A calcium phosphate deposition targeted tumor non-carrier nano-drug and a preparation method thereof

Abstract

The application discloses a calcium phosphate deposition targeted tumor non-carrier nano drug and a preparation method thereof, and relates to the technical field of nano drug preparation. The method comprises the following steps: dissolving a small molecule drug in an organic solvent, dropping the organic solvent into a simulated body fluid, performing calcium phosphate biomimetic mineralization, and performing dialysis to obtain CaP@iBEFT; dissolving lactoferrin in a 4-morpholine ethanesulfonic acid aqueous solution, then adding 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide and N-hydroxysuccinimide, and then adding a 4-morpholine ethanesulfonic acid solution of polyethylene glycol to obtain LP; dissolving the LP and the CaP@iBEFT in water, performing dialysis, and obtaining LP-CaP@iBEFT. The product prepared by the application has a core-shell structure, and has the characteristics of good water solubility, high bioavailability and good safety. The application solves the problems of poor water solubility and low bioavailability of ferroptosis drugs in the prior art.

Description

Technical Field

[0001] This invention relates to the field of nanomedicine preparation technology, specifically to a carrier-free nanomedicine for targeted tumors deposited with calcium phosphate and its preparation method. Background Technology

[0002] In recent years, breast cancer has become one of the leading causes of death worldwide. Currently, the main treatment for breast cancer is traditional chemotherapy, but this method still has limitations, such as low bioavailability, significant drug toxicity and side effects, and damage to the immune system. Therefore, the development of highly effective and low-side-effect new cancer treatments is urgently needed. Researchers are dedicated to developing multifunctional nanomedicine carriers and investigating how to improve the targeting and controlled release efficiency of chemotherapy drugs to ensure accurate and effective delivery to tumor tissues.

[0003] Ferroplasm inducers are novel small chemical molecules that hold promise for breast cancer treatment. They induce the overexpression of reactive oxygen species (ROS) in tumor cells through iron accumulation, thereby disrupting the redox balance and inducing cell death. However, the ROS concentration produced by ferroplasm inducers is often insufficient to cause tumor cell death, and they lack biocompatibility from a molecular perspective, failing to exert their intended antitumor effects in vivo. Therefore, multiple ferroplasm inducers can be used synergistically to enhance their antitumor effects in vivo; however, how to combine multiple ferroplasm inducers to improve antitumor efficacy remains a pressing challenge. Summary of the Invention

[0004] To address the aforementioned technical problems, the present invention aims to provide a carrier-free nanomedicine for targeted tumors deposited with calcium phosphate and its preparation method, thereby solving the problems of poor water solubility and low bioavailability of ferroptosis drugs in the prior art.

[0005] The technical solution of this invention to solve the above-mentioned technical problems is as follows: A method for preparing calcium phosphate-deposited, carrier-free tumor-targeting nanomedicines is provided, comprising the following steps:

[0006] (1) Dissolve the small molecule drug in an organic solvent, stir evenly to obtain reaction solution one, add reaction solution one drop into simulated body fluid for biomineralization, and then dialyze to obtain CaP@iBEFT, which is a carrier-free nanomedicine deposited with calcium phosphate.

[0007] (2) Dissolve lactoferrin in an aqueous solution of 4-morpholinoethanesulfonic acid, then add 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N-hydroxysuccinimide, stir for 1-3 h, then add polyethylene glycol in an aqueous solution of 4-morpholinoethanesulfonic acid, continue stirring for 12-24 h, then dialyze to obtain LP, i.e. polyethylene glycol-modified lactoferrin;

[0008] (3) Dissolve the carrier-free nanomedicine deposited by calcium phosphate obtained in step (1) and the polyethylene glycol-modified lactoferrin obtained in step (2) in water, stir and react for 12-24 hours, and then dialyze to obtain LP-CaP@iBEFT, which is a carrier-free nanomedicine for targeting tumors deposited by calcium phosphate.

