Dimer Aza-BODIPY-based molecule as well as preparation method and application thereof

Abstract

The invention discloses a dimer type Aza-BODIPY based molecule NBDP-Fc and a preparation method thereof, and the dimer type Aza-BODIPY based molecule NBDP-Fc is prepared by coupling an Aza-BODIPY derivative NBDP and 1, 1 '-ferrocenedicarboxylic acid through an esterification reaction under the protection of nitrogen and the action of EDCI / DMAP. The NBDP-Fc molecule disclosed by the invention has relatively strong near-infrared absorption and near-infrared two-region fluorescence, and can be used for in-vivo near-infrared two-region fluorescence imaging; wherein the ferrocene unit can catalyze overexpressed hydrogen peroxide in tumors to generate hydroxyl radicals, so that chemical kinetic treatment is realized; under the irradiation of 808nm laser, the NBDP-Fc nanoparticles show good I-type reactive oxygen species-superoxide anion generation efficiency and photothermal conversion efficiency as high as 45.6%, and the induced local heat effect not only can directly ablate tumors at high temperature, but also can accelerate the Fenton reaction rate and further enhance chemical kinetics therapy; the NBDP-Fc nanoparticles can be used for accurate and efficient near-infrared two-region fluorescence imaging mediated chemical kinetics, I-type photodynamic and photo-thermal combined treatment of deep tumor tissues, and the treatment efficiency and accuracy are improved.

Description

Technical Field

[0001] This invention relates to the field of biofunctional medical materials technology, specifically to a dimer-type Aza-BODIPY-based dye molecule with near-infrared II (NIR-II) fluorescence imaging capability, type I reactive oxygen species (ROS) generation capability, Fenton reaction catalytic activity, and high photothermal conversion efficiency, as well as its preparation method and applications. Background Technology

[0002] Cancer remains one of the major diseases threatening human health, and achieving highly sensitive early diagnosis and efficient, precise treatment has always been an important research direction in medicine and materials science. While existing clinical imaging methods such as positron emission tomography (PET), X-ray computed tomography (X-CT), magnetic resonance imaging (MRI), and ultrasound (US) are widely used, they suffer from insufficient sensitivity, limited spatial resolution, and high costs in early tumor screening, making it difficult to meet the needs of precision medicine development.

[0003] In recent years, phototheranostics for tumors has been considered a promising new strategy to replace or supplement traditional therapies due to its non-invasiveness, real-time monitoring, low side effects, and high selectivity. Within phototheranostics, near-infrared II (NIR-II, 1000–1700 nm) fluorescence imaging has become an important direction for next-generation in vivo imaging due to its deeper tissue penetration, higher spatial resolution, and superior signal-to-noise ratio. However, currently developed NIR-II fluorescent probes generally suffer from low fluorescence quantum efficiency and complex molecular structures.

[0004] In terms of treatment, photothermal therapy (PTT), photodynamic therapy (PDT), and chemodynamic therapy are typical non-invasive treatment methods. PTT relies on photosensitizers to convert near-infrared light energy into heat energy, achieving high-temperature ablation of tumor tissue; however, PTT alone is often insufficient to eradicate tumors. PDT generates reactive oxygen species through the interaction of the excited state of the photosensitizer with molecular oxygen; most photosensitizers are generated through a type II mechanism. 1 O2, but solid tumors are generally in a hypoxic microenvironment, which severely limits the efficacy of type II PDT; in contrast, type I PDT generates superoxide anions (O2) through electron transfer or proton transfer. ·- ·OH radicals are less dependent on oxygen and are particularly suitable for the treatment of hypoxic tumors. Chemokinetic therapy converts overexpressed hydrogen peroxide (H2O2) in the tumor microenvironment into highly toxic ·OH radicals via Fenton or Fenton-like reactions, inducing oxidative damage and apoptosis in cancer cells. This process is highly selective and depends on the unique metabolic characteristics of the tumor microenvironment.

[0005] Despite the rapid development of phototherapy technology, existing systems generally rely on multi-component nanocomposites (such as assemblies of photosensitizers, metal ions, and photothermal agents). These systems, due to their complex composition, ambiguous structure, and poor controllability, face significant challenges in clinical translation. Therefore, constructing a single molecular platform to integrate multiple functions and achieve therapies has become a more ideal direction for technological development. However, this goal presents two core challenges to molecular design: first, the excited-state energy dissipation pathway of the molecule needs to be precisely optimized to ensure it simultaneously meets the synergistic requirements of NIR-II fluorescence imaging, type I photothermolysis (PDT), and photothermolysis (PTT); second, catalytic units need to be precisely embedded in the molecular framework to achieve chemokinetic therapeutic functions.

[0006] However, if the above design steps are not properly controlled, a series of problems can easily arise, causing a single molecular platform to fail to achieve the expected integrated diagnostic and therapeutic effect. For example, an imbalance in the distribution of energy dissipation channels within the molecule may reduce fluorescence brightness or triplet yield, thereby weakening fluorescence imaging or PDT effects. At the same time, the introduced metal catalytic sites are prone to static and dynamic quenching of the excited state of the molecule, further impairing fluorescence imaging and PDT performance. If the ROS reaction pathway and rate of chemokinetic therapy and type I PDT are not matched, premature ROS depletion or non-selective oxidation may occur, damaging the stability of the system. Since chemokinetic therapy is highly sensitive to the tumor microenvironment (such as pH, H2O2 content, reducing substances, etc.), an unreasonable design may lead to insufficient in vivo reaction or material inactivation due to excessive reaction. Summary of the Invention

[0007] The purpose of this invention is to address the shortcomings of existing technologies by constructing a single-molecule platform. The proposed novel dimer-type Aza-BODIPY-based molecule, compared to the monomer, significantly alters the excited-state kinetics of the photosensitizer through intramolecular electronic coupling, making it more prone to generating type I ROS via electron transfer. Furthermore, the introduction of ferrocene groups enables the initiation of the Fenton reaction in the tumor microenvironment, catalyzing the generation of highly cytotoxic hydroxyl radicals. Simultaneously, the dimer structure also endows the device with near-infrared II fluorescence emission and excellent photothermal conversion properties, enabling simultaneous near-infrared II fluorescence imaging-guided chemical kinetics, type I photodynamic therapy, and photothermal combined therapy under 808nm laser irradiation.

[0008] According to a first aspect of the present invention, a dimer Aza-BODIPY-based molecule is provided, which is formed by coupling an Aza-BODIPY-based monomer molecule with 1,1'-ferrocene dicarboxylic acid.

