ROS responsive R848 prodrug as well as preparation method and application thereof

Abstract

The invention provides an ROS responsive R848 prodrug as well as a preparation method and application thereof, and belongs to the technical field of biological medicines. The active oxygen responsive R848 prodrug comprises R848, a maleimide group and an active oxygen responsive thioketal bond connecting the R848 and the maleimide group, the prodrug can be coupled to an albumin molecule through the design, then the prodrug and a photosensitizer Ce6 are self-assembled to form albumin nanoparticles, and after the nanoparticles reach a tumor site, the nanoparticles are self-assembled to form the albumin nanoparticles. When laser irradiates a tumor site, high-concentration active oxygen is locally generated through a photosensitizer to kill tumor cells, meanwhile, active oxygen responsive chemical bonds are broken to release R848 to act on immune cells of the tumor site, photodynamic therapy and immunotherapy are achieved, anti-tumor activity is shown, and the compound has important significance on tumor combined therapy.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, and in particular relates to a ROS-responsive R848 prodrug, its preparation method and application. Background Technology

[0002] R848 (Resiquimod), a small-molecule immune adjuvant, belongs to the imidazoquinoline class of compounds and is an agonist of Toll-like receptors 7 and 8 (TLR7 / 8). Toll-like receptors 7 and 8 (TLR7 / 8) are strongly expressed in the endosomes of immune cells infiltrating the tumor microenvironment. By delivering TLR7 / 8 receptor agonists to the tumor site, immunosuppressive cells in the tumor microenvironment can be regulated, promoting their transformation into antigen-presenting cells, thereby modulating the anti-tumor immune response—a potentially effective tumor immunotherapy. Photodynamic therapy (PDT) utilizes the ability of photosensitizers to generate reactive oxygen species to kill tumor cells. It offers advantages such as non-invasiveness, precise control, good selectivity for tumor sites, minimal impact on healthy tissues, and repeatable treatments, making it a promising new cancer treatment. Numerous studies have shown that PDT can induce immunogenic death in tumor cells, triggering an antigen-specific immune response against tumor cells by releasing tumor-associated antigens and damage-associated molecular patterns. Combining photodynamic therapy with immunotherapy can further enhance tumor immunogenicity and improve the efficacy of tumor treatment.

[0003] As an immune adjuvant, R848, when used alone, cannot fully exert its immune-activating effect, making it difficult to achieve significant tumor treatment efficacy. Furthermore, it suffers from poor water solubility and low bioavailability, and systemic administration may trigger severe inflammatory responses. Although several nanodelivery systems for R848 have been reported, most are made of polymeric materials, and a reactive oxygen species-responsive delivery system based on albumin molecules has not yet been developed. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention proposes a ROS-responsive R848 prodrug, its preparation method, and its application.

[0005] To achieve the above objectives, the present invention provides the following technical solution:

[0006] This invention proposes a ROS-responsive R848 prodrug (R-TK-Mal), comprising R848 and maleimide groups (Mal), and a reactive oxygen species-responsive thioclase bond (TK bond) connecting the two parts, with the specific structural formula as follows:

[0007] .

[0008] This invention also proposes a method for preparing the above-mentioned ROS-responsive R848 prodrug. Under the action of triphosgene, R848 is linked to 2,2'-(propane-2,2-diylbis(thionyl))diethanol via a carbonate bond to obtain an intermediate product. Then, the intermediate product is linked to 4-maleimide butyric acid via an esterification reaction to generate the ROS-responsive R848 prodrug. Specifically, the method includes the following steps:

[0009] (1) R848 was dissolved in dichloromethane, and triphosgene was added. The mixture was stirred under a nitrogen atmosphere, followed by the addition of 2,2'-(propane-2,2-diylbis(thionyl))diethanol. The mixture was stirred continuously, and the resulting mixture was concentrated and purified to obtain a white solid compound with the following structural formula: ;

[0010] (2) The white solid compound was dissolved in dimethylformamide, and then 4-maleimide butyric acid, benzotriazole-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate (BOP) and N,N-diisopropylethylamine (DIEA) were added. The crude product was obtained by stirring at room temperature and then purified to obtain the ROS-responsive R848 prodrug.

