CN117503928A - Polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of medicine for targeted treatment of renal clear cell carcinoma - Google Patents

Polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of medicine for targeted treatment of renal clear cell carcinoma Download PDF

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CN117503928A
CN117503928A CN202311559630.8A CN202311559630A CN117503928A CN 117503928 A CN117503928 A CN 117503928A CN 202311559630 A CN202311559630 A CN 202311559630A CN 117503928 A CN117503928 A CN 117503928A
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heterostructure
red phosphorus
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徐岩
管陈
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Affiliated Hospital of University of Qingdao
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    • A—HUMAN NECESSITIES
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Abstract

The invention provides a polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of a drug for targeted treatment of renal clear cell carcinoma, and belongs to the technical field of nano-drugs. The invention designs a near infrared light response Hittorf red phosphorus (HP) modified polymer carbon nitride heterostructure (PCN@HP) based on a simple Chemical Vapor Deposition (CVD) method. Experimental studies have found that HP nanorods grown vertically on PCN exhibit broad and strong light absorption in the near infrared range. In vitro and in vivo experiments prove that under the irradiation of near infrared light, the tumor inhibition rate reaches 100 percent when the concentration of the suspension is 100 mug/mL. Notably, the complex has no damage to normal tissues and cells, but has a significant targeted inhibition effect on deep ccRCC cells in vivo. In addition, the composite material has good biocompatibility and self-luminous effect, can be completely metabolized in 48 hours in vivo, is a nanoparticle with imaging and PDT/PTT cooperative treatment, and can be used for efficient tumor nanotherapy.

