Aza-boron dipyrromethene dye molecule with near-infrared second-region absorption and preparation method and application of aza-boron dipyrromethene dye molecule

Abstract

The invention provides an aza-BODIPY dye molecule with near-infrared two-region absorption, 4-cyanophenyl is used as an electron acceptor for modification at 1, 7 sites, thiophene-triphenylamine is used as a strong electron donor for modification at 3, 5 sites, an aza-BODIPY dye with extremely strong intramolecular charge transfer is constructed, the absorption spectrum of the aza-BODIPY dye is extended to 1100 nm, and after the aza-BODIPY dye molecule forms an aggregation state, the absorption spectrum of the aza-BODIPY dye molecule has strong absorption spectrum. The aza-BODIPY dye molecule has the advantages that the absorption spectrum of the aza-BODIPY dye molecule is extended to 1250nm, the aza-BODIPY dye molecule has good absorption in an NIR-II region, and the nano-particles prepared by self-assembling the aza-BODIPY dye molecule and the amphiphilic polymer F127 show 46.2% of high photothermal conversion efficiency under the excitation of 1060nm laser, can be used for photoacoustic imaging and photothermal therapy, and can be used for preparing the nano-particles. The problems that tissue penetrability is insufficient and deep tumors cannot be diagnosed and treated in traditional tumor light diagnosis and treatment are solved.

Description

Technical Field

[0001] The present invention relates to the technical field of biomaterials, and in particular to an aza-fluoro-boron fluorescent dye molecule with near-infrared second-region absorption, and a preparation method and application thereof. Background Art

[0002] Cancer is one of the deadliest diseases worldwide, and its high morbidity and mortality rates have driven the advancement of cancer phototherapy. Phototherapy has recently emerged as an effective cancer treatment tool that integrates real-time diagnosis and in situ treatment on a single platform. This method has the advantages of non-invasiveness, high spatiotemporal resolution, and low toxicity. By utilizing the unique properties of light, phototherapy can target and destroy cancer cells without causing significant damage to surrounding healthy tissues. Among them, photothermal therapy (PTT) uses laser irradiation to generate local hyperthermia at the tumor site to ablate cancer cells. Compared with traditional therapies, PTT has the advantages of high specificity, minimal invasiveness, flexible administration, and strong controllability. It is one of the most advanced technologies in the field of nanomedicine. Therefore, it has become a key research topic in tumor treatment.

[0003] Photothermal agents are needed in photothermal therapy. Therefore, various photothermal agents with near-infrared (NIR) absorption have been developed in recent years. At present, research on photothermal agents mainly focuses on absorbing the first near-infrared region (NIR-I, 700-1000nm), but the photon scattering effect of NIR-I light when passing through biological tissues is still large and the penetration depth is limited. Compared with NIR-I photothermal agents, photothermal agents with NIR-II (1000-1700nm) absorption have excellent characteristics such as longer excitation wavelength, deeper tissue penetration, negligible autofluorescence, and less tissue damage. Therefore, they are considered to be ideal materials for photothermal therapy of deep tumors.

[0004] After the photothermal agent is excited by the NIR-II laser, a photothermal effect is generated, causing transient thermoelastic expansion, which leads to the generation of acoustic waves. These photoacoustic (PA) waves can be captured by an ultrasound detector and processed into images using an ultrasound machine. This imaging mode is called photoacoustic imaging (PAI). Therefore, PTT and PAI have homology, and combining NIR-II excitation with PAI-guided PTT is of great significance to current cancer treatment research: first, only a few endogenous molecules in biological systems can be excited by NIR-II light, so NIR-II imaging has a higher signal-to-noise ratio (SNR); second, NIR-II light scatters less than NIR-I light and has a greater tissue penetration depth; finally, the skin's tolerance to NIR-II light is much higher than that of NIR-I light, and NIR-II light is safer. Therefore, it is of great significance to develop exogenous drugs that can effectively respond to NIR-II light to achieve PAI-guided PTT.