[0011] Further, in step (1), the mass ratio of R848 to 2,2'-(propane-2,2-dimethylbis(thionyl))diethanol is 11:10, and the dosage ratio of R848, dichloromethane and triphosgene is 55mg:5mL:37mg.

[0012] Further, in step (2), the mass ratio of the white solid compound, 4-maleimide butyric acid, BOP and DIEA is 22:20:24:7, and the amount ratio of the white solid compound to dimethylformamide is 20mg:1mL.

[0013] This invention also proposes an anti-tumor reactive oxygen species-responsive albumin nanoparticle, the active ingredients of which include the aforementioned ROS-responsive R848 prodrug and the photosensitizer Ce6 (Chlorin e6, dihydroporphyrin e6, is a second-generation porphyrin photosensitizer derived from chlorophyll degradation, and is also one of the most commonly used model molecules in current photodynamic therapy (PDT) and photoimmunotherapy research, with the molecular formula C). 34 H 36 N4O6).

[0014] Furthermore, the mass ratio of the ROS-responsive R848 prodrug to the photosensitizer Ce6 is 8:3.

[0015] Furthermore, the preparation method of the antitumor reactive oxygen species responsive albumin nanoparticles includes the following steps:

[0016] Heme-modified bovine serum albumin and tris(2-carboxyethyl)phosphonic acid hydrochloride (TCEP) were dissolved in ultrapure water, stirred at room temperature, and dialyzed to obtain a reduced albumin solution. The ROS-responsive R848 prodrug and photosensitizer Ce6 were dissolved in dimethyl sulfoxide and added dropwise to the reduced albumin solution. After stirring and ultrafiltration, the antitumor reactive oxygen species-responsive albumin nanoparticles were obtained. The antitumor reactive oxygen species-responsive albumin nanoparticles of this invention can co-load R848 prodrug and photosensitizer Ce6, increasing the accumulation of both drugs at the tumor site. After laser irradiation, the photosensitizer generates high concentrations of singlet oxygen to kill tumor cells, while simultaneously breaking reactive oxygen species-responsive chemical bonds, thereby releasing R848 and achieving photodynamic therapy and immunotherapy, exhibiting antitumor activity.

[0017] The present invention also proposes the application of the above-mentioned ROS-responsive R848 prodrug or antitumor reactive oxygen species-responsive albumin nanoparticles in the preparation of drugs to inhibit tumor growth.

[0018] Compared with the prior art, the present invention has the following advantages and technical effects:

[0019] This invention provides a reactive oxygen species (ROS)-responsive R848 prodrug, comprising R848 and maleimide groups, and an ROS-responsive thioclase bond connecting the two parts. The design of this invention allows the R848 prodrug to be coupled to an albumin molecule, which is then self-assembled with a photosensitizer Ce6 to form albumin nanoparticles. Upon reaching the tumor site, the nanoparticles, when irradiated by laser, generate a high concentration of ROS locally through the photosensitizer, killing tumor cells. Simultaneously, the ROS-responsive chemical bond is broken, releasing R848, which acts on immune cells at the tumor site, achieving photodynamic therapy and immunotherapy, and exhibiting anti-tumor activity. This has significant implications for combined tumor treatment. Attached Figure Description

[0020] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. In the drawings:

[0021] Figure 1 This is a synthetic route diagram of the ROS-responsive R848 prodrug of the present invention;

[0022] Figure 2 The 1H NMR spectrum of product R-TK-Mal from Example 1;

[0023] Figure 3 The results of MALDI-TOF-MS testing of HBSA prepared in Example 2 are shown.

