Chuxin Cai†
a,
Qingming Zhang†b,
Junqiu Ye†a,
Sijia Yao†a,
Qian Lia,
Zhechen Fana,
Sulei Gea,
Yukun Wanga,
Dingyi Xua,
Jianping Zhou*a,
Hao Cheng*a and
Yang Ding*a
aState Key Laboratory of Natural Medicines, Department of Pharmaceutics, China Pharmaceutical University, Nanjing 210009, China. E-mail: dydszyzf@163.com; chenghao@cpu.edu.cn; zhoujianping@cpu.edu.cn
bDepartment of Pharmacy, Jinling Hospital, No. 305 East Zhongshan Road, Nanjing 210002, Jiangsu, People's Republic of China
First published on 17th April 2025
The tumor microenvironment (TME) emerges as a unique challenge to oncotherapy due to its intricate ecosystem containing diverse cell types, extracellular matrix, secreted factors, and neovascularization, which furnish tumor growth, progression, invasion, and metastasis. Graphene oxide (GO)-based materials have garnered increasing attention in cancer therapy owing to their vast specific surface area, flexible lamellar structure, and electronic–photonic properties. Recently, interactions of GO with the TME have been broadly investigated, including trapping biomolecules, catalysis, cancer stem cell targeting, immunoreactions, etc., which inspires combinative therapeutic strategies to overcome TME obstacles. Herein, we summarize TME features, GO modulating various dimensions of the TME, and a TME-triggerable drug delivery system and highlight innovation and merits in combinative cancer therapy based on TME modulation. This review aims to offer researchers deeper insights into the interactions between versatile GO nanomaterials and the TME, facilitating the development of rational and reliable GO-based nanomedicines for advanced oncotherapy.
Interactions of the TME with GO and its derived nanosystem have been extensively studied among various nanomaterials.7 GO's vast specific surface area, flexible lamellar structure, and unique electronic–photonic properties are directly linked to GO's intrinsic TME-modulation activities. For instance, GO's super-large specific surface area could efficiently trap biomolecules like vascular endothelial growth factor (VEGF) for anti-angiogenesis8 and its flexible lamellar structure makes it preferable for internalization by cancer stem cells (CSCs) with lower cellular stiffness to promote CSC differentiation or elimination.9 The electronic–photonic properties endow GO with a capacity for catalysis and photothermal conversion,10 which could generate hydroxyl radicals to aggravate hypoxia and cause hyperthermia for blood perfusion improvement and immunological effects.11 The unique TME could also be utilized for controllable drug delivery system design by introducing TME-responsive moieties on GO nanomaterials.12 A TME-responsive drug delivery strategy is supposed to improve drug selectivity on tumor sites for enhanced efficacy and reduced toxicity and meanwhile regulate TME features, including pH increase,13 hypoxia alleviation, competitive enzyme inhibition, and glutathione depletion for tumor suppression.14 Moreover, abundant oxygen-containing groups and unsaturated bonds of GO make it fit for various modifications.15 The functionalized GO nanomaterials could offer a preferable platform for co-loading multiple drugs in combinative therapies based on TME modulation.
In this review, we summarize the distinctive features of the TME and explore various intrinsic TME-modulation activities of GO nanomaterials. We further discuss GO-derived TME-sensitive drug delivery systems, which perform tumor-specific drug release and combine TME regulation after responsiveness. Taking advantage of GO's amphipathicity and wealthy chemical active groups, we give an overview of multidrug-delivery GO nanomaterials for TME-modulation-based combinative therapies. By summarizing the latest advancements and highlighting the versatile applications of GO nanomaterials in TME modulation and the corresponding combinative therapies, this review aims to provide researchers with deeper insights into the dynamic interactions between GO nanomaterials and the TME, facilitating the development of innovative and reliable GO-based nanosystems for advanced oncotherapy.
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Fig. 1 Schematic illustration of the main cell types, extracellular components in the TME, and hallmarks of the TME. |
Various noncellular components construct the extracellular matrix (ECM) of the TME with a three-dimensional network of proteins and carbohydrates that provides structural support and influences cellular behavior. In the ECM network, cytokines, chemokines, growth factors, and exosomes are among the critical communicators in this network. Cytokines and chemokines modulate immune cell recruitment and immune responses to create an immunosuppressive environment for tumor protection and promotion.24 Growth factors stimulate angiogenesis and cellular proliferation, thereby facilitating the expansion of tumors.25 These signaling molecules create a complex communication network that regulates cell–cell interactions, ECM remodeling, and overall tumor behavior (Fig. 1).
Furthermore, the unique biostructure of the TME contributes to multiple hallmarks that significantly differ from normal tissues.10 One of the critical characteristics of the TME is hypoxia, which results from the rapid proliferation of tumor cells outstripping the supply of oxygen due to poor vascularization.26 This hypoxic condition not only promotes tumor aggressiveness and resistance to therapy but also induces the expression of hypoxia-inducible factors (HIFs) that drive angiogenesis and metabolic reprogramming.27 The hypoxia drives the elevation of the glutathione (GSH) level, which protects tumor cells from oxidative stress and can impair the effectiveness of reactive oxygen species (ROS)-based therapies. Due to the Warburg effect of tumor metabolism, tumor cells rely on glycolysis followed by lactic acid fermentation for energy production; even in the presence of adequate oxygen, an acidic pH often characterizes the TME. This acidification leads to a hostile environment for immune cells and reduces the efficacy of chemotherapeutics.28 Furthermore, the TME exhibits overexpression of specific enzymes such as matrix metalloproteinases (MMPs) and cathepsins, which are involved in extracellular matrix remodeling and facilitating tumor invasion and metastasis. These unique characteristics of the TME provide insight into tumor biology and present significant opportunities for developing targeted and combinative therapeutic strategies.29
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Fig. 2 (A) GO nanomaterials modulate the immunosuppressive microenvironment by macrophage modulation and vaccine adjuvant function. (B) Graphene nanosheets stimulate the secretion of cytokines and chemokines in macrophages. Primary macrophages and macrophage-like RAW264.7 cells were cultured with 20 mg ml−1 graphene for 24 h.40 Reproduced from ref. 40 with permission from Elsevier, copyright (2012). (C) FACS analysis of CD4+ and CD8+ T cell populations after immunization with normal saline, 50 μg OVA, and 25 μg GO-Au@OVA. The lymphocytes were purified from immunized mouse spleens and then incubated with 10 μg mL−1 OVA for 60 h at 37 °C. The cells were then stained with anti-CD3e and anti-CD4 or anti-CD8a antibodies.41 Reproduced from ref. 41 with permission from Ivyspring International, copyright (2020). (D) Schematics showing the design and preparation of GO-APP3/28. (1) A scanning electron microscopy image showing the as-prepared GO. (2) A TEM image negatively stained with uranyl acetate showing the GO-linked protein on the flat region of the GOs. (3–5) False-coloured low-vacuum scanning electron microscopy images showing a fixed T cell (blue) interacting with a piece of GO-APP3/28 (yellow; 3), a dendritic cell (green; 4) and Bead3/28 (red; 5). Scale bars: 30 m (1), 100 nm (2), and 2 μm (3–5).39 Reproduced from ref. 39 with permission from Springer Nature, copyright (2024). |
