Introduction

Carbon dots are nanomaterials with sizes typically less than 10 nm, known for their remarkable optical properties and excellent biocompatibility, which make them suitable candidates for applications in cancer therapy, such as drug and gene delivery, bioimaging and biosensing, and photodynamic and photothermal therapy.1 CDs exhibited stable performance in aqueous solutions and have photoluminescence that varies with the excitation wavelength, distinguishing them from traditional organic dyes and semiconductor quantum dots.2 CDs can be synthesized using various techniques, including microwave, hydrothermal, solvothermal, as well as approaches utilizing natural or synthetic organic precursors.3 Other synthesis techniques, such as ultrasonication, electrochemical oxidation, and pyrolysis, have also been explored to optimize their optical and structural properties for biomedical applications.3,4 However, despite their advantages, CDs have limitations such as low quantum yield (QY), reduced biological functionality, and potential inference from non-specific interactions in detection processes.1 Recent advancements in surface modification and doping strategies have been developed to overcome these challenges, enhancing their fluorescence efficiency, biocompatibility, and targeted delivery for improved diagnostic and therapeutic applications.5

Surface modification techniques have been developed to enhance the targeted delivery of CD-based theranostic platforms.5 A common approach involves conjugating CDs with active targeting molecules (eg, hyaluronic acid (HA),5,6 folic acid (FA),6 lysosomes,7 lipids,8 and protein transferrin9), which which enhance the targeting of drugs (eg, doxorubicin (Dox),10 docetaxel (DTX),11 paclitaxel (PTX),12 epirubicin and temozolomide (EPI-TMZ)9) and genes (pDNA and siRNA).13–20

Recent advances have focused on self-targeting strategies to simplify these systems.21 By using photosensitizers (eg, porphyrin, phthalocyanine, pheophytin22–24), and organic molecules (eg, chitosan,22 hyaluronic acid25) with inherent targeting properties as precursors during CDs synthesis, after the synthesis can preserve the targeting functionality on the CDs surface. This inherent targeting eliminates the need for additional targeting agents and improves the precision of tumor-specific delivery.26 These innovations in surface modification significantly enhance the diagnostic and therapeutic potential of CDs, making them more effective for cancer treatment.

Doping CDs with metals (eg, copper,27 iron,14 hafnium,28 gadolinium,29,30 magnesium31,32) or various heteroatoms (eg, nitrogen,33 sulfur,34 phosphorus,35 boron36) is another effective strategy to enhance their optical properties, making them more suitable for applications in bioimaging and cancer therapy.37 The synthesis of doped CDs involves introducing atomic impurities to adjust the material’s electronic band gap (HOMO-LUMO).27 This process directly impacts their optical properties by increasing the efficiency of light absorption and emission.37 Furthermore, co-doping with multiple elements can result in synergistic effects, enhancing their optical and electronic properties.1 These advances have broadened the applicability of CDs in various biomedical fields, including biosensing, bioimaging, and therapeutic platforms.1

Surface-modified and doped CDs in cancer therapy have shown remarkable potential in improving the effectiveness of both PDT and PTT. PDT involves using photosensitizers (PSs) that generate ROS upon activation by light, leading to oxidative stress and apoptosis in cancer cells.38 Doped CDs, especially those modified with metals or heteroatoms such as phosphorus-nitrogen,33–35 copper,27 tin,39 or manganese,40,41 and selenium42 have shown enhanced photosensitization properties, improving ROS generation and increasing the therapeutic efficacy of PDT. These dopants alter the electronic structure of CDs, enhancing their light absorption capabilities and making them more efficient in generating ROS under light irradiation.43 As a result, these doped CDs function as effective photosensitizers that precisely target and destroy cancerous tissues.

PTT leverages the ability of CDs to convert light energy into heat, leading to the localized ablation of tumor cells.14 Surface-modified or doped CDs are photothermal agents (PTAs), absorbing near-infrared (NIR) light and generating sufficient heat to induce cancer cell death.44 The photothermal conversion efficiency (PCE) of these CDs can be further improved through doping with PTAs (eg, indocyanine green (IR825),45 polydopamine,46 and peptides47) and metals or heteroatoms (eg, nitrogen,48,49 sulfur,50 copper-sulfur,51 and selenium-sulfur52 co-doped), making them highly effective in PTT applications. Combining PDT and PTT in a single treatment platform allows for a synergistic effect, where the heat generated by PTT can enhance oxygen diffusion to the tumor, overcoming hypoxia, a common limitation in PDT. Likewise, the ROS produced in PDT can disrupt cancer cell defenses, making them more susceptible to the thermal effects of PTT.

Surface-modified CDs and their integration into nanocomposites significantly enhance the efficiency of PDT and PTT through a synergistic approach.53 By incorporating CDs with other nanoparticles (eg gold nanorods,54 calcium phosphate nanoparticles10), the photothermal conversion efficiency is markedly improved, leading to increased generation of ROS under laser irradiation. This heightened ROS production, coupled with the heat generated during PTT, results in greater apoptosis of cancer cells.55 Additionally, lipid-coated CDs8 and lysosome-targeted CDs7 facilitate enhanced cellular uptake and provide a protective environment for therapeutic agents, ensuring improved stability and targeted delivery to tumor sites. This lipid encapsulation also mitigates plasma protein adsorption, further amplifying the therapeutic effects of both PDT and PTT while maintaining biocompatibility and reducing toxicity.56 Collectively, these advancements pave the way for more effective and targeted cancer treatments.

