Efficient detection of nitric oxide a biomarker associated with COVID19 via N, P co-doped C60 fullerene: a computational study.

CONTEXT: Coronavirus (COVID-19) is a novel respiratory viral infection, causing a relatively large number of deaths especially in people who underly lung diseases such as chronic obstructive pulmonary and asthma, and humans are still suffering from the limited testing capacity. In this article, a solution is proposed for the detection of COVID-19 viral infections through the analysis of exhaled breath gasses, i.e., nitric oxide, a prominent biomarker released by respiratory epithelial, as a non-invasive and time-saving approach. Here, we designed a novel and low-cost N and P co-doped C60 fullerene-based breathalyzer for the detection of NO gas exhaled from the respiratory epithelial cells. This breathalyzer shows a quick response to the detection of NO gas by directly converting NO to NO2 without passing any energy barrier (0 kcal/mol activation energy). The recovery time of breathalyzer is very short (0.98 × 103 s), whereas it is highly selective for NO sensing in the mixture of CO2 and H2O gasses. The study provides an idea for the synthesis of low-cost (compared to previously reported Au atom decorated nanostructure and metal-based breathalyzer), efficient, and highly selective N and P co-doped C60 fullerene-based breathalyzer for COVID-19 detection. METHODS: The geometries of N and P-doped systems and gas molecules are simulated using spin-polarized density functional theory calculations.

Integrating Explicit and Implicit Fullerene Models into UNRES Force Field for Protein Interaction Studies.

Fullerenes, particularly C60, exhibit unique properties that make them promising candidates for various applications, including drug delivery and nanomedicine. However, their interactions with biomolecules, especially proteins, remain not fully understood. This study implements both explicit and implicit C60 models into the UNRES coarse-grained force field, enabling the investigation of fullerene-protein interactions without the need for restraints to stabilize protein structures. The UNRES force field offers computational efficiency, allowing for longer timescale simulations while maintaining accuracy. Five model proteins were studied: FK506 binding protein, HIV-1 protease, intestinal fatty acid binding protein, PCB-binding protein, and hen egg-white lysozyme. Molecular dynamics simulations were performed with and without C60 to assess protein stability and investigate the impact of fullerene interactions. Analysis of contact probabilities reveals distinct interaction patterns for each protein. FK506 binding protein (1FKF) shows specific binding sites, while intestinal fatty acid binding protein (1ICN) and uteroglobin (1UTR) exhibit more generalized interactions. The explicit C60 model shows good agreement with all-atom simulations in predicting protein flexibility, the position of C60 in the binding pocket, and the estimation of effective binding energies. The integration of explicit and implicit C60 models into the UNRES force field, coupled with recent advances in coarse-grained modeling and multiscale approaches, provides a powerful framework for investigating protein-nanoparticle interactions at biologically relevant scales without the need to use restraints stabilizing the protein, thus allowing for large conformational changes to occur. These computational tools, in synergy with experimental techniques, can aid in understanding the mechanisms and consequences of nanoparticle-biomolecule interactions, guiding the design of nanomaterials for biomedical applications.

Research progress of fullerenes and their derivatives in the field of PDT.

In contemporary studies, the predominant utilization of C60 derivatives pertains to their role as photosensitizers or agents that scavenge free radicals. The intriguing coexistence of these divergent functionalities has prompted extensive investigation into water-soluble fullerenes. The photodynamic properties of these compounds find practical applications in DNA cleavage, antitumor interventions, and antibacterial endeavors. Consequently, photodynamic therapy is progressively emerging as a pivotal therapeutic modality within the biomedical domain, owing to its notable levels of safety and efficacy. The essential components of photodynamic therapy encompass light of the suitable wavelength, oxygen, and a photosensitizer, wherein the reactive oxygen species generated by the photosensitizer play a pivotal role in the therapeutic mechanism. The remarkable ability of fullerenes to generate singlet oxygen has garnered significant attention from scholars worldwide. Nevertheless, the limited permeability of fullerenes across cell membranes owing to their low water solubility necessitates their modification to enhance their efficacy and utilization. This paper reviews the applications of fullerene derivatives as photosensitizers in antitumor and antibacterial fields for the recent years.

A novel fullerene composite material for directional oil control and antioxidant.