[0009] The beneficial effects of this invention are as follows: Carrier-free drugs can be polymerized into highly stable, biocompatible, and tumor microenvironment-responsive nanomedicines through self-assembly and biomineralization, utilizing the physicochemical properties of small molecule drugs or by coating their surfaces with hydrophilic substances. Calcium phosphate (CaP), as a good biomineralizing agent, can form stable nano-formulations. Furthermore, CaP can be released in acidic environments, exhibiting pH responsiveness. The tumor microenvironment (TME) is slightly acidic, allowing carrier-free nanomedicines to release drugs and exert anti-tumor effects upon entering the tumor environment. In addition, the lactoferrin on the surface of this nano-formulation can actively recognize lactoferrin receptors overexpressed on tumor cell membranes, thereby increasing the uptake capacity of tumor cells of carrier-free nanomedicines and enhancing their tumor therapeutic effect.

[0010] This invention prepares CaP@iBEFT by "biomineralizing" a small molecule drug in simulated body fluids, and then encapsulates the CaP@iBEFT surface with polyethylene glycol-modified lactoferrin to obtain a core-shell structured LP-CaP@iBEFT. This calcium phosphate-deposited, carrier-free nanomedicine for targeting tumors has the characteristics of good water solubility, high bioavailability and good safety, and can be used for targeted therapy of triple-negative breast cancer.

[0011] Based on the above technical solution, the present invention can be further improved as follows:

[0012] Furthermore, in step (1), the small molecule drug is an inhibitor of ferroptosis inhibitor 1, buquina, arastin, iron ions, and tannic acid.

[0013] The beneficial effects of adopting the above-mentioned further technical solutions are as follows: Among these five small molecule drugs, the inhibitor of protein 1 (iFSP1) is an inhibitor of the FSP1 pathway, brequinar is an inhibitor of the DHODH pathway, and erastin is an inhibitor of the GPX4 pathway. The above three inhibitors can inhibit the pathways that negatively regulate ferroptosis, thereby indirectly promoting ferroptosis. Iron ions and tannic acid can promote the Fenton reaction and directly induce ferroptosis.

[0014] Furthermore, in step (1), the organic solvent is dimethyl sulfoxide.

[0015] Furthermore, the mass-to-volume ratio of ferroptosis inhibitor 1 inhibitor, buquina, arastin, iron ions, tannic acid, and organic solvent was 0.5-2 mg: 0.5-2 mg: 0.5-2 mg: 0.5-2 mg: 200 μL.

[0016] Furthermore, in step (1), the volume ratio of the organic solvent to the simulated body fluid is 1:15-20.

[0017] Furthermore, the iron ions are ferric chloride.

[0018] Furthermore, in step (2), the total amount of lactoferrin, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N-hydroxysuccinimide, and the mass-to-volume ratio of the aqueous solution of 4-morpholinoethanesulfonic acid are 4-6 mg: 10-11 mg: 1-2 mL.

[0019] Furthermore, in step (2), the concentration of the 4-morpholine ethanesulfonic acid aqueous solution is 45-55 mmol / L.

[0020] Furthermore, in step (2), the mass-to-volume ratio of lactoferrin and polyethylene glycol in 4-morpholinoethanesulfonic acid solution is 4-6 mg: 1-2 mL.

[0021] Further, in step (2), the 4-morpholinoethanesulfonic acid solution of polyethylene glycol is prepared by dissolving 10 mg of polyethylene glycol in 1-2 mL of 4-morpholinoethanesulfonic acid to obtain the 4-morpholinoethanesulfonic acid solution of polyethylene glycol.

[0022] Furthermore, in step (2), the polyethylene glycol is NH2-PEG2000-NH2.

[0023] Furthermore, in step (3), the mass-to-volume ratio of the carrier-free nanomedicine deposited by calcium phosphate, the polyethylene glycol-modified lactoferrin, and water is 1 mg: 1-5 mg: 4-6 mL.

[0024] The present invention also provides carrier-free nanomedicines prepared by the above method.

[0025] Furthermore, in the carrier-free tumor-targeting nanomedicines deposited with calcium phosphate, the loading of ferroptosis inhibitor 1 inhibitor was 5-10 wt%, the loading of buquina was 5-10 wt%, the loading of asterine was 5-10 wt%, the loading of iron ions was 7-15 wt%, and the loading of lactoferrin was 25-35 wt%.