[0005] So far, scholars have become increasingly interested in the development of photothermal agents with NIR-II absorption, and are currently focusing on inorganic materials, including gold nanomaterials, carbon nanomaterials, quantum dot materials, etc., in addition to phthalocyanines, conjugated polymers, organic small molecules, etc. Among them, organic small molecules are considered to be more promising photothermal agents than inorganic materials due to their clear molecular structure, reliable biosafety, reproducibility, and ease of modification. Strategically changing the molecular structure can optimize the photoacoustic performance and regulate the photophysical behavior. Promising platforms such as benzothiadiazole, phthalocyanine, anthracene, and porphyrin have paved the way for NIR-II photoacoustic imaging agents through methods such as π conjugation extension and construction of donor-acceptor (DA) structures. However, organic small molecules with NIR-II absorption are still very limited, so the development of new small molecule photothermal agents with NIR-II absorption is of great significance. Summary of the invention

[0006] The purpose of the present invention is to address the deficiencies in the prior art and provide an aza-fluoroboron fluorescent dye molecule with near-infrared second zone absorption and a preparation method thereof, so as to solve the current problems of scarcity and insufficient diversity of NIR-II absorbing small molecules, and to improve the problems of insufficient tissue penetration and inability to diagnose and treat deep tumors in traditional tumor photodiagnosis and treatment.

[0007] According to the first aspect of the present invention, there is provided an aza-fluoro-boron fluorescent dye molecule having near-infrared second-zone absorption, the aza-fluoro-boron fluorescent dye molecule is denoted as CNTPA, and the chemical structure is shown in Formula I:

[0008]

[0009] According to a second aspect of the present invention, there is provided a method for preparing the aforementioned aza-fluoro-boron fluorescent dye molecule having near-infrared second region absorption, comprising the following steps:

[0010] S1. Under a nitrogen atmosphere, dissolving 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide in an aqueous solution of methanol, and reacting under desired reaction conditions to obtain a first product;

[0011] S2, dissolving the first product obtained in step S1, 4-triphenylamine borate, potassium carbonate and tetrakis(triphenylphosphine)palladium in a mixed solution of toluene and ethanol in sequence, and reacting under desired reaction conditions to obtain a second product;

[0012] S3, dissolving the second product obtained in step S2, nitromethane and triethylamine in methanol, and reacting under desired reaction conditions to obtain a third product;

[0013] S4, dissolving the third product obtained in step S3 and ammonium acetate in n-butanol, and reacting under desired reaction conditions to obtain a fourth product;

[0014] S5. Dissolve the fourth product obtained in step S4 in 1.2-dichloroethane, add N,N-diisopropylethylamine and boron trifluoride etherate, and react under the required reaction conditions to obtain an aza-fluoroboron fluorescent dye molecule, which is recorded as CNTPA.

[0015] As an optional embodiment, in step S1, the molar ratio of 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide is 1:1:0.1, the volume ratio of methanol and water is 4:1, and the required reaction conditions are stirring at room temperature for 18 hours.

[0016] As an optional embodiment, in step S2, the molar ratio of the first product, 4-triphenylamine borate, tetrakis(triphenylphosphine)palladium and potassium carbonate is 1:(1.2-2):(0.03-0.05):(4-6), the volume ratio of toluene and ethanol is 3:1, and the required reaction conditions are to react at a temperature of 100-120°C for 18-24h.

[0017] As an optional embodiment, in step S3, the molar ratio of the second product, nitromethane and triethylamine is 1:5:5, and the required reaction conditions are stirring at a temperature of 75° C. for 24 hours.

[0018] As an optional embodiment, in step S4, the molar ratio of the third product to ammonium acetate is 1.9:109, and the required reaction conditions are heating under reflux at 120° C. for 36 hours.

[0019] As an optional embodiment, in step S5, the molar ratio of the fourth product to N,N-diisopropylethylamine and boron trifluoride etherate is 1:1:16, and the required reaction conditions are heating to 50° C. under a nitrogen atmosphere and stirring for 12-24 hours.

[0020] According to the third aspect of the present invention, there is provided a use of the aforementioned nitrogen-fluorine-boron fluorescent dye molecule with near-infrared second-region absorption in the preparation of a photothermal agent for NIR-II photoacoustic / photothermal imaging-mediated tumor photothermal therapy.

[0021] According to a fourth aspect of the present invention, there is provided a photothermal agent prepared using the aforementioned aza-fluoro-boron fluorescent dye molecules having near-infrared second region absorption.

[0022] As an optional embodiment, the photothermal agent is nanoparticles prepared by self-assembly of CNTPA and amphiphilic polymer F127.