Other applications such as chemodynamic therapy (CDT), sonodynamic therapy (SDT) and gas therapy have been explored to enhance cancer treatment. In CDT, metal-catalyzed reactions generate ROS within the tumor microenvironment, including oxidative stress and selective cancer cell apoptosis.57 Doped CDs, such as Fe3+-decorated CDs and Cu-doped CDs, have been designed to enhance Fenton and Fenton-like reactions, increasing ROS production and tumor-specific cytotoxicity.58 In SDT, ultrasound energy actives sonosensitizers, leading to ROS production through cavitation effects.59 CDs doped with Ti, Cu or Mn as well as CDs integrated into microbubbles, have demonstrated improved ultrasound absorption, enhanced ROS yield, and deep tissue penetration, making SDT a promising non-invasive therapeutic strategy.60 Gas therapy involves the controlled release of therapeutic gases, such as nitric oxide (NO) or hydrogen sulfide (H2S), which modulate cellular pathways and enhance cancer cell apoptosis.61 Surface-modified CDs, including NO-releasing CDs and CDs combined with catalase-mimicking nanozymes, have been developed to generate and regulate gas-mediated tumor suppression.61

This review aims to comprehensively explore and summarize the advancements in the field of surface-modified CDs for both in vitro and in vivo applications in cancer therapy. It will emphasize the significance of surface functionalization and doping strategies in augmenting the therapeutic potential of CDs. Additionally, the review will highlight key applications, including drug delivery, multimodal imaging, PDT, PTT, CDT, SDT and gas therapy. By reviewing studies conducted to date, this work seeks to assess the limitations and strengths of the approaches taken to improve therapeutic outcomes and optimize the design of CDs within targeted cancer therapy, ensuring an absolute equilibrium between therapeutic efficacy, biocompatibility, and safety.

Surface-Modified CDs as a Nanocarrier System

Surface modification of carbon dots (CDs) plays a crucial role in their application as nanocarriers in cancer therapy. Enhancing drug and gene delivery systems through surface modifications allows CDs to improve targeting specificity, biocompatibility, and therapeutic efficiency.4 This section focuses on how surface-modified CDs function as nanocarriers in drug and gene delivery systems, with a detailed discussion of synthesis methods and in vitro and in vivo applications and compares their strengths and limitations (Table 1).

Table 1 Surface Modified CDs as a Nanocarrier Applications

Drug Delivery Systems for Cancer Therapy

Surface-modified CDs have gained attention as ideal drug delivery systems for cancer therapy due to their excellent biocompatibility, small size, and ease of functionalization. These modified CDs can selectively bind to receptors overexpressed on cancer cell surfaces, facilitating the internalization of drugs and improving therapeutic efficacy while minimizing systemic toxicity.67

The drug delivery mechanisms of CDs typically rely on various interactions between the CDs and the drug molecules. The electrostatic interaction is the most common mechanism, where drugs with opposite charges can attach to the surface-modified CDs. For instance, Zeng et al65 synthesized fluorescent CDs with carboxyl-rich surfaces using a hydrothermal method, resulting in a particle size of ≈5 nm that exhibited excellent biocompatibility and low cytotoxicity. Similarly, Yuan et al68 produced CDs from milk hydrothermolysis, achieving an average size of 4 nm and demonstrating high loading efficiency for doxorubicin (DOX) via electrostatic interactions.

Despite the previous results, there are still limitations in targeting drugs and achieving the best interaction between the drug and CDs. CDs are often surface-modified with various ligands, polymers, or targeting agents to enhance this interaction and improve drug delivery. These modifications can improve the selective targeting of cancer cells, enhance solubility, and facilitate drug-controlled release. For instance, J. Li et al63 modified CDs’ surface with a hyaluronic acid (HA) hydrophilic group that helps the electrostatic interaction between CDs and DOX, forming pH-responsive HA-CD@DOX. In a similar study, S. Li et al9 utilized protein transferrin to modify CD’s surface and help the interaction with a cancer drug DOX, enhancing the solubility.

Additionally, the ability of CDs to target the cancer drug docetaxel (DTX) to develop DTX-CDs and paclitaxel (PTX-loaded CDs) and surface modified HA, through covalent bonding; the functional groups, carboxyl and hydroxyl on the surface of CDs can form a covalent bond with drug molecules providing stability in drug loading. Both systems exhibited excellent biocompatibility and enhanced drug delivery with minimized side effects.11,12

The ability to control drug release is a critical feature of CDs-based delivery systems. For instance, S. Wang et al6 reported a pH-responsive release profile with folic acid (FA) to develop an FA-CDs-DOX system, where 79.31% of DOX was released at pH 5.5 compared to just 16.39% at pH 7.4. This differential release mechanism is attributed to the protonation of amino groups in DOX under acidic conditions prevalent in tumor microenvironments, enhancing drug solubility and facilitating targeted therapy. Similarly, Duan et al64 reported modified CDs with HA and DOX, developing a dual-responsive CDs-HA/DOX system that demonstrated a release efficiency of 66% of DOX, showcasing the potential for utilizing both pH and HA responsiveness to enhance therapeutic outcomes. However, Kong et al62 demonstrated 75% of DOX release at pH 7.4 with high accumulation and cytotoxicity of DOX in MCF-7 cancer cells.

In vitro studies have demonstrated the efficacy of surface-modified CDs in drug delivery across various cancer types. Zeng et al65 reported that their DOX-loaded CDs induced a 40% death rate in HepG2 liver carcinoma cells at 2.5 µg/mL concentration while maintaining safety for normal liver cells (HL-7702). In a similar study, Kong et al62 evaluated pharmaceutical activity and cellular toxicity of CDs-DOX on MCF-7 and L929 breast cancer cells with cell viability above 90% at 0.5 μg/, reducing cancer cell growth significantly. The selective targeting capability and cytotoxic effect of CDs improved significantly through surface modification with HA.

J. Li et al63 utilized HA-modified CDs (HA-CDs) to deliver DOX to 4T1 breast cancer cells, achieving high cytotoxicity rates and demonstrating a tumor growth inhibitory rate of 80.23% compared to free DOX. Similarly, Duan et al64 demonstrated the inhibition of growth and migration of CDs-HA-Hep/DOX system dual responsive by HA and pH value in MGC-803 human gastric carcinoma cell line. Additionally, the delivery of drugs for brain cancer has been proved. S. Li et al9 explored transferrin-conjugated CDs (T-CDs) for delivering DOX to brain tumor cells (CHLA-266), achieving greater uptake than non-targeted systems. In a similar study, Hettiarachchi et al66 utilized triple conjugated C-DT with transferrin and drugs (Epirubicin and Temozolomide) to treat pediatric (SJGBM2, CHLA266, CHLA200) and adult (U87) brain tumor cells, and Quiao et al69 proved the high endocytosis rate of C6 brain glioma cells treated with CDs from glucose and L-aspartic acid. This study highlights the importance of functionalizing CDs with targeting ligands to enhance cellular uptake and therapeutic effectiveness in brain tumors.