Description

Polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of medicine for targeted treatment of renal clear cell carcinoma
Technical Field
The invention belongs to the technical field of nano-drugs, and particularly relates to a polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of a drug for targeted treatment of renal clear cell carcinoma.
Background
Renal Cell Carcinoma (RCCs) is one of the most common ten cancers, and is also the most common family of renal tumors. Among all its subtypes, clear cell carcinoma (ccRCC) is the most common sporadic type of renal cancer, accounting for about 75% of all cases, often associated with malignant disease progression and poor therapeutic outcome. At present, although some advanced strategies have been developed for the treatment of RCC, such as immune checkpoint blocking therapies and combination therapy regimens, treatment options for advanced disease have been limited due to the unavoidable toxicity and resistance of newly approved targeted drugs. Currently, surgical excision or ablation remains the treatment of choice for localized ccRCC, but clinical treatment is costly and patient prognosis is poor, thus urgent research into targeted therapies is needed.
The application of the nano technology provides an effective choice for improving the tumor treatment effect. Photothermal and photodynamic therapy has been increasingly used for the treatment of various cancers due to its inherent advantages of low toxicity, low side effects, no drug resistance and repeatable treatment. Although prior studies have reported the treatment of renal clear cell carcinoma based on photothermal and photodynamic therapy, clinical applications have been limited due to high cost of required materials, difficulty in synthesis, insufficient depth of treatment for solid tumors, unclear metabolic conditions, or certain toxicity of materials accumulated in vivo. However, the therapeutic situation of ccRCC is not optimistic, and the development of new and more effective treatments is urgent.
Disclosure of Invention
The invention provides a polymer carbon nitride @ Hittorf red phosphorus heterostructure and application thereof in preparing medicines for targeted therapy of renal transparent cell carcinoma, the heterostructure shows wide and strong light absorption in a near infrared range, has obvious targeted inhibition effect on deep ccRCC cells in vivo, has good biocompatibility and self-luminous effect, can be completely metabolized in 48 hours in vivo, is a nanoparticle with imaging and PDT/PTT cooperative therapy, and can be used for efficient tumor nanotherapy.
In order to achieve the aim, the invention provides application of a polymer carbon nitride @ Hittorf red phosphorus heterostructure in preparing medicines for treating renal clear cell carcinoma.
Preferably, the polymer carbon nitride@Hittorf red phosphorus heterostructure is a composite material prepared by a vapor deposition method, and the structural formula of the composite material is PCN@HP.
Preferably, the composite material is prepared by the following method:
heating melamine for 4 hours to 550 ℃ to prepare a polymer carbon nitride PCN;
grinding PCN and amorphous red phosphorus powder together in a mortar according to a ratio of 2:1;
thereafter, the obtained powder was transferred into a quartz ampoule and sealed with an oxygen-hydrogen flame under a low vacuum condition of-0.09 MPa;
the ampoule is heated to 550 ℃ in a furnace at 450 ℃ at a rate of 2 ℃/min and is kept for 4 hours, then is cooled to 280 ℃ at a rate of 1 ℃/min, when the temperature is reduced to 280 ℃/min, the cooling rate is reduced to 0.1 ℃/min, and after the ampoule is slowly cooled to room temperature at the rate, the capsule is crushed and rinsed by CS2, ethanol and distilled water respectively, and the PCN@HP heterostructure is obtained.
Preferably, in the heterostructure, the HP is anchored vertically on the PCN, with a length of 1-2 μm and a diameter of 40-200 nm.
Preferably, the PCN@HP suspension dose is more than 50% after 10 minutes of light irradiation at near infrared 808nm, and the tumor inhibition rate is 100% when the suspension concentration is 100 μg/mL.
Compared with the prior art, the invention has the advantages and positive effects that:
the invention designs a near infrared light response Hittorf red phosphorus (HP) modified polymer carbon nitride heterostructure (PCN@HP) based on a simple Chemical Vapor Deposition (CVD) method. Experimental studies have found that HP nanorods grown vertically on PCN exhibit broad and strong light absorption in the near infrared range. In vitro and in vivo experiments prove that under the irradiation of near infrared light, the tumor inhibition rate reaches 100 percent when the concentration of the suspension is 100 mug/mL. Notably, the complex has no damage to normal tissues and cells, but has a significant targeted inhibition effect on deep ccRCC cells in vivo. In addition, the composite material has good biocompatibility and self-luminous effect, can be completely metabolized in 48 hours in vivo, is a nanoparticle with imaging and PDT/PTT cooperative treatment, and can be used for efficient tumor nanotherapy.
Drawings
FIG. 1 is a schematic diagram of the PCN@HP synthesis step provided by the embodiment of the invention;
FIG. 2 is a representation of PCN@HP provided by an embodiment of the present invention;
effect of pcn@hp on ccRCC killing in fig. 3 (a): under the irradiation of near infrared light, when the dose of PCN@HP suspension is 60 mug/mL, the tumor inhibition rate of ccRCC reaches more than 50 percent, and when the concentration of the suspension is 100 mug/mL, the tumor inhibition rate reaches 100 percent; (b) PCN@HP has no damaging effect on normal tubular epithelial cells HK-2; (c) The conventional antitumor drugs have the same killing effect on ccRCC and HK-2 cells;
the left panel in fig. 4 is before pcn@hp is treated for ccRCC; the right image shows that the tumor volume is 1cm 3 After PCN@HP is injected into the ccRCC mice for 3 hours, the tumors are rapidly flattened and the volumes are rapidly reduced after the ccRCC mice are irradiated by near infrared light for 10 minutes, so that the ccRCC mice have good tumor cell killing effect;
FIG. 5 is a metabolic diagram of PCN@HP provided by an embodiment of the invention after injection;
FIG. 6 shows the change of biochemical detection index of blood and organ samples after injection of PCN@HP suspension according to an embodiment of the present invention, (a) Scr; (b) BUN; (c) ALT; (d) AST.
Detailed Description
The following description of the technical solutions in the embodiments of the present invention will be clear and complete, and it is obvious that the described embodiments are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
EXAMPLE 1 Synthesis of PCN@HP