[0026] Furthermore, the carrier-free nanomedicine targeting tumors deposited with calcium phosphate has a core-shell structure, with the carrier-free nanomedicine deposited with calcium phosphate as the core and PEGylated lactoferrin as the outer shell.

[0027] Furthermore, the particle size of the calcium phosphate-deposited carrier-free tumor-targeting nanomedicine is 100-110 nm, and the shell thickness is 30-35 nm.

[0028] Furthermore, the calcium phosphate-deposited carrier-free nanomedicine targeting tumors has a particle size of 106 nm and a shell thickness of 33 nm.

[0029] This invention also provides the application of the above-mentioned calcium phosphate deposited carrier-free nanomedicine for targeted tumors in the preparation of targeted tumor drugs.

[0030] The present invention has the following beneficial effects:

[0031] 1. In the LP-CaP@iBEFT prepared in this invention, the nanomedicine comprises ferroptosis inhibitor 1 inhibitor (iFSP1), brequinar, erastin, and iron ions (Fe). 3+ CaP co-assembles with the reducing agent tannic acid (TA); PEGylated lactoferrin (LP) can actively target tumor cells by recognizing lactoferrin receptors on the cell membrane; CaP releases small molecule ferroptosis drugs Eratin, Brequinar, iFSP1, and Fe upon response to an acidic environment. 3+ It and TA lead to an increase in iron-related reactive oxygen species and the accumulation of lipid peroxides, triggering ferroptosis and thus inducing cell death, which has an anti-breast cancer effect.

[0032] 2. In the carrier-free nanomedicine prepared by this invention, CaP has the advantages of good biocompatibility, non-toxicity and pH response.

[0033] 3. In the carrier-free nanomedicine prepared by this invention, the active targeting of lactoferrin (Lf) and the passive targeting of CaP to tumor cells increase the accumulation of drug in tumor tissue, thereby improving drug absorption and efficacy. Compared with other targeting pathways (such as monoclonal antibody targeted therapy), the Lf targeting pathway has advantages such as low cost, low immunogenicity, and stable tumor penetration.

[0034] 4. This invention provides a method for preparing structurally stable and particle-size controllable nanomedicines via calcium phosphate deposition. The newly prepared nanomedicines overcome the poor water solubility of small molecule ferroptosis drugs, improve their bioavailability, and avoid high dosage but low efficiency. Attached Figure Description

[0035] Figure 1 TEM image of LP-CaP@iBEFT prepared in Example 1;

[0036] Figure 2 Silver staining images of LP-CaP@iBEFT and standard lactoferrin prepared in Example 1;

[0037] Figure 3 The image shows the XRD pattern of LP-CaP@iBEFT obtained in Example 1.

[0038] Figure 4 The infrared spectrum of CaP@iBEFT in LP-CaP@iBEFT prepared in Example 1;

[0039] Figure 5 The HPLC chromatogram of the drug in LP-CaP@iBEFT prepared in Example 1 is shown below.

[0040] Figure 6 The release rate of iFSP1 in LP-CaP@iBEFT prepared in Example 1;

[0041] Figure 7 The release rate of Brequinar in LP-CaP@iBEFT prepared in Example 1;

[0042] Figure 8 Erastin release rate in LP-CaP@iBEFT prepared in Example 1;

[0043] Figure 9 The particle size of LP-CaP@iBEFT prepared in Example 1;

[0044] Figure 10 The hydrated particle size distribution of LP-CaP@iBEFT prepared in Example 1 in different solutions is shown.

[0045] Figure 11 This is a diagram illustrating the biosafety of LP-CaP@iBEFT prepared in Example 1.

[0046] Figure 12 This is an in vitro antitumor effect diagram of LP-CaP@iBEFT prepared in Example 1. Detailed Implementation

[0047] The principles and features of the present invention are described below with reference to the accompanying drawings. The examples given are for illustrative purposes only and are not intended to limit the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer should be followed. Reagents or instruments whose manufacturers are not specified are all commercially available products.

