Preparation method of cascade energy transfer type self-luminous nanoparticles and application of cascade energy transfer type self-luminous nanoparticles in induction of pyroptosis of tumor cells

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to a preparation method of cascade energy transfer type self-luminous nanoparticles and application of the cascade energy transfer type self-luminous nanoparticles in induction of pyroptosis of tumor cells. The invention discloses a self-luminous nanoparticle (CC (at) PDC) which is prepared by self-assembling amphiphilic porphyrin lipid (PL), a camptothecin derivative (D2) and a targeting molecule DSPE-PEG2000-CREKA, and meanwhile, oleic acid modified calcium peroxide (OA-CaO2) and bisoxalate (CPPO) are encapsulated. In an acidic tumor microenvironment (TME), CaO2 provides O2 and H2O2, the O2 and H2O2 react with CPPO to trigger camptothecin to emit strong light at 420 nm, and the light can effectively excite porphyrin to generate reactive oxygen species (ROS); meanwhile, camptothecin serving as a chemotherapeutic drug can be slowly released, so that camptothecin enhanced spontaneous photodynamic-chemotherapy combined treatment is realized. The nanoparticles can effectively induce pyroptosis of tumor cells, the problems of poor light penetrability and chemotherapy drug resistance of traditional PDT are solved, a cascade energy transfer mechanism gets rid of dependence on tumor endogenous H2O2 and O2, and the treatment effect is remarkably improved.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a method for preparing cascaded energy transfer type self-luminescent nanoparticles and their application in inducing pyroptosis in tumor cells. Background Technology

[0002] Tumor immunotherapy, as a promising treatment approach, has shown remarkable efficacy in tumor destruction, particularly in inhibiting tumor metastasis. Among these, immune checkpoint blockade (ICB) therapy, based on the PD-1 / PD-L1 regulatory pathway, has achieved significant breakthroughs in the treatment of various advanced cancers by alleviating the immunosuppressive microenvironment. However, its low immunogenicity response rate severely limits its efficacy and widespread application in tumor treatment.

[0003] Pyroptosis, a newly discovered immunogenic cell death mechanism, releases various pro-inflammatory cytokines and alarm factors, triggering a strong antigen-specific immune response. Pyroptosis in just 15% of tumor cells can eliminate the entire tumor, providing a highly promising strategy for enhancing anti-tumor immunity. The occurrence of pyroptosis is closely related to Gasdermin family proteins. The N-terminal domain (GSDMD-N) obtained by Caspase-1 cleavage of gasdermin D (GSDMD) can interact with cell membrane phospholipids, forming pores, disrupting membrane permeability, leading to cell swelling, rupture, and the release of inflammatory contents, significantly enhancing tumor immunogenicity. However, effective methods for inducing pyroptosis are currently lacking. While chemotherapy drugs such as topotecan can induce pyroptosis, they suffer from high toxicity, drug resistance, and poor light penetration in photodynamic therapy (PDT).

[0004] Photodynamic therapy (PDT), as a clinically approved phototherapy method, utilizes light to activate photosensitizers (PSs) and oxygen in tumors to generate reactive oxygen species (ROS). It has advantages such as high specificity, high safety, no drug resistance, and repeatability, and shows great promise in the treatment of malignant tumors. However, the poor penetration of external light into the body's tissues and the problem of tumor hypoxia greatly reduce the efficacy of PDT.

[0005] To address the light-dependent nature of phototherapy (PDT), self-luminescent systems have been developed, including bioluminescence, Cherenkov radiation, and chemiluminescence, among which chemiluminescent resonance energy transfer (CRET) has been extensively studied. It typically utilizes the reaction of hydrogen peroxide (H₂O₂) with high-energy substrates such as bis(oxalate) ester (CPPO) to generate a high-energy intermediate, 1,2-dioxane, which then excites phosphorus sacs (PSs) to emit light and kill tumor cells, showing promising potential in the clinical treatment of deep-seated tumors. However, the low energy transfer efficiency between the energy substrate and the luminescent group often weakens the efficiency of CRET-based PDT, requiring high-dose drugs to achieve effective treatment and thus causing unnecessary toxicity.