Nano-GO was proved to adsorb model antigen OVA with super high efficiency at a OVA to GO ratio of over 5:
1 (w/w). The high aspect ratio of GO allows it to be directly inserted into the cell membrane when in contact with DCs, increasing OVA uptake by about 2.5 times. Besides the lysosomal pathway (the only pathway of OVA), GO-OVA tends to traffic via the cytoplasmic pathway, resulting in the protection of antigen from enzymatic digestion and the cross-presentation of antigen to CD8+T cells, which may be related to the non-lysosomal trafficking pathway of GO into cells.37 Compared with the OVA group, GO-OVA promotes DCs to produce more cytokines and chemokines. Moreover, it significantly increases the expression of major histocompatibility complex 1 (MHC1) molecules, which offers recognition information for CD8+ T in vivo and in vitro.38 Highly efficient and compact adsorption enables GO to change the flat structure to the folded structure, thereby allowing OVA to release slowly, rendering the programmatic activation of specific CD8+ T cells that result in effectively cracking thymoma cells in vivo (the tumor volume decreased by 80%). Based on the potential of antigen absorption, a flexible GO antigen-presenting platform (GO-APP) has been developed to anchor antibodies onto the GO surface. By decorating anti-CD3 (αCD3) and anti-CD28 (αCD28) on graphene oxide (GO-APP3/28), remarkable T cell proliferation has been achieved. Of note, interactions between GO-APP3/28 and T cells closely mimic the in vivo immunological synapses between antigen-presenting cells and T cells. This immunological synapse mimicry shows a high capacity for stimulating T cell proliferation while preserving their multifunctionality and high potency. Meanwhile, GO-APP3/28 enhances CAR gene-engineering efficiency, yielding a more than fivefold increase in CAR T cell production compared with the standard protocol (Fig. 2D).39
In recent years, nano-catalysts for tumor therapy have attracted much interest. Owing to their low cost, high catalytic efficiency, and biocompatibility, carbon-based metal-free catalysts have attracted intense interest for various applications, ranging from energy through environmental to biomedical technologies. Recent studies have validated that functionalized GO nanomaterials with intrinsic catalytic activity could generate ROS in the presence of H2O2.44,45 This ROS production can cause multi-level damage to subcellular components, contributing to cancer cell death. A novel graphene oxide nanoparticle (N-GO) was constructed that could mimic peroxidase-like catalytic activity, specifically in acidic and high hydrogen peroxide (H2O2) environments characteristic of tumors.44 Under such conditions, N-GOs can efficiently convert H2O2 into highly reactive and toxic hydroxyl radicals (HO˙), leading to the necrosis of tumor cells. This targeted effect is advantageous as it avoids damage to normal cells, which reside in a neutral pH environment with lower H2O2 concentrations, where N-GOs instead exhibit catalase-like activity scavenging reactive oxygen species (ROS). In mice models, N-GOs effectively inhibited tumor growth, as evidenced by extensive destruction of tumor cell membranes and nuclei, while showing minimal toxicity to other organs. Blood component analysis revealed no significant differences between N-GO-treated and control groups, confirming the low systemic toxicity of N-GOs. N-GOs offer promising prospects for cancer therapy and diagnostics, combining the benefits of tumor-selective treatment, reduced side effects, and improved tumor cell recognition capabilities. Their ability to selectively target and destroy tumor cells while sparing normal tissue makes them a potentially powerful tool in the fight against cancer.
In the human acute monocytic leukemia cell line (THP-1), GO and vanillin-functionalized GO (V-rGO) exhibited toxic effects on THP-1 cells in a dose-dependent manner. The toxicity manifested as decreased cell viability, proliferation, and increased lactate dehydrogenase (LDH) levels, indicative of cell membrane damage. Furthermore, there was a loss of mitochondrial membrane potential (MMP), decreased ATP content, and cell death.46 The most striking observation was the elevated levels of ROS and lipid peroxidation, causing a redox imbalance within the cells. Oxidative damage led to an increase in malondialdehyde (MDA) levels and a depletion of antioxidants like glutathione (GSH), glutathione peroxidase (GPX), superoxide dismutase (SOD), and catalase (CAT). Notably, V-rGO induced a significantly higher level of ROS in THP-1 cells than GO. The increased ROS production was concentration-dependent, with a 1.5-fold and 3-fold increase in ROS levels observed after 24 hours of exposure to 100 μg mL−1 of GO and V-rGO, respectively. The results suggested that the mechanism of toxicity of V-rGO was more potent than that of GO.44 In addition to the ROS production, lipid peroxidation was measured by quantifying MDA content. Both GO and V-rGO induced lipid peroxidation in THP-1 cells in a dose-dependent manner, further supporting the hypothesis that these graphene materials induced oxidative stress.
Moreover, the defect on GO could enhance the H2O2 catalytic activity. Defect-rich graphene quantum dots (GQDs) have been shown to exhibit high catalytic efficiency in ROS generation, specifically in the H2O2-rich tumor microenvironment.42 The catalytic mechanism of GQDs in generating ROS within the TME was thoroughly investigated, revealing that GQDs possess peroxidase-like activity capable of decomposing H2O2 into highly reactive ˙OH radicals. The generation of ˙OH radicals was confirmed through electron paramagnetic resonance (EPR) spectroscopy using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin-trap, which showed a characteristic signal pattern indicative of ˙OH formation when H2O2 and GQDs were combined. The biological effects of the ROS generated by GQDs were studied in vitro using human breast cancer MCF-7 cells. It was observed that MCF-7 cells continuously consumed ˙OH radicals produced from the GQD-catalyzed H2O2 decomposition.47 Further investigation into the subcellular localization of GQDs revealed their presence in the cytoplasm and nuclei of the cancer cells, indicating successful penetration and aggregation within the cells. These findings suggest that defect-rich GQDs can effectively damage subcellular components, including nuclear structures, due to the ROS they catalytically generate. GO-derived materials open a promising avenue for developing safe and efficient catalytic nanomedicine for cancer treatment (Fig. 3).
