A copper-sulfur co-doped biomass carbon dot CDs1, a preparation method thereof and application thereof in preparation of colorectal cancer treatment drugs

Abstract

The application discloses copper-sulfur co-doped biomass carbon dots CDs1, a preparation method of the CDs1 and application of the CDs1 in preparation of colorectal cancer treatment drugs. The CDs1 is prepared by a hydrothermal method with corn residue as a carbon source, has an ultra-small size (about 0.21 nm lattice stripe) and uniform element distribution (C, O, N, S and Cu). The application proves that the CDs1 releases DAMPs and inflammatory factors (IL-1beta and IL-18) by inducing tumor cell copper death (DLAT oligomerization and FDX1 reduction) and pyroptosis (NLRP3 / cGAS-STING axis activation), remodels a tumor immune microenvironment, and activates CD8+ T cells and M1 macrophages. In-vitro and in-vivo experiments show that the CDs1 significantly inhibits colorectal cancer cell proliferation, migration and tumor growth, and has no obvious liver and kidney toxicity. The CDs1 can be used for preparing green and safe colorectal cancer treatment drugs.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a copper-sulfur co-doped biomass carbon dot CDs1, its preparation method, and its application in the preparation of drugs for colorectal cancer treatment. Background Technology

[0002] Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths worldwide, with a 5-year survival rate of less than 50% for patients with advanced-stage disease. Current treatments include chemotherapy, targeted therapy, and immune checkpoint inhibitors, but microsatellite stable (MSS) patients respond poorly, partly because the apoptosis induced by traditional therapies is "immune silencing," which fails to effectively activate anti-tumor immunity.

[0003] Copper death and pyroptosis, as novel programmed cell death pathways, can induce immunogenic cell death (ICD) accompanied by DAMP exposure and inflammatory cytokine release. However, existing metal ion delivery systems suffer from poor tumor targeting, high toxicity, and complex fabrication processes. Biomass carbon dots (CDs), with their green synthesis, biocompatibility, and metal chelating capabilities, have become ideal nanocarriers, but copper / sulfur co-doped designs for CRC are lacking.

[0004] Therefore, there is an urgent need to develop new, low-toxicity CDs platforms to solve these problems. Summary of the Invention

[0005] Technical problem solved: To address the above-mentioned technical problems, this invention provides a copper-sulfur co-doped biomass carbon dot CDs1, its preparation method, and its application in the preparation of colorectal cancer treatment drugs, which solves the technical problems of poor immune response, high systemic toxicity, and complex preparation of nanocarriers in existing colorectal cancer treatment drugs.

[0006] Technical solution: A copper-sulfur co-doped biomass carbon dot CDs1, which has a quasi-spherical structure, graphitized lattice stripes of 0.21±0.02 nm and uniform distribution of C, O, N, S and Cu.

[0007] Preferably, the oxygen content of CDs1 is not less than 60 at.%, and its FT-IR spectrum is in the range of 500-800 cm⁻¹. -1 It shows Cu-O stretching peaks in the 1050-1250 cm⁻¹ range. -1 The S-related absorption band is shown.

[0008] The preparation method of the above-mentioned copper-sulfur co-doped biomass carbon dots CDs1 is as follows: using corn residue as carbon source, adding Cu source and S source, carrying out hydrothermal reaction, cooling and dialysis purification to obtain the CDs1.

[0009] Preferably, the Cu source is copper chloride and the S source is sodium sulfide.

[0010] Preferably, the hydrothermal reaction is carried out at a temperature of 160-240°C for 4-12 hours.

[0011] The application of the aforementioned copper-sulfur co-doped biomass carbon dots CDs1 in the preparation of drugs for colorectal cancer treatment.

[0012] Preferably, the drug is used to induce copper death in tumor cells, manifested by DLAT oligomerization, decreased expression of FDX1 and LIAS, and downregulation of α-ketoglutarate and pyruvate; to induce pyroptosis in tumor cells by activating the cGAS-STING axis and NLRP3 inflammasome, releasing IL-1β and IL-18; and to remodel the tumor immune microenvironment, including increasing the proportion of CD8+ T cells and promoting M1 macrophage polarization.

[0013] A colorectal cancer treatment drug comprising the CDs1 and a pharmaceutically acceptable carrier or excipient.

[0014] Preferably, the carrier is water, PBS, or liposomes.