As shown in fig. 1, the pcn@hp heterostructure composite material was prepared by a classical vapor deposition method (CVD), and the synthesis process was as follows:
the PCN preparation method comprises the following steps:
the melamine is prepared in a muffle furnace by heating at a speed of 2 ℃/min from room temperature for 4 hours to 550 ℃;
PCN and amorphous red phosphorus powder (commercial red phosphorus purchased from alpha corporation, purity 99.999%) were ground together in a mortar in a ratio of 2:1;
then, the obtained powder was transferred into a quartz ampoule and sealed with an oxygen-hydrogen flame under low vacuum conditions (-0.09 MPa, high purity Ar was charged before evacuation);
the ampoule is heated to 550 ℃ in a furnace at 450 ℃ at a speed of 2 ℃/min and is kept for 4 hours, then is cooled to 280 ℃ at a speed of 1 ℃/min, when the temperature is reduced to 280 ℃/min, the cooling speed is reduced to 0.1 ℃/min, and after the ampoule is slowly cooled to room temperature at the speed, the capsule is crushed and rinsed by CS2, ethanol and distilled water respectively, so that the PCN@HP heterostructure composite material is obtained.
Example 2 characterization of PCN@HP mixture
The morphology and structure of the prepared samples were determined using X-ray diffraction (xRD), uv-vis, raman, scanning electron microscopy (SEM, FEI Magellan 400), transmission electron microscopy (TEM, JEOL JEM-2100F). Wherein xRD is CuK alpha radiation 40kV,30mA; the Raman spectrum data acquisition excitation wavelength is 532nm; transmission electron microscope and scanning electron microscope) voltage was 200kV.
As shown in fig. 2a, the phase composition of pcn@hp was determined using xRD analysis. Pcn@hp exhibited two distinct PCN diffraction peaks at 13.3 ° and 27.7 °, and further exhibited diffraction peaks of HP at 15.4 ° and 34.2 °; consistent with XRD results, raman spectroscopic results (fig. 2 b) showed that pcn@hp simultaneously present characteristic peaks of PCN and HP; the ultraviolet visible spectrum (fig. 2 c) shows that pcn@hp exhibits strong light absorption in the visible and NIR regions. SEM (fig. 2 d) and TEM (fig. 2e, f) were used to evaluate morphology and surface features of the prepared samples. As shown, the HP is anchored vertically to the PCN, with a length of 1-2 μm and a diameter of tens of nanometers. In addition, the [110] crystal plane of HP is located in lattice fringes at a spacing of 0.275 nm.
Example 3 detection of the anti-tumor Effect of PCN@HP
Human tubular epithelial cells (HK-2) and human ccRCC cells (786-O) were from Shanghai cell biology institute of China academy of sciences, respectively. HK-2 cells were cultured in DMEM/F-12 medium containing 10% fetal bovine serum and 1% penicillin-streptomycin solution (100U/mL penicillin, 0.1mg/mL streptomycin), 786-O cells were cultured in RPMI1640 medium at the same concentration of fetal bovine serum and penicillin-streptomycin solution. Both cells were at 37℃and 5% CO 2 Is cultured in a moist environment. HK-2 and 786-O cells were plated in 96-well plates at 1X 10 4 The density of individual cells/wells was cultured. After cell growth densities of 70% were reached, the cells were treated with different concentrations (0, 10, 20, 60, 80 and 100. Mu.g/mL) of PCN@HP and further cultured for 24 hours. The light was then irradiated with a near infrared nir=808 nm laser for 10 minutes in the absence of light. Subsequently, relative cell viability was measured at 450nm after addition of CCK-8 solution to the well plate and incubation at 37 ℃ for 1.5 hours.
As shown in FIG. 3a, the PCN@HP suspension at a dose of 60 μg/mL under near infrared light irradiation has a tumor inhibition rate of more than 50% on ccRCC, and the suspension has a tumor inhibition rate of 100% at a concentration of 100 μg/mL. However, as shown in FIG. 3b, the viability of HK-2 cells remained unchanged under the same conditions. Notably, as shown in fig. 3c, although DOX also exhibits strong tumor killing ability as an antitumor drug widely used clinically; however, significant death of HK-2 cells also resulted at equivalent concentrations compared to PCH@HP, highlighting the superior safety and potential for clinical use of PCN@HP.
Example 4 detection of anti-tumor Effect in PCN@HP in vivo
Male nude mice (4-5 weeks old, 18-20 g) were purchased from Beijing Veitz Lihua Co., ltd, and kept in independently ventilated cages at room temperature of 21.+ -. 2 ℃ and relative humidity of 50.+ -. 15%. The food and water supply is sufficient. After one week of adaptive breeding, 1×10 will be fed 6 786-O cells resuspended in RPMI1640 medium were mixed with Matrigel at a 1:1 ratio and injected into the underarm subcutaneous tissue of male nude mice. When the tumor volume reaches 100mm 3 At this time, the mice were randomly divided into four groups (n=5 per group): PBS, NIR, PCN@HP and PCN@HP-NIR groups. Intratumoral treatment was performed by injecting 1mg/mL (100. Mu.L) of the corresponding liquid. After 3 hours, the tumor site was irradiated with 808nm laser (2.0W/cm 2 ) The mice were irradiated for 10 minutes and tumor sizes were recorded.
As shown in fig. 4, the tumor was reduced immediately after pcn@hp-NIR treatment, indicating that pcn@hp-NIR had a strong tumor growth inhibition effect.
EXAMPLE 5 in vivo Metabolic conditions after PCN@HP injection
The IVIS Spectrum system was used to explore the biodistribution and metabolism of PCN@HP. The major organs of the mice were fluorescent imaged after 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 24 hours and 48 hours of PCN@HP suspension at a dose of 100. Mu.L at a concentration of 1mg/mL by tail vein injection, and PCN@HP metabolism was recorded.
As shown in FIG. 5, the PCN@HP after intravenous injection is mainly metabolized through intestinal tracts, and according to the change of fluorescence intensity, the PCN@HP starts to be excreted 2 hours after the intravenous injection, and the PCN@HP reaches a peak after 3 hours, and can be completely metabolized 48 hours, so that the PCN@HP nanoparticle has no risk of accumulation in vivo and has potential clinical application value.
EXAMPLE 6 biological safety detection of PCN@HP
Toxicity in vivo experiment: liver and kidney function was assessed in 50C 57BL6J mice, including 25 males and 25 females, and PCN@HP suspension (1 mg/mL, 200. Mu.L) was administered by intraperitoneal injection. These mice were randomly divided into five groups, each group consisting of 5 males and 5 females. In addition, untreated mice 10 served as control group. Blood and organ samples were collected 1 day, 3 days, 7 days, 14 days and 28 days after injection of pcn@hp suspension for biochemical detection.
As shown in fig. 6a-d, the essential indicators of kidney function (Scr, BUN) and key biomarkers of liver function (AST, ALT) did not change significantly over time compared to the control group. These findings suggest the biosafety of pcn@hp treatment, underscores its potential for clinical use.