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Fig. 3 (A) GO nanomaterials modulate hypoxia in cancer cells and generate oxygen in normal cells. (B) Effects of GO and V-rGO on anti-oxidant markers: glutathione (GSH), glutathione peroxidase (GPx), super oxide dismutase (SOD), and catalase (CAT). THP-1 cells were treated with GO (50 μg mL−1), V-rGO (50 μg mL−1), and AgNPs (10 μg mL−1) for 24 h. (C) GO (20–100 μg mL−1) induces reactive oxygen species (ROS) generation. (D) V-rGO (20–100 μg mL−1) induces lipid peroxidation.48 Reproduced from ref. 48 with permission from MDPI, copyright (2019). (E) Catalytic performance of GQDs analysed using the Michaelis–Menten kinetics profile.49 Reproduced from ref. 49 with permission from Springer, copyright (2021). |
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Fig. 4 (A) GO nanomaterials can target soft CSCs selectively and inhibit CSCs. (B) GO nanomaterials kill CSCs and remodel the tumor microenvironment. (C) CSCs are resistant to free Dox but sensitive to N-GQD@Dox. Bulk tumor cells and CSCs were treated with DMSO (control), N-GQDs, free Dox, and N-GQD@Dox for 48 h, respectively. Cell viability was measured using the MTS assay. n = 3. N-GQDs@Dox is accumulated in the tumor tissues rather than the major organs. (D) The major organs (heart, liver, spleen, lungs, and kidneys) and the generated tumors.54 Reproduced from ref. 54 with permission from Elsevier, copyright (2021). (E) GDYO exhibits anti-lymphoma effects in vivo and the morphology of tumors after GDYO treatment. (F) Flow cytometric analysis of inflammatory cytokines affecting the frequency of Tregs and CSCs in vitro.55 Reproduced from ref. 55 with permission from Elsevier, copyright (2022). |
A recent study further demonstrates that the GO-like nanostructure of graphdiyne oxide (GDYO) nanosheets could also show CSC targeting and elimination activities.51 GDYO has been shown to effectively combat lymphoma by targeting and eliminating cancer stem cells (CSCs), which are often referred to as the “seeds” of cancer, while concurrently remodeling the TME, likened to the “soil” in which cancer grows. The mechanism of action involves GDYO significantly reducing Mif-Ackr3 signaling from the tumor to cancer-associated fibroblasts (CAFs), decreasing the levels of inflammatory cytokines within the CAF cells, and lowering the frequency of Treg cells and CSCs.56 As a result, the growth of lymphoma is inhibited. The long-term safety profile of GDYO has also been assessed, demonstrating that it does not adversely affect mouse survival even when administered for extended periods, up to 20 months. This underscores the biosafety and biocompatibility of GDYO, suggesting its potential application in clinical medicine to treat lymphoma.
Carbon quantum dots (CQDs) can serve as a physiological niche for stem cells, influencing their formation and differentiation.57 While much work has been done on stem cell imaging with CQDs, less attention has been paid to their interaction with stem cells and their potential to selectively guide differentiation towards a specific lineage.54 CQDs interact with stem cells in various ways, influencing their differentiation potential. They can modulate the expression of critical genes and proteins involved in the differentiation process, acting as signaling molecules that steer stem cells toward specific lineages. For instance, CQDs have been shown to upregulate osteogenic markers such as alkaline phosphatase (ALP), RUNX2, OCN, and BSP in bone marrow mesenchymal stem cells (MSCs), indicating the promotion of osteogenic differentiation. Specific microRNAs like miR-2861 can further enhance this effect, which promotes osteoblast differentiation (Fig. 5).53
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Fig. 5 (A) GO nanomaterials promote the differentiation of CSCs.50 Reproduced from ref. 50 with permission from American Chemical Society, copyright (2022). (B) Immunostaining of MSCs seeded for 15 days in osteogenic differentiation medium on different substrates. MSCs growing on Si/SiO2, PDMS, and PET without graphene show OCN negative staining. Once these substrates are coated with graphene, cells are positive for OCN, indicating osteogenic differentiation. Scale bars are 100 μm.59 Reproduced from ref. 59 with permission from BioMed Central, copyright (2014). |
The novel mechanism by which the GO nanomaterial promotes CSC differentiation is also investigated. This research article discusses bioactive graphene quantum dots (GQDs) to target and eradicate drug-resistant cancer stem cells (CSCs), critical in cancer metastasis and recurrence.54 Conventional chemotherapy fails to eliminate these CSCs effectively due to inadequate interfacial inhibition effects. However, GQDs can self-insert into the DNA major groove (MAG) sites within cancer cells, enhancing interfacial inhibition significantly.58 Transcription factors, which regulate gene expression at MAG sites, are affected by this targeted action of GQDs, resulting in the downregulation of cancer stem genes like ALDH1, Notch1, and Bmi1. The nano-scale interface inhibition mechanism of GQDs reverses multidrug resistance (MDR) in cancer cells by inhibiting the MDR1 gene expression. When GQDs are applied at non-toxic concentrations (1/4 of the half-maximal inhibitory concentration, IC50), they act as MDR reversers. The efficacy of this approach allows for substantial blockage of CSC-mediated migration, invasion, and metastasis of cancer cells. GQDs can also sensitize clinical cytotoxic agents, improving their effectiveness in combination chemotherapy.57
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Fig. 6 Schematic illustration of GO-derived PTT for activating anti-tumor responses via immunogenic cell death (ICD) and M2 tumor-associated macrophage elimination. |
GO-based PTT directly kills tumor cells and induces immunogenic cell death (ICD). During ICD, damage-associated molecular patterns (DAMPs) such as calreticulin (CRT) translocation, high-mobility group box 1 (HMGB1) release, and heat shock protein (HSP) efflux are observed. These DAMPs collectively activate antigen-presenting cells (APCs), triggering subsequent immune processes.68 Hydrophilic Fe3O4 nanoparticles were combined with rGO through electrostatic adsorption, significantly promoting DC maturation up to 30.2% under 808 nm laser irradiation.69 The GO-arginine-soy lecithin nanogenerator (GO-Arg-SL) for PTT and gas therapy (GT) effectively releases l-Arg in the cytoplasm of tumor cells, which was then catalyzed by inducible nitric oxide synthase (iNOS) to produce a large amount of nitric oxide (NO). The released NO promoted the maturation of antigen-presenting cells (DCs) and caused a significant increase in CD8+CD4−CD3+ in the spleen. Mature DCs can activate CTLs, stimulating the innate immune system and enhancing anti-tumor immunity. Tumors were significantly eliminated due to the enhanced anti-tumor effect of PTT and GT.70 Nanoparticles coated with granulocyte-macrophage colony-stimulating factor (GM-CSF@CS/GO) were synthesized, which could cause the photothermal destruction of tumors, release a large number of tumor-associated antigens and significantly promote the recruitment of DC cells and the production of anti-tumor cytokines.71 Transdermal delivery of paclitaxel (DTX) and 1-methyl-D-tryptophan (1MT) (D-1/GH) based on heparin-modified graphene oxide (GH) enhances the therapeutic effect of chemotherapy and PTT while inducing immunogenic cell death (ICD), thereby recruiting immune cells, reversing the immunosuppressive tumor microenvironment, and enhancing immune response (Fig. 7).72