[0023] It can be seen from the above technical scheme of the present invention that the nitrogen-fluoroboron fluorescent dye molecule with near-infrared region II absorption proposed in the present invention has a DA structure, and is modified at the 1,7 positions by 4-cyanophenyl as an electron acceptor and at the 3,5 positions by thiophene-triphenylamine as a strong electron donor to construct a nitrogen-fluoroboron fluorescent dye with extremely strong intramolecular charge transfer, and its absorption spectrum extends to 1100nm (located in the near-infrared region II), and after it forms an aggregated state, the absorption spectrum extends to 1250nm (located in the near-infrared region II), and has good absorption in the NIR-II region. The nanoparticles prepared by self-assembly of the nitrogen-fluoroboron fluorescent dye molecule and the amphiphilic polymer F127 show a high photothermal conversion efficiency of 46.2% under 1060nm laser excitation, and can be used for photoacoustic imaging and photothermal therapy. As a new NIR-II small molecule photothermal agent, it has good application prospects in the precise diagnosis and treatment of cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 It is a synthetic reaction scheme diagram of CNTPA of the present invention.

[0025] Figure 2 The CNTPA of the present invention is 1 H-NMR spectrum.

[0026] Figure 3 is a mass spectrum of CNTPA according to an example of the present invention.

[0027] Figure 4 It is a dynamic light scattering particle size distribution test chart of the CNTPA nanoparticles of the present invention.

[0028] Figure 5 Graph 1 is the ultraviolet absorption spectrum of the CNTPA nanoparticles in water according to the present invention.

[0029] Figure 6 Graph 1 is a fitting curve of the light-to-heat conversion efficiency of the CNTPA nanoparticles of the present invention.

[0030] Figure 7 Relative survival rate of mouse breast cancer cells (4T1) incubated with different concentrations of CNTPA nanoparticles for 24 hours under light or dark conditions.

[0031] Figure 8 This is the NIR-II photoacoustic imaging image of the CNTPA nanoparticles of the present invention in a breast cancer model mouse.

[0032] Fig. 9 This is a photothermal imaging image of the CNTPA nanoparticles exemplified in the present invention at the tumor site of a breast cancer model mouse.

[0033] Fig.10Tumor volume change curve of the CNTPA nanoparticles exemplified in the present invention during the treatment of breast cancer mouse model. DETAILED DESCRIPTION

[0034] In order to better understand the technical content of the present invention, specific embodiments are given and described as follows in conjunction with the accompanying drawings.

[0035] Various aspects of the present invention are described in this disclosure with reference to the accompanying drawings, in which many illustrative embodiments are shown. The embodiments of the present disclosure are not necessarily intended to include all aspects of the present invention. It should be understood that the various concepts and embodiments introduced above, as well as those described in more detail below, can be implemented in any of many ways.

[0036] Generally speaking, aza-fluoro-boron fluorescent dyes modify the aza-fluoro-boron fluorescent matrix (as shown in Formula II) at the four sites 1, 3, 5, and 7 to construct a D-π-A structure and promote intramolecular charge transfer (ICT), thereby promoting the red shift of the ultraviolet absorption / fluorescence emission peak.

[0037]

[0038] However, most donors and acceptors are not able to red-shift the ultraviolet absorption spectrum of nitrogen-fluorinated boron fluorescent dyes to the second near-infrared region (1000-1700nm) due to poor modification sites or insufficient electron-pushing / pulling capabilities.

[0039] To date, although there have been many studies on nitrogen-azo-fluorine-boron fluorescent dye molecules with near-infrared second region fluorescence emission, there are fewer nitrogen-azo-fluorine-boron fluorescent dye molecules with near-infrared second region absorption.

[0040] Therefore, the present invention provides an aza-fluoro-boron fluorescent dye molecule with near-infrared second-zone absorption. This dye has no fluorescence emission but exhibits a high photothermal conversion efficiency and can be used for photoacoustic imaging and photothermal therapy.

[0041] Aza-Fluoro-Boronic Dye Molecules with Near-Infrared Second Region Absorption

[0042] In an exemplary embodiment of the present invention, a nitrogen-fluorine-boron fluorescent dye molecule having near-infrared second-zone absorption is provided, and the nitrogen-fluorine-boron fluorescent dye molecule is denoted as CNTPA, and the chemical structure is shown in Formula I:

[0043]

[0044] The nitrogen-alumina-fluorine-boron fluorescent molecules exemplified above have a donor-π-donor (D-π-A) structure and have good absorption in the NIR-II region.