[0009] The chemical structural formula of the Aza-BODIP-based monomer molecule is shown in Formula I, and is denoted as NBDP;

[0010]

[0011] The structural formula of the dimer type Aza-BODIPY molecule is shown in Formula II, and is denoted as NBDP-Fc;

[0012]

[0013] According to a second aspect of the present invention, a method for preparing a dimer-type Aza-BODIPY-based molecule as described above is provided, comprising the following steps:

[0014] Under a nitrogen atmosphere, 4-hydroxyacetophenone and 4-cyanobenzaldehyde were dissolved in anhydrous ethanol, and then KOH was added. Under the first reaction conditions, an aldol condensation reaction was carried out to obtain the first compound.

[0015] The first compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and a Michael addition reaction was carried out under the second reaction conditions to give the second compound.

[0016] Under a nitrogen atmosphere, 4-iodoacetophenone and 4-diphenylaminobenzaldehyde were dissolved in anhydrous ethanol, then KOH was added, and an aldol condensation reaction was carried out under the third reaction conditions to give the third compound.

[0017] The third compound, cyanide ketone, and iodide ketone were dissolved in anhydrous N,N-dimethylformamide and reacted under the fourth reaction conditions to obtain the fourth compound;

[0018] The fourth compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and a Michael addition reaction was carried out under the fifth reaction conditions to give the fifth compound.

[0019] Under a nitrogen atmosphere, the second compound, the fifth compound, and ammonium acetate were dissolved in n-butanol medium and reacted under the sixth reaction conditions to obtain the sixth compound;

[0020] Under a nitrogen atmosphere, the sixth compound and boron trifluoride diethyl ether were placed in 1,2-dichloroethane solvent, with N,N-diisopropylethylamine as the organic base, and reacted under the seventh reaction conditions to obtain the fluorinated borate product NBDP.

[0021] Under a nitrogen atmosphere, NBDP and 1,1'-ferrocene dicarboxylic acid (Fc-2COOH) were dissolved in dichloromethane, with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI) as the carboxyl activator and 4-dimethylaminopyridine (DMAP) as the reaction catalyst. The reaction was carried out under the eighth reaction condition to obtain the dimer type Aza-BODIPY-based molecule NBDP-Fc.

[0022] As an optional implementation method, the molar ratio of 4-hydroxyacetophenone, 4-cyanobenzaldehyde and KOH is 1:(0.5-1.5):(2-3); the first reaction conditions include: reacting at a temperature of 30℃-35℃ for 18h-24h;

[0023] The molar ratio of the first compound, nitromethane, and triethylamine is 1:(8-12):(8-12); the second reaction conditions include: reacting at 70℃-80℃ for 24-30 hours.

[0024] As an optional implementation method, the molar ratio of 4-iodoacetophenone, 4-diphenylaminobenzaldehyde and KOH is 1:(0.5-1.5):(1-2); the third reaction conditions include: reacting at a temperature of 30℃-35℃ for 18h-24h.

[0025] As an optional implementation, the molar ratio of the third compound, cyanide ketone, and iodide ketone is 1:(1.5-2.5):(0.3-1); the fourth reaction conditions include: reacting at a temperature of 120℃-150℃ for 24h-30h;

[0026] The molar ratio of the fourth compound, nitromethane, and triethylamine is 1:(4-6):(4-6); the fifth reaction conditions include: reacting at 70℃-80℃ for 12h-24h.

[0027] As an optional implementation, the molar ratio of the second compound, the fifth compound, and ammonium acetate is 1:(0.5-1.5):(55-65); the sixth reaction conditions include: reacting at a temperature of 125℃-135℃ for 24h-30h.

[0028] As an optional implementation, the molar ratio of the sixth compound, boron trifluoride ether, and N,N-diisopropylethylamine is 1:(15-20):(10-11); the seventh reaction conditions include reacting at a temperature of 45°C-55°C for 12-16 hours.

[0029] As an optional implementation method, the molar ratio of Fc-2COOH, EDCI, DMAP, and NBDP is 1:(2-3):(0.2-0.3):(2.5-3.5); the eighth reaction conditions include: reacting at a temperature of 30℃-35℃ for 18h-24h.

[0030] In a third aspect of the present invention, the aforementioned dimer-type Aza-BODIPY-based molecule is provided for the preparation of phototherapeutic reagents for near-infrared II fluorescence / photoacoustic imaging-mediated tumor chemokinetics, type I photodynamic therapy, and photothermal combined therapy.

[0031] In a fourth aspect of the present invention, a phototherapy reagent prepared using the aforementioned dimer-type Aza-BODIPY-based molecule is provided.

[0032] As can be seen from the above technical solutions of the present invention, the dimeric Aza-BODIPY-based molecule proposed in this invention, through molecular engineering design, achieves multifunctional integration of the Aza-BODIPY-based dimer molecule, exhibiting unique advantages in high-sensitivity imaging and efficient combined treatment of deep tumors, and providing a novel and translatable solution for precision tumor diagnosis and treatment. Its advantages are as follows:

[0033] (1) Innovative dimer molecular design: By coupling NBDP with 1,1'-ferrocene dicarboxylic acid, a dimer molecular structure is constructed, avoiding the complex assembly process of traditional nanocomposite systems and realizing the multifunctional integration of a single molecular platform.

[0034] (2) Excellent optical properties: The molecules exhibit strong NIR absorption and bright NIR-II fluorescence, which is beneficial for accurate imaging of deep tissues.

[0035] (3) Synergistic effect of multimodal therapy: The ferrocene unit can catalyze the generation of H2O2 into ·OH, realizing efficient chemokinetic therapy; the Aza-BODIPY skeleton has good type I ROS generation ability and still has efficient therapeutic activity in hypoxic environment; it has a photothermal conversion efficiency of up to 45.6% under 808nm laser irradiation, effectively realizing PTT and accelerating Fenton reaction, forming positive feedback to enhance therapy.

[0036] (4) Advantages of integrated diagnosis and treatment: Integrating NIR-II imaging, type I PDT, chemokinetic therapy and PTT into a single molecular platform, avoiding complex multi-component assembly systems, and greatly improving the system's stability, controllability and clinical translation potential.

[0037] The dimer-type Aza-BODIPY-based molecule of this invention has a well-defined molecular structure, a simple synthesis process, and low raw material costs. The nanoparticles prepared from this molecule have uniform particle size, good stability, strong near-infrared II fluorescence emission, good type I ROS generation ability, and a high photothermal conversion efficiency of 45.6%. As a single molecular system, this phototherapy reagent enables multimodal combined therapy of tumor chemokinetics / type I photodynamic therapy / photothermal therapy guided by single-wavelength laser-triggered near-infrared II fluorescence / photoacoustic dual-modal imaging. Attached Figure Description

[0038] Figure 1 This is a schematic diagram of the synthesis reaction of NBDP-Fc according to the present invention.

