NO / acid two-factor activated near-infrared light excited iridium (III) complex photosensitizer as well as preparation method and application of NO / acid two-factor activated near-infrared light excited iridium (III) complex photosensitizer

Abstract

The invention discloses a NO / acid two-factor activated near-infrared light excited iridium (III) complex photosensitizer as well as a preparation method and application thereof, and belongs to the technical field of biomedical materials. The NO / acid two-factor activated near-infrared light excited iridium (III) complex photosensitizer provided by the invention is a class I photosensitizer without near-infrared light absorption (no absorption is realized when the wavelength is greater than 650nm), and can be activated into a near-infrared absorption iridium (III) complex (class II photosensitizer) in the presence of NO and acid at the same time; under the excitation of near-infrared light, the iridium (III) complex photosensitizer (class II photosensitizer) activated by NO / acid double factors has efficient photodynamic performance and photothermal conversion performance, and part of photophysical performance and killing performance to tumor cells can be improved.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biomedical materials, and particularly relates to a NO / acid dual-factor activated near-infrared light-excited iridium (III) complex photosensitizer, a preparation method and application thereof. BACKGROUND

[0002] Photodynamic therapy (PDT) is an efficient treatment method, which uses a combination of photosensitizers, light and oxygen to produce cytotoxic reactive oxygen species (ROS) to eliminate cancer cells. As a clinically recognized non-invasive treatment, PDT has the advantages of precise controllability and negligible multi-drug resistance compared with traditional chemotherapy or surgery in cancer treatment. However, due to the phototoxicity of adjacent normal tissues, its use is still limited. In addition, in order to reach the concentration required for tumor treatment, photosensitizers need to be repeatedly administered, which has the risk of photosensitizer overdose, which may cause systemic side effects.

[0003] Early traditional photosensitizers (such as porphyrin derivatives) rely on direct activation of continuous light sources, although they have mature safety and clinical applicability, but their non-specific photoactivation and dependence on long-wavelength light result in limited tissue penetration depth and easy induction of normal tissue phototoxicity. Activated photosensitizers significantly optimize the precision and safety of traditional photosensitizers in PDT, which selectively activate and produce ROS by responding to tumor microenvironment-specific signals (such as acidity, high concentration of reduced glutathione, etc.), and only start the photodynamic effect in the targeted area, thereby greatly reducing the phototoxicity to normal tissues. However, single-factor activation mechanism is easily disturbed by biological marker heterogeneity, and there is still a risk of false positive / negative. Further developed dual-factor activated photosensitizers enhance tumor recognition specificity by integrating dual biological markers (such as pH / enzyme cooperative response), but most systems are still limited by short-wavelength excitation light sources, making it difficult to achieve effective treatment of deep lesions. Recently, iridium (III) complex dual-factor activated photosensitizers have attracted attention due to their excellent photophysical properties (such as high singlet oxygen quantum yield, long excited state lifetime, etc.) and precise dual-target response capability. However, their excitation wavelengths are mostly concentrated in the visible light range, and the tissue penetration ability is insufficient, which restricts their application potential in in vivo tumor treatment. Therefore, developing a new photosensitizer system with high selective dual-activation characteristics and near-infrared light response has become a key direction to break through the current technical bottleneck. SUMMARY

[0004] The present application aims to provide a NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer, a preparation method and application thereof. The NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer provided by the present application is a type I photosensitizer without near-infrared light absorption (no absorption above 650 nm), which can be activated into an iridium (III) complex (type II photosensitizer) with near-infrared absorption under the condition of the simultaneous presence of NO and acid; under the excitation of near-infrared light, the NO / acid dual-factor activated iridium (III) complex photosensitizer (type II photosensitizer) has high photodynamic performance and photothermal conversion performance, and can improve part of the photophysical performance and the killing performance on tumor cells. The NO / acid dual-factor activated near-infrared light excited iridium (III) complex provided by the present application aims to solve the problems of traditional single-factor activated false positives and short excitation light of iridium (III) complexes applied in vivo.

[0005] To achieve the above-mentioned purpose, the present application provides the following technical solutions.