[0048] iFSP1 (ferroptosis inhibitor 1 inhibitor), Brequinar, Erastin, FeCl3 (ferric ions), TA (tannic acid)

[0049] The manufacturer of the simulated body fluid is Solarbio.

[0050] Example 1:

[0051] A carrier-free nanomedicine for tumor targeting via calcium phosphate deposition is prepared by the following steps:

[0052] (1) Dissolve 0.5 mg each of small molecule drugs iFSP1, Brequinar, Erastin, FeCl3 and TA in 200 μL of organic solvent dimethyl sulfoxide (DMSO), stir evenly by ultrasonication to obtain reaction solution one, add reaction solution one dropwise to 4 mL of simulated body fluid (SBF) for biomineralization, and then dialyze at 1000 kDa to obtain CaP@iBEFT, i.e. calcium phosphate deposited carrier-free nanomedicine;

[0053] (2) Take a beaker, dissolve 5 mg of lactoferrin (Lf) in 1.5 mL of 4-morpholinoethanesulfonic acid (MES) aqueous solution (concentration of 50 mmol / L, pH of 9), and then add 10.75 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS). Stir magnetically at room temperature for 2 h. Dissolve 10 mg of polyethylene glycol (PEG) in 1.5 mL of 4-morpholinoethanesulfonic acid (MES) to prepare a 4-morpholinoethanesulfonic acid solution of polyethylene glycol. Add it to the beaker and continue to stir magnetically for 24 h. Then dialyze at 1000 kDa to obtain LP, i.e., polyethylene glycol-modified lactoferrin.

[0054] (3) Dissolve 1 mg of calcium phosphate-deposited carrier-free nanomedicine (CaP@iBEFT) obtained in step (1) and 2.5 mg of polyethylene glycol-modified lactoferrin (LP) obtained in step (2) in 5 mL of water, stir magnetically for 24 h, and then dialyze at 1000 kDa to obtain LP-CaP@iBEFT, which is calcium phosphate-deposited carrier-free nanomedicine targeting tumors.

[0055] Example 2:

[0056] A carrier-free nanomedicine for tumor targeting via calcium phosphate deposition is prepared by the following steps:

[0057] (1) Dissolve 2 mg each of small molecule drugs iFSP1, Brequinar, Erastin, FeCl3 and TA in 200 μL of organic solvent dimethyl sulfoxide (DMSO), stir evenly by ultrasonication to obtain reaction solution one, add reaction solution one drop into 4 mL of simulated body fluid (SBF) for biomineralization, and then dialyze at 1000 kDa to obtain CaP@iBEFT, i.e. calcium phosphate deposited carrier-free nanomedicine;

[0058] (2) Take a beaker and dissolve 4 mg of lactoferrin (Lf) in 1 mL of 4-morpholinoethanesulfonic acid (MES) aqueous solution (concentration of 45 mmol / L, pH of 9). Then add 10 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS). Stir magnetically at room temperature for 1 h. Dissolve 10 mg of polyethylene glycol (PEG) in 1 mL of 4-morpholinoethanesulfonic acid (MES) to prepare a 4-morpholinoethanesulfonic acid solution of polyethylene glycol. Add it to the beaker and continue to stir magnetically for 12 h. Then dialyze at 1000 kDa to obtain LP, i.e., polyethylene glycol-modified lactoferrin.

[0059] (3) Dissolve 1 mg of calcium phosphate-deposited carrier-free nanomedicine (CaP@iBEFT) obtained in step (1) and 1 mg of polyethylene glycol-modified lactoferrin (LP) obtained in step (2) in 4 mL of water, stir magnetically for 12 h, and then dialyze at 1000 kDa to obtain LP-CaP@iBEFT, which is calcium phosphate-deposited carrier-free nanomedicine targeting tumors.

