1,8-naphthalimide-based photosensitizer and its phototherapy applications

Abstract

The application discloses a photosensitizer based on 1,8-naphthalimide and phototherapy application thereof, which is synthesized from 1,8-naphthalimide and acetylpyrrole as raw materials, through phosphorus oxychloride condensation, and then fluoroboration, and the synthesis method is very simple, and separation and purification are convenient. The compound has long-wavelength absorption characteristics due to its extended conjugated structure, and the optical characteristics enable the compound to be effectively applied to the field of photodynamic therapy.

Description

Technical Field

[0001] This invention relates to a photosensitizer based on 1,8-naphtholactamide, and more specifically, to a photosensitizer based on 1,8-naphtholactamide and its phototherapy application. Background Technology

[0002] Photodynamic therapy (PDT) is a therapeutic technique that uses photosensitizers to generate reactive oxygen species (ROS) upon excitation by light of a specific wavelength to kill tumor cells. This technology offers advantages such as high selectivity, minimal invasiveness, and repeatable treatments, showing great promise for applications in cancer therapy. However, existing photosensitizers still face many challenges in practical applications, such as insufficient light penetration depth, poor targeting to tumor tissue, and low singlet oxygen quantum yield, which limit their therapeutic efficacy.

[0003] 1,8-Naphtholactamimides have attracted widespread attention in the field of photosensitizers due to their excellent photophysicochemical properties. These compounds possess large conjugated systems, allowing for the modulation of absorption wavelengths through molecular structure modification; they also exhibit high singlet oxygen quantum yields and good photostability. In recent years, researchers have developed a series of 1,8-naphtholactamimide derivatives by introducing different functional groups, significantly enhancing their photodynamic therapy effects. For example, by introducing an electron donor-acceptor structure onto the 1,8-naphtholactamimide backbone, a redshift in absorption wavelength and an improvement in ROS generation efficiency were achieved.

[0004] However, existing 1,8-naphtholactam photosensitizers still have some problems that need to be solved: (1) some compounds have poor water solubility, which affects their transport and distribution in the body; (2) their stability in the tumor microenvironment needs to be improved. Therefore, developing novel 1,8-naphtholactam photosensitizers with good water solubility and excellent photostability is of great significance for promoting the clinical application of photodynamic therapy. Summary of the Invention

[0005] The main objective of this invention is to provide a photosensitizer based on 1,8-naphthoylimide and its phototherapy application. The technical solution of this invention is as follows:

[0006] A photosensitizer based on 1,8-naphtholide and its phototherapy application, wherein the chemical structural formula of the compound is as follows:

[0007]

[0008] The substituent R is selected from any one of chlorine, methoxy, and ethoxy. The chemical structural formula of the compound is:

[0009] Any one of them.

[0010] The method for synthesizing the photosensitizer based on 1,8-naphtholide includes the following synthetic route:

[0011]

[0012] The method includes the following steps:

[0013] (1) Add 1,8-naphthoimide, acetylpyrrole, and toluene to a reaction flask at room temperature, dissolve by sonication, heat, add phosphorus oxychloride, and obtain the reaction solution;

[0014] (2) The reaction solution obtained in step (1) was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0-9.0 with saturated sodium hydroxide solution. After extraction with a dichloromethane-water system, the solvent was removed by rotary evaporation. The resulting residue Np-Cl was subjected to fluorination reaction with boron trifluoride diethyl ether complex in the presence of triethylamine. The reaction product was purified by silica gel column chromatography to obtain the target compound Np-BOP.

[0015] (3) The compound Np-BOP obtained in step (2) was placed in a reaction flask and stirred at room temperature with methanol or ethanol as solvents to obtain Np-OMe and Np-OEt respectively.

[0016] The order of adding reactants in step (1) is compound 1, compound 2, toluene, and phosphorus oxychloride; the ratio of compound 1 to phosphorus oxychloride is 1:1 to 3. The order and ratio of adding reactants are optimized. Adding reactants (i.e., compound 1 and compound 2) first ensures sufficient contact between the main reactants, adding solvent later facilitates control of concentration and temperature, and adding catalyst last can effectively regulate the reaction rate. The 1:1 to 3 ratio range can ensure complete reaction and avoid side reactions. Changing the order or ratio of adding reactants may lead to incomplete reaction, increased by-products, or runaway reaction.

[0017] The heating temperature in step (1) is 25~120℃ and the heating time is 3~5 hours.

[0018] The feeding sequence in step (2) is compound Np-Cl, triethylamine, and boron trifluoride ether; the feeding ratio of compound Np-Cl to triethylamine and boron trifluoride ether is 1:5:5.