[0039] Figure 2 This is an example of NBDP-Fc in this invention. 1H-NMR spectrum.

[0040] Figure 3 This is an example of NBDP-Fc in this invention. 13 C-NMR spectrum.

[0041] Figure 4 This is the mass spectrum of NBDP-Fc as an example of the present invention.

[0042] Figure 5 This is a dynamic light scattering particle size distribution test diagram of NBDP-Fc nanoparticles as an example of the present invention.

[0043] Figure 6 This is the ultraviolet absorption and fluorescence emission spectrum of the NBDP-Fc nanoparticle aqueous solution as an example of the present invention.

[0044] Figure 7 The graph shows the generation of hydroxyl radicals by NBDP-Fc nanoparticles through the Fenton reaction under different conditions, using TMB as a probe.

[0045] Figure 8 This is a test diagram of total ROS generation of NBDP-Fc nanoparticles, as exemplified by this invention.

[0046] Figure 9 This invention exemplifies NBDP-Fc nanoparticles O2. ·- Generate a test graph.

[0047] Figure 10 This is a test graph of the photothermal conversion efficiency of NBDP-Fc nanoparticles as an example of the present invention.

[0048] Figure 11 The relative survival rates of different concentrations of NBDP-Fc nanoparticles incubated with mouse breast cancer cells (4T1) for 24 hours under light or dark conditions are compared.

[0049] Figure 12 This is an in vivo fluorescence imaging image of NBDP-Fc nanoparticles in a breast cancer model nude mouse, as exemplified by this invention, using NIR-II.

[0050] Figure 13 This is an in vivo photoacoustic imaging image of NBDP-Fc nanoparticles in a breast cancer model nude mouse, as an example of the present invention.

[0051] Figure 14 This is a photothermal imaging image of NBDP-Fc nanoparticles as an example of the present invention at the tumor site in a breast cancer model mouse.

[0052] Figure 15 This is a graph showing the changes in tumor volume in a mouse breast cancer model during 16 days of treatment. Detailed Implementation

[0053] To better understand the technical content of the present invention, specific embodiments are described below in conjunction with the accompanying drawings.

[0054] Various aspects of the invention are described in this disclosure with reference to the accompanying drawings, in which numerous illustrative embodiments are shown. The embodiments of this disclosure are not necessarily intended to encompass all aspects of the invention. It should be understood that the various concepts and embodiments described above, as well as those described below in more detail, can be implemented in any of a number of ways.

[0055] Dimeric Aza-BODIPY-based molecules

[0056] In an exemplary embodiment of the present invention, a dimer Aza-BODIPY-based molecule is provided, which is formed by coupling Aza-BODIPY-based monomer molecules with 1,1'-ferrocene dicarboxylic acid.

[0057] The chemical structural formula of the Aza-BODIP-based monomer molecule is shown in Formula I, denoted as NBDP;

[0058]

[0059] The structural formula of the dimer type Aza-BODIPY molecule is shown in Formula II, denoted as NBDP-Fc;

[0060]

[0061] The dimer-type Aza-BODIPY-based molecule of this invention, through a dimer-ferrocene single-molecule platform combined with excited-state regulation of electron transfer, solves the problems of excited-state quenching, energy channel conflict, and microenvironment compatibility in traditional Aza-BODIPY small molecule superimposed chemokinetic therapy at the structural level, allowing PDT, PTT, and chemokinetic therapy to mutually enhance each other; specifically as follows:

[0062] (i) By covalently embedding ferrocene into the Aza-BODIPY dimer framework, a single-molecule endogenous catalytic site is constructed, which enables the iron source and the photosensitive core to combine under controllable electronic coupling and spatial distance, thus avoiding disordered quenching caused by external metals.

[0063] (ii) By leveraging the dimer configuration and donor-acceptor coupling to enhance charge transfer characteristics, it thermodynamically and kinetically favors the type I electron transfer pathway. At the same time, through the design of an intramolecular "barrier / spacer", it reduces the excessive quenching of fluorescence and triplet state by ferrocene, while retaining near-infrared II imaging and type I PDT efficiency.

[0064] (III) Utilizing the localized heating brought about by the 45.6% photothermal conversion efficiency (PCE) under 808nm laser, photothermal therapy (PTT) can be directly achieved, while also increasing the Fenton reaction rate constant to amplify the yield of hydroxyl radicals (·OH). Furthermore, the superoxide anion (O2) generated by type I PDT... ·- Hydrogen peroxide (H2O2) serves as a supplementary raw material for chemokinetic therapy, forming...

[0065] (iv) Ferrocene sites are more likely to undergo Fe(II) / Fe(III) cycling in the acidic / hydrogen peroxide microenvironment of tumors, and the endogenous coupling at the molecular level reduces the dependence on exogenous metal ions and carriers, thereby improving in vivo reproducibility and safety window.

[0066] (v) By covalently controlling the precise distance and matching the energy level / potential (making the Rehm-Weller free energy more conducive to electron transfer to oxygen and peroxides rather than ground state heat dissipation), the imaging brightness, type I ROS yield and photothermal release can coexist and be adjusted.

[0067] preparation

[0068] Combination Figure 1 As shown, the method for preparing the dimer-type Aza-BODIPY-based molecule of the present invention, which possesses near-infrared II fluorescence, type I ROS generation capability, and high photothermal conversion efficiency, uses NBDP, 1,1'-ferrocene dicarboxylic acid (Fc-2COOH), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI), and 4-dimethylaminopyridine (DMAP) as raw materials, according to... Figure 1 The reaction is prepared using the reaction route described above.

[0069] In an exemplary embodiment of the present invention, the method for preparing the aforementioned dimer-type Aza-BODIPY-based molecule includes the following steps:

[0070] Under a nitrogen atmosphere, 4-hydroxyacetophenone (compound 1) and 4-cyanobenzaldehyde (compound 2) were dissolved in anhydrous ethanol, and then KOH was added. Under the first reaction conditions, an aldol condensation reaction was carried out to obtain the first compound (compound 3).

[0071] The first compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and a Michael addition reaction was carried out under the second reaction conditions to give the second compound (compound 4);

[0072] Under a nitrogen atmosphere, 4-iodoacetophenone (compound 5) and 4-diphenylaminobenzaldehyde (compound 6) were dissolved in anhydrous ethanol, then KOH was added, and an aldol condensation reaction was carried out under the third reaction conditions to give the third compound (compound 7).

[0073] The third compound, cyanide ketone, and iodide ketone were dissolved in anhydrous N,N-dimethylformamide and reacted under the fourth reaction conditions to obtain the fourth compound (compound 8).