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Fig. 7 (A) Schematic showing the synthesis of GGIC and its immunostimulatory effect. (B) PBS, CpG ODNs, GGI, and GGIC were injected intratumorally in a single dose, followed by 5 min NIR irradiation or without NIR irradiation. (C) In vivo photothermal tumor heating. IR thermal images of tumor-bearing mice exposed to the NIR laser after injection with PBS, GGI, or GGIC. (D) The volumetric changes in tumor size and changes with time in body weight were achieved from mice injected with PBS, CpG ODNs, GGI, and GGIC. Data are presented as mean ± SD of five mice per group. (E–J) Histopathological studies of tissue organs of (E) PBS + NIR, (F) GGI, (G) CpG ODNs, (H) GGIC, (I) GGI + NIR, and (J) GGIC + NIR in the CT26 tumor-bearing mouse model.73 Reproduced from ref. 73 with permission from Elsevier, copyright (2014). |
The anti-tumor immunity triggered by PTT is severely hindered by the dynamic amplification of programmed cell death protein ligand 1 (PD-L1) and indoleamine 2,3-dioxygenase 1 (IDO-1) in tumors. To fully unleash the efficiency of PTT, blocking the PD-L1 and IDO-1 pathways is essential to enhance effective anti-tumor immunity.74,75 Folic acid-coupled rGO loaded with IDO inhibitors (PEG-rGO-FA-IDOi) effectively blocks tryptophan catabolism and enhances T cell proliferation while simultaneously blocking PD-L1, and combining PTT, IDO inhibition, and anti-PD-L1 blockade counters immune suppression effectively. This approach enhances the presence of tumor-infiltrating lymphocytes, including CD45+ leukocytes, CD4+ and CD8+ T cells, and NK cells. It also inhibits the immunosuppressive activity of regulatory T cells (Tregs) and boosts the production of interferon-gamma (INF-γ) (Fig. 8).76 Based on rGO, the chemotherapeutic drug mitoxantrone (MTX) and the transforming growth factor β (TGF-β) inhibitor SB-431542 (SB) were loaded onto rGO-PEG through π–π stacking interactions. rGO-based PTT provides an immunogenic antigen source, forming an in situ vaccine with rGO acting as an adjuvant. The use of SB changed the tumor microenvironment, enhancing the therapeutic effect of MTX-induced chemotherapy and rGO-based PTT. The infiltration of tumor-specific cytotoxic CD8+ lymphocytes increases while the infiltration of distal Tregs decreases, providing a synergistic chemo–immuno–photothermal effect.77 GO-loaded nanoparticles with SNX-2112 and folic acid (GFS) guide LTPTT by initially inhibiting the AKT pathway, subsequently inducing the over-activation of autophagy, thereby reducing the expression of PD-L1. The results indicate that GFS-guided LTPTT can diminish tumor cell immune escape and immunosuppression. It enables T cells to regain their natural immune function, leading to the destruction of tumor cells. This process is facilitated by the combined effect of PTT and drug treatment, which activates cell autophagy and enhances immune function.78 Moreover, GO loaded with immune-related substances can synergize with PTT to activate the immune response at the tumor site. GO modified with polyethylene glycol (PEG) and polyethyleneimine (PEI) could compress CpG oligonucleotide sequences effectively to generate a GGIC nanosystem. The cellular uptake of GGIC is significantly enhanced after GO-mediated photothermal heating, effectively enhancing intracellular delivery of CpG. Subsequently, Toll-like receptor 9 (TLR9) recognizes CpG, releasing various pro-inflammatory cytokines such as tumor necrosis factor TNF-α and interleukin IL-6 for immune stimulation enhancement.79 Collectively, the interaction between heat shock and thermal immunity is significant at the cellular biology level and provides new strategies and mechanisms for the immunotherapy of tumors. In this way, the increase in heat shock proteins helps cells cope with thermal stress and effectively activates the immune system, enhancing the immune response to tumors and providing an essential target for tumor therapy.
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Fig. 8 (A) A multifunctional IDO inhibitor (IDOi)-loaded reduced graphene oxide (rGO)-based nanosheets (IDOi/rGO nanosheets) with the properties of directly killing tumor cells under laser irradiation and triggering anti-tumor immune responses in situ. (B) PEG-rGO-FA-IDOi can induce DC activation in vivo after irradiation with NIR; mature DCs induced increased expressions of CD86 and CD80. (C) Individual tumor growth for a primary tumor (1#) and a distant tumor (2#). (D) Treatment with the combined PEG-rGO-FA-IDOi-mediated PTT plus PD-L1 antibody led to a remarkably increased proportion of CD45 leukocytes, CD4 T cells, and CD8 T cells in the distant tumors and approximately 1.8- to 4-fold enhancement compared to treatment with the single PEG-rGO-FA-IDOi-mediated PTT.76 Reproduced from ref. 76 with permission from American Chemical Society, copyright (2019). |
The results indicated that the uptake of FITC-PEG-GO did not induce polarization of macrophages towards the M1 pro-inflammatory phenotype. Instead, it promoted a controlled balance between M1 and M2 phenotypes and slightly shifted towards the M2 reparative phenotype involved in tissue repair, ensuring an appropriate immune response to these nanosheets.86 However, macrophages treated with IL-4 would show M2 polarization, while post GO-mediated PTT, the RAW264.7 cell line exhibited more polarization towards M1 and less towards M2. These findings suggest that GO itself does not possess the capability to influence TAMs in the tumor microenvironment, and PTT is a crucial factor for mediating TAM's immune function. Through thermal stress, GO-mediated PTT affects the function of M2 macrophages, potentially inducing their transformation to the M1 type, thereby inhibiting tumor progression (Fig. 9).87 The effects of GO and rGO on primary bone marrow-derived macrophages (BMDM) and the immortalized macrophage cell line (J774A.1) were also evaluated in previous studies. They found that rGO, compared to GO, induced higher levels of TNF-α and IL-6 production, indicating a more robust pro-inflammatory response in macrophages. These studies demonstrate the potential application of GO and its derivatives in promoting macrophage polarization toward the M1 phenotype.88 Additionally, leveraging the activities of M2 macrophages can synergize with PTT. A multifunctional, biologically active scaffold comprising hydrated CePO4 nanorods and chitosan (CS) forms a CePO4/CS/GO structure. During PTT, the released Ce3+ ions promote the polarization of macrophages towards the M2 phenotype. Utilizing M2 macrophages to secrete vascular endothelial growth factor (VEGF) and arginase-1 (Arg-1) fosters angiogenesis and induces bone regeneration. This approach optimizes adjuvant therapy following tumor surgery and improves prognosis.89