[0019] The beneficial effects of this invention are as follows:

[0020] (1) The compound of this invention is a photosensitizer based on 1,8-naphtholactamimide. Phototoxicity experiments have verified that this photosensitizer can efficiently generate reactive oxygen species (ROS) under specific wavelengths of light, exhibiting significant phototoxicity to HeLa tumor cells. This photosensitizer shows low toxicity to normal cells under light-free conditions, demonstrating good biocompatibility, which is beneficial for improving the safety and selectivity of phototherapy. Simultaneously, this photosensitizer exhibits selective accumulation characteristics in tumor cells, and combined with the spatiotemporal controllability of light irradiation, it can achieve precise tumor-targeted therapy. The molecular design based on the 1,8-naphtholactamimide skeleton gives this photosensitizer good photostability and modifiability, providing an important lead compound for the development of novel, highly efficient photodynamic therapy drugs. These characteristics indicate that this photosensitizer has significant application value and clinical translational potential in the field of tumor photodynamic therapy.

[0021] (2) The synthesis reaction conditions of the photosensitizer based on 1,8-naphthoimide described in this invention are easy to control, the product purification operation is simple, and it has universal applicability. Attached Figure Description

[0022] Figure 1 This is the proton NMR spectrum of the compound Np-BOP obtained in Example 6.

[0023] Figure 2 This is the proton NMR spectrum of the compound Np-OMe obtained in Example 7.

[0024] Figure 3 This is the proton NMR spectrum of the compound Np-OEt obtained in Example 8.

[0025] Figure 4 The UV absorption (a) and fluorescence spectra (b) of the compound Np-BOP obtained in Example 6 are shown.

[0026] Figure 5 The UV absorption (a) and fluorescence spectra (b) of the compound Np-OMe obtained in Example 7 are shown.

[0027] Figure 6 The UV absorption (a) and fluorescence spectra (b) of the compound Np-OEt obtained in Example 8 are shown.

[0028] Figure 7 This is a single crystal image of the compound Np-BOP obtained in Example 6.

[0029] Figure 8 These are cell live / dead staining images of the compound Np-BOP obtained in Example 6.

[0030] Figure 9 These are cell live / dead staining images of the compound Np-OMe obtained in Example 7. Detailed Implementation

[0031] The present invention will be further illustrated below with reference to the embodiments, but the scope of protection of the present invention is not limited to the scope described in the embodiments.

[0032] Example 1

[0033] Weigh out 1,8-naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 327.39 mg, 3.0 mmol), and dissolve them in 25 mL of toluene. Place the reaction system at room temperature and slowly add phosphorus oxychloride (275 μL, 3.0 mmol) dropwise with magnetic stirring. Maintain the reaction at room temperature for 5 hours. Monitor the reaction progress by thin-layer chromatography (TLC); no reaction occurred.

[0034] Example 2

[0035] Weigh out 1,8-naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 327.39 mg, 3.0 mmol), and dissolve them in 25 mL of toluene. Heat the reaction mixture in an oil bath to 120 °C, and slowly add phosphorus oxychloride (275 μL, 3.0 mmol) dropwise with magnetic stirring. Monitor the reaction progress by thin-layer chromatography (TLC, developing solvent: PE / DCM = 3:1, v / v). After 3 hours, compound 2 was completely reacted, while a small amount of compound 1 remained. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After three extractions with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain the intermediate Np-Cl (125.13 mg, 0.45 mmol). The intermediate was fluorinated with a boron trifluoride diethyl ether complex (319 μL, 2.25 mmol) in the presence of triethylamine (313 μL, 2.25 mmol). The product was purified by silica gel column chromatography (eluent: PE / DCM = 5:1, v / v) to give the target compound Np-BOP as a black solid with a green metallic luster, in 15% yield.

[0036]

[0037] Example 3

[0038] 1,8-Naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 491.10 mg, 4.5 mmol) were weighed and dissolved in 25 mL of toluene. The reaction system was heated to 120 °C in an oil bath, and phosphorus oxychloride (275 μL, 3.0 mmol) was slowly added dropwise with magnetic stirring. The reaction progress was monitored by thin-layer chromatography (TLC, developing solvent: PE / DCM = 3:1, v / v). After 3 hours, compound 2 reacted completely, and compound 1 reacted substantially completely. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After extraction three times with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain intermediate Np-Cl (208.55 mg, 0.75 mmol). The intermediate was fluorinated with a boron trifluoride diethyl ether complex (531 μL, 3.75 mmol) in the presence of triethylamine (521 μL, 3.75 mmol). The product was purified by silica gel column chromatography (eluent: PE / DCM = 5:1, v / v) to give the target compound Np-BOP as a black solid with a green metallic luster, in a yield of 25%. When the amount of compound 1 was increased by 1.5 mmol relative to that in Example 2, its yield increased by 10%.