[0045] Preparation method

[0046] Combination Figure 1 As shown, the exemplary method for preparing the aza-fluoroboron fluorescent dye molecule with near-infrared second-region absorption of the present invention is to use 4-cyanobenzaldehyde and 2-acetyl-5-bromothiophene as the initial raw materials, according to Figure 1 The reaction route is used for reaction preparation.

[0047] As an optional example, the preparation method of the nitrogen-fluorine-boron fluorescent dye molecule having near-infrared second-region absorption specifically includes the following steps:

[0048] S1. Under a nitrogen atmosphere, dissolving 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide in an aqueous solution of methanol, and reacting under desired reaction conditions to obtain a first product;

[0049] S2, dissolving the first product obtained in step S1, tetrakis(triphenylphosphine)palladium, potassium carbonate, and 4-triphenylamine borate in a mixed solution of toluene and ethanol, and reacting under desired reaction conditions to obtain a second product;

[0050] S3, dissolving the second product obtained in step S2, nitromethane and triethylamine in methanol, and reacting under desired reaction conditions to obtain a third product;

[0051] S4, dissolving the third product obtained in step S3 and ammonium acetate in n-butanol, and reacting under desired reaction conditions to obtain a fourth product;

[0052] S5. Dissolve the fourth product obtained in step S4 in 1.2-dichloroethane, add N,N-diisopropylethylamine and boron trifluoride etherate, and react under the required reaction conditions to obtain an aza-fluoroboron fluorescent dye molecule, which is recorded as CNTPA.

[0053] As an optional example, in step S1, the molar ratio of 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide is 1:1:0.1, the volume ratio of methanol and water is 4:1, and the required reaction conditions are stirring at room temperature for 18 hours.

[0054] In a preferred example, after the reaction in step S1 is completed, the reaction solution is filtered and the precipitate is washed with cold methanol to obtain a yellow solid, which is the first product.

[0055] As an optional example, in step S2, the molar ratio of the first product, 4-boric acid triphenylamine, tetrakis(triphenylphosphine)palladium and potassium carbonate is 1:(1.2-2):(0.03-0.05):(4-6), the volume ratio of toluene and ethanol is 3:1, and the required reaction conditions are to react at a temperature of 100-120°C for 18-24h.

[0056] In a preferred example, after the reaction in step S2 is completed, the reaction solution is cooled to room temperature, extracted with dichloromethane and water, and then purified by column chromatography using dichloromethane and petroleum ether as eluents and a silica gel column to obtain a red solid, which is the second product.

[0057] As an optional example, in step S3, the molar ratio of the second product, nitromethane, and triethylamine is 1:5:5, and the required reaction conditions are stirring at a temperature of 75° C. for 24 hours.

[0058] In a preferred example, after the reaction in step S3 is completed, the reaction solution is cooled to room temperature, neutralized with dilute hydrochloric acid to neutrality, and then extracted with dichloromethane to obtain a yellowish brown crude product. Finally, column chromatography is performed on a silica gel column using dichloromethane petroleum ether as an eluent to obtain a light yellow oily product, which is the third product.

[0059] As an optional example, in step S4, the molar ratio of the third product to ammonium acetate is 1.9:109, and the required reaction conditions are heating under reflux at 120° C. for 36 hours.

[0060] In a preferred example, after the reaction in step S4 is completed, the reaction solution is cooled to room temperature and then filtered, and the precipitate is washed with cold methanol multiple times to obtain a dark green solid, which is the fourth product.

[0061] As an optional example, in step S5, the molar ratio of the fourth product to N,N-diisopropylethylamine and boron trifluoride etherate is 1:1:16, and the required reaction conditions are heating to 50° C. under a nitrogen atmosphere and stirring for 12-24 hours.

[0062] In a preferred example, after the reaction in step S5 is completed, the reaction solution is cooled to room temperature, extracted with dichloromethane, and the solvent is removed by distillation under reduced pressure. The crude product is purified by column chromatography to obtain dark green solid CNTPA.

[0063] As an optional example, the aforementioned preparation method further includes: after the reaction is completed, removing the solvent in the reaction mixture, and purifying the obtained crude product by column chromatography to obtain a dark green solid CNTPA.