[0060] Example 3:

[0061] A carrier-free nanomedicine for tumor targeting via calcium phosphate deposition is prepared by the following steps:

[0062] (1) Dissolve 1 mg each of small molecule drugs iFSP1, Brequinar, Erastin, FeCl3 and TA in 200 μL of organic solvent dimethyl sulfoxide (DMSO), stir evenly by ultrasonication to obtain reaction solution one, add reaction solution one drop into 3 mL of simulated body fluid (SBF) for biomineralization, and then dialyze at 1000 kDa to obtain CaP@iBEFT, i.e. calcium phosphate deposited carrier-free nanomedicine;

[0063] (2) Take a beaker and dissolve 6 mg of lactoferrin (Lf) in 2 mL of 4-morpholinoethanesulfonic acid (MES) aqueous solution (concentration of 55 mmol / L, pH of 9). Then add 11 mg of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS). Stir magnetically at room temperature for 3 h. Dissolve 10 mg of polyethylene glycol (PEG) in 2 mL of 4-morpholinoethanesulfonic acid (MES) to prepare a 4-morpholinoethanesulfonic acid solution of polyethylene glycol. Add it to the beaker and continue to stir magnetically for 20 h. Then dialyze at 1000 kDa to obtain LP, i.e., polyethylene glycol-modified lactoferrin.

[0064] (3) Dissolve 1 mg of calcium phosphate-deposited carrier-free nanomedicine (CaP@iBEFT) obtained in step (1) and 5 mg of polyethylene glycol-modified lactoferrin (LP) obtained in step (2) in 6 mL of water, stir magnetically for 20 h, and then dialyze at 1000 kDa to obtain LP-CaP@iBEFT, which is calcium phosphate-deposited carrier-free nanomedicine targeting tumors.

[0065] Comparative Example 1:

[0066] A rice-based drug, the preparation method of which includes the following steps: the small molecule drug does not contain TA, and the rest is the same as in Example 1.

[0067] Ultimately, it was impossible to produce carrier-free nanomedicines targeting tumors with calcium phosphate deposition.

[0068] Test case

[0069] The following tests were performed on the LP-CaP@iBEFT prepared in Example 1.

[0070] I. The LP-CaP@iBEFT prepared in Example 1 was subjected to TEM, silver staining, and X-ray diffraction analysis. The results are shown in the figure. Figure 1-3 .

[0071] Depend on Figure 1 It is known that the calcium phosphate-deposited carrier-free nanomedicine for targeting tumors prepared in this invention has a distinct core-shell structure, with the calcium phosphate-deposited carrier-free nanomedicine (CaP@iBEFT) as the core and PEG-modified lactoferrin (LP) as the shell. The nanomedicine particle size is about 106 nm, and the protein layer thickness is about 33 nm.

[0072] Depend on Figure 2 It can be seen that (1 is Lf, 2 is LP-CaP@iBEFT), LP-CaP@iBEFT and standard Lf show protein bands at the same positions, indicating that LP was successfully loaded;

[0073] Depend on Figure 3 It can be seen that LP-CaP@iBEFT has the crystal angle of CaP, indicating that CaP has been successfully mineralized.

[0074] II. The LP-CaP@iBEFT prepared in Example 1 was subjected to infrared spectroscopy detection, and the results are shown in the figure. Figure 4 .Depend on Figure 4It is evident that the calcium phosphate-deposited, carrier-free tumor-targeting nanomedicine (LP-CaP@iBEFT) prepared in this invention exhibits absorption peaks of characteristic functional groups iFSP1, Brequinar, Erastin, FeCl3, and TA, indicating the successful loading of these drugs. Simultaneously, the presence of iron can be confirmed by determining the iron content using inductively coupled plasma optical emission spectrometry (ICP-OES), further demonstrating the successful loading of FeCl3.