[0039]

[0040] Example 4

[0041] 1,8-Naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 654.78 mg, 6 mmol) were weighed and dissolved in 25 mL of toluene. The reaction system was heated to 120 °C in an oil bath, and phosphorus oxychloride (275 μL, 3.0 mmol) was slowly added dropwise with magnetic stirring. The reaction progress was monitored by thin-layer chromatography (TLC, developing solvent: PE / DCM = 3:1, v / v). After 3 hours, compound 2 reacted completely, and compound 1 reacted substantially completely. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After extraction three times with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain intermediate Np-Cl (216.89 mg, 0.78 mmol). The intermediate was fluorinated with a boron trifluoride diethyl ether complex (553 μL, 3.90 mmol) in the presence of triethylamine (543 μL, 3.90 mmol). The product was purified by silica gel column chromatography (eluent: PE / DCM = 5:1, v / v) to give the target compound Np-BOP as a black solid with a green metallic luster, in a yield of 26%. Increasing the amount of compound 1 by 1.5 mmol compared to Example 3 did not significantly change the yield.

[0042]

[0043] Example 5

[0044] 1,8-Naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 491.10 mg, 4.5 mmol) were weighed and dissolved in 25 mL of toluene. The reaction system was heated to 120 °C in an oil bath, and phosphorus oxychloride (550 μL, 6.0 mmol) was slowly added dropwise with magnetic stirring. The reaction progress was monitored by thin-layer chromatography (TLC, developing solvent: PE / DCM = 3:1, v / v). After 3 hours, compound 2 reacted completely, and compound 1 reacted substantially completely. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After extraction three times with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain intermediate Np-Cl (258.60 mg, 0.93 mmol). The intermediate was fluorinated with a boron trifluoride diethyl ether complex (659 μL, 4.65 mmol) in the presence of triethylamine (647 μL, 4.65 mmol). The product was purified by silica gel column chromatography (eluent: PE / DCM = 5:1, v / v) to give the target compound Np-BOP as a black solid with a green metallic luster, in a yield of 31%. The yield increased by 6% when the amount of phosphorus oxychloride was increased by 3 mmol compared to Example 3.

[0045]

[0046] Example 6

[0047] 1,8-Naphthoimide (compound 1, 507.55 mg, 3.0 mmol) and acetylpyrrole (compound 2, 491.10 mg, 4.5 mmol) were weighed and dissolved in 25 mL of toluene. The reaction system was heated to 120 °C in an oil bath, and phosphorus oxychloride (825 μL, 9.0 mmol) was slowly added dropwise with magnetic stirring. The reaction progress was monitored by thin-layer chromatography (TLC, developing solvent: PE / DCM = 3:1, v / v). After 3 hours, compound 2 reacted completely, and compound 1 reacted substantially completely. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After extraction three times with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain intermediate Np-Cl (300.30 mg, 1.08 mmol). The intermediate was fluorinated with a boron trifluoride diethyl ether complex (765 μL, 5.40 mmol) in the presence of triethylamine (751 μL, 5.40 mmol). The product was purified by silica gel column chromatography (eluent: PE / DCM = 5:1, v / v) to give the target compound Np-BOP as a black solid with a green metallic luster, in a yield of 36%. The yield increased by 5% when the amount of phosphorus oxychloride was increased by 3 mmol compared to Example 5.

[0048] A black solid Np-BOP (3.26 mg, 0.01 mmol) with a green metallic luster was dissolved in 1 mL of dichloromethane to prepare a mother liquor. 6 μL of the mother liquor was then added to 3 mL each of toluene, dichloromethane, ethyl acetate, acetone, and methanol. The absorption wavelengths of the compound were measured. Figure 4 The maximum absorption wavelength was used to excite the compound to obtain its emission wavelength. Figure 4 This compound has a long absorption wavelength, with a double absorption peak between 400-700 nm. Irradiation with a laser in this wavelength range will cause it to generate reactive oxygen species (ROS), thereby killing tumor cells.

[0049]

[0050] Example 7

[0051] Compound Np-BOP (326.06 mg, 1.0 mmol) was weighed and dissolved in a mixed solvent of 20 mL methanol and 5 mL dichloromethane. The reaction system was stirred at room temperature, and the reaction progress was monitored by thin-layer chromatography (TLC, eluent: PE / DCM = 2:1, v / v). After 5 hours, TLC showed that compound Np-BOP had completely reacted. The reaction solution was concentrated by rotary evaporation and purified by silica gel column chromatography (eluent: PE / DCM = 4:1, v / v) to give a pink solid product Np-OMe (167.50 mg), with a yield of 52%.