[0006] One of the technical solutions of the present application provides a NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer, the structure formula of which is as follows:

[0007]

[0008] In the above formula, the C^N ligand is The X group is

[0009] R in the X group is any one of R1-R5:

[0010]

[0011] The second technical solution of the present application provides a NO / acid dual-factor activated iridium (III) complex photosensitizer, the structure formula of which is as follows:

[0012]

[0013] In the above formula, the C^N ligand is The X group is

[0014] R in the X group is any one of R1-R5:

[0015]

[0016] The third technical solution of the present application provides a preparation method of the above-mentioned NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer, comprising the following steps:

[0017] Mixing compound A, hydrazine hydrate, solvent and catalyst, reacting under inert atmosphere to obtain the NO / acid dual factor activated near-infrared light excited iridium (III) complex photosensitizer;

[0018] The structural formula of the compound A is as follows:

[0019]

[0020] The compound 1 of the present application is synthesized by referring to the method disclosed in Chinese invention patent CN118930586A.

[0021] Optionally, the solvent is ethanol.

[0022] Preferably, the catalyst is palladium-carbon catalyst (Pd / C).

[0023] Preferably, the temperature of the reaction is 70-90 DEG C, and the time is 8-14 h.

[0024] The fourth technical scheme of the present application provides a preparation method of the above-mentioned NO / acid dual factor activated iridium (III) complex photosensitizer, comprising the following steps:

[0025] Mixing the above-mentioned iridium (III) complex photosensitizer, NO donor and acid liquid, and reacting under inert atmosphere to obtain the NO / acid dual factor activated iridium (III) complex photosensitizer.

[0026] Optionally, the NO donor is sodium nitrite (NaNO2).

[0027] Optionally, the pH value of the acid liquid ranges from 5.4 to 6.4.

[0028] Preferably, the time of the reaction is 2-4 h.

[0029] The fifth technical scheme of the present application provides an application of the above-mentioned NO / acid dual factor activated near-infrared light excited iridium (III) complex photosensitizer in preparing photosensitizer in photodynamic therapy.

[0030] The sixth technical scheme of the present application provides an application of the above-mentioned NO / acid dual factor activated iridium (III) complex photosensitizer in preparing photosensitizer in photodynamic therapy.

[0031] The present application has the following beneficial technical effects:

[0032] The application provides a kind of NO / acid double-factor activated near-infrared light excited iridium (III) complex photosensitizer and its activator.The application utilizes D-A-D structure nitrogen nitrogen ligand desulfurization to obtain o-phenylenediamine group, regulates host ligand structure, and constructs "NO / acid" activated iridium (III) complex photosensitizer.With the increase of the conjugated structure of the complex, the absorption gradually red shifts, and it is found that it appears absorption at 600-750 nm, which shows that the complex can be activated by NO in acid environment.The selected longest absorption wavelength Ir 3 is subjected to depth performance research, and Ir 3-NO is obtained after NO treatment of Ir 3.The experimental results show that the maximum absorption peak of Ir 3-NO is at 600-750 nm, which shows good selectivity and quantification of NO, and shows high sensitivity.Ir 3-NO NPs are obtained by preparing Ir 3-NO into nanoparticles using DSPE-mPEG2000 (distearoyl phosphatidyl ethanolamine-polyethylene glycol 2000), which has good water solubility and biocompatibility.In vitro test shows that Ir 3-NO NPs have good photothermal response under 730 nm laser excitation, and can produce 1 O2 and O2 ·- Cell experiments show that Ir 3-NONPs have good cell dark toxicity and phototoxicity, and generate reactive oxygen species to kill cells in cells.The photosensitizer provided by the application realizes double-factor activation, long-wavelength excitation and photothermal synergistic enhancement, and shows great potential in the field of tumor phototherapy, and has very broad application prospect. BRIEF DESCRIPTION OF DRAWINGS

[0033] Figure 1 The matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 1 in Example 1.

[0034] Figure 2 The nuclear magnetic resonance hydrogen spectrum of Ir 1 in Example 1.

[0035] Figure 3 The matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 1-NO in Example 1.

[0036] Figure 4 The matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 2 in Example 2.

[0037] Figure 5 The nuclear magnetic resonance hydrogen spectrum of Ir 2 in Example 2.

[0038] Figure 6 The matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 2-NO in Example 2.

[0039] Figure 7Matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 3 in Example 3.