[0015] Preferably, the drug is an injectable preparation with a dosage of 25-50 mg / kg; the drug can be used in combination with chemotherapy drugs or targeted drugs.

[0016] Beneficial effects: The principle of this invention is as follows: CDs1 utilizes the oxygen-containing functional groups in corn residue to chelate Cu. 2+ S doping enhances stability. It disrupts mitochondrial metabolism (TCA cycle imbalance) via a copper-dependent mechanism, inducing copper death; mtDNA leakage activates the cGAS-STING axis, driving NLRP3 inflammasome-mediated pyroptosis. This process was validated by Western blot, TEM, and metabolomics, with ROS levels increasing in a dose-dependent manner.

[0017] Compared with the prior art, the present invention has the following beneficial effects: 1. CDs1 is a green synthetic material with high biocompatibility and no significant hepatotoxicity or nephrotoxicity, making it suitable for in vivo application.

[0018] 2. By synergistically inducing ICD through copper death and pyroptosis, the immune microenvironment is remodeled, thereby improving the therapeutic response to MSS-type CRC.

[0019] 3. The preparation process is simple and low-cost, utilizing agricultural waste to achieve high-value applications.

[0020] 4. Combined use with chemotherapy drugs enhances synergistic effects, providing a new treatment strategy for immune-cold tumors. Attached Figure Description

[0021] Figure 1 TEM, AFM, and EDS characterization of CDs1; Figure 2 Effects of CDs1 on CRC cell proliferation, migration, and invasion; Figure 3 : Schematic diagram of the mechanism of CDs1-induced copper death (DLAT oligomerization, ROS changes); Figure 4 : Morphological and molecular evidence of CDs1-induced pyroptosis; Figure 5 Evaluation of the in vivo tumor-suppressive effect and safety of CDs1; Figure 6 : The regulation of the tumor immune microenvironment by CDs1. Detailed Implementation

[0022] The present invention will be further described below with reference to the accompanying drawings and specific embodiments. Experimental methods in the following embodiments, unless otherwise specified, employ conventional techniques in this technical field or follow the conditions recommended by the manufacturer; reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0023] Example 1: Synthesis and Characterization of CDs1 Corn residue was used as the carbon source, and Cu (CuCl2) and S (Na2S) sources were added. The mixture was reacted in a hydrothermal reactor at 180℃ for 6 hours. After cooling, CDs1 was purified by dialysis. TEM showed that CDs1 exhibited a monodisperse, ultrasmall size with lattice fringes of approximately 0.21 nm; AFM confirmed a quasi-spherical structure. Figure 1 a). EDS mapping shows that C, O, N, S, and Cu are uniformly distributed, with an oxygen content of ~63 at.% ( Figure 1 b). FT-IR spectra show Cu-O stretching peaks (500-800 cm⁻¹). -1 ) and S-related absorption band (1050-1250 cm) -1 () Figure 1 c). XPS full spectrum confirmed the Cu2p and S2p peaks, and the shifts of the C1s and O1s peaks indicated an adjustment of the electronic structure ( Figure 1 d). The results show that CDs1 was successfully synthesized and has good stability.

[0024] Example 2: Inhibitory effect of CDs1 on CRC cells In CRC cell lines such as HCT116 and RKO, treatment with CDs1 (25-200 μg / mL) for 24 h resulted in a dose-dependent decrease in cell viability, with high concentration groups showing a viability rate <50%. Figure 2 a); weak inhibition of normal NCM460 cells ( Figure 2 b). EdU and colony formation assays showed that CDs1 reduced cell proliferation ( Figure 2c, d). Transwell showed a reduction in migration / invasion ( Figure 2 e). Western blot showed downregulation of N-cadherin and vimentin (e). Figure 2 f). Cell cycle analysis showed S phase arrest ( Figure 2 g). Combining with chemotherapy drugs such as 5-FU and targeted drugs such as Cetuximab further reduces viability ( Figure 2 h). The results showed that CDs1 effectively inhibited the development and progression of CRC cells.

[0025] Example 3: CDs1-induced copper death mechanism CDs1 treatment increases intracellular Cu levels ( Figure 3 a). Western blot showed decreased esterified DLAT / DLST and decreased FDX1 / LIAS (a). Figure 3 b). DLAT oligomerization ( Figure 3 c), enhanced mitochondrial colocalization ( Figure 3 d). Metabolomics showed that α-ketoglutarate was downregulated and succinate was upregulated ( Figure 3 e).