[0052] A mother liquor was prepared by dissolving a rose-red solid Np-OMe (3.26 mg, 0.01 mmol) in 1 mL of dichloromethane. 6 μL of this mother liquor was then added to 3 mL each of toluene, dichloromethane, ethyl acetate, acetone, methanol, and dimethyl sulfoxide. The absorption wavelengths of the compound were measured. Figure 5 The maximum absorption wavelength was used to excite the compound to obtain its emission wavelength. Figure 5 This compound has a long absorption wavelength, with a double absorption peak between 400-700 nm. Irradiation with a laser in this wavelength range will cause it to generate reactive oxygen species (ROS), thereby killing tumor cells.

[0053]

[0054] Example 8

[0055] Compound Np-BOP (322.06 mg, 1.0 mmol) was weighed and dissolved in 25 mL of ethanol. The reaction system was stirred at room temperature, and the reaction progress was monitored by thin-layer chromatography (TLC, eluent: PE / DCM = 2:1, v / v). After 4 hours, TLC showed that compound Np-BOP had completely reacted. The reaction solution was concentrated by rotary evaporation and purified by silica gel column chromatography (eluent: PE / DCM = 4:1, v / v) to give an orange-red solid product Np-OEt (218.50 mg), with a yield of 65%.

[0056] The orange-red solid product Np-OEt (3.36 mg, 0.01 mmol) was dissolved in 1 mL of dichloromethane to prepare a mother liquor. 6 μL of the mother liquor was then added to 3 mL each of toluene, dichloromethane, ethyl acetate, acetone, methanol, and dimethyl sulfoxide, and the absorption wavelength of the compound was measured. Figure 6 The maximum absorption wavelength was used to excite the compound to obtain its emission wavelength. Figure 6 This compound has a long absorption wavelength, with a double absorption peak between 400-700 nm. Irradiation with a laser in this wavelength range will cause it to generate reactive oxygen species (ROS), thereby killing tumor cells.

[0057]

[0058] Example 9 Cell viability staining experiment of compound Np-BOP

[0059] Cell viability assays were performed using HeLa cell lines. Cells in good growth condition were cultured at 1.0 × 10⁶ cells per well. 5Cells were seeded at a density of 1000 cells / well in six-well plates and incubated overnight at 37°C in a 5% CO2 incubator. Four treatment groups were set up: (1) Control group (fresh culture medium only); (2) Light-only group (635 nm, 1.0 W / cm²). 2 (3) Np-BOP treatment group (medium containing Np-BOP); (4) Np-BOP combined with light treatment group. After changing the corresponding medium, the cells were incubated for another 4 hours. Then, the cells in each group were stained with 10 μL Calein-AM (1.0 mM, live cell staining) and 10 μL PI (1.0 mM, dead cell staining), respectively. The cells in groups 2 and 4 were irradiated with 635 nm laser (1.0 W / cm²) after staining. 2 After incubation for another 30 minutes, all groups were observed and photographed using a fluorescence microscope (Olympus, BX51). The results showed that the control group had the highest cell survival rate; a small number of cells died in the light-only group; some cells died in the Np-BOP-only treatment group; and in the Np-BOP combined with light treatment group, almost all cancer cells were killed, indicating that Np-BOP has a significant photodynamic therapy effect under 635 nm laser irradiation.

[0060] Example 10 Cell viability staining experiment of compound Np-OMe

[0061] Cell viability assays were performed using HeLa cell lines. Cells in good growth condition were cultured at 1.0 × 10⁶ cells per well. 5 Cells were seeded at a density of 1000 cells / well in six-well plates and incubated overnight at 37°C in a 5% CO2 incubator. Four treatment groups were set up: (1) Control group (fresh culture medium only); (2) Light-only group (635 nm, 1.0 W / cm²). 2 (3) Np-OMe treatment group (medium containing Np-OMe); (4) Np-OMe combined with light treatment group. After changing the corresponding medium, the cells were incubated for another 4 hours. Subsequently, the cells in each group were stained with 10 μL Calein-AM (1.0 mM, live cell staining) and 10 μL PI (1.0 mM, dead cell staining), respectively. The cells in groups 2 and 4 were irradiated with 635 nm laser (1.0 W / cm²) after staining. 2 After incubation for another 30 minutes, all groups were observed and photographed using a fluorescence microscope (Olympus, BX51). The results showed that the control group had the highest cell survival rate; a small number of cells died in the light-only group; some cells died in the Np-OMe-only treatment group; and in the Np-OMe combined with light treatment group, almost all cancer cells were killed, indicating that Np-OMe has a significant photodynamic therapy effect under 635 nm laser irradiation.