[0074] The fourth compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and Michael addition reaction was carried out under the fifth reaction conditions to give the fifth compound (compound 9);

[0075] Under a nitrogen atmosphere, the second compound, the fifth compound, and ammonium acetate were dissolved in n-butanol medium and reacted under the sixth reaction conditions to obtain the sixth compound (compound 10);

[0076] Under a nitrogen atmosphere, the sixth compound and boron trifluoride diethyl ether were placed in 1,2-dichloroethane solvent, with N,N-diisopropylethylamine as the organic base, and reacted under the seventh reaction conditions to obtain the fluorinated borate product NBDP.

[0077] Under a nitrogen atmosphere, NBDP and 1,1'-ferrocene dicarboxylic acid (Fc-2COOH) were dissolved in dichloromethane, with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI) as the carboxyl activator and 4-dimethylaminopyridine (DMAP) as the reaction catalyst. The reaction was carried out under the eighth reaction condition to obtain the dimer type Aza-BODIPY-based molecule NBDP-Fc.

[0078] In an optional example, the molar ratio of 4-hydroxyacetophenone, 4-cyanobenzaldehyde and KOH is 1:(0.5-1.5):(2-3), and is particularly preferred to be 1:1:2; the first reaction conditions include reacting at a temperature of 30°C-35°C for 18-24 hours.

[0079] In an optional example, the molar ratio of the first compound, nitromethane, and triethylamine is 1:(8-12):(8-12), particularly preferably 1:10:10; the second reaction conditions include reacting at a temperature of 70°C-80°C for 24-30 hours.

[0080] In an optional example, the molar ratio of 4-iodoacetophenone, 4-diphenylaminobenzaldehyde and KOH is 1:(0.5-1.5):(1-2), and is particularly preferred to be 1:1:1; the third reaction conditions include reacting at a temperature of 30°C-35°C for 18-24 hours.

[0081] In an optional example, the molar ratio of the third compound, cyanide ketone, and iodide ketone is 1:(1.5-2.5):(0.3-1), and is particularly preferred to be 1:2:0.5; the fourth reaction conditions include reacting at a temperature of 120°C-150°C for 24-30 h.

[0082] In an optional example, the molar ratio of the fourth compound, nitromethane, and triethylamine is 1:(4-6):(4-6), particularly preferably 1:5:5; the fifth reaction conditions include reacting at a temperature of 70°C-80°C for 12-24 hours.

[0083] In an optional example, the molar ratio of the second compound, the fifth compound, and ammonium acetate is 1:(0.5-1.5):(55-65), particularly preferably 1:1:60; the sixth reaction conditions include reacting at a temperature of 125°C-135°C for 24-30 hours.

[0084] In an optional example, the molar ratio of the sixth compound, boron trifluoride ether, and N,N-diisopropylethylamine is 1:(15-20):(10-11), particularly preferably 1:16:11; the seventh reaction conditions include reacting at a temperature of 45°C-55°C for 12-16 hours.

[0085] In an optional example, the molar ratio of Fc-2COOH, EDCI, DMAP, and NBDP is 1:(2-3):(0.2-0.3):(2.5-3.5), and is particularly preferred to be 1:2:0.25:3; the eighth reaction conditions include reacting at a temperature of 30℃-35℃ for 18h-24h.

[0086] In some optional examples, the aldol condensation reaction is carried out under the first reaction conditions. After the reaction is completed, the crude product is acidified with 2 mol / L hydrochloric acid until a solid precipitates out. After filtration and washing with methanol, a pale yellow solid compound 3 is obtained.

[0087] In some optional examples, a Michael addition reaction occurs under the second reaction conditions. After the reaction is complete, the solvent in the reaction mixture is removed, and the crude product is purified by column chromatography to give a white solid compound 4.

[0088] In some optional examples, the aldol condensation reaction is carried out under the third reaction condition. After the reaction is completed, the crude product is filtered and washed with methanol to obtain an orange solid compound 7.

[0089] In some optional examples, the reaction is carried out under the fourth reaction condition. After the reaction is complete, the solvent in the reaction mixture is removed, and the crude product is purified by column chromatography to give yellow compound 8.

[0090] In some optional examples, the Michael addition reaction occurs under the fifth reaction condition. After the reaction is complete, the solvent in the reaction mixture is removed, and the crude product is purified by column chromatography to give white compound 9.

[0091] In some optional examples, the reaction is carried out under the sixth reaction condition. After the reaction is complete, the crude product is washed with cold ethanol and filtered to obtain a black solid compound 10.

[0092] In some optional examples, the reaction is carried out under the seventh reaction condition. After the reaction is complete, the solvent in the reaction mixture is removed, and the crude product is purified by column chromatography to obtain a black solid NBDP.

[0093] In some optional examples, the reaction is carried out under the eighth reaction condition. After the reaction is complete, the solvent in the reaction mixture is removed, and the crude product is purified by column chromatography to obtain a dark blue metallic solid NBDP-Fc.

[0094] In an optional example, the solvent in the reaction mixture is removed by extracting the reaction mixture with dichloromethane and removing the solvent by vacuum distillation.

[0095] application

[0096] In another exemplary embodiment of the present invention, the application of the aforementioned dimer-type Aza-BODIPY-based molecule in the preparation of phototherapeutic reagents for near-infrared II fluorescence / photoacoustic imaging-mediated tumor chemical kinetics, type I photodynamic therapy, and photothermal combined therapy is also provided.

[0097] In another exemplary embodiment of the present invention, a phototherapy reagent prepared using the aforementioned dimer-type Aza-BODIPY-based molecule is also provided. This reagent is formed by the self-assembly of NBDP-Fc and poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) triblock copolymer (F-127) into nanoparticles, which have both near-infrared II fluorescence imaging and chemical kinetics / type I photodynamic / photothermal combined therapy capabilities. It can catalyze the generation of highly toxic hydroxyl radicals from hydrogen peroxide overexpressed in the tumor microenvironment, exhibiting excellent chemical kinetic performance. At the same time, under 808nm laser excitation, the reagent exhibits strong near-infrared II fluorescence emission and can efficiently generate superoxide anions, while achieving a photothermal conversion efficiency of up to 45.6%, thus possessing both near-infrared II imaging and multimodal therapy functions.

[0098] To facilitate better understanding, the present invention will be further illustrated below with several specific examples, but the preparation process is not limited to these examples, and the content of the present invention is not limited to these examples.

[0099] Unless otherwise specified, the materials in the examples are prepared according to existing methods or purchased directly from the market.