[0040] Figure 8 Nuclear magnetic resonance hydrogen spectrum of Ir 3 in Example 3.

[0041] Figure 9 Matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 3-NO in Example 3.

[0042] Figure 10 Absorption spectra of Ir 1, Ir 2, Ir 3 in different solvents and emission spectra in water in Example 4; wherein (a) is the absorption spectrum of Ir 1 in different solvents, (b) is the absorption spectrum of Ir 2 in different solvents, (c) is the absorption spectrum of Ir 3 in different solvents, (d) is the emission spectrum of Ir 1 in water, (e) is the emission spectrum of Ir 2 in water, (f) is the emission spectrum of Ir 3 in water.

[0043] Figure 11 Absorption spectra of Ir 1, Ir 2, Ir 3, Ir 1-NO, Ir 2-NO, Ir 3-NO in water in Example 5, wherein (a) is the absorption spectrum of Ir 1 and Ir 1-NO, (b) is the absorption spectrum of Ir 2 and Ir 2-NO, (c) is the absorption spectrum of Ir 3 and Ir 3-NO.

[0044] Figure 12 Absorption spectra of Ir 3 in different pH conditions in NO environment (a) and absorption spectra of Ir 3 in different NO concentration conditions in pH=5.4 environment (b) in Example 6.

[0045] Figure 13 Absorbance of Ir 3 after reacting with different ions in acidic environment in Example 7.

[0046] Figure 14 Changes of relative emission intensity at 525 nm in active oxygen indicator DCFH aqueous solution with time after 730 nm laser irradiation of Ir 3-NO NPs under normoxic condition (a) and hypoxic condition (b) in Example 8.

[0047] Figure 15 Changes of relative emission intensity at 525 nm in O2 ·- indicator DHR123 aqueous solution with time after 730 nm laser irradiation of Ir 3-NO NPs under normoxic condition (a) and hypoxic condition (b) in Example 9.

[0048] Figure 16Temperature change curves of aqueous solutions of Ir 3-NO NPs with different concentrations as a function of time under 730 nm laser irradiation (a) and temperature change curve of 100 μΜ concentration as a function of time during five heating-cooling cycles (b) for Example 10.

[0049] Figure 17 Results of toxicity test of photosensitizer Ir 3-NO NPs on 4T1 cancer cells under normoxic (a) and hypoxic (b) conditions, in the dark and under 730 nm laser irradiation for Example 11.

[0050] Figure 18 Activation mechanism of photosensitizers Ir 1, 2, 3. DETAILED DESCRIPTION

[0051] The specific embodiments of the present application will now be described in detail with specific reference being made to the figures. It is to be understood that the description given herein is only for the purpose of illustrating preferred embodiments of the present application and is not intended to limit the present application in any way.

[0052] It should be noted that matters described herein which are not described in detail are routine operations in the art and are not the focus of the present application.

[0053] In addition, for numerical ranges recited herein, every intervening value between the upper and lower limits is specifically contemplated. In any stated range, every narrower range that falls within the broader range is also specifically contemplated. The upper and lower limits of these smaller ranges can independently be included or excluded in the smaller ranges.

[0054] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application.

[0055] As used herein, the terms "comprise", "comprising", "include", "including", "have", "having" and the like are open-ended terms that are intended to mean including, but not limited to.

[0056] Normal temperature and room temperature in the embodiments of the present application both refer to temperature of 25±2 °C.

[0057] All raw materials in the embodiments of the present application are commercially available.

[0058] Example 1

[0059] (1) Refer to the method disclosed in Chinese invention patent CN118930586A to synthesize compound 1.

[0060] (2) Preparation of Ir 1, the reaction formula is as follows:

[0061]

[0062] Compound 1 (150 mg, 0.085 mmol), Pd / C (15 mg) were added to a two-necked flask, protected by nitrogen, dissolved in 5 mL of ethanol, 0.5 mL of hydrazine hydrate at room temperature was added, and refluxed at 80°C for 12 h. After the reaction was completed, it was cooled to room temperature, filtered, and the solvent was evaporated. Purification was performed by column chromatography (CH2Cl2:CH3OH = 20:1, v / v). Finally, the product was obtained as a yellow solid Ir 1 182 mg (yield 58%).