[0062] Example 11: Solubility determination of compounds Np-BOP, Np-OMe, and Np-OEt

[0063] Excess Np-BOP, Np-OMe, and Np-OEt were weighed out at 1.0 g each and placed in 9 sets of containers. Each set corresponded to a combination of temperature (20°C, 40°C, 60°C) and water volume (25 mL, 50 mL, 100 mL) in an orthogonal experimental design. After adding the specified volume of deionized water, the containers were kept at the corresponding temperature and shaken at 200 rpm for 24 hours until dissolution equilibrium was reached. The saturated solution was then rapidly filtered, and the concentration of each compound in the filtrate was determined by HPLC. The solubility per unit volume (mg / mL) was calculated as shown in Table 1.

[0064] Table 1. Solubility of Np-BOP, Np-OMe, and Np-OEt

[0065]

[0066] The above embodiments are merely preferred technical solutions of the present invention and should not be considered as limitations on the present invention. The embodiments and features described in these embodiments can be arbitrarily combined without conflict. The scope of protection of the present invention should be limited to the technical solutions described in the claims, including equivalent substitutions of the technical features described in the claims. That is, equivalent substitutions and improvements within this scope are also within the scope of protection of the present invention.

Claims

1. A photosensitizer based on 1,8-naphtholactamide, characterized in that, The chemical structural formula of the compound is: The substituent R is selected from any one of chlorine, methoxy, and ethoxy.

2. The method for synthesizing a photosensitizer based on 1,8-naphtholide according to claim 1, characterized in that, The method includes the following synthesis path: Wherein, the substituent R is selected from either methoxy or ethoxy; (1) Weigh 3.0 mmol of 1,8-naphthyl lactimide and 4.5 mmol of acetylpyrrole, dissolve them in 25 mL of toluene, heat the reaction system in an oil bath to 120 °C, and slowly add 6.0 mmol of phosphorus oxychloride under magnetic stirring. The reaction process was monitored by thin-layer chromatography (TLC). The developing solvent was PE / DCM = 3:1, v / v. After 3 hours, compound 2 was completely reacted and compound 1 was basically completely reacted. The reaction solution was concentrated by rotary evaporation, dissolved in dichloromethane, and the pH was adjusted to 8.0 with saturated sodium hydroxide solution. After extraction three times with a dichloromethane-water system, the solvent was removed by rotary evaporation to obtain intermediate Np-Cl 0.93 mmol. (2) The Np-Cl obtained in step (1) was subjected to fluorination reaction with boron trifluoride diethyl ether complex in the presence of triethylamine. The reaction product was purified by silica gel column chromatography to obtain the target compound Np-BOP. (3) Using the compound Np-BOP obtained in step (2) as a reactant, and using methanol or ethanol as a solvent, the reaction was stirred at room temperature to obtain Np-OMe and Np-OEt respectively.

3. The method according to claim 2, characterized in that, In step (1), the molar ratio of 1,8-naphthoimide, acetylpyrrole, and phosphorus oxychloride is 1:1~2:1~3.

4. The method according to claim 2, characterized in that, The heating temperature in step (1) is 25~120℃ and the heating time is 3~5 hours.

5. The method according to claim 2, characterized in that, In step (2), the ratio of compound Np-Cl, triethylamine, and boron trifluoride diethyl ether is 1:5:

5.

6. The method according to claim 2, characterized in that, In step (1), the order of feeding is compound 1, compound 2, toluene, and phosphorus oxychloride; in step (2), the order of feeding is compound Np-Cl, triethylamine, and boron trifluoride ether.

7. The use of the photosensitizer based on 1,8-naphthoylimide according to claim 1 or the photosensitizer based on 1,8-naphthoylimide prepared by the method according to any one of claims 2-6 in the preparation of a medicament for photodynamic therapy of cancer tumor cells.

8. The use according to claim 7, characterized in that, The cancerous tumor cells mentioned are human cervical cancer cells, which are selected from HeLa cells.

9. The use according to claim 7, characterized in that, The 1,8-naphthoimide-based photosensitizer generates a large amount of reactive oxygen species (ROS) under laser irradiation within the absorption wavelength range of the photosensitizer.

10. The use according to claim 9, characterized in that, The photosensitizer described herein has an absorption wavelength range of 400-700 nm.

Images