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Fig. 9 (A) GP-mediated PTT affects the function of M2 macrophages through heat stress and may induce their conversion to M1, thereby inhibiting tumor progression. It involves the polarization of macrophages by heating and regulates their anti-tumor ability. (B) The IL-4 group showed significantly increased levels of the surface protein CD206, and the NIR + IL-4 group showed considerably decreased expressions of CD206. (C) The expression of M2-related genes in the IL-4 group was significantly increased, whereas the expression of these M2-related genes was sharply inhibited after PTT. (D) The NIR group could not repair the scar after 24 h of culture, and its ability to repair scratches was the worst. (E) The macrophage supernatants treated with NIR could inhibit the invasion and migration of tumor cells. (F) PTT showed an excellent anti-tumor effect in vivo via macrophage polarization.87 Reproduced from ref. 87 with permission from Elsevier, copyright (2020). |
Researchers have modified and altered GO to enhance its therapeutic efficacy against tumors in response to this unique environment. These modifications aim to leverage the distinctive redox characteristics of the tumor microenvironment, creating more effective cancer treatment strategies. In the TME, the levels of reduced GSH, a crucial antioxidant, are typically elevated. GSH neutralizes ROS and other free radicals, protecting cancer cells from oxidative stress damage.98,99 Therefore, the responsive design targeting GSH is considered a feasible strategy. Disulfide bonds are essential in stabilizing protein structures in normal human tissues. They break in the presence of a high level of GSH. Disulfide bonds are, therefore, a classical means to target high GSH levels in the TME.100 A novel approach utilizing a hyaluronic acid (HA)-modified, oxidation–reduction-sensitive graphene oxide nanosheet (HSG) was developed for targeted and rapid delivery to tumor cells. HA was conjugated to GO via disulfide bonds, conferring active targeting capabilities to the graphene oxide-based system. Within the intracellular environment, the disulfide bonds of HA were cleaved by the overexpressed GSH, detaining conjugated HA fragments from the GO surface and accelerating the release of DOX (Fig. 11).101 The arginine–glycine–aspartic acid (RGD) peptide and amine-functionalized polyethylene glycol (6ARM-PEG-NH2, PEG-NH2) were employed to functionalize graphene oxide. A disulfide bond was introduced at one end of PEG, and DOX was covalently attached, constructing a pH and GSH dual-responsive nano-drug delivery system (RGD–GO–PEG–S–S–DOX). DOX exhibited rapid release in a PBS solution with pH 5.50 and a GSH concentration of 10 mM.102 Moreover, magnetic GO nanosheets were constructed by coupling cystamine-functionalized GO nanosheets with Fe3O4 nanoparticles.103 The magnetic GO nanosheets were loaded with DOX at a high drug/carrier ratio of 1 to create a redox-responsive delivery system for controllable chemotherapy. In drug release evaluation, the maximum drug release after 10 days was only 17.9% without GSH addition, while the maximum drug release reached 51.7% in the presence of GSH. The magnetic GO delivery system achieves highly selective drug release at tumor sites and high drug-loading efficiency, offering a novel strategy for precision chemotherapy.
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Fig. 11 Scheme illustration of DOX released from HSG-DOX, which contained disulfide. (A) Glutathione triggers HA detachment and rapid release of DOX in the cytoplasm. (B) Based on the redox sensitivity of HSG-DOX, it can significantly improve the killing ability of DOX against MDA-MB-231 and HELF cells. The system has significant anti-tumor activity in vivo. (C) UV–vis spectra of the GO, GO-COOH, HSG, and HCG. (D) Measurement of the temperature of water, GO, GO-COOH, HSG, and HCG under NIR irradiation (808 nm, 5 W cm−2). (E) CLSM images of the PCI effect of HSG and HCG based on their photothermal ability. (F) CLSM images of MDA-MB-231 cells incubated with free DOX, HSG-DOX, HCG-DOX, and HA + HSG-DOX over time.101 Reproduced from ref. 101 with permission from Wiley, copyright (2017 ). |
The basis of carrier response mechanisms based on redox reactions is ROS levels in tumor tissues. H2O2 is the most typical ROS. H2O2 has a variety of catalytic activities and has been widely explored for the activation conditions of drug release.104 The ability of nanoscale graphene oxide particles (N-GO) to mimic peroxidase-like catalytic activity was investigated in the tumor microenvironment characterized by weak acidity and high levels of ROS. This dual-responsive behavior significantly and effectively inhibited tumor growth. In the neutral microenvironment of normal cells, N-GO exhibited peroxidase-like activity similar to catalase, converting H2O2 into oxygen and water, thereby protecting normal cells from harm. These biofunctions hold crucial implications for the diagnosis and prognosis of cancer.105 Similarly, a unique H2O2-triggered nanoenzyme-catalyzed photoacoustic imaging (PAI) contrast agent was developed utilizing a nanoscale graphene quantum dot fragment (GQDzyme) as the core, coupled with 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). Using nasopharyngeal carcinoma as a model, in the presence of elevated H2O2 expression at the tumor site, GQDzyme converted ABTS into its oxidized form. The oxidized ABTS exhibited strong NIR absorbance, making it an ideal PAI contrast agent for precise imaging at the tumor site and assisting in diagnostics.106
GO-based nanocarriers exhibit a unique pH-responsive drug release mechanism in the TME, primarily attributed to the π–π stacking interactions between the drug and the GO surface. Research indicates that these interactions remain relatively stable under neutral or mildly alkaline conditions.112 However, the release of drugs like DOX, which are conjugated to GO through π–π stacking, is controlled by the degree of carboxyl group protonation on the nanocarrier, which is influenced by the pH of different environments. In acidic media, most carboxyl groups on GO are protonated, altering the charge state and weakening the electrostatic interaction between the nanocarrier and DOX, leading to drug release.113 Additionally, under acidic conditions, the structure of GO may partially disintegrate or expand, further facilitating drug release from its surface. This characteristic renders GO-based nanocarriers particularly suitable for tumor microenvironment-specific drug delivery, enhancing therapeutic efficacy while reducing toxicity to normal tissues (Fig. 12).