Claims (5)

1. The application of the polymer carbon nitride@Hittorf red phosphorus heterostructure in preparing medicaments for targeted treatment of renal clear cell carcinoma.
2. The use according to claim 1, wherein the polymer carbon nitride @ Hittorf red phosphorus heterostructure is a composite material prepared by a vapor deposition method and has the structural formula PCN @ HP.
3. The use according to claim 2, wherein the composite material is prepared by the following method:
heating melamine for 4 hours to 550 ℃ to prepare a polymer carbon nitride PCN;
grinding PCN and amorphous red phosphorus powder together in a mortar according to a ratio of 2:1;
thereafter, the obtained powder was transferred into a quartz ampoule and sealed with an oxygen-hydrogen flame under a low vacuum condition of-0.09 MPa;
the ampoule is heated to 550 ℃ in a furnace at 450 ℃ at a rate of 2 ℃/min and is kept for 4 hours, then is cooled to 280 ℃ at a rate of 1 ℃/min, when the temperature is reduced to 280 ℃/min, the cooling rate is reduced to 0.1 ℃/min, and after the ampoule is slowly cooled to room temperature at the rate, the capsule is crushed and rinsed by CS2, ethanol and distilled water respectively, and the PCN@HP heterostructure is obtained.
4. Use according to any one of claims 1-3, characterized in that in the heterostructure the HP is anchored vertically on the PCN, with a length of 1-2 μm and a diameter of 40-200 nm.
5. Use according to any one of claims 1-3, characterized in that the pcn@hp suspension dose is above 50% tumor inhibition at 60 μg/mL after 10 minutes of light irradiation at near infrared 808nm and 100% tumor inhibition at a suspension concentration of 100 μg/mL.
CN202311559630.8A 2023-11-21 2023-11-21 Polymer carbon nitride@Hittorf red phosphorus heterostructure and application thereof in preparation of medicine for targeted treatment of renal clear cell carcinoma Pending CN117503928A (en)

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