[0024] Figure 4The results of particle size potential of CR@HBSA prepared in Example 2 are shown on the left, where the particle size distribution results are on the left and the potential and PDI test results are on the right.

[0025] Figure 5 The UV absorption spectra of the two free drugs (R-TK-Mal, Ce6) and CR@HBSA prepared in Example 2;

[0026] Figure 6 The graph shows the singlet oxygen generation capacity of DPBF mother liquor, Ce6 photosensitizer system, and CR@HBSA system.

[0027] Figure 7 The cumulative drug release curves of R848 in the light-exposed group (CR@HBSA(+)) and the non-light-exposed group (CR@HBSA) are shown.

[0028] Figure 8 The results of cytotoxicity experiments on 4T1 cells by CR@HBSA and photosensitizer Ce6 in the light-illuminated and non-light-illuminated groups are shown.

[0029] Figure 9 The growth curves of tumor volume in 4T1 tumor-bearing mice in different treatment groups are shown. Detailed Implementation

[0030] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0031] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0032] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0033] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be apparent to those skilled in the art. This specification and embodiments are merely exemplary.

[0034] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0035] This invention provides a ROS-responsive R848 prodrug (R-TK-Mal), comprising R848 and maleimide groups (Mal), and a reactive oxygen species-responsive thioclase bond (TK bond) connecting the two parts. The specific structural formula is as follows:

[0036] .

[0037] This invention also proposes a method for preparing the above-mentioned ROS-responsive R848 prodrug. Under the action of triphosgene, R848 is linked to 2,2'-(propane-2,2-diylbis(thionyl))diethanol via a carbonate bond to obtain an intermediate product. Then, the intermediate product is linked to 4-maleimide butyric acid via an esterification reaction to generate the ROS-responsive R848 prodrug. The specific steps include:

[0038] (1) Dissolve R848 in dichloromethane and add triphosgene. Stir under a nitrogen atmosphere, then add 2,2'-(propane-2,2-diylbis(thionyl))diethanol and continue stirring. Concentrate and purify the resulting mixture to obtain a white solid compound.

[0039] (2) Dissolve the white solid compound in dimethylformamide, then add 4-maleimide butyric acid, benzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate (BOP) and N,N-diisopropylethylamine (DIEA), and stir at room temperature to obtain the crude product. After purification, the ROS-responsive R848 prodrug is obtained.

[0040] The structural formula of R848 is as follows: ;

[0041] The structural formula of 2,2'-(propane-2,2-diylbis(thionidyl))diethanol is: ;

[0042] The structural formula of the white solid compound is: ;

[0043] The structural formula of 4-maleimidebutyric acid is: .

[0044] An exemplary method for preparing a ROS-responsive R848 prodrug is as follows:

[0045] (1) Accurately weigh 110 mg of R848 and dissolve it in 10 mL of dichloromethane (DCM). Add 74 mg of triphosgene (bis(trichloromethyl) carbonate) and stir for 1 h at 0 °C under a nitrogen atmosphere. Then add 100 mg of 2,2'-(propane-2,2-diylbis(thionyl))diethanol and continue stirring for 1 h. Concentrate the mixture and purify it by reverse-phase column chromatography (C18 column, in the first minute of elution, the proportion of acetonitrile is increased from 5% to 95%; then maintain the proportion and elute for 1 minute. The mobile phase contains 0.01 wt% trifluoroacetic acid) to obtain a white solid compound 1.

[0046] (2) Accurately weigh 100 mg of white solid compound 1, dissolve it in 5 mL of dimethylformamide (DMF), then add 110 mg of 4-maleimide butyric acid, 120 mg of BOP and 35 mg of DIEA, stir at room temperature for 2 h, and purify the crude product by reversed-phase column chromatography (C18 column, in the first 8 minutes of elution, the proportion of acetonitrile is increased from 5% to 95%; then maintain the proportion for 7 minutes of elution. The mobile phase contains 0.01 wt% trifluoroacetic acid) to obtain the ROS-responsive R848 prodrug (R-TK-Mal).