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Fig. 12 (A) GCE/siRNA was prepared from GO-COOH, functionalization with chitosan (GO-CS), immobilization with anti-EpCAM (GCE), and then mixed with survivin-siRNA (GCE/siRNA) in aqueous solution. (B) After targeting tumor tissues, GCE could deliver survivin-siRNA into cells successfully, and survivin-siRNA could be released from GCE/siRNA, exhibiting the anti-tumor effect. (C) Confocal images were obtained by treating the cells with PBS, naked siRNA, GCE/siRNA, and Lipo/siRNA; siRNA was labeled with the FAM fluorescent molecule (green) to show the amount of siRNA in cells and Hoechst 33![]() ![]() |
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Fig. 13 Illustration of hypothesized mechanisms involved in sequential activation of anticancer therapeutic effects triggered by selective sensitization of imaging in a tumor microenvironment. Structures of CP (A) and IP (B). Cy5.5 in IP and CP is not fluorescent upon anchoring to GO. (C) CPGO/BF does not contain MMP-cleavable peptides and remains quenched in the tumor microenvironment with the therapeutic peptide BF hidden in the PEGylated probe. IPGO/BF contains the MMP-cleavable sequence therapeutic peptide BF anchored to GO nanosheets; the PEG cloud shields the functional activity of BF. In the tumor microenvironment, MMP-responsive de-shielding of the PEGylated imaging probe activates imaging and causes cascading exposure of BF on GO. (D) In the bloodstream, the fluorescence of Cy5.5 derivatives on GO nanosheets is quenched. The presence of PEG blocks the apoptotic activity of the therapeutic peptide. GO nanosheets accumulate in tumor tissues. (E) In the tumor microenvironment, IP on GO is cleaved by MMPs, resulting in selective fluorescence activation. (F) Release of PEGylated Cy5.5 subsequently triggers the exposure of the apoptotic anticancer peptide BF into tumor cells. (G) The sequential exposure of BF on GO results in anti-tumor activity. (H) SCC7 tumor-bearing mice were intravenously injected with free BF, GO/BF, CPGO/BF, or IPGO/BF three times at 2 d intervals (injection times indicated by arrows). Tumor volume was measured periodically for 21 d after tumor inoculation. (I) Representative tumors from each group at day 21. (J) Weights of tumors for each group at day 21 (*P < 0.05; **P < 0.01). (K) For determining the immunohistochemistry of tumor tissues, SCC7 tumor-bearing mice were intravenously injected with free BF, GO/BF, CPGO/BF, or IPGO/BF three times at 2 d intervals. On day 21, after tumor inoculation, tumor tissues were sectioned for anti-PCNA antibody immunostaining (upper panels) and the TUNEL assay (lower panels). Scale bar = 100 μm. (L and M) Summary data comparing the number of proliferating (L) and apoptotic (M) cells (***P < 0.005). The results are the mean ± SD of five independent experiments.120 Reproduced from ref. 120 with permission from Elsevier, copyright (2019). |
Tumor vasculature-targeted nanomedicines represent an innovative and precise strategy in cancer therapy, aiming to improve efficacy while minimizing systemic toxicity. Tumors cannot survive without a constant supply of nutrients and oxygen, which they obtain through their network of blood vessels. Targeted therapies that work by inhibiting the development of these blood vessels could effectively starve the tumor and prevent its growth.128 Tumor vasculature instead of tumor cell targeting is more desirable for graphene-based nanomaterials since the targets are immediately accessible upon intravenous injection, and extravasation is not required to achieve tumor targeting.
Previous studies explored reduced graphene oxide (RGO) for tumor vasculature targeting in a breast cancer model. RGO was conjugated with the anti-CD105 antibody (anti-CD105), where CD105 is overexpressed on proliferating tumor endothelial cells. The RGO conjugate, 64Cu-RGO-anti-105, exhibited excellent stability and specificity for tumor endothelial cells in vivo. Serial PET imaging revealed rapid uptake by tumor endothelial cells, peaking at 3 h post-injection, with sustained levels over time. The results demonstrated the efficacy of RGO for tumor vasculature targeting and imaging. Apart from imaging applications, nano-GO conjugated with monoclonal antibodies against follicle-stimulating hormone receptor (FSHR) has been used for tumor vasculature targeting and inhibition in metastatic breast cancer. By focusing on angiogenic markers on the tumor vasculature, researchers can achieve more effective delivery of nanomaterials directly to the tumor site, bypassing the need for extravasation. In this work, GO nanoparticles were functionalized with a monoclonal antibody (mAb) against FSHR, a particular marker found abundantly in both primary and metastatic tumors. The resulting GO nano-conjugates, measuring approximately 120 nm in diameter, were radiolabeled with 64Cu to enable their visualization using positron emission tomography (PET) imaging. The imaging function allowed real-time monitoring of the conjugates’ distribution and tumor accumulation. The findings demonstrated that the GO conjugates accumulated significantly in metastatic tumor nodules within the lungs. The conjugates’ ability to selectively target tumor vasculature and deliver therapeutic agents efficiently makes them attractive candidates for image-guided therapeutic delivery in metastatic breast cancer treatment (Fig. 14).
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Fig. 14 (A) GO nanomaterials modulate tumor angiogenesis by tumor vasculature targeting and VEGF adsorption. (B) Inhibition of angiogenesis by BSA-GO in the chick chorioallantoic membrane (CAM). Dotted circles represent the area of the CAM under cellulose filter paper discs. The relative branch numbers of the corresponding CAMs were quantified by counting the blood vessel branch points in each image. (C) The CAM vasculature was treated with PBS (control), NC (RGD+), NC (RGD−), SFB-loaded GCRP-NC, and free SFB on day 14 for 3 days. The total vessel network length (Tot. length), the number of branching points (Nb. branches), the number of nodes (Nb. nodes), and the number of junction points (Nb. junctions). Reproduced from ref. 32 with permission from Elsevier, copyright (2016). (D) The stereomicroscopy images of the CAM vessels. (E) Quantitative analysis using NIH ImageJ with the angiogenesis analyzer plugin. The total vessel network length (Tot. length), the number of branching points (Nb. branches), the number of nodes (Nb. nodes), and the number of junction points (Nb. junctions).130 Reproduced from ref. 130 with permission from Elsevier, copyright (2022). Data are expressed as mean ± SD (n = 6). Notes: *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; NS, not significant. |
Another study discussed the development of a targeted drug delivery system using graphene quantum dots (GQDs) and poly(D,L-lactide-co-glycolide) (PLGA) nanocomposites to deliver sorafenib (SFB) for angiogenesis inhibition.129 SFB, a multi-tyrosine kinase inhibitor, effectively suppresses tumor progression and angiogenesis but has limitations due to its narrow therapeutic window and adverse side effects. To enhance SFB's delivery and reduce toxicity, researchers have functionalized nanocomposites with an integrin-targeting ligand, the RGD peptide. Integrin receptors, particularly the αvβ3 subtype, are overexpressed in tumor tissues and play a role in angiogenesis and metastasis. The RGD peptide binds to these receptors, facilitating selective drug transfer to cancer cells. The targeted delivery system effectively suppresses angiogenesis, making it a promising drug delivery system for poorly water-soluble therapeutic agents like SFB. The nanocomposites’ unique therapeutic and bioimaging properties make them a valuable tool for improving the curative effect of angiogenesis therapies.