[0100] Example 1

[0101] according to Figure 1The synthesis was carried out using the following synthetic route, and the specific process is as follows:

[0102] [Synthesis of Compound 3]

[0103] Compound 1 (1.36 g, 10 mmol) and compound 2 (1.31 g, 10 mmol) were dissolved in anhydrous ethanol (50 mL). Then, KOH (1.12 g, 20 mmol) was added, and the reaction was continued with stirring at room temperature for 12 hours. The precipitate was filtered, washed with ethanol, and then dried under vacuum to give compound 3 (1.89 g, 75%) as a pale yellow solid.

[0104] 1 H NMR (400MHz, DMSO-d6): δppm 10.50 (s, 1H), 8.11-8.06 (m, 5H), 7.92 (d, J = 8.0Hz, 2H), 7.71 (d, J = 16.0Hz, 1H), 6.92-6.89 (m, 2H).

[0105] 13 C NMR (100MHz, DMSO-d6): δppm 186.95, 162.61, 140.54, 139.55, 132.73, 131.45, 129.33, 128.81, 125.50, 118.74, 115.51, 115.47, 112.05.

[0106] The structural formula of compound 3 can be determined as shown in Formula III:

[0107]

[0108] [Synthesis of Compound 4]

[0109] Compound 3 (1.25 g, 5 mmol) was added to a 250 mL double-necked flask containing 25 mL of anhydrous methanol. Then, under a nitrogen atmosphere, nitromethane (1.25 mL, 50 mmol) and triethylamine (3.50 mL, 50 mmol) were added dropwise sequentially, and the mixture was refluxed at 75 °C for 12 hours. After the reaction was complete, the mixture was cooled to room temperature, acidified to pH 3 with 2 mol / L hydrochloric acid, extracted with dichloromethane, and purified by column chromatography using ethyl acetate / petroleum ether (1:1) as eluent to give a white solid, compound 4 (2.11 g, 68%).

[0110] 1H NMR (400MHz, CDCl3): δppm 7.84(d,J=12.0Hz,2H),7.63(d,J=8.0Hz,2H),7.42(d,J=8.0Hz,2H),6.87(d,J=8.0Hz,2H),6.19(s, 1H), 4.85 (dd, J=8.0Hz, 12.0Hz, 1H), 4.70 (dd, J=8.0Hz, 12.0Hz, 1H), 4.28 (m, 1H), 3.45-3.34 (m, 2H).

[0111] 13 C NMR (100MHz, DMSO-d6): δppm 195.11,170.47,162.51,146.27,132.45,130.67,129.18,127.99,118.80,115.40,110.27,79.17,59.88,20.78,14.11.

[0112] The structural formula IV of compound 4 can be determined as shown in the formula:

[0113]

[0114] [Synthesis of Compound 7]

[0115] Compound 5 (4.90 g, 0.02 mol) and compound 6 (5.47 g, 0.02 mol) were dissolved in anhydrous ethanol (50 mL), followed by the addition of KOH (1.12 g, 0.02 mol), and the reaction was continued with stirring at room temperature for 24 hours. The precipitate was filtered, washed with anhydrous ethanol, and then dried under vacuum to give an orange solid, compound 7 (7.01 g, 70%).

[0116] 1 H NMR (400MHz, CDCl3) δppm 7.85(d,J=8.0Hz,2H),7.76(s,1H),7.71(d,J=8.0Hz,2H),7.48(d,J=8.0Hz,2H),7.33-7.29(m,5H),7.16-7.09(m,6H),7.02(d,J=8.0Hz,2H).

[0117] 13 C NMR (100MHz, CDCl3): δppm 189.66, 150.37, 146.70, 145.30, 137.89, 137.80, 129.83, 129.52, 127.46, 125.54, 124.23, 121.35, 118.59, 100.15.

[0118] The structural formula V of compound 7 can be determined as shown in the formula:

[0119]

[0120] [Synthesis of Compound 8]

[0121] Compound 7 (4.01 g, 8 mmol) and cuprous cyanide (1.43 g, 16 mmol) were dissolved in N,N-dimethylformamide (15 mL). Then, under a nitrogen atmosphere, cuprous iodide (0.06 g, 0.3 mmol) was added, and the mixture was heated to 130 °C for 24 hours. After cooling to room temperature, 100 mL of water was added to the crude product, and the mixture was extracted with dichloromethane. The extract was then purified by column chromatography using dichloromethane / petroleum ether (1:1) as the eluent to give a yellow solid, compound 8 (1.92 g, 60%).

[0122] 1 H NMR(400MHz,DMSO-d6)δppm 8.25(d,J=12.0Hz,2H),8.04(d,J=8.0Hz,2H),7.75(dd,J=8.0Hz,16.0Hz,4H),7.40-7.36(m,4H),7.18-7.11(m,6H),6.90(d,J=12.0Hz,2H).

[0123] 13 C NMR (100MHz, CDCl3): δppm 189.02,150.73,146.52,146.45,142.02,132.39,130.06,129.56,128.72,126.93,125.68,124.43,121.02,118.12,115.52.

[0124] The structural formula VI of compound 8 can be determined as shown in the formula:

[0125]

[0126] [Synthesis of Compound 9]

[0127] Compound 8 (1.60 g, 4 mmol) was added to a 250 mL double-necked flask containing anhydrous methanol (25 mL). Then, under a nitrogen atmosphere, nitromethane (1.22 g, 20 mmol) and triethylamine (2.02 g, 20 mmol) were added dropwise, and the mixture was refluxed for 12 hours. After the reaction was complete, the mixture was cooled to room temperature, adjusted to pH 3 with 4 mol / L hydrochloric acid, extracted with dichloromethane, and purified by column chromatography with ethyl acetate / petroleum ether (1:6) to give a white solid compound 9 (1.01 g, 55%).

[0128] 1 H NMR(400MHz,DMSO-d6)δppm 8.04(q,J=8.0Hz,24.0Hz,4H),7.30-7.26(m,6H),7.02(t,J=16.0Hz,2H),6.91(dd,J=8.0Hz,12.0Hz,6H) ,4.96(dd,J=4.0Hz,12.0Hz,1H),4.82(dd,J=12.0Hz,16.0Hz,1H),4.02–3.94(m,1H),3.63–3.51(m,2H).

[0129] 13 C NMR (100MHz, CDCl3): δppm 195.79,147.60,147.36,139.25,132.59,131.77,129.30,128.45,128.09,124.55,123.46,123.21,117.73,116.76,79.41,41.85,38.56.