BACKGROUND: There are very few cosmetic ingredients that can target oil control and extend the wear time. Fullerenes have been reported to have excellent antioxidant capacity and a variety of biological activities, such as quenching free radicals, inhibiting lipid peroxidation, and promoting lipid flocculation. OBJECTIVE: The purpose of applying foundation makeup on the face is to make the skin color even, but the secretion and oxidation of skin oil will make the makeup mottled and dull. In order to solve this problem, a fullerene composite material that can directionally absorb oil and resist oil oxidation has been developed. METHODS: Fullerenes and hydroxyapatite composite was prepared by high pressure homogenization under alkaline condition. The indicated morphology and structure were characterized by SEM, UV-Vis, Raman, and XRD. The oil absorption capacity was determined by adding the C60-hydroxyapatite composite to a mixed solution of hexane and oil, shaking for 1 h, filtering, analyzed by GC-MS, and calculating the oil absorption by external standard method. Artificial sebum was prepared by adding different mass of water and oleic acid to screen the optimum ratio. C60-hydroxyapatite mixture and C60-hydroxyapatite composite were added to the artificial sebum to test the oil-absorbing capacity of the materials. The hydroxyl radical scavenging ability of C60-hydroxyapatite composite containing different fullerene contents was measured by X-band ESR spectroscopy, and the long-term radical scavenging ability of the composites was tested in comparison with VC. Antioxidant experiment is adding C60-hydroxyapatite composite material, and hydroxyapatite to oleic acid, then the UV light irradiation is aimed to accelerate the oxidation of oleic acid. Oleic acid act as a control group, and make the detection of oleic acid peroxide value after 7 days. The safety of the materials was tested by using culture media to soak the C60-hydroxyapatite composite for 24 h and then used to culture cells. RESULTS: The characterization of SEM, UV-Vis, Raman, and XRD showed that fullerene clusters were dispersed on the surface of hydroxyapatite stably, and they formed a stable composite. The adsorption rates of C60-hydroxyapatite composites for oleic acid, phenyl trimethicone, caprylic capric glyceride, isooctyl palmitate, mineral oil, olive oil, and dimethicone were 60.5%, 9.3%, 9.15%, 5.24%, 2.94%, 1.01%, and 0%, respectively. The flocculation amount of artificial sebum was 5.9 g per gram of C60-hydroxyapatite mixture and 24.2 g per gram of C60-hydroxyapatite composite. C60-hydroxyapatite composites have excellent quenching ability for hydroxyl radicals. When the fullerene content is 1, 2, 3, and 4 mg/kg, the quenching rates are 25.02%, 39.57%, 49.75%, and 62.24%, respectively. The quenching effect was enhanced with the increase of fullerene content, and it had strong long-term antioxidant properties. It can also be proved that C60-hydroxyapatite composites have strong antioxidant capacity through antioxidant experiments. The peroxide value of oleic acid on Day 0 was 2.8, and after 7 days of UV irradiation, the peroxide values of blank control, hydroxyapatite group, C60-hydroxyapatite composite containing 0.5% and 1% fullerenes four groups of materials were 8.02 meq O2/kg, 7.98 meq O2/kg, 7.11 meq O2/kg, and 6.87 meq O2/kg, respectively. The cell activity was 20.94% and 99.2% after the cells were cultured for 24 h using C60-hydroxyapatite composite and hydroxyapatite extracts, respectively, and the addition of fullerene was able to significantly increase the cell activity. CONCLUSION: Fullerene hydroxyapatite complex has excellent directional oil absorption characteristics, which can effectively remove free radicals and reduce skin oil oxidation.

Host-Guest Mechanism via Induced Fit Fullerene Complexation in Porphin Receptor to Probe Salivary Alpha-Amylase in Dental Caries for Clinical Applications.

Salivary α-amylase is the most abundant protein of human saliva that potentially binds to streptococcus and other bacteria via specific surface-exposed α-amylase-binding proteins and plays a significant role in caries development. The detection of α-amylase in saliva can be used as a bioindicator of caries development. Herein, a facile strategy has been applied, tailoring the photochemical properties of 5,10,15,20-tetrakis(4-hydroxyphenyl)-21H,23H-porphine (TPPOH) and the fullerene C60 complex. The fluorescence emission of TPPOH is quenched by starch-coated fullerene C60 via charge-transfer effects, as determined by UV absorption and fluorescence spectroscopic studies. The starch-coated C60 has been thoroughly characterized via Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), field emission scanning electron microscopy (FE-SEM), optical microscopy, thermal gravimetric analysis (TGA), static water contact angle measurements, and zeta potential measurements. The analytical response of the assay showed a linear fluorescent response in α-amylase concentrations ranging from 0.001-0.1 Units/mL, with an LOD of 0.001 Units/mL. The applicability of the method was tested using artificial saliva with quantitative recoveries in the range 95-100%. The practicability of the procedure was verified by inspecting saliva samples of real clinical samples covering all age groups. We believe that the proposed method can serve as an alternative analytical method for caries detection and risk assessment that would also minimize the cost of professional preventive measures and treatments.