[0047] See the detailed synthesis route diagram. Figure 1 .

[0048] This invention also proposes an antitumor reactive oxygen species responsive albumin nanoparticle (CR@HBSA), the active ingredients of which include the above-mentioned ROS-responsive R848 prodrug and photosensitizer Ce6.

[0049] In a preferred embodiment of the present invention, the mass ratio of ROS-responsive R848 prodrug to photosensitizer Ce6 is 8:3.

[0050] In a preferred embodiment of the present invention, the preparation method of antitumor reactive oxygen species responsive albumin nanoparticles includes the following steps:

[0051] Heme-modified bovine serum albumin and TCEP were dissolved in ultrapure water, stirred at room temperature, and dialyzed to obtain a reduced albumin solution. ROS-responsive R848 prodrug and photosensitizer Ce6 were dissolved in dimethyl sulfoxide and added dropwise to the reduced albumin solution. After stirring and ultrafiltration, antitumor reactive oxygen species-responsive albumin nanoparticles were obtained. The antitumor reactive oxygen species-responsive albumin nanoparticles of this invention can co-load R848 prodrug and photosensitizer Ce6, increasing the accumulation of both drugs at the tumor site. After laser irradiation, the photosensitizer generates high concentrations of singlet oxygen to kill tumor cells, while simultaneously breaking reactive oxygen species-responsive chemical bonds, thereby releasing R848 and achieving photodynamic therapy and immunotherapy, exhibiting antitumor activity.

[0052] This invention also proposes the application of the above-mentioned ROS-responsive R848 prodrug or antitumor reactive oxygen species-responsive albumin nanoparticles in the preparation of drugs to inhibit tumor growth.

[0053] In all embodiments of the present invention, the room temperature is "25±2℃".

[0054] All raw materials used in the embodiments of this invention were purchased commercially available. R848 was purchased from MedChemExpress, photosensitizer Ce6 was purchased from Shanghai Yuanye Biotechnology Co., Ltd., heme was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd., and bovine serum albumin was purchased from Sigma-Aldrich.

[0055] The technical solution of the present invention will be further illustrated by the following embodiments.

[0056] Example 1

[0057] A method for preparing a ROS-responsive R848 prodrug includes the following steps:

[0058] (1) Accurately weigh 110 mg of R848 and dissolve it in 10 mL of DCM. Add 74 mg of triphosgene and stir for 1 h at 0 °C under a nitrogen atmosphere. Then add 100 mg of 2,2'-(propane-2,2-diylbis(thioalkyldiyl))diethanol and continue stirring for 1 h. Concentrate the mixture and purify it by reverse-phase column chromatography (C18 column, in the first minute of elution, the proportion of acetonitrile is increased from 5% to 95%; then the proportion is maintained for elution for 1 minute. The mobile phase contains 0.01 wt% trifluoroacetic acid) to obtain white solid compound 1.

[0059] (2) Accurately weigh 100 mg of white solid compound 1, dissolve it in 5 mL of dimethylformamide, then add 110 mg of 4-maleimide butyric acid, 120 mg of BOP and 35 mg of DIEA, stir at room temperature for 2 h, and purify the crude product by reversed-phase column chromatography (C18 column, in the first 8 minutes of elution, the proportion of acetonitrile is increased from 5% to 95%; then maintain the proportion for 7 minutes of elution. The mobile phase contains 0.01 wt% trifluoroacetic acid) to obtain the ROS-responsive R848 prodrug (R-TK-Mal).