[0130] The structural formula VII of compound 9 can be determined as shown in the formula:

[0131]

[0132] [Non-NBDP Synthesis]

[0133] Compound 4 (0.62 g, 2 mmol) and compound 9 (0.83 g, 2 mmol) were dissolved in anhydrous n-butanol (15 mL), followed by the addition of ammonium acetate (9.25 g, 120 mmol). The reaction was carried out at 50 °C for 24 hours under a nitrogen atmosphere. Finally, the reaction was cooled to room temperature, the resulting precipitate was filtered, washed with cold ethanol, and dried under vacuum to give a black solid, compound 10, which was used directly as an intermediate in the next step. The structural formula of compound 10 is shown in Formula VIII:

[0134]

[0135] Compound 10 (0.62 g, 1 mmol) was dispersed in anhydrous 1,2-dichloroethane (15 mL), and then N,N-diisopropylethylamine (DIPEA) (1.85 mL, 10 mmol) and boron trifluoride diethyl ether (48% BF3Et2O, 3.5 mL, 14 mmol) were added dropwise under a nitrogen atmosphere. The mixture was heated to 50 °C and reacted for 24 hours. After cooling to room temperature, it was extracted with dichloromethane. Finally, it was purified by column chromatography with silica gel using dichloromethane / petroleum ether (2:1) as the eluent to give a black solid NBDP (0.13 g, 18%).

[0136] 1 H NMR (400MHz, CDCl3) δppm 8.12(q,J=8.0Hz,4H),8.03(d,J=8.0Hz,2H),7.88(d,J=12.0Hz,2H),7.76(d,J=8.0Hz,2H),7.66(d,J=8.0H z,2H),7.37(t,J=8.0Hz,4H),7.23-7.15(m,7H),7.08(d,J=8.0Hz,2H),6.96(d,J=8.0Hz,2H),6.85(s,1H).

[0137] The structural formula IX of NBDP can be determined as shown in the formula:

[0138]

[0139] [Synthesis of NBDP-Fc]

[0140] In a dry 250 mL double-necked flask, 1,1'-ferrocene dicarboxylic acid (0.14 g, 0.3 mmol), EDCI (0.09 g, 0.4 mmol), and DMAP (0.02 g, 0.2 mmol), along with 10 mL of anhydrous dichloromethane, were added. The mixture was reacted at 35 °C for 180 min under a nitrogen atmosphere, followed by the addition of NBDP (0.23 g, 0.4 mmol). The reaction mixture was stirred at room temperature under a nitrogen atmosphere for 12 h. After the reaction was complete, 100 mL of H₂O was added, and the mixture was extracted with dichloromethane. The solvent was removed by vacuum distillation. Finally, the crude product was purified by silica gel chromatography (ethyl acetate / petroleum ether = 1:1) to give a deep blue metallic solid NBDP-Fc (0.19 g, 57.2%).

[0141] like Figure 2 As shown, 1H NMR(400MHz,C2D2Cl4-d2)δppm 8.09(d,J=8.0Hz,2H),7.98(d,J=12.0Hz,2H),7.80-7.72(m,12H),7.37-7.29(m,12H),7.2 1-7.17(m,16H),7.08(d,J=8.0Hz,2H),6.98(s,1H),6.82(s,1H),5.17(s,4H),4.63(s,4H).

[0142] like Figure 3 As shown, 13 C NMR (100MHz, CDCl3): δppm 167.54,157.52,156.64,152.84,150.13,146.17,145.71,145.30,144.65,138.60,135.53,132.37,131.50,130.87,130.36,129.78,12 9.69,128.97,127.12,125.83,125.01,124.44,122.14,120.54,119.98,118.70,118.42,116.15,113.84,111.74,72.75,72.51,72.09.

[0143] like Figure 4 As shown, MALDI-TOF mass spectrometry (m / z): calculated for C 104 H 64 B2F4FeN 12 O4[M+H] + :1699.542; found,1699.566.

[0144] Combination Figures 2-4 As shown, the molecular structure of NBDP-Fc can be determined to be Formula I.

[0145] Example 2

[0146] [Preparation of NBDP-Fc Nanoparticles]

[0147] NBDP-Fc nanoparticles were prepared via a nanoprecipitation method, as detailed below:

[0148] NBDP-Fc molecules (1 mg) (prepared according to the method of Example 1) and F127 (10 mg) were dissolved in tetrahydrofuran (1 mL) and sonicated for 5 minutes.

[0149] Then, the solution was quickly injected into deionized water (10 mL) and sonicated for 1 hour. After stirring and evaporating to completely remove tetrahydrofuran, it was filtered with a 0.22 μm filter to obtain a clear and transparent dark blue NBDP-Fc nanoparticle solution, which was stored at 4 °C for later use.

[0150] like Figure 5 As shown, the dynamic light scattering particle size distribution test results of NBDP-Fc nanoparticles show that the hydrated particle size is 47 nm, indicating that the size of the nanoparticles meets the requirements of enhanced permeability and retention (EPR).

[0151] like Figure 6 As shown, the absorption spectrum of NBDP-Fc nanoparticles in water covers the range of 400 nm to 1000 nm and shows two absorption bands with absorption peaks at 646 nm and 781 nm, respectively. The absorption peak at 781 nm extends to 1000 nm, and the emission peak is located at 1080 nm, which is in the near-infrared II region. This indicates that the nanoparticles can be used for near-infrared II fluorescence imaging of in vivo tumors.

[0152] Example 3

[0153] [Test of hydroxyl radical generation performance of NBDP-Fc nanoparticles]

[0154] The ability of NBDP-Fc NPs to generate ·OH by consuming hydrogen peroxide was detected using 3,3',5,5'-tetramethylphenyldiamine (TMB) as a hydroxyl radical probe.

[0155] TMB (1 μL, 0.1 M) and hydrogen peroxide (5 μL, 30%) were added to NBDP-Fc NPs (150 μL, 100 ppm). The illumination group involved exposing the mixed solution to an 808 nm laser (1.0 W / cm²). 2 Irradiated for 10 minutes. The absorbance of TMB at 652 nm was monitored using a UV-Vis spectrophotometer. The following experiments were performed simultaneously: TMB + H₂O₂, TMB + H₂O₂ + NBDP NPs solution group, TMB + H₂O₂ + NBDP-Fc NPs solution group, and TMB + H₂O₂ + NBDP-Fc NPs + L solution group. "+L" indicates the irradiated group. After the reaction, the UV absorbance of the reaction system was measured using a UV spectrophotometer.

[0156] like Figure 7 As shown, NBDP-Fc NPs can consume hydrogen peroxide to generate hydroxyl radicals, and this ability is even stronger after irradiation with an 808nm laser.

[0157] This indicates that the thermal effect generated by the NBDP-Fc NPs of the present invention after light irradiation can promote the Fenton reaction to generate more hydroxyl radicals, which directly proves that the heat generated by the photothermal effect accelerates the Fenton reaction and promotes chemokinetic therapy.