Molecular modelling of fullerene C60 functionalized by nitric oxide for use in biological environment.

The unique potential of fullerene C60 for various biological applications has ignited significant interest. However, its inherent non-polarity poses a critical challenge for its effective integration within biological systems. This study delves into the intricate physicochemical characteristics of the innovative [C60 + NO] complex using density functional theory and time-dependent density functional theory. The computational analyses encompass molecular charge, surface electrostatic potential, and dipole moment evaluations. Impressively, the dipole moment of the [C60 + NO] complex significantly increases to 12.92 D. Meticulous surface analysis reveals a subtle interplay between molecular structures, indicating weak interactions. The analysis of the absorption spectrum unveils a noteworthy red-shift of 200 nm subsequent to complex formation. To elucidate the electron transfer mechanisms, we explore photo-induced electron transfer through CAM-B3LYP. This exploration elucidates intricate pathways governing electron transfer, with complementary insights gleaned from Marcus theory's outputs, especially the Gibbs free energy of electron transfer. Changes in the physicochemical properties of approaching C60 and NO molecules reveal interesting results compared to separate molecules. These findings resonate profoundly in the context of potential biological and pharmaceutical utilization. With implications for the biomedical area, the outcomes linked to the [C60 + NO] complex kindle optimism for pioneering biomedical applications.

C60-based Multivalent Glycoporphyrins Inhibit SARS-CoV-2 Specific Interaction with the DC-SIGN Transmembrane Receptor.

Since WHO has declared the COVID-19 outbreak a global pandemic, nearly seven million deaths have been reported. This efficient spread of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is facilitated by the ability of the spike glycoprotein to bind multiple cell membrane receptors. Although ACE2 is identified as the main receptor for SARS-CoV-2, other receptors could play a role in viral entry. Among others, C-type lectins such as DC-SIGN are identified as efficient trans-receptor for SARS-CoV-2 infection, so the use of glycomimetics to inhibit the infection through the DC-SIGN blockade is an encouraging approach. In this regard, multivalent nanostructures based on glycosylated [60]fullerenes linked to a central porphyrin scaffold have been designed and tested against DC-SIGN-mediated SARS-CoV-2 infection. First results show an outstanding inhibition of the trans-infection up to 90%. In addition, a deeper understanding of nanostructure-receptor binding is achieved through microscopy techniques, high-resolution NMR experiments, Quartz Crystal Microbalance experiments, and molecular dynamic simulations.

Water-soluble endohedral metallofullerenes: new horizons for biomedical applications.

Endohedral metallofullerenes (EMFs) offer a safe avenue to manipulate metals important to biomedical applications such as MRI contrast, X-ray contrast, radiolabeling, radiotherapy, chemotherapy, and the control of inflammation by scavenging reactive oxygen species (ROS). Moreover, functionalizing the double bonds on the surface of EMFs modifies their solubility, supramolecular behaviour, binding, targeting characteristics, and physical properties. While most existing water-soluble derivatives possess a statistical mixture of appended functional groups, progress has been made in creating molecularly-precise derivatives with a defined number of surface functional groups, leading to potentially more nuanced control of their behaviour and properties. Further elucidation of the structure-function relationships of these materials is expected to enhance their utility in biomedical applications and possibly broaden their use in diverse areas of science and technology.

Investigation of Magnetic Electrodes in Conducting Polymeric Materials: Electrochemical Properties of a Fullerene[C60 ]-Pd Polymer and Iron Oxide Magnetic Nanocomposite.