[0006] Studies have found that doping certain dyes into chemiluminescence systems can produce CRET, significantly amplifying the intensity of chemiluminescence. Chemotherapy drugs such as camptothecin (CPT) and doxorubicin (DOX) are themselves dyes, capable of energy transfer and synergistically enhancing the efficiency of autoluminescent photodynamic therapy (PDT). Furthermore, PDT can reduce chemotherapy resistance by degrading drug resistance proteins. In addition, chemically designed micro / nanoparticles can be precisely targeted to organelles using subcellular targeting strategies, significantly improving photodynamic effects or local drug accumulation efficiency. Therefore, the combination of PDT and chemotherapeutic dyes may induce stronger and more effective apoptosis; however, there are currently few reports on the mutual amplification of these two combinations to initiate effective pyroptosis.

[0007] Based on the above considerations, we developed a cascaded energy transfer type self-luminescent nanoparticle (CC@PDC). This nanoparticle consists of an amphiphilic porphyrin lipid (PL), a camptothecin derivative (D2), and the targeting molecule DSPE-PEG. 2000 -CREKA is self-assembled and encapsulates oleic acid-modified calcium peroxide (OA-CaO2) and CPPO. By targeting fibrin deposition areas in the tumor microenvironment, it enhances drug accumulation at the tumor site. In the acidic tumor microenvironment, OA-CaO2 self-supplyes H2O2 and O2, reacting with CPPO to trigger D2 luminescence. Through intraparticle cascade energy transfer, PL generates reactive oxygen species, while D2 is slowly released as a chemotherapeutic drug, achieving a chemotherapeutic-enhanced autoluminescent photodynamic-chemotherapy combination that effectively induces pyroptosis in tumor cells. When CC@PDC is used in combination with PD-L1 antibodies, it can elicit excellent anti-tumor immune effects, providing a new strategy for the treatment of deep in situ and metastatic tumors. Summary of the Invention

[0008] The purpose of this invention is to provide a method for preparing cascaded energy transfer type self-luminescent nanoparticles. This addresses the problems of existing self-luminescent photodynamic therapy (PDT) which relies on endogenous H2O2 and O2, has low energy transfer efficiency, insufficient induction of tumor cell pyroptosis, and limited anti-tumor effects.

[0009] Another objective of this invention is to provide the application of the above-mentioned cascaded energy transfer type self-luminescent nanoparticles in inducing pyroptosis of tumor cells and in tumor therapy.

[0010] The structure of the cascaded energy transfer type self-luminous nanoparticles (CC@PDC) described in this invention is shown in the attached figure. Figure 1 As shown.

[0011] The cascaded energy transfer type self-luminescent nanoparticles (CC@PDC) of this invention are self-assembled from amphiphilic porphyrin lipids (PL), camptothecin derivatives (D2), and the targeting molecule DSPE-PEG2-CREKA, while simultaneously encapsulating oleic acid-modified calcium peroxide (OA-CaO2) (synthesized according to the method in the literature: Chemical Engineering Journal 2024, 494, 152400) and CPPO. Among them, D2, PL, DSPC, and DSPE-PEG... 2000 The molar ratio of -CREKA is 10:40:40:10 (lipid mixture). OA-CaO2 dissolved in chloroform and CPPO dissolved in ethanol (OA-CaO2 and CPPO mass ratio 1:1) are added to the above lipid mixture at a drug-lipid mass ratio of 1:2.

[0012] The amphiphilic porphyrin lipid (PL) is a porphyrin-grafted lipid. The synthesis steps of the porphyrin lipid are described in Chinese Patent (ZL.201010222238.0). PL has fluorescence and the ability to generate singlet oxygen, and can be used as a photosensitizer (PSs) to avoid quenching effects caused by aggregation.

[0013] The preparation method of the camptothecin derivative (D2) is based on the patent application (application number: 201611231246.5). It consists of two camptothecin (CPT) molecules with good overlap of emission spectrum and PL absorption, connected to a pentaerythritol backbone. It serves as an intraparticle luminescent resonance energy transfer medium and can also play a cytotoxic role as a chemotherapy drug.

[0014] Among them, the target molecule DSPE-PEG 2000 - The preparation method of CREKA is described in the reference (Theranostics 2017, 7(5): 1062-1071). CREKA (Cys-Arg-Glu-Lys-Ala) is a pentapeptide that can target fibrin deposition areas in the tumor microenvironment and enhance the enrichment of nanoparticles at the tumor site.