[0063] The matrix-assisted laser desorption ionization time-of-flight mass spectrum of Ir 1 is shown in Figure 1 :

[0064] The MALDI-TOF-MS m / z = 1733.066 [M-PF6 - ] + .

[0065] The nuclear magnetic resonance hydrogen spectrum of Ir 1 is shown in Figure 2 :

[0066] 1 H NMR (500 MHz, Chloroform-d) δ (ppm): 9.17 (d, J = 8.4 Hz, 2H), 8.38 (s, 2H), 8.20 (dq, J = 18.8, 9.2, 8.7 Hz, 4H), 8.06 (d, J = 7.8 Hz, 2H), 7.65 (d, J = 6.0 Hz, 2H), 7.52 (dq, J = 28.4, 11.0, 8.9 Hz, 6H), 7.29-7.19 (m, 14H), 7.14-7.02 (m, 18H), 6.85 (d, J = 7.9 Hz, 2H), 6.70 (d, J = 17.1 Hz, 2H), 6.63 (d, J = 8.2 Hz, 2H), 6.26 (s, 4H), 2.45-2.18 (m, 6H), 1.41 (d, J = 39.8 Hz, 4H), 1.01-0.62 (m, 18H).

[0067] (3) Preparation of Ir 1-NO, the reaction formula is as follows:

[0068]

[0069] Ir 1 (100 mg, 0.053 mmol) and NaNO2(18.3 mg, 0.265 mmol) were placed in a dry two-necked flask under nitrogen protection, 1 mL CH3COOH was added, and then a mixture of CH3CH2OH:H2O = 1:1 (v / v) 4 mL was added, and stirred at room temperature for 2 h. CH2Cl2:CH3OH = 20:1 (v / v) was used to track the reaction progress, and after the reaction was completed, the remaining CH3COOH was neutralized with a saturated solution of sodium bicarbonate, and purified with CH2Cl2 / CH3OH = 20:1 (v / v) to obtain a brown-yellow solid Ir 1-NO 79 mg (yield 79%).

[0070] The matrix-assisted laser desorption ionization time-of-flight mass spectrum of Ir 1-NO is shown in Figure 3

[0071] MALDI-TOF-MS m / z = 1744.61 [M-PF6 - ] + .

[0072] Example 2

[0073] (1) Compound 2 was synthesized according to the method disclosed in Chinese invention patent CN118930586A.

[0074] (2) Preparation of Ir 2, the reaction formula is as follows:

[0075]

[0076] Compound 2 (180 mg, 0.096 mmol) and Pd / C (18 mg) were added to a dry two-necked flask under nitrogen protection, 8 mL of ethanol was added, and then 0.5 mL of hydrazine hydrate at room temperature was added, and refluxed at 80°C for 12 h. After the reaction was completed, it was cooled to room temperature, and after the solvent was evaporated, it was purified by column chromatography (CH2Cl2:CH3OH = 60:1, v / v). Finally, orange solid Ir 2 295 mg (yield 60%) was obtained.

[0077] The matrix-assisted laser desorption ionization time-of-flight mass spectrum of Ir 2 is shown in Figure 4

[0078] MALDI-TOF-MS m / z = 1743.680 [M-PF6 - ] + .

[0079] The nuclear magnetic resonance hydrogen spectrum of Ir 2 is shown in Figure 5

[0080] 1 ​​​H NMR (500 MHz, Chloroform-d) d (ppm): 9.35 (d, J = 8.7 Hz, 2H), 9.10 (d, J = 8.2 Hz, 2H), 8.07 (s, 2H), 7.90-7.77 (m, 8H), 7.68 (d, J = 7.9 Hz, 2H), 7.53 (dt, J = 22.3, 11.2 Hz, 4H), 7.27 (d, J = 29.6 Hz, 12H), 7.17-6.94 (m, 22H), 6.76 (s, 2H), 6.56 (s, 2H), 6.22 (d, J = 8.4 Hz, 2H), 2.33 (dd, J = 31.1, 8.1 Hz, 6H), 1.45 (d, J = 56.3 Hz, 4H), 1.08-0.77 (m, 18H).