[0026] Example 4: CDs1-induced pyroptosis mechanism CDs1 treatment resulted in cell swelling and pyroptosis vesicles. Figure 4 a). Increased LDH release and elevated PI-positive cell count ( Figure 4 b, c). Western blot showed upregulation of NLRP3, Cleaved-caspase 1, and GSDMD, and release of IL-1β / IL-18 (b, c). Figure 4 d, e, f). mtDNA leakage activates cGAS-STING ( Figure 4 (g, h). The results show that S doping enhances pyrolysis induction.

[0027] Example 5: In vivo tumor suppression and safety of CDs1 In the RKO / CT26 subcutaneous tumor model, tail vein injection of CDs1 (25 / 50 mg / kg) significantly reduced tumor volume / weight. Figure 5 ac). Ki67 decreased, TUNEL positivity increased ( Figure 5 d, e). No weight loss ( Figure 5 f). H&E showed no organ damage ( Figure 5 g). Serum ALT / AST levels are normal ( Figure 5 h). The results indicate that CDs1 is safe and effective in vivo.

[0028] Example 6: Regulation of the tumor immune microenvironment by CDs1 Single-cell transcriptome analysis showed enrichment of M2 macrophages and depletion of CD8+ T cells in the tumor. Figure 6 CDs1 treatment upregulates CD86+ macrophages, downregulates CD206+, and increases the ratio of CD4+ / CD8+ T cells. Figure 6 The results indicate that CDs1 remodels the immune microenvironment and activates anti-tumor immunity.

[0029] The present invention has been described above by way of example. It should be noted that any simple modifications, alterations or other equivalent substitutions that can be made by those skilled in the art without creative effort without departing from the core of the present invention fall within the protection scope of the present invention.

Claims

1. A copper-sulfur co-doped biomass carbon dot CDs1, characterized in that, The CDs1 has a quasi-spherical structure, graphitized lattice fringes of 0.21±0.02 nm, and a uniform distribution of C, O, N, S, and Cu.

2. The copper-sulfur co-doped biomass carbon dot CDs1 according to claim 1, characterized in that, The oxygen content of CDs1 is not less than 60 at.%, and its FT-IR spectrum is in the range of 500-800 cm⁻¹. -1 It shows Cu-O stretching peaks in the 1050-1250 cm⁻¹ range. -1 The S-related absorption band is shown.

3. The method for preparing copper-sulfur co-doped biomass carbon dots CDs1 according to claim 1, characterized in that, The preparation method is as follows: using corn residue as a carbon source, adding Cu and S sources, carrying out a hydrothermal reaction, and then purifying by dialysis after cooling to obtain the CDs1.

4. The method for preparing copper-sulfur co-doped biomass carbon dots CDs1 according to claim 3, characterized in that, The Cu source is copper chloride, and the S source is sodium sulfide.

5. The method for preparing copper-sulfur co-doped biomass carbon dots CDs1 according to claim 3, characterized in that, The hydrothermal reaction is carried out at a temperature of 160-240℃ for 4-12 hours.

6. The use of the copper-sulfur co-doped biomass carbon dots CDs1 as described in claim 1 in the preparation of drugs for colorectal cancer treatment.

7. The application of the copper-sulfur co-doped biomass carbon dots CDs1 according to claim 6 in the preparation of colorectal cancer therapeutic drugs, characterized in that, The drug is used to induce copper death in tumor cells, manifested by DLAT oligomerization, decreased expression of FDX1 and LIAS, and downregulation of α-ketoglutarate and pyruvate; to induce pyroptosis in tumor cells by activating the cGAS-STING axis and NLRP3 inflammasome, releasing IL-1β and IL-18; and to remodel the tumor immune microenvironment, including increasing the proportion of CD8+ T cells and promoting M1 macrophage polarization.

8. A drug for treating colorectal cancer, characterized in that, Includes CDs1 as described in claim 1 and pharmaceutically acceptable carriers or excipients.

9. A colorectal cancer treatment drug according to claim 8, characterized in that, The carrier is water, PBS, or liposomes.

10. A colorectal cancer treatment drug according to claim 9, characterized in that, The drug is an injectable preparation with a dosage of 25-50 mg / kg; the drug can be used in combination with chemotherapy drugs or targeted drugs.

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