A composite of iron oxide magnetic nanoparticles and coordination fullerene polymer (C60 Pd3 )n is formed by chemical deposition of spherical polymer nanoparticles on iron oxide magnetic nanoparticles in benzene containing C60 and Pd(0) complex. The composition of the composite can be controlled by the amount of magnetite and concentration of polymerization precursors as well as the time of polymerization. The magnetic composite material Fe3 O4 -γFe2 O3 /(C60 Pd3 )n is used as a model system to investigate its deposition on a magnetic electrode and its electrochemical properties. The iron oxide magnetic nanoparticles ensure both the magnetic activity of the composite and its nanostructured morphology. Both of these factors are responsible for the enhancement of the electrochemical activity of the polymer phase forming the composite in comparison to the pure polymer material deposited on the same magnetic electrode. In the magnetic field of the electrode, the composite undergoes permanent and strong bonding with the surface of the electrode. The nanostructured morphology of the Fe3 O4 -γFe2 O3 /(C60 Pd3 )n composite also provides very good capacitive properties.

Water-Soluble Fullerene Monoderivatives for Biomedical Applications.

Monoderivatives of fullerenes functionalized with hydrophilic groups make them water soluble, while preserving the hydrophobic fullerene cage. This class of molecules have intriguing biomedical applications, including drug delivery, photodynamic therapy (PDT), antiviral and antimicrobial activity and reactive oxygen species (ROS)-scavenging abilities. In this Concept we discuss the synthesis and biomedical applications of water-soluble fullerene monoderivatives and their biological behavior based on their structures.

Potential Medical Use of Fullerenols After Two Decades of Oncology Research.

Fullerenes are carbon molecules that are found in nature in various forms. They are composed of hexagonal and pentagonal rings that create closed structures. Almost 4 decades ago, fullerenes were identified in the form of C60 and C70, and following the award of the Nobel Prize in Chemistry for this discovery in 1996, many laboratories started working on their water-soluble derivatives that could be used in different industries, including pharmaceutical industries. One of the first fullerene forms that was the focus of different research groups was fullerenol, C60(OH)n (n = 2-44). Both in-vitro and in-vivo studies have shown that polyhydroxylate fullerene derivatives can potentially be used as either antioxidative agents or cytostatics (depending on their co-administration, forms, and concentration/dose) in biological systems. The current review aimed to present a critical view of the potential applications and limitations of fullerenols in oncology, as understood from the past 2 decades of research.

In vitro and in silico studies of the antiviral activity of polyhydrated fullerenes against influenza A (H1N1) virus.

As of today, influenza viruses remain a relevant target for the development of antiviral compounds due to their rapid evolution and acquisition of the resistance to existing drugs. Fullerene derivatives have already shown the ability to successfully interact with viruses, and polyhydrated fullerenes (or fullerenols) are particularly attractive due to their compatibility with biological fluids and low toxicity. Therefore, the goal of this work was to study the effect of two batches of a mixture of polyhydrated fullerenes with a mass ratio of 78.1% C60/C70 and 21.9% C76/C78/C84 on the influenza A (H1N1) virus. It was determined that the mixture of fullerenols, along with the low toxicity, showed high antiviral activity with a decrease in the viral infectious titer up to 4 orders of magnitude. In addition, studied fullerenols did not affect the hemagglutination process and did not show any significant prophylactic activity. With the help of molecular docking and molecular dynamics simulation, the likely target of fullerenols' action was determined-the binding site of the RNA primer of the viral RNA-dependent RNA polymerase. Therefore, we assume that the high antiviral effect of polyhydrated fullerenes on influenza A virus is related to their interaction with the viral RNA polymerase.

C60 adduct with L-arginine as a promising nanomaterial for treating cerebral ischemic stroke.

The work aimed to investigate the biocompatibility and biological activity of the water-soluble fullerene adduct C60-Arg. It was found that the material is haemocompatible, is not cyto- and genotoxic, possesses pronounced antioxidant activity. Additionally, this paper outlines the direction of application of water-soluble fullerene adducts in the creation of neuroprotectors. It has been suggested that a putative mechanism of the protective action of the C60-Arg adduct is associated with its antioxidant properties, the ability to penetrate the blood-brain barrier, and release nitrogen monoxide as a result of the catabolism of L-arginine residues, which promote vascular relaxation. The action of the C60-Arg adduct was compared with the action of such an antioxidant as Edaravone, which is approved in Japan for the treatment of ischemic and haemorrhagic strokes.

(Ro)vibrational Spectroscopic Constants, Lifetime and QTAIM Evaluation of Fullerene Dimers Stability.