[0081] (3) Preparation of Ir 2-NO, the reaction scheme is as follows:

[0082]

[0083] Ir 2 (100 mg, 0.051 mmol) and NaNO2(17.5 mg, 0.254 mmol) were placed in a dry two-necked flask under nitrogen protection, 1 mL of CH3COOH was added, and then a mixture of CH3CH2OH:H2O = 1:1 (v / v) 4 mL was added, and stirred at room temperature for 2 h. CH2Cl2:CH3OH = 20:1 (v / v) was used to track the reaction progress, and after the reaction was completed, the remaining CH3COOH was neutralized with a saturated sodium bicarbonate solution, and purified with CH2Cl2:CH3OH = 20:1 (v / v) to obtain brown-yellow solid Ir 2-NO 72 mg (yield 71%).

[0084] The matrix-assisted laser desorption ionization time-of-flight mass spectrum of Ir 2-NO is shown in Figure 6 :

[0085] MALDI-TOF-MS m / z = 1856.233 [M-PF6 - ] + .

[0086] Example 3

[0087] (1) Compound 3 was synthesized according to the method disclosed in Chinese invention patent CN118930586A.

[0088] (2) Preparation of Ir 3, the reaction scheme is as follows:

[0089]

[0090] Compound 3 (150 mg, 0.076 mmol), Pd / C (15 mg) were added into a dry two-necked flask, protected by nitrogen, 8 mL of ethanol and 0.5 mL of hydrazine hydrate at room temperature were added, and refluxed at 80 °C for 12 h. After the reaction was completed, it was cooled to room temperature, and the solvent was evaporated. CH2Cl2:CH3OH = 60: 1 (v / v) column chromatography purification. After vacuum drying, the orange-red product Ir 3 was obtained 375 mg (yield 63%). 2: CH3OH = 60: 1 (v / v) column chromatography purification. After vacuum drying, the orange-red product Ir 3 was obtained 375 mg (yield 63%).

[0091] The matrix-assisted laser desorption ionization time-of-flight mass spectrum of Ir 3 is shown in Figure 7 :

[0092] The MALDI-TOF-MS m / z = 1845.395 [M-PF6 - ] + .

[0093] The nuclear magnetic resonance hydrogen spectrum of Ir 3 is shown in Figure 8 :

[0094] 1 H NMR (500 MHz, Chloroform-d) δ (ppm): 9.44 (d, J = 7.3 Hz, 2H), 9.06 (d, J = 7.0 Hz, 2H), 8.77 (d, J = 6.3 Hz, 2H), 8.47 (d, J = 8.4 Hz, 2H), 8.12 (d, J = 7.8 Hz, 2H), 7.99-7.89 (m, 6H), 7.60 (s, 2H), 7.52-7.43 (m, 4H), 7.27-7.01 (m, 32H), 6.80-6.36 (m, 10H), 2.42-2.10 (m, 6H), 1.41-1.17 (m, 16H), 0.88 (s, 6H).

[0095] (3) Preparation of Ir 3-NO, the reaction formula is as follows:

[0096]

[0097] Ir 3 (100 mg, 0.049 mmol) and NaNO2(16.9 mg, 0.245 mmol) were placed in a dry two-necked flask, protected by nitrogen, 1 mL of CH3COOH was added, and 4 mL of CH3CH2OH:H2O = 1:1 (v / v) mixture was added, and stirred at room temperature for 2 h. CH2Cl2:CH3OH = 20:1 (v / v) was used to track the reaction progress, and after the reaction was completed, the remaining CH3COOH was neutralized with a saturated sodium bicarbonate solution, and CH2Cl2:CH3OH = 20:1 (v / v) was used to run a large plate purification, and the brown-yellow solid Ir 3-NO was obtained 68 mg (yield 67%).

[0098] The matrix assisted laser desorption ionization time-of-flight mass spectrum of Ir 3-NO is shown in Figure 4. Figure 9

[0099] MALDI-TOF-MS m / z = 1956.304 [M-PF6 - ] + .

[0100] Example 4

[0101] The absorption spectra of iridium complex photosensitizers Ir 1, Ir 2, Ir 3 in different solvents and the emission spectra in water were tested:

[0102] 1980 μL of solvent was placed in a cuvette, 20 μL of iridium complex photosensitizer DMSO solution with a concentration of 1 x 10 -3 M was added to the above solvent, and another 2 mL of solvent was added to another cuvette as a reference, and the absorption and emission spectra were tested using a UV-Vis spectrophotometer and a fluorescence spectrophotometer, respectively.