[0015] The preparation method of the cascaded energy transfer type self-luminous nanoparticles (CC@PDC) of the present invention includes the following steps:

[0016] (1) Add a certain proportion of DSPC, PL, and DSPE-PEG 2000 -CREKA is dissolved in ethanol to obtain a lipid solution. OA-CaO2 dissolved in chloroform and CPPO dissolved in ethanol (OA-CaO2 and CPPO mass ratio 1:1) are added to the above lipid solution at a drug-lipid mass ratio of 1:2.

[0017] (2) A uniform thin film was formed at the bottom of a round-bottom flask by using the thin film hydration method. After adding a certain amount of aqueous solution, the film was ultrasonically dispersed in a water bath for 3 minutes to obtain uniformly dispersed nanoparticles.

[0018] (3) Dialyze the nanoparticles obtained in step (2) using an 8000-14000KD dialysis bag for 2-4 hours to remove residual organic solvents, and store them in a 4℃ refrigerator for later use to obtain cascaded energy transfer type self-luminous nanoparticles (CC@PDC).

[0019] The cascaded energy transfer type self-luminescent nanoparticles (CC@PDC) described in this invention have unique properties in the acidic tumor microenvironment (TME): CaO2 provides O2 and H2O2, which react with CPPO to trigger camptothecin derivative (D2) to emit strong light at 420 nm. This light emission can effectively excite porphyrin lipids (PL) to produce reactive oxygen species (ROS). At the same time, D2 can be slowly released as a chemotherapeutic drug, realizing camptothecin-enhanced self-luminescent photodynamic-chemotherapy.

[0020] Characterization revealed that the nanoparticles exhibited a uniformly dispersed nanostructure as observed by transmission electron microscopy. Malvern-Zeta assays showed that their particle size and zeta potential conformed to the characteristic parameters of nanoparticles. High-resolution electron microscopy revealed a uniform distribution of elements such as OA-CaO2. Specific absorption and fluorescence characteristic peaks were detected by ultraviolet and fluorescence spectrophotometers. IVIS imaging system analysis indicated that the luminescence intensity was optimal at pH 5.0, and that the luminescence characteristics were dose-dependent and H2O2 concentration-dependent.

[0021] The cascaded energy transfer type self-luminescent nanoparticles (CC@PDC) described in this invention can effectively induce pyroptosis in tumor cells. The mechanism lies in achieving highly efficient photodynamic-chemical combined action through cascaded energy transfer, triggering Gasdermin D-mediated pyroptosis, releasing pro-inflammatory cytokines, and stimulating a systemic immune response. When used in combination with PD-L1 antibodies, it can further enhance the anti-tumor immune effect, effectively inhibiting deep in situ tumor growth and metastasis, providing a new and effective strategy for tumor treatment. Attached Figure Description

[0022] Figure 1 This is a structural diagram of the cascaded energy transfer type self-luminous nanoparticles (CC@PDC) described in this invention; Figure 2 The image is a transmission electron microscope image of CC@PDC in specific embodiment 1, showing its uniform spherical morphology; Figure 3 This is a particle size distribution and potential diagram of CC@PDC in specific embodiment 2; Figure 4 This is an elemental distribution diagram of CC@PDC in specific embodiment 2, showing the uniform distribution of elements such as Ca, Cl, O, and C; Figure 5These are the ultraviolet absorption and fluorescence emission spectra of PL and D2 in specific embodiment 3; Figure 6 This is a luminescence characteristic diagram of CC@PDC in specific embodiment 3, where A is the dose-dependent luminescence intensity, B is the H2O2 concentration-dependent luminescence intensity, and C is the luminescence intensity at different pH values. Figure 7 This is a graph showing the H2O2 production capacity of CC@PDC at different pH levels in specific embodiment 4; Figure 8 This is a diagram showing the O2 generation capability of CC@PDC in specific embodiment 4; Figure 9 This is the SOSG fluorescence intensity diagram of CC@PDC in specific embodiment 5; Figure 10 This is a laser confocal image of CC@PDC and CC@PD being taken up by tumor cells in specific embodiment 6. Figure 11 This is a graph showing the results of Western blot detection of pyroptosis-related protein expression in specific embodiment 7; Figure 12 These are the tumor growth curves of each group of tumor-bearing mice in Specific Example 8, comparing the anti-tumor effects of different treatment regimens. Figure 13 This is a diagram showing the results of CC@PDC inducing the body's immune response in specific embodiment 9. Detailed Implementation

[0023] The following detailed descriptions will help to understand the present invention, but do not limit the scope of the invention.

[0024] Example 1: Preparation of cascaded energy transfer type self-luminescent nanoparticles (CC@PDC).