[0075] III. The LP-CaP@iBEFT prepared in Example 1 was lyophilized into powder, reconstituted with methanol, and the contents of iFSP1, Brequinar, and Erastin in the carrier-free nanomedicine were determined under the same chromatographic analysis conditions (see Table 1). The results are shown in Table 1. Figure 5 .Depend on Figure 5 It can be seen that iFSP1, Brequinar, and Erastin were successfully loaded;

[0076] Table 1 HPLC conditions

[0077]

[0078] Another sample of LP-CaP@iBEFT prepared in Example 1 was digested overnight at 70°C, diluted with primary water, and the iron content in the carrier-free nanomedicine was determined by ICP-OES, confirming the successful loading of FeCl3. The encapsulation efficiency and drug loading of iFSP1, Brequinar, Erastin, and FeCl3 in the carrier-free nanomedicine are shown in Table 2.

[0079] Table 2 Encapsulation efficiency and drug loading

[0080]

[0081] IV. pH-responsive release characteristics of CaP

[0082] The LP-CaP@iBEFT prepared in Example 1 was prepared into an aqueous solution. 1 mL of the sample was placed in a 3500 Mw dialysis bag, and then the dialysis bag was immersed in different pH environments (pH = 7.4, simulating the normal physiological environment; pH = 6.5, simulating the tumor microenvironment; pH = 5.0, simulating the lysosomal environment) and shaken at 37°C. The contents of iFSP1, Brequinar, and Erastin in 1 mL of buffer solution outside the dialysis bag at different time intervals were measured by HPLC. The results are shown in [Figure number missing]. Figure 6-8 .

[0083] Depend on Figure 6-8 It can be seen that within the same time interval, the release of iFSP1, Brequinar, and Erastin increases as the pH value decreases.

[0084] Simultaneously, the particle size variation of carrier-free nanomedicines under different pH conditions was determined using a Malvern laser particle size analyzer. The results are shown in [Figure number missing]. Figure 9 .

[0085] Depend on Figure 9 It can be seen that as pH decreases, the mineralized framework of CaP gradually disintegrates, water and particle size decrease, and the material gradually becomes heterogeneous.

[0086] V. Stability of carrier-free nanomedicines

[0087] The LP-CaP@iBEFT prepared in Example 1 was used to prepare carrier-free nanomedicine solutions with primary aqueous solution, PBS solution, and DMEM solution, respectively. Water content and particle size were continuously measured at 1, 3, 5, 7, 9, 11, 13, and 15 days. The results are shown in [Figure number missing]. Figure 10 .

[0088] Depend on Figure 10 It can be seen that the water content and particle size of the carrier-free nanomedicine prepared by the present invention did not change significantly in different buffer solutions within 15 days, indicating that the carrier-free nanomedicine prepared by the present invention has good stability.

[0089] VI. Biosafety of Carrier-Free Nanomedicines

[0090] HUVEC cells (human umbilical vein endothelial cells) were inoculated into 96-well plates, with each well containing approximately 4 × 10⁶ cells. 3 Cells were cultured for 24 h and then injected with LP-CaP@iBEFT at concentrations of 0, 15.625, 31.25, 62.5, 125, 250, 500, and 1000 μg / mL. After 24 h of culture, the cells were washed with PBS, and CCK-8 assay solution (v / v: 10%) without fetal bovine serum (FBS) was added. Cells were then cultured at 37°C for another 2.5 h before detection. Cell viability was measured at 450 nm using an enzyme-linked immunosorbent assay (ELISA) analyzer. Results are shown below. Figure 11 .

[0091] Depend on Figure 11 It can be seen that LP-CaP@iBEFT has no obvious killing effect on HUVEC cells, indicating that the carrier-free nanomedicine prepared in this invention is non-toxic to normal cells and has biosafety.

[0092] VII. Antitumor Effects of Carrier-Free Nanomedicines

[0093] 4T1 cells (mouse breast cancer cells) were inoculated into 96-well plates, so that each well contained approximately 4 × 10⁶ cells. 3Cells were cultured for 24 h and then injected with LP-CaP@iBEFT at concentrations of 0, 15.625, 31.25, 62.5, 125, 250, 500, and 1000 μg / mL. After 24 h of culture, the cells were washed with PBS, and CCK-8 assay solution (v / v: 10%) without fetal bovine serum (FBS) was added. Cells were then cultured at 37°C for another 2.5 h before detection. Cell viability was measured at 450 nm using an enzyme-linked immunosorbent assay (ELISA) analyzer. Results are shown below. Figure 12 .