[0064] As an optional example, the method for removing the solvent from the reaction mixture is: extracting the reaction mixture with dichloromethane, and removing the solvent by distillation under reduced pressure.

[0065] application

[0066] In another exemplary embodiment of the present invention, there is also provided a use of the aforementioned nitrogen-fluoroboron fluorescent dye molecule with near-infrared second-zone absorption in the preparation of a photothermal agent for NIR-II photoacoustic / photothermal imaging-mediated tumor photothermal therapy.

[0067] In another exemplary embodiment of the present invention, a photothermal agent prepared using the aforementioned aza-fluoro-boron fluorescent dye molecule having near-infrared second region absorption is also provided.

[0068] As an optional example, the aforementioned photothermal agent is nanoparticles prepared by self-assembly of CNTPA and amphiphilic polymer F127.

[0069] As an optional example, the photothermal conversion efficiency of the aforementioned photothermal agent under 1060nm laser excitation is 46.2%.

[0070] The aza-fluoro-boron fluorescent dye of the present invention has a clear molecular structure, a simple synthesis process, and a low cost of synthetic raw materials. The nanoparticles prepared from the molecule have uniform particle size, good stability, and high NIR-II excitation photothermal conversion efficiency, which is higher than that of most NIR-II region absorbing small molecules reported so far.

[0071] The photothermal agent prepared using the aforementioned near-infrared second-zone nitrogen-fluoroboron fluorescent dye molecules with high photothermal conversion efficiency exhibits a high photothermal conversion efficiency of 46.2% under 1060nm laser excitation, and can be used for precise and efficient NIR-II photoacoustic / photothermal imaging-mediated photothermal therapy of deep tumor tissues, reducing tumor treatment damage and side effects and improving treatment efficiency and accuracy.

[0072] For better understanding, the present invention is further described below in conjunction with several specific examples, but the preparation process is not limited thereto, and the content of the present invention is not limited thereto.

[0073] Unless otherwise specified, the materials in the examples were prepared according to existing methods or directly purchased from the market.

[0074] Example 1

[0075] [Synthesis of CNTPA]

[0076] Synthesis of product 1:

[0077] In a dry 100mL double-necked flask, 4-cyanobenzaldehyde (3.931g, 30mmol), 2-acetyl-5-bromothiophene (6.152g, 30mmol) and potassium hydroxide (0.168g, 3mmol) were added and dissolved in a mixed solution of methanol and water (40mL:10mL, 50mL in total). The mixture was stirred at room temperature for 18 hours. After the reaction was completed, the mixture was filtered and the precipitate was washed with cold methanol to obtain a yellow solid product 1 (6.864g, 72.2%).

[0078] 1H NMR (400MHz, CDCl3): δppm 7.799 (d, J = 15.2Hz, 1H), 7.716 (s, 4H), 7.613 (d, J = 4.4Hz, 1H), 7.376 (d, J = 16Hz, 1H), 7.179 (d, J = 4Hz, 1H).

[0079] Synthesis of product 2:

[0080] In a dry 100mL double-necked flask, product 1 (6.334g, 20mmol), 4-boric acid triphenylamine (6.938g, 24mmol), potassium carbonate (11.040g, 80mmol) and tetrakis(triphenylphosphine)palladium (0.692g, 0.6mmol) were added in sequence, dissolved in a mixed solution of toluene and ethanol (42mL:14mL, 56mL in total), stirred at 100°C for 24 hours, and after the reaction was completed, cooled to room temperature, extracted with dichloromethane and water, and finally purified by column chromatography using dichloromethane and petroleum ether (1:1, volume ratio) as eluent using a silica gel column to obtain a red solid product 2 (5.023g, 52.1%).

[0081] 1H NMR (400MHz, CDCl3): δppm 7.829, 7.721 (s, 4H), 7.549 (d, J = 9.2Hz, 2H), 7.505 (d, J = 16Hz, 1H), 7.320-7.281 (m, 5H), 7.151-7.058 (m, 8H).

[0082] Synthesis of product 3:

[0083] The product 2 (4.821 g, 10 mmol), triethylamine (5.095 g, 50 mmol), nitromethane (3.051 g, 50 mmol) and potassium hydroxide (0.561 g, 10 mmol) were added to a 100 mL double-necked flask containing 30 mL of methanol, heated at 75 ° C for 24 hours, cooled to room temperature, neutralized with 2M dilute hydrochloric acid to neutrality, and then extracted with dichloromethane to obtain a yellow-brown crude product. Finally, column chromatography was performed on a silica gel column using dichloromethane and petroleum ether (2:1, volume ratio) as the eluent to obtain a light yellow oily product 3 (2.211 g, 40.7%).