[0158] Example 4

[0159] [Photodynamic Performance Test of NBDP-Fc Nanoparticles]

[0160] 2,7-Dichlorodihydrofluorescein (DCFH) and dihydrorhodamine 123 (DHR 123) were used as the total ROS and O2, respectively. ·- Specific probes were used to detect the ROS generation capacity of NBDP NPs and NBDP-Fc NPs.

[0161] The probes were thoroughly dissolved in aqueous solutions of NBDP NPs and NBDP-Fc NPs, respectively, and analyzed using an 808 nm laser (0.1 W / cm²). 2 Irradiation was performed for 150 seconds, with fluorescence data recorded every 30 seconds. Simultaneously, the same procedure was performed using separate aqueous solutions of DCFH and DHR 123 as control groups.

[0162] like Figures 8-9 As shown, under irradiation by an 808nm laser, the monomeric NBDP NPs produced almost no ROS, while the dimer NBDP-Fc NPs exhibited significant type I ROS (O2). ·- ) generates capacity.

[0163] This indicates that the NBDP-Fc NPs of the present invention generate ROS through a type I photodynamic pathway, and that the dimer structure can promote ROS generation. The aqueous solution of the NBDP-Fc nanoparticles of the present invention exhibits excellent O2 content. ·- Generation capacity, O2 ·- It can be converted into hydrogen peroxide in the body, which provides a key reaction substrate for the chemokinetic treatment process, thereby promoting chemokinetic therapy.

[0164] Example 5

[0165] [Photothermal Performance Test of NBDP-Fc Nanoparticles]

[0166] Using an 808nm laser (1W / cm) 2 Irradiate 1 mL of NBDP-Fc nanoparticle aqueous solution (100 μg / mL) and 1 mL of deionized water. After heating for 10 minutes, allow the solution to cool naturally for 12 minutes to return to room temperature. Calculate the photothermal conversion efficiency of the NBDP-Fc nanoparticle aqueous solution using the cooling period.

[0167] like Figure 10 As shown, the photothermal conversion efficiency of the NBDP-Fc nanoparticle aqueous solution of the present invention is 45.6%.

[0168] Example 6

[0169] [Cytotoxicity assay of NBDP-Fc nanoparticles]

[0170] 4T1 cells were loaded at 5.0 × 10⁶ cells per well. 3 Cells were seeded at densities of [number] cells per well in two 96-well plates and incubated at 37°C in a dark environment with 5% CO2 for 24 hours. Different concentrations of NBDP-Fc nanoparticles (0, 5, 10, 15, 20, 30, 40, 60 μg / mL) were then added to the plates. -1 Add the nanoparticles to the well plate and incubate for 12 hours (two well plates are used here, each with a different concentration of NBDP-Fc nanoparticles).

[0171] After incubation for 12 hours, the sample was subjected to an 808nm laser (1W / cm²) at room temperature. 2 One well of the plate was irradiated for 3 minutes per well, while the other well was incubated in the dark to test the cytotoxicity of NBDP-Fc nanoparticles.

[0172] After illumination, the two wells were incubated in the dark for 12 hours. Then, 200 μL of MTT solution was added to each well and the cells were incubated in the dark for 4 hours. The culture medium was removed, and 200 μL of dimethyl sulfoxide was added to dissolve the blue-purple formazan crystals. The absorbance at 490 nm was read using a microplate reader, and the viability of 4T1 cells under different concentrations of NBDP-Fc nanoparticles was calculated. The cell viability was calculated as: (Average absorbance of the treatment group / Average absorbance of the control group) × 100%.

[0173] like Figure 11 As shown, under dim light, because the ferrocene unit can act as a catalyst for the Fenton reaction, catalyzing the generation of hydroxyl radicals from hydrogen peroxide, causing oxidative damage to cells, the cell viability decreases gradually with increasing nanomaterial concentration. When the concentration of NBDP-Fc nanoparticles reaches 60 μg / mL... -1 The survival rate of 4T1 cells remained at around 60%; however, under light conditions, only 40 μg / mL... -1 The NBDP-Fc nanoparticles were able to kill nearly 90% of 4T1 cells, demonstrating that the NBDP-Fc nanoparticles of this invention exhibit excellent biocompatibility and high photocytotoxicity in the absence of light exposure, and can significantly kill cancer cells under light exposure. This result clearly shows that the cytotoxicity of the three-mode combined therapy is far superior to that of single chemokinetic therapy.

[0174] Example 7

[0175] [NIR-II fluorescence imaging of NBDP-Fc nanoparticles in mice]

[0176] NBDP-Fc nanoparticles (200 μL, 500 μg mL) were added. -1 The tumor was injected into mice via the tail vein, and fluorescence images of the tumor site were recorded at different time points (10 min, 3, 6, 9, 12, 20, 24 hours).

[0177] like Figure 12 As shown, after intravenous injection of NBDP-Fc nanoparticles into mice, the fluorescence signal at the tumor site gradually increased over time, reaching its maximum value at around 20 hours, and then gradually weakened.

[0178] Example 8

[0179] [Photoacoustic imaging of NBDP-Fc nanoparticles in mice]

[0180] NBDP-Fc nanoparticles (200 μL, 500 μg mL) were added. -1 The tumor was injected into mice via the tail vein, and photoacoustic images of the tumor site were recorded at different time points (pre, 3, 6, 9, 12, 16, and 20 hours before injection).

[0181] like Figure 13 As shown, after intravenous injection of NBDP-Fc nanoparticles into mice, the photoacoustic signal at the tumor site gradually increased over time, reaching its maximum value at around 20 hours.

[0182] As can be seen from Examples 7 and 8, the NBDP-Fc nanoparticles of the present invention can be effectively enriched at the tumor site, with the optimal diagnostic window being 20 hours after drug injection, enabling high-resolution imaging of deep tissues and precise tumor localization.

[0183] Example 9

[0184] [In vivo photothermal imaging of NBDP-Fc nanoparticles in mice]

[0185] NBDP-Fc nanoparticles (200 μL, 500 μg mL) were added. -1 The tumor site was recorded at different time points (0, 1, 2, 3, 4, 5 minutes) by intravenous injection of either PBS buffer (200 μL) into mice via the tail vein.

[0186] like Figure 14As shown, under irradiation by an 808nm laser, the tumor temperature of mice in the PBS group only increased to 40.0℃ within five minutes, while the tumor temperature of mice in the NBDP-Fc nanoparticle group rapidly increased to 48.2℃ within five minutes, indicating that the NBDP-Fc nanoparticles of the present invention can be effectively used for photothermal therapy of mouse tumors.