In vivo studies further validate the efficacy of targeted drug delivery using CDs. Zheng et al21 demonstrated that their CD-Asp system effectively targeted C6 glioma cells in orthotopic models, with the highest fluorescence intensity observed at 15 minutes post-injection. This rapid accumulation indicates efficient targeting and potential for real-time monitoring. Zeng et al65 and S. Wang et al6 evaluated CDs-DOX in liver cancer cells of nude white mice. The HepG2 tumors reduced 50% of the volume in their first 24 hours and 72.13% after 20 days of treatment.

Additionally, J. Li et al63 evaluated the effectiveness of DOX in 4T1 tumor-bearing BALB/C mice, which were intravenously injected with ICG-loaded HA-CD@p-CBA-DOX. The results showed a tumor growth inhibitory rate of 80% compared to free DOX. These studies confirmed that the pH-responsive release mechanism significantly contributed to the selective accumulation of DOX in tumor tissues while minimizing effects on healthy cells.

The targeted drug delivery capabilities of carbon dots are supported by numerous studies demonstrating their effectiveness across different cancer types through tailored synthesis methods and functionalization strategies. The combination of in vitro and in vivo studies highlights the potential of modified carbon dots as ideal carriers for anticancer drugs like DOX, providing a promising avenue for improving therapeutic efficacy while minimizing side effects. Despite their efficiency in drug release, CDs may have a limited capacity for loading large amounts of therapeutic agents. Moreover, the surface modification process, especially with polymers and targeting ligands, can be complex and may affect the stability of the CDs during storage or in biological systems.

Gene Therapy for Cancer Therapy

Gene therapy involves introducing genetic material into cells to correct or inhibit the expression of disease-related genes. This process requires an efficient delivery system to protect the genetic material and ensure its uptake by the target cells. Surface-modified CDs have emerged as promising candidates for gene delivery due to their ability to condense genetic material and protect it from degradation while offering efficient cellular uptake and gene transfection.70

CDs serve as carriers for plasmid DNA (pDNA), small interfering RNA (siRNA), or other nucleic acids in gene delivery for cancer therapy. The main mechanisms through which CDs deliver genes are electrostatic interactions. Luo et al14 synthesized iron-doped carbon dots (Fe@CDs) functionalized with a dendritic lipopeptide (PEG-RLS). This modification improved the stability of CDs and facilitated plasmid DNA (pDNA) targeting gene delivery. The positive charges on surface-modified CDs interact with negatively charged nucleic acids, facilitating efficient binding and condensation of the genetic material. The resulting nanohybrid system exhibited a diameter of 77 nm and demonstrated effective gene transfection in vitro, with fluorescence intensity indicating a two-fold increase in green fluorescent protein (GFP) expression upon laser irradiation compared to controls (see Figure 1A and B).

Figure 1 PEG-RLS/Fe@CDs: (A) schematic mechanism of cancer nanotheranostic platform for gene delivery, PTT and bioimaging applications. Reproduced from Luo T, Nie Y, Lu J, et al. Iron doped carbon dots based nanohybrids as a tetramodal imaging agent for gene delivery promotion and photothermal-chemodynamic cancer synergistic theranostics. Mater Des. 208. © 2021 The Authors. Published by Elsevier Ltd. Creative Commons CC-BY-NC-ND license.14 (B) In vivo photo-enhanced therapeutic effect of PEG-RLS/Fe@CDs under 660 nm (0.5 W/cm2 for 5 min) in 4T1 breast cancer, tumor growth inhibition after different treatments, tumor tissues after treatment. HP-CDs/DNA: (C) Zeta potential of free DNA, HP-CDs and HP-CDs/DNA complex, identification of transfection efficiency of HP-CDs, fluorescent images of gene expression obtained by a confocal laser scanning microscope in HEK-293T cells transfected with DNA/HP-CDs complexes, (D) Schematic synthesis and biomedical applications of HP-CDs. Reproduced from Zhang M, Zhao X, Fang Z, et al. Fabrication of HA/PEI-functionalized carbon dots for tumor targeting, intracellular imaging and gene delivery. RSC Adv. 2017;7:3369–3375. This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence.15

Similarly, M. Zhang et al15 developed hyaluronate and polyethylenimine (HA-PEI) functionalized carbon dots (HP-CDs) through a bottom-up synthesis method. These HP-CDs exhibited excellent dispersibility and biocompatibility, allowing efficient internalization into cancer cells via CD44 receptor-mediated endocytosis. After binding on nucleic acids, CDs are internalized by cells through endocytosis, which allows for the delivery of genes into the cytoplasm and eventual gene expression. In vitro transfection experiments using HEK-293T human embryonic kidney cells showed that HP-CDs could effectively condense pDNA, enhancing gene expression in MGC-803 gastric cancer cells (see Figure 1C and D).

Moreover, CDs can be surface-modified to carry interfering RNAs (siRNAs). S. Kim et al16 developed a CD-based siRNA approach and delivered into breast cancer cells for the first time. Similarly, H. J. Wang et al17 constructed amphiphilic carbon dots (ACDs) by conjugating hydrophobic alkyl epoxide to PEI-derived CDs, achieving a high transfection efficiency of approximately 96.7% of siRNA in A549 lung cancer cells. The ACDs exhibited superior cellular uptake compared to traditional transfection agents like Lipofectamine 2000, indicating their potential as effective gene carriers and causing cell apoptosis.

Surface modifications help protect genetic material from degradation and enhance cellular uptake, allowing for effective gene silencing or expression in cancer cells. X. Yang et al18 synthesized positively charged CDs using PEI and FA, facilitating selective imaging and transfection of folate receptor-positive HeLa cells. The C-dots demonstrated a significant ability to target pDNA due to electrostatic interactions, achieving a transfection efficiency of 42.63% in HeLa cells. This study underscores the importance of targeting specific receptors for enhanced gene delivery. In a related study, Zhao et al19 reported the development of CDs modified with PEG and PEI to deliver pDNA, developing CDs-PEG-PEI/pDNA complexes that exhibited pH-responsive charge reversal, enhancing cellular uptake in mildly acidic tumor microenvironments. This system achieved high transfection efficiency in MCF-7 breast cancer cells, with a notable % tumor growth inhibition rate of 70% observed in vivo. The study highlights the potential of pH-responsive systems for targeted gene therapy.