[0084] 1H NMR (400MHz, CDCl3): δppm 7.646(d,J=8Hz,2H),7.619(d,J=4.8Hz,1H),7.483(d,J=9.2Hz,2H),7.429(d,J=8Hz,2H),7.317-7.272(m,3H),7.721(d,J=6.8H z,1H),7.138-7.028(m,9H),4.873(dd,J=6Hz,12.8Hz,1H),4.731(dd,J=8Hz,12.8Hz,1H),4.821(m,1H),3.355(d,J=6.8Hz,2H).

[0085] Synthesis of product 4:

[0086] The product 3 (2.065 g, 3.8 mmol) and ammonium acetate (16.803 g, 218 mmol) were dissolved in a double-necked flask containing 100 mL of n-butanol, heated under reflux at 120 °C for 36 hours, cooled to room temperature and filtered, and the precipitate was washed with cold methanol several times to obtain a dark green solid (0.978 g, 52.6%).

[0087] Synthesis of the final product CNTPA:

[0088] Product 4 (998 mg, 1 mmol) was added to a dry 100 mL double-necked flask and dissolved in 1,2-dichloromethane (20 mL). N, N-diisopropylethylamine (1.292 g, 1 mmol) and boron trifluoride etherate (2.271 g, 16 mmol) were then added. The mixture was heated to 50 °C under a nitrogen atmosphere. After reacting for 12 hours, the mixture was cooled to room temperature. The reaction mixture was extracted with dichloromethane and the solvent was removed by distillation under reduced pressure. The crude product was purified by column chromatography (silicon dioxide, dichloromethane: petroleum ether = 3:1, volume ratio) to obtain dark green solid CNTPA (88 mg, 8.2%).

[0089] like Figure 2 As shown, 1 H NMR (400MHz, CDCl3): δppm 8.241(d,J=4.0Hz,2H),7.988(d,J=8.8Hz,4H),7.571(d,J=8.4Hz,4H),7.490(d, J=8.4Hz,4H),7.343-7.291(m,8H),7.158-7.086(m,16H),7.043(d,J=8.8Hz,4H).

[0090] like Figure 3 As shown, MALDI-TOF mass spectrum (m / z):calcd for C 66 H42 BF2N7S2[M] + :1045.3004; found,1045.2656.

[0091] Combination Figure 2-3 As shown, the molecular structure of CNTPA can be determined as Formula I:

[0092]

[0093] Example 2

[0094] [Preparation of CNTPA nanoparticles]

[0095] CNTPA nanoparticles were prepared by nanoprecipitation method as follows:

[0096] CNTPA molecules (1 mg) (prepared according to the method of Example 1) and Pluronic F-127 (10 mg) were dissolved in tetrahydrofuran (1 mL) and sonicated at a frequency of 40 KHz for 15 minutes.

[0097] Then, the solution was quickly injected into deionized water (10 mL) under ultrasonic conditions and ultrasonication was continued for 30 minutes. After 24 hours of stirring and volatilization to completely remove tetrahydrofuran, it was filtered with a 0.22 μm filter to obtain a clear and transparent dark blue CNTPA nanoparticle solution, which was stored in a refrigerator at 4°C for later use.

[0098] like Figure 4 As shown in the dynamic light scattering particle size distribution test results of CNTPA nanoparticles, the hydrated particle size is 79nm, which meets the requirements of high permeability and retention effect.

[0099] like Figure 5 As shown, CNTPA nanoparticles have a maximum absorption peak at 929 nm in water, and the absorption peak tail extends to 1300 nm.

[0100] Example 3

[0101] [Photothermal performance test of CNTPA nanoparticles]

[0102] The 1060 nm laser (1 W cm -2 ) was irradiated with an aqueous solution of CNTPA nanoparticles (1 mL, 50 μg / mL) and deionized water (1 mL). The temperature was raised for 10 minutes and then naturally cooled for 10 minutes to return to room temperature. The photothermal conversion efficiency of the aqueous solution of CNTPA nanoparticles was calculated using the cooling section.

