs demonstrate a safe profile, prolonged blood half-life, and rapid excretion in mice, which are the characteristics favorable for in vivo biomedical applications.

Introduction

Fluorescent imaging in the conventional near-infrared (NIR) window (NIR-I, 750–900 nm) is a widely used imaging modality for disease detection and analysis of functional biomolecules in vivo [1], [2], [3]. Its popularity comes largely from the fact that it utilizes a nonionizing portion of the electromagnetic spectrum and thus provides a safety profile that is unattainable by other popular imaging modalities such as X-ray and conventional computed tomography (CT) that utilize ionizing radiations. Also significantly, the wavelength window of NIR-I imaging falls in the regions of low energy absorption by water, oxyhemoglobin, and deoxyhemoglobin which are the major contributors to the poor penetration depth of optical imaging using visible light [4]. Despite the significant advantages, NIR-I imaging suffers from the drawback of background autofluorescence by endogenous chromophores in the body and yet further improvement in tissue penetration depth beyond those provided by NIR-I imaging is also highly desirable [5], [6], [7], [8]. Since the light scattering experienced by photons when they interact with matter is inversely proportional to their wavelength, imaging using lights with wavelengths longer than those of NIR-I may reduce the light scattering and thereby increase the tissue penetration depth and detection sensitivity [9], [10]. These considerations prompted the research of seeking imaging techniques that operate in a region of the electromagnetic wavelength spectrum known as the second near-infrared window or NIR-II (1000–1700 nm) [11], [12], [13], [14], [15].

Several materials have been investigated to serve as NIR-II contrast agents for imaging biological systems. These include semiconductor quantum dots (InSb, Ag₂S, and Ag₂Se) and rare-earth metal nanoparticles (NPs) [15], [16], [17]. However, the potential heavy metal toxicity associated with these NPs or quantum dots poses health risks when used in vivo [18]. Biocompatible metal-free single-walled carbon nanotubes (SWCNTs) are an alternative to heavy metal-based materials to serve as NIR-II contrast agent [11], [19]. But SWCNTs are hydrophobic and lack functional groups, and require sophisticated surface modifications to render their surfaces hydrophilic to prevent them from aggregation when used in vivo [20], [21]. Further, all the above-mentioned NIR II fluorophores, upon injected intravascularly, eventually accumulate and largely retain in the organs of the reticuloendothelial systems such as the liver and spleen for a long period of time and only small amount of them can reach cells or tissues of interest, an undesirable behavior that hinders their clinical translation [21].

More recently, small organic molecules such as IR-PEG and CH1055 were developed as NIR-II agents; they were surface-modified with polyethylene glycol (PEG) to improve their hydrophilicity, and with various tumor targeting ligands to enhance their ability to target tumors. These small organic molecules have demonstrated the ability to enable the real-time imaging of organs, tissues, blood vessels, and tumors in the NIR-II window and are rapidly cleared from the body through renal excretion [22], [23], [24]. Despite these encouraging results, however, organic fluorophores are generally poorly soluble in aqueous media due to their aromatic structures, usually lack chemical stability, and are incapable of providing long-term visualization and multiplexing. They possess only limited functional groups for conjugation of biomolecules for various applications. In addition, none of these organic fluorophores or nanoparticles with NIR-II capability has demonstrated as a theranostic agent that can serve both diagnostic and therapeutic purposes.

Here we introduce a new NIR-II contrast agent with a number of desirable characteristics that have not been unattainable by existing NIR-II contrast agents, including high chemical stability and photostability, hydrophilicity compatible with aqueous solutions, and procession of multiple functional groups that requires no additional surface modification for chemical conjugation. The nitrogen and boron dual-doped graphene quantum dots (N-B-GQDs) were synthesized via a one-pot process. We showed that N-B-GQDs demonstrate an NIR-II photoluminescence emission (950–1100 nm) and can serve as NIR-II imaging agent for the visualization of internal organs and blood vessels as demonstrated in wide-type mice. Furthermore, N-B-GQDs efficiently absorb and convert NIR light into heat effectively, which kills cancer cells in vitro and reduces tumor growth as demonstrated in a xenograft tumor mouse model.