In vivo studies provide critical insights into the therapeutic potential of CDs for gene delivery. Luo et al14 developed Fe-doped CDs (Fe@CDs) and modified the surface with PEG to enhance the biocompatibility and delivery of genes. The delivery system PEG/Fe@CDs demonstrated 80% tumor growth inhibition in 4T1 breast cancer models, with the delivery of green fluorescent protein (EGFP) gene expression that achieves fluorescence intensity signals indicating effective accumulation at tumor sites over time (see Figure 1B). The results suggest that this multifunctional delivery system could be an effective platform for photothermal-responsive gene therapy. According to H. J. Wang et al17 demonstrated that Amphiphilic CDs (ACDs) were developed as vectors to target siRNA. ACDs showed promising results in delivering siRNA to A549 lung cancer cells. The study reported enhanced apoptosis rates due to effective gene silencing, further validating the potential of ACDs as efficient gene delivery vehicles. Ghosh et al20 developed CDs functionalized with poly(amidoamine) (PAMAM) dendrimers for triple-negative breast cancer (TNBC). Their CD-PAMAM system demonstrated low toxicity even at high concentrations and improved gene transfection efficiency, making it a promising candidate for TNBC therapy.

Multimodal-Imaging in Cancer Therapy

Multimodal imaging is a crucial aspect of cancer diagnostics, offering real-time monitoring and visualization of tumor cells. Surface-modified CDs and heteroatoms/metals-doped CDs are pivotal for enhancing imaging capabilities due to their tunable fluorescence, high quantum yield, and biocompatibility (Table 2).19 This section focuses on using surface-modified and doped CDs for tumor detection and bioimaging, discussing their synthesis methods, mechanisms, and performance in vitro and in vivo studies.

Table 2 Multimodal Imaging Applications of Surface-Modified Carbon Dots

Surface Modified-CDs for Tumor Detection

Fluorescence imaging leverages the photoluminescence (PL) properties of CDs, which originate from quantum confinement effects and surface defect states.74 The reduced size of CDs (<10 nm) restricts the motion of electrons and holes, resulting in size-dependent emission. Additionally, functional groups such as –COOH, –OH, and –NH2 on the CD surface enhance fluorescence by introducing trap states, that influence electron–hole recombination.75

For instance, Wang et al17 synthesized amphiphilic carbon dots (ACDs) by conjugating alkyl epoxide to CDs. The alkyl epoxide reacts with amine (–NH2) or hydroxyl (–OH) groups on the CD surface through epoxide ring-opening reactions, forming covalent bonds. This process produces CDs with hydrophobic alkyl chains and hydrophilic functional groups (–OH, –COOH, –NH2) on the surface. The amphiphilic modification enhances interaction with cell membranes and improves cellular uptake, enabling efficient delivery to the target sites. These ACDs exhibited bright fluorescence in A549 lung cancer cells and demonstrated effective cellular imaging.

Similarly, Yang et al18 developed positively charged carbon dots by functionalizing them with folic acid (FA) and polyethyleneimine (PEI). These CDs showed selective imaging of folate receptor-positive HeLa cells. FA binds specifically to folate receptors (FRα), which are overexpressed on cancer cells, facilitating receptor-mediated endocytosis. Meanwhile, the positive charges on PEI interact electrostatically with negatively charged phospholipids in the cell membrane, promoting cellular uptake. This resulted in efficient intracellular fluorescence signals, demonstrating potential for real-time imaging applications.

In both studies, surface functionalization played a critical role in modulating the electronic bandgap, thereby influencing excitation wavelength and emission intensity.75 Hydrophilic groups improved surface passivation by reducing nonradiative recombination and enhancing fluorescence. Furthermore, the amphiphilic structure of ACDs facilitates penetration through lip bilayers, enabling cytoplasmic localization. Amine (–NH2) groups of PEI and FA contributed to covalent bonding with targeting agents, improved surface passivation, and provided positive charges for electrostatic interactions. Additionally, the cationic charges of PEI enhanced interaction with negatively charged cell membranes, promoting intracellular delivery and localized fluorescence.

In vivo applications of functionalized carbon dots have shown promising results for tumor imaging and monitoring therapeutic efficacy. Zheng et al21 synthesized highly fluorescent CD-Asp from L-aspartic acid, which showed selective imaging of C6 glioma cells in orthotopic models. The highest fluorescence intensity was observed 15 minutes post-injection, indicating rapid tumor accumulation, a critical feature for real-time imaging during therapeutic interventions. Similarly, Li et al9 reported the use of transferrin-conjugated CDs to deliver doxorubicin to brain tumor cells (CHLA-266). The conjugated system demonstrated significantly higher uptake in tumor tissues compared to normal cells, enabling precise monitoring of drug distribution via fluorescence imaging.

Moreover, Zhao et al19 developed a pH-responsive system (CDs-PEG-PEI/pDNA) that exhibited size-shrinkage in response to the acidic tumor microenvironments, promoting enhanced accumulation and penetration into tumor sites. This was confirmed through in vivo imaging of MCF-7 tumor-bearing mice, highlighting the system’s adaptability to physiological conditions. These studies underscore the versatility of CDs, which can be functionalized with ligands such as FA, or hyaluronic acid (HA) to target overexpressed receptors on cancer cells. Such ligand-mediated targeting enhances imaging specificity and reduces off target effects on healthy tissues.

The in vivo findings complement the in vitro studies discussed previously, demonstrating how surface functionalization of CDs not only enhances fluorescence properties but also enables precise cellular and tissue targeting. By leveraging their tunable surface chemistry and size, CDs hold promise as multifunctional agents for real-time tumor imaging, targeted drug delivery, and monitoring therapeutic outcomes.