[0060] The 1H NMR spectrum of product R-TK-Mal from Example 1 is shown below. Figure 2 The 1H NMR data are as follows: 1H NMR (400 MHz, DMSO-d6) δ 8.65 (d, J = 4.0 Hz 1H), 8.14 (d, J = 4.0 Hz 1H), 7.76-7.74 (m, 1H), 7.66-7.65 (m, 1H), 6.98 (s, 2H), 4.81 (brs, 3H), 4.37 (t, J = 6.8 Hz, 2H), 4.15 (t, J = 6.8 Hz, 1H), 3.70-3.67 (m, 5H), 3.41 (t, J = 6.8 Hz, 3H), 2.94 (t, J = 6.8 Hz, 2H), 2.84 (t, J = 6.8 Hz, 2H). 2H), 2.29 (t, J = 6.8Hz, 2H), 1.74-1.71 (m, 2H), 1.60 (s, 6H), 1.23-1.10 (m, 9H).

[0061] Example 2

[0062] A method for preparing antitumor reactive oxygen species responsive albumin nanoparticles (CR@HBSA) includes the following steps:

[0063] (1) Preparation of heme-modified bovine serum albumin: 4 mg of hemin (heme chloride) was dissolved in 4 mL of 5 mM NaOH solution, and then 200 μL of a solution containing 19 mg of EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide) and 17 mg of NHS (N-hydroxysuccinimide) was added. The mixture was stirred at room temperature for 1 h. Subsequently, 4 mL of bovine serum albumin solution with a concentration of 40 mg / mL was added, and the mixture was heated and stirred at 37 °C for 12 h. The mixture was dialyzed for 48 h to obtain heme-modified bovine serum albumin (HBSA).

[0064] The successful preparation of HBSA was verified by MALDI-TOF-MS, and the results are shown in [Figure number missing]. Figure 3 It can be seen that the charge-to-weight ratio of heme-modified bovine serum albumin is significantly higher than that of bovine serum albumin, indicating that heme has been successfully covalently bound to bovine serum albumin.

[0065] (2) 10 mg of heme-modified bovine serum albumin and 6 mg of TCEP were dissolved in 2 mL of ultrapure water and stirred at room temperature for 1 h. The reaction solution was dialyzed in ultrapure water for 24 h to obtain a reduced albumin solution. Then, 1.6 mg of R-TK-Mal prepared in Example 1 and 0.6 mg of photosensitizer Ce6 were dissolved in 200 μL of DMSO and added dropwise to the reduced albumin solution. The mixture was stirred for 1 h and ultrafiltered 3 times to remove free drugs to obtain CR@HBSA.

[0066] Performance testing

[0067] (1) Particle size characterization

[0068] Particle size, potential, and polydispersity index (PDI) were determined using a laser particle size analyzer. The particle size and potential results of the CR@HBSA prepared in Example 2 are shown below. Figure 4 The left side shows the particle size distribution results, and the right side shows the potential and PDI test results. It can be seen that the hydrated particle size of CR@HBSA is 62.5 ± 2.2 nm, the particle size distribution is uniform, and the particles are electronegative.

[0069] (2) Ultraviolet absorption spectrum

[0070] 1 mg of R-TK-Mal prepared in Example 1 and 0.5 mg of photosensitizer Ce6 (hereinafter referred to as Ce6) were respectively dissolved in 1 mL of DMSO to prepare drug stock solutions, which were then diluted 20 times with ultrapure water. The UV absorption spectra of the two free drugs (R-TK-Mal, Ce6) and CR@HBSA prepared in Example 2 were measured. The results are shown in the figure. Figure 5 It can be seen that CR@HBSA has the same characteristic peaks as Ce6 and R-TK-Mal, indicating that the two drugs were successfully loaded.

[0071] (4) In vitro singlet oxygen generation capacity

[0072] Using DPBF (1,3-diphenylisobenzofuran) as a singlet oxygen probe, 120 μL of DPBF stock solution (10 μM) was added to 20 mL of PBS solution, followed by the addition of photosensitizer Ce6 or CR@HBSA from Example 3, to achieve a concentration of 2 μg / mL. A 660 nm laser (100 mW / cm²) was then used. 2After irradiating DPBF mother liquor, Ce6 photosensitizer system (Ce6(+)), and CR@HBSA system (CR@HBSA(+)) for different times, the UV absorption spectra from 350 to 550 nm were recorded. The rate of change of absorbance at 410 nm was used to reflect the ability to generate singlet oxygen. The results are shown in [Figure number missing]. Figure 6 It can be seen that CR@HBSA can generate a large amount of singlet oxygen after laser irradiation.