The iconic caged shape of fullerenes gives rise to a series of unique chemical and physical properties; hence a deeper understanding of the attractive and repulsive forces between two buckyballs can bring detrimental information about the structural stability of such complexes, providing significant data applicable for several studies. The potential energy curves for the interaction of multiple van der Waals buckyball complexes with increasing mass were theoretically obtained within the DFT framework at ωB97xD/6-31G(d) compound model. These potential energy curves were employed to estimate the spectroscopic constants and the lifetime of the fullerene complexes with the Discrete Variable Representation and with the Dunham approaches. It was revealed that both methods are compatible in determining the rovibrational structure of the dimers and that they are genuinely stable, i.e., long-lived complexes. To further inquire into the nature of such interaction, Bader's QTAIM approach was applied. QTAIM descriptors indicate that the interactions of these closed-shell systems are dominated by weak van der Waals forces. This non-covalent interaction character was confirmed by the RDG analysis scheme. Indirectly, QTAIM also allowed us to confirm the stability of the non-covalent bonded fullerene dimers. Our lifetime calculations have shown that the studied dimers are stable for more than 1 ps, which increases accordingly with the number of carbon atoms.

Interactions between modified fullerenes and proteins in cancer nanotechnology.

Fullerenes have numerous properties that fill the gap between small molecules and nanomaterials. Several types of chemical reaction allow their surface to be ornamented with functional groups designed to change them into 'ideal' nanodelivery systems. Improved stability, and bioavailability are important, but chemical modifications can render them practically soluble in water. 'Buckyball' fullerene scaffolds can interact with many biological targets and inhibit several proteins essential for tumorigeneses. Herein, we focus on the inhibitory properties of fullerene nanomaterials against essential proteins in cancer nanotechnology, as well as the use of dedicated proteins to improve the bioavailability of these promising nanomaterials.

Trichalcogenasupersumanenes and its concave-convex supramolecular assembly with fullerenes.

Synthesis of buckybowls have stayed highly challenging due to the large structural strain caused by curved π surface. In this paper, we report the synthesis and properties of two trichalcogenasupersumanenes which three chalcogen (sulfur or selenium) atoms and three methylene groups bridge at the bay regions of hexa-peri-hexabenzocoronene. These trichalcogenasupersumanenes are synthesized quickly in three steps using an Aldol cyclotrimerization, a Scholl oxidative cyclization, and a Stille type reaction. X-ray crystallography analysis reveals that they encompass bowl diameters of 11.06 Å and 11.35 Å and bowl depths of 2.29 Å and 2.16 Å for the trithiasupersumanene and triselenosupersumanene, respectively. Furthermore, trithiasupersumanene derivative with methyl chains can form host-guest complexes with C60 or C70, which are driven by concave-convex π ⋯ π interactions and multiple C-H ⋯ π interactions between bowl and fullerenes.

Metalloporphyrin reduced C70 fullerenes as adsorbents and detectors of ethenone; A DFT, NBO, and TD-DFT study.

In the present work, the structure and electronic properties of Ti-, Cr-, Fe-, Ni-, Zn-, and Cu-inserted in porphyrin-reduced C70 fullerenes (TM-PIC70Fs) and their interactions with the ethenone were studied using DFT, NBO, and TD-DFT at CAM-B3LYP/6-31G(d) level of theory. 2.89-3.83 and 4.02-4.56 eV were obtained for the HOMO-LUMO gap energies and work functions of TM-PIC70Fs, respectively, compared with 3.76 and 4.54 eV for PIC70F. Among considered TM-PIC70Fs, the adsorption of the ethenone on Ti-PIC70F appreciably changed the HOMO-LUMO energy gap and work function. Consequently, Ti-PIC70F may be used as the ethenone's electronic conductivity and work function types sensor. According to calculated UV-visible spectra, the ethenone adsorption may change the color of Fe- and Ti-PIC70Fs. Therefore, they can be used as color-changing sensors of ethenone. In addition, Ti-, Cr-, Fe-, and Zn-PIC70Fs can be employed as suitable adsorbents of ethenone. Among proper sensors and adsorbents of ethenone, Cr-, Fe-, and Zn-PIC70Fs may be recovered and reused.

Improvement in photodynamic activity by a porphyrin-fullerene composite system in lipid membranes.