GO nanomaterials have been studied for their potential to induce apoptosis in tumor-associated macrophages (TAMs), which play a critical role in the tumor microenvironment by supporting tumor growth and suppressing immune responses. Research has demonstrated that functionalized graphene and graphene oxide can induce apoptosis in macrophages through various mechanisms. Oxidative stress, disruption of the cellular cytoskeleton, and interference with cellular functions are among the pathways through which graphene-based materials affect macrophages. The interaction between graphene and macrophages can produce reactive oxygen species (ROS), which can overwhelm the cell's antioxidant defenses, leading to oxidative damage and eventual cell death.136 Moreover, GO can induce autophagy in macrophages in a concentration-dependent manner, as evidenced by the appearance of autophagic vacuoles and activation of autophagic marker proteins. This autophagy is regulated by TLR4, TLR9, and downstream adaptor proteins MyD88, TRIF, and TRAF6, highlighting the connection between autophagy and TLR signaling.137
GO nanomaterials could activate macrophages via toll-like receptor (TLR)-mediated and nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB)-related signaling pathways, leading to the secretion of cytokines such as interleukin-1 alpha (IL-1α), interleukin-6 (IL-6), interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-α), and granulocyte-macrophage colony-stimulating factor (GM-CSF), as well as chemokines like monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein-1 alpha (MIP-1α), macrophage inflammatory protein-1 beta (MIP-1β), and regulated upon activation, normal T-cell expressed and secreted (RANTES).138 This activation can contribute to the creation of a pro-inflammatory microenvironment. GO nanomaterials have been shown to modulate the tumor microenvironment by enhancing the infiltration of immune cells and stimulating anti-tumor immunity. GO can activate macrophages to release pro-inflammatory cytokines, which recruit and activate immune cells, contributing to tumor eradication.139 Furthermore, GO nanomaterials have been used to deliver anticancer drugs directly to tumors, improving treatment efficacy while minimizing systemic toxicity. A multifunctional nanoplatform for efficient chemo-gene combination cancer therapy is developed based on platinum-functionalized nano-GO. This system leverages GO's tumor-targeted accumulation and sustained drug release to significantly reduce nonspecific toxicity of cisplatin (CisPt). Furthermore, the platform achieves synergistic chemo-gene therapy via co-delivery of the chemotherapeutic agent CisPt and anti-miR-21. This approach simultaneously addresses drug resistance and systemic toxicity, thereby providing a novel paradigm for precision cancer therapy.140 Moreover, the nanosheets (FU/GO-PEG-GE11) made of GO were functionalized with GE11, an effective ligand for the epidermal growth factor receptor (EGFR), and loaded with the anticancer drug 5-fluorouracil. The targeting nanosystem was substantially more cytotoxic in vitro than non-targeting nanohybrids.141 According to in vivo data, targeted nanocomposites exhibited a potent anticancer effect while having almost no systemic adverse effects in mice with subcutaneous cervical cancer. Notably, the lateral size of GO nanomaterials could significantly impact macrophage activation. A series of GO samples with varying lateral sizes were constructed from the same starting material to focus specifically on the effect of size.132 They discovered that larger GO sheets exhibited stronger adsorption on the plasma membrane but were less prone to phagocytosis. The features led to more robust interactions with toll-like receptors (TLRs) and more potent activation of the NF-κB pathway. More miniature GO sheets, on the other hand, were more readily taken up by cells. Consequently, more significant GO promoted greater M1 polarization, which is associated with increased production of inflammatory cytokines and recruitment of immune cells.
To relieve hypoxia in the TME, a novel strategy was designed to enhance chemo-photodynamic therapy's efficacy through a smart nanosystem.46 This system is based on manganese dioxide (MnO2)-doped GO nanosheets, which have been engineered to simultaneously address two significant limitations in cancer therapy: tumor hypoxia and high levels of the GSH antioxidant. In this work, the MnO2 component imparts three fundamental properties. Firstly, it catalyzes the decomposition of H2O2 into oxygen, alleviating the hypoxic conditions characteristic of tumors. Secondly, it depletes the intracellular GSH levels, making tumor cells more susceptible to chemotherapeutic agents. Thirdly, MnO2 generates Mn2+ ions, which can participate in a Fenton-like reaction, further contributing to the anti-tumor activity. This work not only achieves chemo-photodynamic synergistic therapy but also significantly enhances therapeutic efficacy by modulating the TME, thereby providing a novel strategy for developing intelligent nanodrug delivery systems.
Analogously, gold nanoparticles are also hybridized with GO nanosheets for oxygen generation. A polypyrrole (PPy)-based multifunctional nanocomposite, abbreviated as PGPAI, has been developed for dual-modal imaging and enhanced synergistic phototherapy against cancer cells.142 This composite is composed of PPy nanoparticles, graphene oxide (GO) sheets, polyethylene glycol (PEG) chains, gold nanoparticles (Au NPs), and IR820 molecules. Notably, the Au NPs possess catalase-like activity, enabling them to decompose hydrogen peroxide (H2O2), which is upregulated in tumor environments, to produce oxygen. This oxygen production enhances the efficacy of oxygen-dependent PDT.
In current oncological research, PTT mediated by GO has garnered widespread attention as an innovative treatment strategy. Studies have demonstrated that the application of GO in PTT significantly influences the biological behavior of tumor cells.145 However, the substantial local temperature increase in the tumor microenvironment can induce thermal stress, leading to the heat shock effect. Heat shock is a biological response where cells react to high temperatures or other external stressors, serving as an adaptive mechanism to environmental stress. Closely associated with cellular stress, cells undergo heat shock in response to elevated temperatures, hypoxia, or exposure to toxins as a protective measure.146 A characteristic feature of this response is the increased expression of heat shock proteins (HSPs). HSPs, a group of proteins whose expression is upregulated in response to heat stress, assist in maintaining proper folding and stability of cellular proteins, preventing aggregation of damaged proteins. In PTT, this local thermal stress triggers the production of intracellular HSPs as a natural cellular protective response. During PTT treatment of cancer, the expression of HSPs can rapidly confer protection to cancer cells and reduce the efficacy of PTT, leading to insufficient apoptosis and tumor recurrence.147 In conventional PTT, the surrounding normal tissue of the tumor site may suffer from high-temperature burns. Therefore, conducting mild-temperature photothermal therapy (MTPTT) at lower temperatures is a promising approach for tumor treatment. However, in MTPTT, the overexpression of HSPs increases the thermal tolerance of tumor cells, thereby limiting its therapeutic effect.148
The temperature increase caused by PTT may induce more significant heat shock protein Hsp70 activity. With more intense treatment, the expression of the Hsp70 gene increases to counteract the therapeutic effect. Thus, the expression of the Hsp70 gene may be used as an indicator of treatment efficacy because it is upregulated under extreme thermal stress.149 Based on reduced graphene oxide (rGO), IR780 was covalently conjugated to rGO. The system showed an eightfold increase in HSP70 protein synthesis compared to the control group. This time-dependent upregulation of HSP70 protein expression confirms the mechanism of near-infrared light-induced PTT, making HSP70 a potential target for enhancing the efficacy of PTT.150 HSP90 is significantly believed to interact with AKT (protein kinase B). In many tumor cells, HSP90 is overexpressed and promotes tumor growth and survival by stabilizing essential signaling proteins such as AKT.20 Therefore, inhibiting the activity of HSP90 has been viewed as a potential anti-tumor treatment strategy, which can reduce the stability and activity of AKT, thereby inhibiting the growth and survival of tumor cells.