[0094] Depend on Figure 12 It can be seen that LP-CaP@iBEFT has a significant killing effect on 4T1 cells, indicating that the carrier-free nanomedicine prepared in this invention is toxic to tumor cells and has an anti-tumor effect.

[0095] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for preparing a calcium phosphate-deposited, tumor-targeting, carrier-free nanodrug, characterized by, The method comprises the following steps: (1) dissolving a small molecule drug in an organic solvent, stirring to obtain a reaction solution I, adding the reaction solution I dropwise into a simulated body fluid, carrying out biomimetic mineralization, and then dialysis to obtain CaP@iBEFT, i.e. calcium phosphate deposited carrier-free nanodrug; (2) dissolving lactoferrin in 4-morpholine ethanesulfonic acid aqueous solution, then adding 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide and N-hydroxysuccinimide, stirring for 1-3 h, then adding 4-morpholine ethanesulfonic acid solution of polyethylene glycol, continuing to stir for 12-24 h, and then dialysis to obtain LP, i.e. pegylated lactoferrin; (3) dissolving the calcium phosphate deposited carrier-free nanodrug prepared in step (1) and the pegylated lactoferrin prepared in step (2) in water, stirring for 12-24 h, and then dialysis to obtain LP-CaP@iBEFT, i.e. calcium phosphate deposited tumor-targeted carrier-free nanodrug; In step (1), the small molecule drug is ferroptosis inhibitor 1 inhibitor, buquinolate, alarisatin, iron ion and tannic acid.

2. The method for preparing carrier-free tumor-targeting nanomedicines by calcium phosphate deposition according to claim 1, characterized in that, The mass-volume ratio of ferroptosis inhibitor 1 inhibitor, buquinolate, alarisatin, iron ion, tannic acid and organic solvent is 0.5-2 mg:0.5-2 mg:0.5-2 mg:0.5-2 mg:0.5-2 mg:200 μL.

3. The method for preparing carrier-free tumor-targeting nanomedicines by calcium phosphate deposition according to claim 1, characterized in that, In step (1), the volume ratio of organic solvent and simulated body fluid is 1:15-20.

4. The method for preparing carrier-free tumor-targeting nanomedicines by calcium phosphate deposition according to claim 1, characterized in that, In step (2), the mass-volume ratio of total amount of lactoferrin, 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide and N-hydroxysuccinimide, 4-morpholine ethanesulfonic acid aqueous solution is 4-6 mg:10-11 mg:1-2 mL.

5. The method for preparing carrier-free tumor-targeting nanomedicines by calcium phosphate deposition according to claim 1, characterized in that, In step (2), the mass-volume ratio of lactoferrin and 4-morpholine ethanesulfonic acid solution of polyethylene glycol is 4-6 mg:1-2 mL.

6. The method of claim 1 or 5, wherein the calcium phosphate deposited tumor-targeting nano-carrier-free nanodrug is prepared by the following steps of: (a) mixing a calcium phosphate compound and a tumor-targeting ligand to form a mixture; (b) adding a drug to the mixture; (c) adding a solvent to the mixture; and (d) drying the mixture. In step (2), the 4-morpholine ethanesulfonic acid solution of polyethylene glycol is prepared by the following method: dissolving 10 mg of polyethylene glycol in 1-2 mL of 4-morpholine ethanesulfonic acid to obtain a 4-morpholine ethanesulfonic acid solution of polyethylene glycol.

7. The method for preparing carrier-free tumor-targeting nanomedicines by calcium phosphate deposition according to claim 1, characterized in that, In step (3), the mass-volume ratio of calcium phosphate deposited carrier-free nanodrug, pegylated lactoferrin and water is 1 mg:1-5 mg:4-6 mL.

8. The calcium phosphate deposited tumor-targeted carrier-free nanodrug prepared by the method according to any one of claims 1-7.

9. The use of the calcium phosphate deposited tumor-targeted carrier-free nanodrug according to claim 8 in the preparation of breast cancer targeted tumor drugs.

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