Heteroatoms/Metals Doped Carbon Dots for Bioimaging and MRI Applications

When CDs are irradiated with excitation light, they can fluoresce at specific wavelength. Their PL can be tuned by altering their size, surface states or through electrochemical doping with heteroatoms (eg nitrogen, sulfur) or metal ions (eg copper, gadolinium).75,76 Heteroatom doping enhances the QY of CDs by modifying their electronic structure. For example, doping with elements like copper (Cu) or boron (B) introduces intermediate energy levels or alters the HOMO-LUMO gap, increasing the efficiency of photon absorption.75,77

J. Wang et al27 developed copper-doped carbon dots (Cu-CDs) with a high fluorescence QY of 24.4%. Cu doping minimizes nonradioactive decay caused by surface defects while enhancing radiative decay pathways. These Cu-CDs demonstrated significant photoinduced cytotoxicity against HeLa cervical cancer cells and SH-SY5Y neuroblastoma multicellular spheroids. Their bright fluorescence and low cytotoxicity make them promising candidates for fluorescence imaging in cancer diagnostics. Additionally, Cu doping enhances red and near-infrared (NIR) fluorescence by increasing light absorption in lower-energy regions.

Similarly, Shen et al71 synthesized boron doped CDs (B-CDs) with excellent multicolor fluorescence imaging capabilities, as demonstrated in HeLa cells. These B-CDs exhibited a high QY of 22% and showed potential for cell imaging agents due to their solid-state PL properties and good biocompatibility. Boron doping introduces electron-deficient sites in the CDs, which narrow the bandgap, enhancing light absorption and shifting fluorescence emission toward longer wavelengths (eg green, red, or NIR regions). This modification improves radiative recombination of electron–hole pairs, significantly increasing the QY. Furthermore, B-CDs effectively penetrate HeLa cancer cells via endocytosis, facilitated by their small size (<10 nm) and surface functional groups. Once internalized, B-CDs exhibit bright fluorescence under excitation, enabling visualization of intracellular structures. These attributes make B-CDs valuable tools for imaging applications in cancer research and diagnostics.

Doped CDs combine fluorescence with other imaging modalities, such as magnetic resonance imaging (MRI), photoacoustic imaging (PA), and computed tomography (CT), providing more comprehensive approaches to tumor detection. MRI relies on the excitation and subsequent relaxation of nuclear spins in the presence of magnetic field to generate detailed images. CDs doped with paramagnetic ions (eg gadolinium (Gd3+), manganese (Mn2+), boron (B)) or metals (eg, iron (Fe3+)) function as T1 or T2 contrast agents by enhancing proton relaxation rates.77

Gd3+ or Mn2+ dopants in CDs introduce unpaired electrons that interact with the nuclear spins of surrounding water protons. These dopants reduce the longitudinal relaxation time (T1), making tissues appear brighter in MRI scans while also creating local magnetic field that proton relaxation. For example, Du et al29 synthesized Gd-doped CDs (Gd-CDs) with excellent biocompatibility and a T1 relaxivity rate of 6.45 mM−1s−1, making them suitable for magnetic resonance/fluorescence (MR/FI) bimodal imaging. In another study, Jiao et al30 developed AS1411-Gd-CDs by conjugating Gd-CDs with AS1411 aptamers, which target nucleonic overexpressed on various adenocarcinoma cells, including breast and renal cancers. This functionalization improved targeting specificity and significantly enhanced MRI signal intensity in 4T1 tumor cells compared to non-targeted Gd-CDs (see Figure 2A–C). These findings demonstrate the potential of Gd-CDs to enhance MR imaging contrast while maintaining low cytotoxicity, thereby facilitating imaging-guided cancer treatment.

Figure 2 AS1411-Gd-CDs: (A) Schema of preparation of AS1411-Gd-CDs and FL/MR-guided PTT of tumor, (B) In vitro MR imaging properties, T1-weighted MR images of 4 T1 and NIH-3T3 cells treated with Gd-CDs. The red circles indicated the tumor cells, (C) In vivo fluorescence and PR imaging tests of 4 T1 tumor-bearing mice model pre- and post-injection of Gd-CDs and AS1411-Gd-CDs at various time intervals. The red circles represent the tumor area. Mn-CDs-NH: (D) MRI images of tumor -bearing mice treated with Gadovist or Mn-CDs-NH, (E) Metastatic investigation of lungs with MRI imaging. PEG-RLS/Fe@CDs: (F) In vitro tetramodal imaging in 4T1 cells by confocal fluorescence imaging, T1-weighted magnetic resonance (MR) images with different concentrations of PEG-RLS/Fe@CDs, (G) Real-time in vivo fluorescence images under 660 nm light irradiation (0.5 W/cm2). The white circles show the position of the tumor. (A,C,D) Reproducced from Jiao M, Wang Y, Wang W, et al. Gadolinium doped red-emissive carbon dots as targeted theranostic agents for fluorescence and MR imaging guided cancer phototherapy. Chem Eng J. 2022;440. © 2022 The Author(s). Published by Elsevier B.V. Creative Commons CC-BY-NC license.30 (E,F,G) Reproduced from Tiron A, Stan CS, Luta G, et al. Manganese-doped n-hydroxyphthalimide-derived carbon dots—theranostics applications in experimental breast cancer models. Pharmaceutics. 2021;13. © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license.72

Similarly, Mn-doped (Mn-CDs) have shown promise as dual-modal imaging agents for both MRI and optical imaging. Ji et al31 demonstrated that Mn-CDs provided significant T1 contrast enhancement in MRI imaging and enabled precise intraoperative localization of glioma tumors in mouse brain models.32 Tiron et al72 further reported Mn-CDs with a T1 relaxivity of 9.96 mM−1 s−1, highlighting their efficacy as contrast agents for combined T1-weighted MR/fluorescence imaging. In vivo studies using breast cancer animal models revealed that Mn-CDs generated strong MRI signals, with predominant uptake in the liver, kidneys, peritoneal cavity, and heart. Tumor regions exhibited intense MRI signals, as visualized through rainbow color maps for enhanced differential contrast (see Figure 2D and E). These studies underscore the potential of Mn-CDs for precise imaging and effective clearance through renal pathways, making them highly suitable for imaging and therapeutic applications. Additionally, H. Wang et al36 introduced boron-doped CDs (B-CDs) as alternative T1 contrast agents. Although B-CDs demonstrated lower magnetization compared to Gd3+ or Mn2+-based agents, they offer a novel approach to bimodal bioimaging applications. Boron doping introduces unpair electrons into the CDs structure, generating localized paramagnetic centers. These centers interact with the magnetic field and surrounding water protons, facilitating their relaxation. While Gd³⁺ and Mn²⁺ effectively enhance relaxation times, boron doping within the π-conjugated carbon network of CDs creates a distinctive paramagnetic environment with notable relaxation efficiency.