To maximize PTT efficiency of graphene oxide (GO), a primary strategy involves suppressing heat shock protein (HSP)-mediated thermoresistance. This was achieved by non-covalent loading of the HSP90 inhibitor NVP-AUY922 onto GO nanosheets, constructing a multifunctional GO/NVP-AUY922 nanoplatform. The nanosystem synergistically combines GO's photothermal conversion capability with NVP-AUY922's HSP90 inhibition, effectively disrupting the cytoprotective heat shock response and enhancing tumor ablation under near-infrared (NIR) irradiation. This system was applied for photothermal therapy in HeLa cells, significantly inhibiting the expression of HSP90 in cancer cells, thereby helping to overcome the thermal stress resistance of tumor cells.151 SNX-2112, another Hsp90 inhibitor, primarily functions by binding to the N-terminal ATP binding site of Hsp90. HSP90 inhibitor SNX2112 was loaded onto the surface of GO. By non-covalently binding chitosan (CHI) onto the GO surface to improve biocompatibility and grafting hyaluronic acid (HA) as a targeting ligand for CD44, they achieved specific recognition of tumor cells and enhanced efficiency in anti-tumor drug delivery. Under NIR irradiation, the GO-CHI-HA/SNX-2112 nanosystem released SNX-2112 molecules at the tumor site through a photothermal effect. This photothermal therapy mechanism enhances the drug's efficacy by raising the temperature at the tumor site, simultaneously synergizing to reverse drug resistance, effectively inhibiting and killing A549 lung cancer cells while exhibiting lower toxicity to normal cells.152
Simultaneously targeting HSP as a therapeutic endpoint, combined with PTT and other treatment modalities such as chemotherapy, immunotherapy, and PDT, is anticipated to enhance PTT's efficacy synergistically. Elevated temperatures induced by PTT can augment blood flow, facilitating increased intracellular transport and accumulation of chemotherapeutic agents within tumor cells. Additionally, complementary treatments may render cancer cells more susceptible to PTT, effectively improving therapeutic outcomes and prognosis. A stimulus-responsive scaffold composed of poly(acrylic acid)-grafted polylactic acid (PAA-g-PLLA) modified GO with gambogic acid (GA) and polycaprolactone (PCL) was constructed and further pre-implanted with adipose-derived stem cells (ADSCs). GA, a chemical small molecule inhibitor of heat shock protein synthesis, enhances the therapeutic effect of MTPTT. Under mild-temperature photothermal conditions, tumor cells were directly killed, and ADSCs also survived better and differentiated into adipocytes, thus promoting the formation of new fatty tissue. This method is efficient in treating tumors and provides strong support for postoperative tissue regeneration (Fig. 15).153 Additionally, some methods can circumvent the protective mechanism of HSPs. A mitochondria-targeted low-temperature photothermal therapy (LTPTT) nanocomposite was constructed based on berberine derivatives (BD). This nanocomposite, composed of PEG-modified GO and BD combined with tumor-targeting folic acid (FA), effectively accumulates in the mitochondria of osteosarcoma cells, achieving enhanced mitochondrial-targeted LPTT effects. It directly activates the mitochondria's intrinsic apoptotic pathway, overcoming the tumor thermal resistance caused by HSPs in MTPTT.68
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Fig. 15 (A) Schematic illustration of the therapeutic process of an ADSC-loaded GO-GA-polymer scaffold with the pH-triggered dual release of GO and glycyrrhetinic acid (GA). (B) Under NIR irradiation, the tumor site temperature of tumor-bearing mice implanted with the GO-GA-polymer scaffold significantly increased compared to those implanted with PCL and GO-polymer scaffolds. (C) The GO-GA-polymer scaffold significantly promotes the adhesion of ADSCs under NIR irradiation, enhances the expression of adipogenic genes such as PPAR-γ, C/EBP-α, adiponectin, etc., ultimately stimulating the adipogenic differentiation of ADSCs and promoting the formation of adipose tissue in vivo, making it an ideal candidate material for achieving both photothermal tumor therapy and adipose tissue regeneration.153 Reproduced from ref. 153 with permission from Wiley, copyright (2019). |
Recent studies have highlighted the efficacy of GO-based nanomedicines in altering the TME to create a less hospitable environment for tumor growth. GO's ability to scavenge ROS can normalize the redox balance within the TME, mitigating inflammation and disrupting the conditions that support tumor proliferation. Its anti-angiogenic effects contribute to tumor regression by inhibiting the formation of new blood vessels necessary for tumor sustenance. Moreover, GO's interaction with immune cells can convert the immunosuppressive TME into one that promotes immune cell infiltration and activation, thereby enhancing the body's natural defenses against cancer. The utilization of GO in cancer theranostics has expanded to include photothermal therapy. GO acts as a photothermal converter to generate heat upon light exposure, effectively destroying cancer cells while minimizing damage to surrounding healthy tissues. This approach can be augmented with other modalities, such as chemotherapy and immunotherapy, to provide a more comprehensive treatment strategy. Furthermore, GO's capacity for targeted drug delivery, facilitated by its responsive surface modifications, allows for the precise shuttling of drugs directly to the tumor site, improving therapeutic efficacy and reducing side effects.
GO demonstrates innovative potential in oncotherapy due to its two-dimensional structure, high specific surface area, abundant oxygen-containing functional groups, and excellent photothermal conversion efficiency. As a nanocarrier, GO enables the loading and tumor-targeted delivery of various drug molecules including chemotherapeutics, nucleic acids, immunomodulators etc. Meanwhile, the intrinsic PTT and PDT capabilities of GO could induce effective tumor ablation upon near-infrared irradiation. The first strictly controlled human exposure clinical trial indicates that inhaling GO causes no short-term adverse effects on lung or cardiovascular function (NCT03659864). The positive outcomes of the clinical study provide essential evidence for GO biosafety validation. However, future development of GO-based nanomedicines still faces various critical challenges. Long-term biocompatibility and in vivo metabolic pathways of GO remain unclear. To address the challenge, GO biodistribution and metabolic products could be tracked using the isotope labelling method. Chemical structure modification of GO is supposed to be an effective approach for improving biocompatibility. Biodegradable chemical bonds could be introduced into the 2D structure skeleton. Optimization of surface modifications to reduce immunogenicity and toxicity becomes imperative, such as a stable protein corona. On the other hand, large-scale production processes of GO suffer from issues of batch-to-batch variability and residual impurities. These variations critically impair GO's performance in biomedical applications, especially its biosafety. Future investigations should prioritize the development of green synthesis techniques to address process-induced variations and ensure production consistency. Additionally, clinical translation of GO nanomaterials necessitates GLP/GMP-compliant validation and comprehensive pharmacodynamic, pharmacokinetic, and toxicological profiling.
Footnote |
† These authors contributed equally to this work. |
This journal is © The Royal Society of Chemistry 2025 |