[0025] CC@PDC nanoparticles were prepared using a thin-film hydration method. Specifically, camptothecin derivative (D2), amphiphilic porphyrin lipid (PL), DSPC, and the targeting molecule DSPE-PEG were weighed in a molar ratio of 10:40:40:10. 2000 -CREKA was dissolved in ethanol to obtain a lipid solution; oleic acid-modified calcium peroxide (OA-CaO2) was dissolved in chloroform and CPPO was dissolved in ethanol (OA-CaO2 and CPPO mass ratio 1:1). These were added to the lipid solution at a drug-lipid mass ratio of 1:2 and stirred until homogeneous. The mixture was transferred to a round-bottom flask, and the organic solvent was removed by rotary evaporation to form a uniform film at the bottom of the flask. An appropriate amount of deionized water was added, and the mixture was ultrasonically dispersed in a water bath for 3 minutes to obtain a uniformly dispersed nanoparticle suspension. The suspension was transferred to a dialysis bag (molecular weight cutoff 8000-14000 KD) and dialyzed in deionized water for 2-4 hours to remove residual organic solvent. It was then stored at 4°C for later use, yielding cascaded energy transfer type self-luminescent nanoparticles (CC@PDC) (see attached). Figure 1 ).

[0026] Example 2: Morphological and physicochemical characterization of CC@PDC.

[0027] The CC@PDC suspension prepared in Example 1 was dropped onto a copper grid, stained with phosphotungstic acid, and its morphology was observed using a transmission electron microscope (TEM, JEM 1400). The results are attached. Figure 2 As shown, CC@PDC is spherical, uniformly dispersed, and has a particle size of approximately 100 nm.

[0028] The above CC@PDC suspension was dispersed in an aqueous solution, and the particle size and zeta potential were investigated using Malvern assay over 7 days. The results are shown in the attached figure. Figure 3 As shown, the average particle size over 7 days was approximately 120 nm, and the zeta potential was approximately -20 mV, demonstrating that CC@PDC has good stability and providing a physicochemical basis for subsequent applications.

[0029] The elemental distribution of CC@PDC was observed using a high-resolution electron microscope, as shown in the attached figure. Figure 4 As shown, elements such as Ca, Cl, O, and C are uniformly distributed in the nanoparticles, confirming the effective encapsulation of OA-CaO2 and CPPO.

[0030] Example 3: Optical performance test of CC@PDC.

[0031] The spectra of D2 and PL were detected using a UV spectrophotometer and a fluorescence spectrophotometer, as shown in the attached figure. Figure 5 As shown, the ultraviolet absorption peak of PL is at 425 nm; when the excitation wavelength is 365 nm, the characteristic fluorescence peak of D2 is detected at 430 nm. The two overlap well, which is a key factor in the CRET process and confirms the possibility of energy transfer.

[0032] Take CC@PDC and test different doses (PL: 0.05-0.5 mg / mL) (see attached). Figure 6 A) Different H2O2 concentrations (20-100mM) (see attached) Figure 6 B) and luminescence intensity at different pH values ​​(5.0-7.4) (see attached image) Figure 6 C). The results showed that the luminescence intensity increased with increasing dose and H2O2 concentration, reaching a peak at pH 5.0, indicating that it has pH-responsive luminescence properties.

[0033] Example 4: Test of H2O2 and O2 generation capacity of CC@PDC.

[0034] CC@PDC suspension was placed in EP tubes and then incubated in PBS at pH 5.0, 6.5, and 7.4 at different time points using an H2O2 kit. Results are attached. Figure 7As shown, the amount of H2O2 generated gradually increases as the pH value decreases, and the H2O2 release is the highest at pH 5.0 for NP, confirming the ability of tumor acidic TME to trigger H2O2 generation in CC@PDC.

[0035] O2 generation was detected using a dissolved oxygen meter. CC@PDC was placed in deoxygenated PBS (pH=5.0) and oxygen concentration changes were monitored under sealed conditions. (See attached image.) Figure 8 As shown, oxygen production exhibits a significant concentration dependence, indicating that OA-CaO2 can effectively provide O2.

[0036] Example 5: ROS generation verification of CC@PDC.