[0103] like Figure 6As shown, the light-to-heat conversion efficiency of the CNTPA nanoparticle aqueous solution of the present invention is 46.2%.

[0104] Example 4

[0105] [Cytotoxicity experiment of CNTPA nanoparticles]

[0106] 4T1 cells were plated at 4.0 × 10 3 The cells were seeded in two 96-well cell culture plates at a density of 100 μg / mL and incubated at 37°C in a 5% CO2 dark environment for 24 hours.

[0107] In one of the wells, different concentrations of CNTPA nanoparticles (0, 5, 10, 15, 20, 25, 30 μg mL -1 ) were added to the well plate and incubated for 4 hours. Then, the cells were illuminated with a 1060 nm laser (1 W cm -2 ) irradiation, irradiation for 4 minutes per well, to test the cellular phototoxicity of CNTPA nanoparticles. After the illumination is completed, the well plate is incubated for 12 hours under dark light conditions, and 200 μL of MTT solution is added to each well and incubated in the dark for 4 hours. Remove the well plate culture medium, add 200 μL of dimethyl sulfoxide to dissolve the blue-purple formazan crystals, read the absorbance value at 490nm with an enzyme reader, and calculate the survival rate of 4T1 cells under different concentrations of CNTPA nanoparticles. The cell survival rate calculation formula is: average absorbance of the treatment group / average absorbance of the control group × 100%.

[0108] Another well plate was incubated in the dark, and different concentrations of CNTPA nanoparticles (0, 10, 20, 30, 40, 50, 60 μg mL -1 ) were added into the well plate and incubated for 12 h, and the cytotoxicity of CNTPA nanoparticles was tested by MTT assay.

[0109] like Figure 7 As shown in Figure 2, under dim light, even at a concentration of 60 μg mL -1 , the survival rate of 4T1 cells can still be maintained above 90%; however, under light conditions, only 25 μg mL -1 CNTPA nanoparticles can kill nearly 90% of 4T1 cells.

[0110] The above experiments show that the CNTPA nanoparticles of the present invention have good biocompatibility and phototoxicity.

[0111] Example 5

[0112] [NIR-II photoacoustic imaging of CNTPA nanoparticles in mice]

[0113] CNTPA nanoparticles (100 μL, 500 μg mL -1 ) were injected into mice through the tail vein, and photoacoustic images of the tumor site were recorded at different time points (pre, 8, 16, 20, 24 and 36 hours before injection).

[0114] like Figure 8 As shown, after mice were intravenously injected with CNTPA nanoparticles, the photoacoustic signal at the tumor site gradually increased with time, reached a maximum value at 24 hours, and then gradually weakened.

[0115] The above experiments can illustrate that the CNTPA nanoparticles of the present invention can effectively accumulate at the tumor site and achieve NIR-II photothermal imaging.

[0116] Example 6

[0117] [Photothermal imaging of CNTPA nanoparticles in mice]

[0118] CNTPA nanoparticles (100 μL, 500 μg mL -1 ) or PBS buffer (100 μL) was injected into mice through the tail vein, and the photothermal images of the tumor site at different time points (0, 1, 2, 3, 4, and 5 minutes) were recorded. Fig. 9 shown.

[0119] The above experiments show that after the CNTPA nanoparticles of the present invention accumulate in the tumor site, they can effectively cause the temperature of the tumor site to rise under light, and can be used for photothermal therapy.

[0120] Example 7

[0121] [Photothermal therapy experiment of CNTPA nanoparticles in mice]

[0122] 4T1 cancer cells were injected subcutaneously to establish a 4T1 tumor-bearing mouse model. When the tumor volume reached 100 mm 3 After that, you can carry out subsequent experiments.

[0123] The 4T1 tumor-bearing mice were randomly divided into three groups (PBS+L, NPs, and NPs+L, 5 mice in each group, L represents laser irradiation). The mice in the PBS+L group were intravenously injected with PBS (100 μL), and the mice in the NPs group and NPs+L group were intravenously injected with CNTPA nanoparticles (100 μL, 500 μg mL -1 24 h after injection, the tumor sites of mice in the PBS+L group and NPs+L group were illuminated with 1060 nm laser (1 W cm -2) for 5 minutes. After treatment, the tumor size and mouse body weight were measured every 2 days and recorded continuously until the 14th day.