Porphyrin-fullerene composite systems are attracting great attention as photodynamic agents; however, water-soluble derivatives are still scarce. Herein, we prepared noncovalently a lipid membrane-incorporated porphyrin-fullerene composite system with relative stability in aqueous solution. As in the case of porphyrin-fullerene composite systems in nonpolar solvents, efficient formation of singlet oxygen occurred via photoinduced energy transfer between porphyrin and fullerene as the predominant pathway in the photodynamic activity under the hydrophobic conditions of the lipid membranes, resulting in enhanced photodynamic activity toward Colon26 and HeLa cells compared with the individual porphyrin and fullerene components. Furthermore, the porphyrin-fullerene composite system exhibited high selectivity toward HeLa cells over normal mouse fibroblast L929 cells.

Simultaneous Ring Contraction and Expansion Reaction: Electrosynthesis of Heterocycle-Fused Fulleroids and Photovoltaic Application.

Simultaneous electrochemical ring contraction and expansion reactions remain unexplored to date. Herein, the reductive electrosynthesis of heterocycle-fused fulleroids from fullerotetrahydropyridazines and electrophiles in the presence of a trace amount of oxygen has been achieved with concurrent ring contraction and ring expansion. When trifluoroacetic acid and alkyl bromides are employed as electrophiles, heterocycle-fused fulleroids with a 1,1,2,6-configuration are regioselectively formed. In contrast, heterocycle-fused fulleroids with a 1,1,4,6-configuration are regioselectively produced as two separable stereoisomers if phthaloyl chloride is used as the electrophile. The reaction proceeds through multiple steps of electroreduction, heterocycle ring-opening, oxygen oxidation, heterocycle contraction, fullerene cage expansion, and nucleophilic addition. The structures of these fulleroids have been determined by spectroscopic data and single-crystal X-ray diffraction analyses. The observed high regioselectivities have been rationalized by theoretical calculations. Representative fulleroids have been applied in organic solar cells as the third component and exhibit good performance.

Identification and Biological Evaluation of a Water-Soluble Fullerene Nanomaterial as BTK Kinase Inhibitor.

Introduction: Thanks to recent advances in synthetic methodology, water-soluble fullerene nanomaterials that interfere with biomolecules, especially DNA/RNA and selected proteins, have been found with tremendous potential for applications in nanomedicine. Herein, we describe the synthesis and evaluation of a water-soluble glycine-derived [60]fullerene hexakisadduct (HDGF) with T h symmetry, which is a first-in-class BTK protein inhibitor. Methods: We synthesized and characterized glycine derived [60]fullerene using NMR, ESI-MS, and ATR-FT-IR. DLS and zeta potential were measured and high-resolution transmission electron microscopy (HRTEM) observations were performed. The chemical composition of the water-soluble fullerene nanomaterial was examined by X-ray photoelectron spectrometry. To observe aggregate formation, the cryo-TEM analysis was carried out. The docking studies and molecular dynamic simulations were performed to determine interactions between HDGF and BTK. The in vitro cytotoxicity was evaluated on RAJI and K562 blood cancer cell lines. Subsequently, we examined the induction of cell death by autophagy and apoptosis by determining the expression levels of crucial genes and caspases. We investigated the direct association of HDGF on inhibition of the BTK signalling pathway by examining changes in the calcium levels in RAJI cells after treatment. The inhibitory potential of HDGF against non-receptor tyrosine kinases was evaluated. Finally, we assessed the effects of HDGF and ibrutinib on the expression of the BTK protein and downstream signal transduction in RAJI cells following anti-IgM stimulation. Results: Computational studies revealed that the inhibitory activity of the obtained [60]fullerene derivative is multifaceted: it hampers the BTK active site, interacting directly with the catalytic residues, rendering it inaccessible to phosphorylation, and binds to residues that form the ATP binding pocket. The anticancer activity of produced carbon nanomaterial revealed that it inhibited the BTK protein and its downstream pathways, including PLC and Akt proteins, at the cellular level. The mechanistic studies suggested the formation of autophagosomes (increased gene expression of LC3 and p62) and two caspases (caspase-3 and -9) were responsible for the activation and progression of apoptosis. Conclusion: These data illustrate the potential of fullerene-based BTK protein inhibitors as nanotherapeutics for blood cancer and provide helpful information to support the future development of fullerene nanomaterials as a novel class of enzyme inhibitors.