[0037] SOSG probe (10 μM) was mixed with PBS or CC@PDC (pH = 5.0-7.4), and fluorescence spectra were detected at different time points (excitation 488 nm, emission 525 nm). Results are attached. Figure 9 As shown, the SOSG fluorescence intensity in the PBS group did not change significantly, while in the CC@PDC nanoparticle groups at pH 6.5 and 5.0, the SOSG fluorescence intensity gradually increased over time in a pH-dependent manner. 1 O2 production increases as pH decreases, confirming... 1 O2 is continuously generated.

[0038] Example 6 illustrates the effect of CC@PDC on targeting tumor cells. The uptake of CC@PD (non-targeted) and CC@PDC (targeted) was observed using confocal microscopy, as shown in the attached figure. Figure 10 As shown. 4T1 cells were seeded in confocal culture dishes and incubated overnight. Then, nanoparticles (PL concentration 25 μg / mL) were co-incubated with the cells for 2, 4, 8, and 12 hours. Lysosomes were labeled with the Lyso Tracker Green DND-26 probe, and cell nuclei were labeled with Hoechst. Within 12 hours, the fluorescence of PL and D2 in 4T1 cells gradually increased over time, indicating a gradual increase in endocytosis. In the CC@PDC group (see attached...) Figure 10 The fluorescence intensities of PL and D2 in group A) were significantly higher than those in group CC@PD at the same time point (see attached image). Figure 10 B) indicates that cells in the CC@PDC group took up more drug, possibly due to DSPE-PEG. 2000- CREKA binds to the tumor ECM and fibrin deposition regions highly expressed by tumor cells, enabling NP to accurately locate the tumor site.

[0039] Example 7: In vitro experiment on CC@PDC-induced pyroptosis of tumor cells.

[0040] Mouse breast cancer 4T1 cells were seeded into 6-well plates (1×10⁻⁶ cells per well). 5 Cells / well were used to treat different groups, and after 24 hours of incubation, the expression levels of pyroptosis-related proteins were detected by Western blot. Results are attached. Figure 11 As shown, intracellular expression of N-GSDMD in the CC@PDC group was upregulated by 1.54-fold, while NLRP3 and Cleaved-Caspase were upregulated by 2.42-fold and 16.3-fold, respectively, indicating the presence of pyroptosis, and that this process was successfully triggered via the Caspase-1 / GSDMD pathway.

[0041] Example 8: In vivo animal experiment on the inhibition of tumor growth by CC@PDC. A 4T1 tumor-bearing BALB / c mouse model was established and randomly divided into 6 groups: PBS group, PD-L1 antibody group, CC@PC group, PDC group, CC@PDC group, and CC@PDC+PD-L1 antibody group. Administered via tail vein injection, CC@PDC at a dose of 5 mg / kg and PD-L1 antibody at a dose of 100 μg / mouse, every 2 days via intraperitoneal administration.

[0042] Tumor volume was measured every two days (formula: 1 / 2 × length × width²), and the results are attached. Figure 12 As shown, compared with the control group, CC@PC exhibited a weaker tumor-suppressive effect, which is attributed to the limited tumor-suppressive effect of ROS generated by PL stimulation alone. The PDC group achieved better tumor suppression through the release of chemotherapeutic drugs via camptothecin. The αPD-L1 group showed some degree of tumor suppression; αPD-L1 induces a systemic immune response by inhibiting immune checkpoints, enhancing T cell activity, and influencing the tumor microenvironment. The CC@PDC group, possessing both chemotherapeutic and PDT functions, exhibited significant tumor-suppressive effects. The combination of CC@PDC and αPD-L1 further enhanced the therapeutic effect, demonstrating the potential of this approach in improving anti-tumor immunity and treatment outcomes.

[0043] Example 9: Testing of tumor immune effects.

[0044] The isolated tumor tissue and draining lymph nodes were analyzed by flow cytometry. CD4+ in the CC@PDC+αPD-L1 group tumors was analyzed. + T cell infiltration level and CD8 + T cell infiltration rates reached 67.1% and 22.4%, respectively, which were higher than those in the CC@PDC group (CD4+). + T: 47.9%, CD8 + T: 16.0% or αPD-L1 alone (CD4) + T: 20.5%, CD8 + T: 13.2% (with appendix) Figure 13 A, D, E).

[0045] CD80 and CD86 markers were used to assess the maturity of dendritic cells (DCs) in lymph nodes. Compared with the PBS group (13.7%), the levels of mature DCs in tumor-draining lymph nodes were increased to some extent in other groups. The proportion of mature dendritic cells in the CC@PDC group reached approximately 59.1%, and increased to 87.6% after binding to αPD-L1, indicating that it significantly enhanced antigen presentation capacity and improved the overall activation level of the immune system (see appendix). Figure 13 B,F).