[0124] like Fig.10 As shown in the data, after one treatment, the tumor growth in the NPs+L group was inhibited, and the tumor volume gradually decreased over time, and was completely eliminated on the 10th day after treatment, with no recurrence within 14 days. However, the tumor volume in the PBS group, PBS+L group, and NPs group increased by 3 to 4 times on the 14th day after treatment.

[0125] The above experiments show that the CNTPA nanoparticles of the present invention have good photothermal therapy effects.

[0126] Combined with the above tests, it can be proved that the photothermal agent prepared by the nitrogen-fluoroboron fluorescent NIR-II dye molecules of the present invention can be used for accurate and efficient NIR-II photoacoustic / photothermal imaging-mediated photothermal therapy of deep tumor tissues, reducing tumor treatment damage and side effects and improving treatment efficiency and accuracy.

[0127] Although the present invention has been disclosed as above with preferred embodiments, it is not intended to limit the present invention. A person with ordinary knowledge in the technical field to which the present invention belongs may make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention shall be determined by the definition of the claims.

Claims

1. An aza-fluorine-boron fluorescent dye molecule with near-infrared second region absorption, characterized in that: The aza-fluoro-boron fluorescent dye molecule is denoted as CNTPA, and its chemical structure is shown in Formula I:

2. A method for preparing an aza-fluoro-boron fluorescent dye molecule having near-infrared second region absorption as claimed in claim 1, characterized in that: The following steps are involved: S1. Under a nitrogen atmosphere, dissolving 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide in an aqueous solution of methanol, and reacting under desired reaction conditions to obtain a first product; S2, dissolving the first product obtained in step S1, 4-triphenylamine borate, potassium carbonate and tetrakis(triphenylphosphine)palladium in a mixed solution of toluene and ethanol in sequence, and reacting under desired reaction conditions to obtain a second product; S3, dissolving the second product obtained in step S2, nitromethane and triethylamine in methanol, and reacting under desired reaction conditions to obtain a third product; S4, dissolving the third product obtained in step S3 and ammonium acetate in n-butanol, and reacting under desired reaction conditions to obtain a fourth product; S5. Dissolve the fourth product obtained in step S4 in 1.2-dichloroethane, add N,N-diisopropylethylamine and boron trifluoride etherate, and react under the required reaction conditions to obtain an aza-fluoroboron fluorescent dye molecule, which is recorded as CNTPA.

3. The preparation method according to claim 2, characterized in that: In the step S1, the molar ratio of 4-cyanobenzaldehyde, 2-acetyl-5-bromothiophene and potassium hydroxide is 1:1:0.1, the volume ratio of methanol and water is 4:1, and the required reaction conditions are stirring at room temperature for 18 hours.

4. The preparation method according to claim 2, characterized in that: In step S2, the molar ratio of the first product, 4-triphenylamine borate, tetrakis(triphenylphosphine)palladium and potassium carbonate is 1:(1.2-2):(0.03-0.05):(4-6), the volume ratio of toluene and ethanol is 3:1, and the required reaction conditions are to react at a temperature of 100-120°C for 18-24h.

5. The preparation method according to claim 2, characterized in that: In the step S3, the molar ratio of the second product, nitromethane and triethylamine is 1:5:5, and the required reaction conditions are stirring at a temperature of 75° C. for 24 hours.

6. The preparation method according to claim 2, characterized in that: In step S4, the molar ratio of the third product to ammonium acetate is 1.9:109, and the required reaction conditions are heating under reflux at 120° C. for 36 hours.

7. The preparation method according to claim 2, characterized in that: In the step S5, the molar ratio of the fourth product to N,N-diisopropylethylamine and boron trifluoride etherate is 1:1:16, and the required reaction conditions are heating to 50° C. under a nitrogen atmosphere and stirring for 12-24 hours.

8. Use of the aza-fluoroboron fluorescent dye molecule with near-infrared second-zone absorption as claimed in claim 1 in the preparation of a photothermal agent for tumor photothermal therapy mediated by NIR-II photoacoustic / photothermal imaging.

9. A photothermal agent prepared by using the aza-fluoro-boron fluorescent dye molecule with near-infrared second-zone absorption as claimed in claim 1.

10. The photothermal agent according to claim 9, characterized in that: The photothermal agent is nanoparticles prepared by self-assembly of CNTPA and amphiphilic polymer F127.

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