Iron-doped CDs can shorten transverse relaxation time (T2), Fe3+ ions cause magnetic inhomogeneities, shortening transverse relaxation time (T2) and darkening tissues in MRI. In a study, iron doped-CDs (Fe@CDs), was developed by solvothermal method.14 The Fe@CDs provided longitudinal relaxivity (r1) = 1.25 mm⁻¹s⁻¹, indicating they are effective T2 contrast agent in vivo (Figure 2F and G). Fe²+doped carbon dots showed significant enhancement of proton relaxation rates because its paramagnetic properties were greatly enhanced by these ions.

Photoacoustic (PA) imaging leverages the ability of CDs to convert absorbed light energy into localized heat through nonradioactive relaxation processes, enabling high-resolution imaging. Dopants like Fe2+ and Cu2+ reduce the bandgap, enhance defect states, and improve photothermal conversion efficiency, enabling effective photoacoustic (PA) imaging. The localized heat induces thermoelastic expansion in surrounding tissues, generating acoustic waves that are detected by ultrasound sensors and converted into high-resolution PA images.78 In a study, Fe@CDs were synthesized and evaluated in 4T1 cancer cells, demonstrating strong acoustic signal generation and potential for multimodal imaging (PA/fluorescence). Fe doping narrowed the bandgap and enhanced NIR absorption (650–900), improving photothermal properties.14 Similarly, Liu et al51 reported that Cu doped CDs (Cu-CDs) provided precise tumor boundary delineation due to their strong PA signals, aiding in MFC-7 breast cancer cells. Copper dopants modify the electronic structure of CDs, enhancing NIR absorption and photothermal efficiency. The rapid localized heating caused by Cu-CDs leads to thermoelastic expansion of tumor tissues, generating ultrasonic pressure waves. The intensity of these waves is directly correlated with the photothermal efficiency of Cu-CDs, significantly improved by Cu doping.

CT imaging relies on the attenuation of X-rays by high atomic number (Z) elements. Common contrast agents (iodixanol) show inherent defects and poor sensitivity. Based on this, Su et al28 synthesized hafnium-doped CDs (HfCDs) as a contrast agent, showcasing robust stability and excellent water solubility. In vivo studies demonstrated that HfCDs could provide immediate contrast enhancement at tumor sites within one-minute post-injection into H22 orthotopic liver tumor-bearing mice. The HU (Hounsfield units) CT value of the tumor region increased from 114.0 hU (0 min) to 296.0 hU at 1 min post-administration. This rapid imaging capability underscores the potential of HfCDs as contrast agents for in vivo CT for clinical diagnosis. In a similar study, CDs were synthesized with iohexol (contrast agent) as a precursor and cetuximab was targeted on the surface of CDs to form I-CQDs-C225. The contrast system showed a low cytotoxicity effect in H23 and HLF cancer cells, with cell viability at about 80%, and CT image brightness of I-CQDs increased significantly at 635 hU, compared with iodixanol (commercial contrast agent).73 These studies proved that surface modified-CDs can be used for cellular multicolor imaging and CT imaging due to their low toxicity, excellent biocompatibility, and good X-ray attenuation capacity. (Table 2)

PDT and PTT for Cancer Treatment Surface-Modified CDs in PDT for Cancer Therapy

Surface-modified CDs have emerged as promising nanomaterials for PDT and PTT, two light-based cancer treatment strategies. PDT relies on the activation of PSs by light to produce ROS that destroys cancer cells. PTT involves converting light energy into heat, which causes localized hyperthermia and ablates tumors.27,38 The functionalization of CDs with targeting agents, photosensitizers, or metal ions can significantly enhance their efficacy in PDT and PTT by improving selective accumulation, ROS generation, and thermal conversion efficiency.79 The following sections will explore the application of surface-modified CDs in PDT and PTT, focusing on their roles as nanocarrier of PSs, doped CDs as a PSs for enhanced therapeutic effects, and the use of PSs as precursors in the synthesis of CDs. Through this discussion, we aim to provide insights into the mechanisms and advantages of these approaches, emphasizing their potential in advancing cancer treatment strategies.

CDs have gained recognition as effective nanocarriers for PSs in PDT. Their unique structural features facilitate their application in drug delivery systems, including a π-conjugated core and a surface rich in hydrophilic functional groups such as –NH2, –OH, and –COOH. CDs have the following advantages as they function as nanocarriers for molecular PSs: water solubility and biocompatibility, stability and dispersibility of organic PSs in aqueous solution, and the large two-photon absorption cross-section (TPACS) enables CDs to have excellent two-photon excitation properties which can effectively kill cancer cells in deep tissues.

The central mechanism of PDT using surface-modified CDs involves the generation of singlet oxygen (1O2) or other ROS upon light activation. In a pioneering study by Sun et al,80 the PS chlorin e6 (Ce6) was targeted onto amino-rich red emissive carbon dots (RCDs) to form Ce6-RCDs. This innovative approach synergistically enhanced PTT and PDT under a single near-infrared (NIR) laser at 671 nm. The study reported an impressive photothermal conversion efficiency of 46%, enabling effective cancer treatment at a reduced laser power density of 0.50 W/cm2. In vitro experiments demonstrated that Ce6-RCDs exhibited significantly higher cytotoxicity against MCF-7 and 4T1 breast cancer cells than free Ce6 or RCDs alone. In vivo studies further confirmed the enhanced therapeutic efficacy, with multimodal imaging capabilities including fluorescence, photoacoustic, and photothermal imaging, facilitating real-time monitoring of the therapy process.