[0046] Furthermore, the proportion of immunosuppressive T regulatory (Treg) cells within the tumor was significantly reduced in the CC@PDC+αPD-L1 group. Compared to 13.7% in the control group, the proportion of Treg cells in the CC@PDC+αPD-L1 group was only 0.73%, effectively reversing the immunosuppressive microenvironment within the tumor (see appendix). Figure 13 C,G). The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A cascaded energy transfer type self-luminous nanoparticle, characterized in that, Composed of amphiphilic porphyrin lipids (PL), camptothecin derivative (D2), and the targeting molecule DSPE-PEG 2000 -CREKA self-assembles and internally encapsulates oleic acid-modified calcium peroxide (OA-CaO2) and CPPO; wherein D2, PL, DSPC, and DSPE-PEG are present. 2000 The molar ratio of -CREKA is 10:40:40:

10.

2. The cascaded energy transfer type self-luminous nanoparticles according to claim 1, characterized in that, By replacing DSPE-PEG 2000 -CREKA is a DSPE-PEG 2000 Non-targeted control nanoparticles were obtained, or Cy5.5-labeled nanoparticles were obtained by doping DSPE-Cy5.5 into the phospholipid component at a mass ratio of 3%.

3. The cascaded energy transfer type self-luminescent nanoparticle according to claim 1, wherein the chemical structural formula of PL is as follows:

4. The cascaded energy transfer type self-luminescent nanoparticle according to claim 1, wherein the chemical structural formula of D2 is as follows:

5. The cascaded energy transfer type self-luminescent nanoparticles according to claim 1, wherein the DSPE-PEG 2000 The chemical structural formula of -CREKA is as follows:

6. The method for preparing cascaded energy transfer type self-luminescent nanoparticles according to claim 1, comprising the following steps: (1) Add a certain proportion of DSPC, PL, and DSPE-PEG 2000 -CREKA is dissolved in ethanol to obtain a lipid solution. OA-CaO2 dissolved in chloroform and CPPO dissolved in ethanol (OA-CaO2 and CPPO mass ratio 1:1) are added to the above lipid solution at a drug-lipid mass ratio of 1:

2. (2) A uniform thin film was formed at the bottom of a round-bottom flask by using the thin film hydration method. After adding an aqueous solution, the film was ultrasonically dispersed in a water bath for 3 minutes to obtain uniformly dispersed nanoparticles. (3) Use a dialysis bag with 8000-14000KD and dialyze for 2-4 hours to remove residual organic solvents; (4) The system obtained above is transferred into a centrifuge tube to obtain cascaded energy transfer type self-luminous nanoparticles.

7. The cascaded energy transfer type self-luminous nanoparticles according to claim 1, characterized in that, It exhibits pH responsiveness in acidic environments and can provide O2 and H2O2 through OA-CaO2. It reacts with CPPO to trigger D2 to emit strong light at 420 nm, and this light emission can excite PL to produce reactive oxygen species (ROS). At the same time, D2 can slowly release camptothecin to achieve combined therapy.

8. The cascaded energy transfer type self-luminous nanoparticles according to claim 1, characterized in that, Transmission electron microscopy revealed that the morphology of the particles was a uniformly dispersed nanostructure. Malvern analysis showed that the particle size and zeta potential were consistent with the characteristic parameters of nanoparticles, with an average particle size of about 100 nm and a potential of ~-15 mV. Furthermore, a uniform distribution of elements such as OA-CaO2 could be observed using a high-resolution electron microscope.

9. The cascaded energy transfer type self-luminous nanoparticles according to claim 1, characterized in that, It exhibits optimal luminescence intensity at pH 5.0, and the luminescence intensity is dose-dependent and H2O2 concentration-dependent. It can also generate corresponding amounts of H2O2 and O2 in different pH environments. The generation of ROS can be verified by SOSG probe and Rhodamine B degradation assay.

10. The application of the cascaded energy transfer type self-luminescent nanoparticles as described in claim 1 in the preparation of a drug that induces pyroptosis in tumor cells, characterized in that, These nanoparticles achieve self-luminescent photodynamic-chemotherapy through cascaded energy transfer, effectively inducing pyroptosis in tumor cells. When used in combination with PD-L1 antibodies, they can enhance anti-tumor and immune effects.

Images