[0103] Figure 10 The absorption spectra of iridium complex photosensitizers Ir 1, Ir 2, Ir 3 in different solvents and the emission spectra in water were tested:

[0104] Figure 10 It is shown that Ir 1-3 have moderate intensity absorption peaks and wide absorption bands in the region of 420-550 nm, indicating that the material can effectively utilize visible light. The emission peak of Ir 3 is at 725 nm, in the near infrared region. The large Stokes shift and the emission in the near infrared region indicate the potential application of Ir 3 as a photosensitizer in photodynamic therapy.

[0105] Example 5

[0106] The absorption spectra of iridium complex photosensitizers Ir 1, Ir 2, Ir 3, Ir 1-NO, Ir 2-NO, Ir 3-NO in water were tested:

[0107] The test method is the same as Example 4.

[0108] Figure 11 ​Absorption spectra of Ir 1, Ir 2, Ir 3, Ir 1-NO, Ir 2-NO, Ir 3-NO in water in Example 5, wherein (a) is the absorption spectra of Ir 1 and Ir 1-NO, (b) is the absorption spectra of Ir 2 and Ir 2-NO, (c) is the absorption spectra of Ir 3 and Ir 3-NO.

[0109] Figure 11 It is shown that the absorbance of the photosensitizer after activation at 600-750 nm is increased compared with that before activation of the NO / acid double factor, indicating that Ir 1-3 has a good response to NO under acidic conditions. Among them, Ir 3-NO has the strongest absorption, and the maximum is 700 nm.

[0110] Example 6

[0111] Investigation of the activation conditions of Ir 3:

[0112] Ir 3 (final concentration 1×10 -5 M) was added to a buffer solution containing 1×10 -4 M NO (from NaNO2), and the pH of the buffer solution was 5.4, 6.4, and 7.4, respectively. The absorption spectrum was measured after stirring for 12 h.

[0113] Ir 3 (final concentration 1×10 -5 M) was added to a buffer solution containing different concentrations of NO (concentrations were 0 μM, 2 μM, 4 μM, 6 μM, 8 μM, and 10 μM, respectively, from NaNO2) and pH 5.4, and the absorption spectrum was tested after stirring for 12 h.

[0114] Figure 12 Absorption spectra of Ir 3 under different pH conditions in the presence of NO in Example 6 (a) and absorption spectra of Ir 3 under different NO concentration conditions at pH=5.4 (b).

[0115] Figure 12 It is shown that only in the presence of NO and acid (pH less than 7) can Ir 3 be activated.

[0116] Example 7

[0117] Investigation of the selectivity of Ir 3 under acidic conditions:

[0118] Ir 3 (final concentration 10 μM) was added to a buffer solution containing NO (NO2 - ) (15 μM), H2O2 (500 μM), HClO (50 μM), · OH (100 μM), 1 O2 (100 μM), Cys (500 μM), NO3 -(50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h. - (50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h. + (50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h. 2+ (50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h. + (50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h. 3+ (50 μM), ONOOH (50 μM), GSH (1 mM), Na2S (500 μM), CaCl2(500 μM), KCl (500 μM), and FeCl3(500 μM) in pH = 5.6 buffer were tested for absorbance at 700 nm after stirring for 2 h.

[0119] Figure 13 Absorbance of Ir 3 after reacting with different ions in acidic environment in Example 7.

[0120] Figure 13 It shows that Ir 3 has high selectivity to NO under acidic conditions, indicating that the photosensitizer can be activated in the cancer tissue environment characterized by elevated levels of NO and lower pH.

[0121] Example 8

[0122] Preparation of Ir 3-NO into nanoparticles (Ir 3-NO NPs):

[0123] Dissolve 1 mg of Ir-3 and 20 mg of DSPE-mPEG2000 in 1 mL of THF, ultrasonic treatment for 1 min, pour the mixture into 10 mL of water, ultrasonic treatment for 5 min, and place in a fume hood to remove THF by volatilization under stirring at room temperature. Purify by filtration with a 0.22 μm ultrafiltration filter to obtain Ir 3-NO NPs.