Similarly, Kim et al81 developed pH-sensitive carbon dots modified with Ce6 (Ce6@IDCDs). These CDs were designed to release Ce6 (80%) specifically in acidic tumor microenvironments (pH 6.5), triggering immunogenic cell death through PDT upon 671 nm laser irradiation. In vitro studies revealed that the treatment led to increased expression levels of CD80 and CD86 (amplifying T cell activation), indicating successful dendritic cell maturation and immune activation. The in vivo results mirrored these findings, showcasing significant antitumor effects in CT26 mouse colon cancer cells as evidenced by reduced tumor volumes and increased activation of natural killer (NK) cells, CD4+, and CD8+ T cells.

CDs show a broad absorption and tunable emission spectrum that can be exploited in a FRET (Forster Resonance Energy Transfer) system. CDs serve as energy donors to photosensitizers via FRET in some systems, transferring light energy to excite the photosensitizer, producing ROS generation. Wu et al82 introduced a novel CQDs@PtPor system through electrostatic interactions between positively charged tetraplatinated porphyrin photosensitizer complex (PtPor) and negatively charged carbon quantum dots (CQDs). The FRET process allowed CQDs to transfer light energy to PtPor, resulting in enhanced 1O2 production compared to pure PtPor. This system demonstrated improved PDT efficacy under xenon lamp irradiation, showcasing the potential of CQDs as effective nanocarriers for PSs and enhancing cytotoxicity effect (see Figure 3). In a similar study, magnesium and nitrogen-doped carbon dots (Mg, N-doped CDs) were synthesized as a nanocarrier for Ce6, significantly enhancing FRET efficiency due to their high fluorescence QY and proximity to the PS. The resulting Mg, N-doped CDs-Ce6 system demonstrated improved therapeutic efficacy in both in vitro and in vivo models.83

Figure 3 CQDs@Pt: (A) Schematic illustration of synthesis of CQDs@Pt, (B) TEM images and corresponding size distribution histograms and (C) FT-IR spectra of CQDs@Pt, (D) Cell viability of CQDs@Pt in HeLa cells after treatment, (E) Singlet oxygen of CQDs@PtPor, (F) confocal fluorescence microscopy images of HeLa cells under 405 nm excitation after treatment with CQDs, PtPor and CQDs@PtPor. And (G) PL Spectra of CQDs@Pt. Reproduced from Wu F, Yue L, Su H, et al. Carbon Dots @ platinum porphyrin composite as theranostic nanoagent for efficient photodynamic cancer therapy. Nanoscale Res Lett. 13. © The Author(s). 2019, corrected publication January 2019 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License.82

A pioneer strategy to enhance the water solubility of hydrophobic PSs is targeting the surface of carbon dots. For instance, riboflavin has been targeted on CDs surface. Riboflavin, having a diol group, was covalently linked with CD, promoting the killing of cancer cells of B1610 murine melanoma cells and 4T1 breast cancer cells in the presence of laser irradiation (365 nm, 60 mW/cm2).84,85 Similarly, Choi et al86 developed folic acid-modified carbon dots loaded with zinc phthalocyanine (ZnPc), enhancing water solubility and tumor-targeting ability. The CD-PEG-FA/ZnPc nanoparticles produced 1O2 effectively under 660 nm laser irradiation (0.3 W/cm2, 20 min), demonstrating significant therapeutic effects in vitro against A549 human lung cancer cells. The conjugation of FA to CD significantly improved the active tumor-targeting capability of the ZnPc delivery vehicle to folate receptor-overexpressing tumors.

Doped CDs are increasingly recognized for their dual role as photosensitizers and nanocarriers in PDT. By incorporating heteroatoms such as nitrogen, phosphorus, sulfur, and metals like copper and tin, the properties of CDs can be fine-tuned to enhance their ability to generate ROS under light irradiation. These doped CDs offer improved photodynamic effects by increasing 1O2 production, light absorption, and biocompatibility, making them powerful agents in cancer therapy.

In a study, Zhao et al35 synthesized red-emitting nitrogen and phosphorus co-doped carbon dots (R-CDs) using phosphoric acid and o-phenylenediamine through a hydrothermal method. R-CDs could generate 1O2 and have an absolute fluorescence QY of 15% in water. In vitro studies demonstrated that R-CDs effectively inhibited the proliferation of A549 lung cancer cells under 532 nm light irradiation (100 mW/cm2), showing promise as a standalone PDT agent. Furthermore, in vivo experiments on tumor-bearing mice indicated significant tumor reduction, confirming R-CDs’ potential as a PS and effective cancer treatment. Similarly, Jia et al87 prepared self-assembled carbon dot nanospheres (CDNS) that produced 1O2 under 671-nm laser irradiation with a QY of 45%. The CDNS showed superior cytotoxic effects against 4T1 cancer cells compared to conventional CDs. In vivo fluorescence imaging indicated effective accumulation at tumor sites, significantly reducing tumor volume after treatment. These studies highlight the potential of surface-modified CDs as efficient photosensitizers capable of generating ROS while also providing imaging capabilities (Table 3).

Table 3 Photodynamic and Photothermal Therapy Applications of Surface-Modified Carbon Dots

Metal-doped CDs can generate ROS depending on the dose, metal speciation, and exposure route. For instance, J. Wang et al27 developed copper-doped carbon dots (Cu-CDs) using acrylic acid and copper nitrate as precursors. The Cu-CDs exhibited a 1O2 QY of 36% under LED light irradiation (400–700 nm). In vitro studies revealed that Cu-CDs effectively induced apoptosis in HeLa cancer cells through ROS generation. In vivo experiments demonstrated substantial tumor growth inhibition in mice treated with Cu-CDs, indicating their potential as effective photosensitizers for PDT. In a similar study, Sun et al33 investigated nitrogen-doped carbon dots for their application in oral squamous cell carcinoma (OSCC). The CDs demonstrated significant ROS production under 660 nm laser irradiation, leading to effective apoptosis through mitochondrial pathways. Both in vitro and in vivo studies confirmed that these nitrogen-doped CDs promoted DNA damage and inhibited tumor growth. Hu et al39 also synthesized tin-doped carbon dots (Sn@CDs) that exhibited a high 1O2 quantum yield of 58.3% with low cytotoxicity toward healthy cells. In vitro studies confirmed that Sn@CDs effectively reduced the viability of 4T1 cancer cells upon irradiation with LED light (490 nm, 40 mW/cm2). The results from both in vitro and in vivo experiments indicated significant tumor reduction of HepG2 liver cancer, demonstrating the promise of Sn@CDs as effective photosensitizers for PDT applications.