[0073] (5) Reactive oxygen species release capacity

[0074] The CR@HBSA samples from Example 2 were divided into an illuminated group (CR@HBSA(+)) and a non-illuminated group (CR@HBSA), with three parallel samples in each group. The illuminated group was subjected to 660 nm laser irradiation (100 mW / cm²). 2 CR@HBSA solution was placed in a dialysis bag (MWCO = 3.5 kDa) and immersed in 10 mL of PBS solution (containing 0.5% Tween 80) for 5 min. The bag was placed in a 37℃ constant temperature shaker. At fixed time points, 2.5 mL of the solution outside the dialysis bag was taken and an equal volume of PBS solution (containing 0.5% Tween 80) was added as release medium. The OD value at 320 nm was measured using a UV spectrophotometer, and the cumulative drug release of R848 was calculated. The cumulative drug release of R848 in the light-exposed group (CR@HBSA(+)) and the non-light-exposed group (CR@HBSA) were compared. The results are shown in the figure. Figure 7 It can be seen that CR@HBSA generates a large amount of reactive oxygen species under laser irradiation. The reactive oxygen species break the thioclase bond and release R848, achieving a release rate of nearly 80% after 48 hours.

[0075] (6) Evaluation of in vitro and in vivo antitumor effects

[0076] The in vitro and in vivo antitumor effects of free drug and reactive oxygen species responsive albumin nanoparticles (CR@HBSA) co-loaded with R-TK-Mal and photosensitizer Ce6 (Example 2) were evaluated. The specific procedures are as follows:

[0077] 4T1 mammary cancer cells from mice were obtained from the Cell Resource Center of the Shanghai Institute of Biotechnology, Chinese Academy of Sciences. Female BALB / c nude mice (4-5 weeks old) were purchased from Spiefer Beijing Biotechnology Co., Ltd. 1×10-1 cells were injected into the lower right back of female BALB / c mice (4-5 weeks old). 6 A mouse model of subcutaneous breast cancer was established using 4T1 cells.

[0078] Cytotoxicity experiment procedure:

[0079] 4T1 cells were seeded in 96-well plates at 5 × 10⁶ cells per well. 3Individual samples were cultured for 12 h and then divided into four treatment groups: Ce6, Ce6+L, CR@HBSA, and CR@HBSA+L. After 4 h of incubation, the light-treated group was irradiated with a 660 nm laser (100 mW / cm²). 2 Cells were cultured for 24 h (2 min). Cell viability was then measured using a CCK-8 assay kit. The cytotoxicity results of CR@HBSA and photosensitizer Ce6 on 4T1 cells in the light-treated and non-light-treated groups are shown below. Figure 8 As can be seen, CR@HBSA exhibits enhanced cytotoxicity compared to free drug.

[0080] 4-5 week old female BALB / c mice were injected with 1×10⁻⁶ scalp irradiation on the right back. 6 4T1 cells were used when the mouse tumor volume reached 100 mm. 3 At approximately 10:00 PM, participants were randomly divided into 5 groups, with 5 biological replicates per group. The groups were: I (PBS), II (Ce6+L), III (R-TK-Mal), IV (CR+L), and V (CR@HBSA+L), with II, IV, and V being the light exposure groups. Administered intravenously on days 0, 5, and 10. PBS was administered in 150 μL volumes. The doses of CR and CR@HBSA were the same as the free drug groups, with Ce6 at 2 mg / kg and R-TK-Mal at 6 mg / kg. The light exposure groups received 660 nm laser irradiation (100 mW / cm²) 6 h after administration. 2 (10 min). Measure the length and width of the tumor every two days using calipers and calculate the tumor volume using the formula: Tumor volume = 1 / 2 * length * width. 2 .