[0187] Example 10

[0188] [Tumor Therapy Experiment with NBDP-Fc Nanoparticles]

[0189] 4T1 tumor-bearing mouse models were established by subcutaneous injection of 4T1 cancer cells until the tumor volume reached 100 mm. 3 After moving left and right, subsequent experiments can be conducted.

[0190] 4T1 tumor-bearing mice were randomly divided into three groups (PBS+L, NPs, and NPs+L, n=5 per group, where L represents laser irradiation). Mice in the PBS+L group were intravenously injected with PBS (200 μL), while mice in the NPs and NPs+L groups were intravenously injected with NBDP-Fc nanoparticles (200 μL, 500 μg / mL). -1 Twenty hours after injection, the tumor sites of mice in the PBS+L group and NPs+L group were treated with an 808nm laser (1W / cm²). 2 Irradiate for 5 minutes. After one treatment, the tumor size is measured every 2 days until day 16.

[0191] like Figure 15 As shown, after one treatment, the tumors in the NPs+L group were completely eliminated on the 4th day after treatment and there was no recurrence within 16 days. In contrast, the tumor volume in the PBS+L group and the NPs group increased by 3 to 5 times on the 16th day after treatment. This indicates that the NBDP-Fc nanoparticles of the present invention have significant anti-tumor effects under laser irradiation, demonstrating their potential for combined chemical kinetics, type I photodynamic therapy and photothermal therapy.

[0192] The above tests show that the dimer-type Aza-BODIP-based molecule of the present invention not only exhibits strong near-infrared II fluorescence, but also can efficiently generate superoxide anions, while achieving a photothermal conversion efficiency of up to 45.6%. This single-molecule reagent can be used for precise and efficient near-infrared II fluorescence / photoacoustic imaging-mediated chemical kinetics, type I photodynamic therapy and photothermal multimodal combined therapy in deep tumor tissues, reducing tumor treatment damage and side effects, and improving treatment efficiency and accuracy.

[0193] While the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the invention. Those skilled in the art can make various modifications and refinements without departing from the spirit and scope of the invention. Therefore, the scope of protection of the present invention shall be determined by the claims.

Claims

1. A dimer-type Aza-BODIPY-based molecule, characterized in that, This dimer-type Aza-BODIPY molecule is formed by coupling Aza-BODIP molecule with 1,1'-ferrocene dicarboxylic acid. The chemical structural formula of the Aza-BODIP-based monomer molecule is shown in Formula I, and is denoted as NBDP; The structural formula of the dimer type Aza-BODIPY molecule is shown in Formula II, and is denoted as NBDP-Fc; 2. A method for preparing the dimer-type Aza-BODIPY-based molecule as described in claim 1, characterized in that, Includes the following steps: Under a nitrogen atmosphere, 4-hydroxyacetophenone and 4-cyanobenzaldehyde were dissolved in anhydrous ethanol, and then KOH was added. Under the first reaction conditions, an aldol condensation reaction was carried out to obtain the first compound. The first compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and a Michael addition reaction was carried out under the second reaction conditions to give the second compound. Under a nitrogen atmosphere, 4-iodoacetophenone and 4-diphenylaminobenzaldehyde were dissolved in anhydrous ethanol, then KOH was added, and an aldol condensation reaction was carried out under the third reaction conditions to give the third compound. The third compound, cyanide ketone, and iodide ketone were dissolved in anhydrous N,N-dimethylformamide and reacted under the fourth reaction conditions to obtain the fourth compound; The fourth compound and nitromethane were placed in methanol solvent, with triethylamine as the organic base, and a Michael addition reaction was carried out under the fifth reaction conditions to give the fifth compound. Under a nitrogen atmosphere, the second compound, the fifth compound, and ammonium acetate were dissolved in n-butanol medium and reacted under the sixth reaction conditions to obtain the sixth compound; Under a nitrogen atmosphere, the sixth compound and boron trifluoride diethyl ether were placed in a 1,2-dichloroethane solvent, with N,N-diisopropylethylamine as the organic base, and reacted under the seventh reaction conditions to give the fluorinated borate product NBDP. Under a nitrogen atmosphere, NBDP and 1,1'-ferrocene dicarboxylic acid (Fc-2COOH) were dissolved in dichloromethane, with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDCI) as the carboxyl activator and 4-dimethylaminopyridine (DMAP) as the reaction catalyst. The reaction was carried out under the eighth reaction condition to obtain the dimer type Aza-BODIPY-based molecule NBDP-Fc.

3. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of 4-hydroxyacetophenone, 4-cyanobenzaldehyde and KOH is 1:(0.5-1.5):(2-3); the first reaction conditions include: reacting at 30℃-35℃ for 18h-24h; The molar ratio of the first compound, nitromethane, and triethylamine is 1:(8-12):(8-12); the second reaction conditions include: reacting at 70℃-80℃ for 24-30 hours.

4. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of 4-iodoacetophenone, 4-diphenylaminobenzaldehyde and KOH is 1:(0.5-1.5):(1-2); the third reaction conditions include: reacting at 30℃-35℃ for 18h-24h.

5. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of the third compound, cyanide ketone, and iodide ketone is 1:(1.5-2.5):(0.3-1); the fourth reaction conditions include: reacting at 120℃-150℃ for 24-30 hours. The molar ratio of the fourth compound, nitromethane, and triethylamine is 1:(4-6):(4-6); the fifth reaction conditions include: reacting at 70℃-80℃ for 12h-24h.

6. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of the second compound, the fifth compound, and ammonium acetate is 1:(0.5-1.5):(55-65); The sixth reaction conditions include: reacting at a temperature of 125℃-135℃ for 24-30 hours.

7. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of the sixth compound, boron trifluoride ether, and N,N-diisopropylethylamine is 1:(15-20):(10-11); the seventh reaction conditions include: reacting at 45℃-55℃ for 12-16 hours.

8. The method for preparing the dimer-type Aza-BODIPY-based molecule according to claim 2, characterized in that, The molar ratio of Fc-2COOH, EDCI, DMAP, and NBDP is 1:(2-3):(0.2-0.3):(2.5-3.5); the eighth reaction conditions include: reacting at a temperature of 30℃-35℃ for 18-24 hours.

9. The use of the dimer-type Aza-BODIPY-based molecule of claim 1 in the preparation of phototherapy reagents for near-infrared II fluorescence / photoacoustic imaging-mediated tumor chemokinetics, type I photodynamic therapy, and photothermal combined therapy.

10. A phototherapy reagent prepared using the dimer-type Aza-BODIPY-based molecule as described in claim 1.

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