Heteroatoms doped CDs, such as N-CDs, Se-N-CDs, and S, N-CDs, have been developed as PS for PDT in cancer therapy.40,42 Li et al90 investigated nitrogen-doped CDs (NCDs) synthesized via solvothermal transformation of coal resources, achieving a 1O2 QY of 19%. Their findings suggest that NCDs possess significant potential as photosensitizers for PDT due to their ability to generate ROS effectively. However, the combination of sulfur and nitrogen-doped CDs (S, N-CDs) can enhance the generation of ROS in hypoxic tumor environments. Bai et al34 developed sulfur and nitrogen co-doped carbon dots (S, N-CDs) using a hydrothermal method, which showed improved therapeutic efficiency compared to single doping methods. These multifunctional heteroatom-doped CDs produced 1O2 with a QY of 27% and demonstrated effective photothermal conversion capabilities. In a similar study, Xu et al42 designed selenium- and nitrogen-doped carbon dots (Se/N-CDs) as PSs and used RNA as a carrier to enhance cellular uptake near the nucleus during laser treatment. However, this study reported lower  1O2 production (10.6%) compared to other PSs, indicating challenges in their efficiency relative to established agents.

These findings underscore the potential of doped carbon dots as versatile photosensitizers capable of generating ROS independently under light irradiation, offering promising avenues for improving photodynamic therapy outcomes across various cancer types. Many doped CDs, such as Sn@CDs and Cu-CDs, exhibit low cytotoxicity towards healthy cells while maintaining strong ROS-generating capabilities against cancer cells. This selective cytotoxicity is crucial for minimizing side effects in clinical applications.

Integrating organic PSs as precursors in synthesizing CDs has garnered significant attention in PDT. While PSs such as diketopyrrolopyrrole (DPP), 5,10,15,20-tetraphenylporphyrin (TPP), and phthalocyanine exhibit excellent photostability and high 1O2 QY, their poor water solubility often leads to aggregation in aqueous environments, which diminishes their efficacy in generating 1O2. PSs, as a carbon source, can develop multifunctional CDs with improved ROS generation, quantum yields, and water solubility. He et al22 synthesized blue-emitting diketopyrrolopyrrole-based CDs (DPP-CDs) using DPP and chitosan as precursors. This combination improved the water solubility and biocompatibility of the resulting CDs and retained their ability to produce 1O2 with a QY of 27.6%. In vitro experiments demonstrated that DPP-CDs effectively killed HepG2 cancer cells under 540-nm laser irradiation (15 mW/cm2 for 10 min). Additionally, in vivo studies on tumor-bearing mice showed significant inhibition of tumor growth, confirming the therapeutic potential of DPP-CDs as effective PSs. In similar research, Y. Li et al23 developed porphyrin-based carbon dots (TPP-CDs) through a solvothermal method using TPP and chitosan as precursors. The TPP-CDs exhibited strong blue fluorescence and improved water solubility due to the abundant hydroxyl groups on their surface. TPP-CDs demonstrated superior 1O2 generation to pure TPP, indicating that the CDs enhanced the PS’s performance in PDT applications. These findings suggest that TPP-CDs can be effective therapeutic agents against H22 hepatocarcinoma cells under 625 nm light irradiation (60 mW/cm2 for 1 h).

The CDs synthesized with PSs as a precursor could exhibit hydrophobic qualities. To improve the solubility of PSs, Wen et al24 synthesized red-emitting CDs using pheophytin as a carbon source through a microwave-assisted hydrothermal method. These CDs were then assembled with DSPE-PEG2000 to enhance hydrophilicity through hydrophobic-hydrophobic interactions. The resulting CDs demonstrated a high 1O2 QY of 62% under 671-nm laser irradiation (0.1 W/cm2 for 10 min), indicating strong ROS generation capacity. The CDs effectively killed 4T1 breast cancer cells, highlighting the advantages of NIR-induced PDT. In vivo studies on tumor-bearing mice showed efficient accumulation of these CDs at tumor sites, enabling deep tissue penetration and effective PDT. Tumor growth was significantly inhibited, further confirming the efficacy of PS-derived CDs. The combination maintained efficient 1O2 generation while exhibiting strong NIR fluorescence properties, making them suitable for deep tissue PDT applications.

Hydrophobic photosensitizers, such as porphyrins and phthalocyanines, frequently experience low water solubility, which restricts their usage in biological environments. The utilization of these molecules as precursors for synthesizing CDs enhances solubility and expands their applicability in aqueous biological systems. Although PS-derived CDs demonstrate high singlet oxygen quantum yields, the precise quantum yield may differ depending on the synthesis process and the type of the photosensitizer employed.

Surface Modification-CDs in PTT

PTT is a promising cancer treatment modality that leverages the ability of PTAs to convert light energy into heat, effectively destroying cancer cells. CDs have gained attention for their dual role as nanocarriers and PTAs in PTT, particularly when modified to enhance their optical properties and therapeutic efficacy. Choi et al45 developed pH/redox-responsive CDs loaded with indocyanine green IR825 through π-π stacking and hydrophobic interactions. In this complex, the fluorescence of IR825 was quenched by the CDs. Upon reaching the acidic tumor microenvironment, the imine and disulfide bonds were cleaved, leading to the release of IR825 and activation of fluorescence. Under 808-nm laser irradiation, the released IR825 demonstrated effective photothermal effects, indicating the potential for controlled release in mice bearing MDA-MB-231 breast cancer tumors and enhanced therapeutic outcomes. In a similar study, polydopamine-folate carbon dots (PFCDs) were developed as theranostic nanocarriers for