[0081] The growth curves of tumor volume in 4T1 tumor-bearing mice in different treatment groups are shown in the figure. Figure 9 It can be seen that both the free drug R-TK-Mal and the Ce6 light irradiation group showed certain anti-tumor effects. The combination of the two drugs can further enhance the anti-tumor effect. This is because reactive oxygen species responsive albumin nanoparticles can increase the accumulation of drugs in tumors and show the strongest anti-tumor effect.

[0082] The above are merely preferred embodiments of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A ROS-responsive R848 prodrug, characterized in that, The structural formula is: 。 2. A method of preparing the ROS-responsive R848 prodrug of claim 1, wherein, The intermediate product is obtained by connecting R848 and 2,2'-(propane-2,2-diylbis(sulfanediyl))diethanol through a carbonate bond under the action of triphosgene, and then the intermediate product is connected with 4-maleimide butyric acid through an esterification reaction to generate the ROS-responsive R848 prodrug.

3. The method of claim 2, wherein the ROS-responsive R848 prodrug is prepared by, The method comprises the following steps: (1) R848 was dissolved in dichloromethane and added with triphosgene under stirring in a nitrogen atmosphere, then 2,2'-(propane-2,2-diylbis(sulfanediyl))diethanol was added and stirring was continued, the resulting mixture was concentrated and purified to obtain a white solid compound, i.e. an intermediate product, of formula ; (2) The intermediate product is dissolved in dimethylformamide, 4-maleimide butyric acid, benzotriazol-1-yloxytris(dimethylamino) phosphonium hexafluorophosphate and N,N-diisopropylethylamine are added, and the mixture is stirred at room temperature to obtain a crude product, and the ROS-responsive R848 prodrug is obtained after purification.

4. The method of claim 3, wherein the ROS-responsive R848 prodrug is prepared by, In step (1), the mass ratio of the R848 and 2,2'-(propane-2,2-diylbis(sulfanediyl))diethanol is 11:10, and the amount ratio of the R848, dichloromethane and triphosgene is 55 mg:5 mL:37 mg.

5. The method of claim 3, wherein the ROS-responsive R848 prodrug is prepared by the method comprising the steps of: In step (2), the mass ratio of the intermediate product, 4-maleimide butyric acid, benzotriazol-1-yloxytris(dimethylamino) phosphonium hexafluorophosphate and N,N-diisopropylethylamine is 22:20:24:7, and the amount ratio of the white solid compound and dimethylformamide is 20 mg:1 mL.

6. An antitumor active oxygen-responsive albumin nanoparticle, characterized by, The active ingredient comprises the ROS-responsive R848 prodrug of claim 1 and the photosensitizer Ce6.

7. The antitumor active oxygen-responsive albumin nanoparticle according to claim 6, wherein, The mass ratio of the ROS-responsive R848 prodrug and the photosensitizer Ce6 is 8:

3.

8. The antitumor active oxygen-responsive albumin nanoparticle according to claim 6, wherein, The preparation method of the anti-tumor active oxygen-responsive albumin nanoparticle comprises the following steps: Heme-modified bovine serum albumin and tris(2-carboxyethyl)phosphine are dissolved in ultrapure water, and after stirring at room temperature, a reduced albumin solution is obtained by dialysis; the ROS-responsive R848 prodrug and the photosensitizer Ce6 are dissolved in dimethyl sulfoxide, and then added dropwise into the reduced albumin solution; the mixture is stirred and ultrafiltrated to obtain the anti-tumor active oxygen-responsive albumin nanoparticle.

9. Use of the ROS-responsive R848 prodrug of claim 1 or the anti-tumor active oxygen-responsive albumin nanoparticle of any one of claims 6-8 in the preparation of a drug for inhibiting tumor growth.

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