[0124] Test the change of relative emission intensity of Ir 3-NO NPs at 525 nm in the active oxygen indicator DCFH aqueous solution over time after 730 nm laser irradiation under normoxic and hypoxic conditions:

[0125] Use the commercial fluorescence indicator 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) to detect the total active oxygen production of Ir 3-NO NPs in aqueous solution under 730 nm (1 W·cm -2 ) laser irradiation. DCFH-DA is activated into DCFH by the general method, and the concentration of DCFH after activation is 40 μM.

[0126] Add 20 μL of Ir 3-NO NPs (1 × 10 -3 M) solution to 1980 μL of DCFH solution, deoxygenate the solution, and use 730 nm (1 W·cm -2) laser irradiation, the emission value was tested every 30 s, for a total of 5 min, and the control solution was not added with Ir3-NO NPs. The emission intensity of DCFH at 525 nm was measured using a fluorescence spectrophotometer.

[0127] Another group of solutions was not subjected to oxygen removal treatment, and the above steps were repeated.

[0128] Figure 14 The relative emission intensity of Ir3-NO NPs at 525 nm in the active oxygen indicator DCFH aqueous solution under normal oxygen conditions (a) and anaerobic conditions (b) after 730 nm laser irradiation in Example 8 was measured.

[0129] Figure 14 It was shown that the fluorescence intensity of Ir3-NO NPs was rapidly enhanced and more than 12 times the initial value after 5 min of laser irradiation, indicating that Ir3-NO NPs could rapidly generate ROS after laser irradiation. The fluorescence intensity also increased significantly under anaerobic conditions. Ir3-NO NPs can effectively generate ROS under both normal oxygen and anaerobic conditions.

[0130] Example 9

[0131] The relative emission intensity of Ir3-NO NPs (preparation method same as Example 8) at 525 nm in the O2 ·- indicator DHR123 aqueous solution under normal oxygen and anaerobic conditions after 730 nm laser irradiation was tested:

[0132] Dihydrorhodamine 123 (DHR123) was used as a fluorescent probe to monitor the generation of superoxide anion (O2 ·- ) in an aqueous solution. The probe itself has no fluorescence characteristics in the reduced state, and when it undergoes a specific oxidation reaction with O2 ·- , it is converted into oxidized rhodamine 123 with strong green fluorescence emission ability. This change in fluorescence intensity is positively correlated with the amount of O2 ·- generated, and can be quantitatively detected by fluorescence spectroscopy.

[0133] Ir3-NO NPs and DHR123 were added to a PBS solution, the concentration of Ir3-NO NPs was 10 μM, and the concentration of DHR123 was 10 μM. The solution was subjected to oxygen removal treatment, and the emission spectrum of the solution was measured using a fluorescence spectrometer. 730 nm (1 W·cm -2 ) laser irradiation, the emission value was tested every 30 s, for a total of 5 min, and the control solution was not added with Ir3-NO NPs. The fluorescence intensity at 525 nm was recorded for studying the level of O2· - generated.

[0134] Another group of solutions was not subjected to oxygen removal treatment, and the above steps were repeated.

[0135] Figure 15 The relative emission intensity at 525 nm in the DHR123 aqueous solution was measured over time. ·- The relative emission intensity at 525 nm in the DHR123 aqueous solution was measured over time.

[0136] Figure 15 It was shown that the fluorescence intensity of DHR123 mixed with Ir 3-NO NPs was significantly enhanced under light irradiation, which indicated that Ir 3-NO NPs could generate O2 ·- under laser irradiation in both anoxic and normoxic conditions. Therefore, Ir 3-NO NPs have the potential to be an effective photosensitizer for PDT tumor treatment under hypoxic conditions.

[0137] Example 10

[0138] The temperature change of the solution of Ir 3-NO NPs (prepared according to the method of Example 8) under 730 nm laser irradiation was tested:

[0139] The Ir 3-NO NPs were prepared into solutions with concentrations of 0 μM, 10 μM, 25 μM, 50 μM, and 100 μM, respectively, and irradiated with a 730 nm (1 W·cm -2 ) laser. The temperature change of the solution was recorded every 30 s with an infrared sensitive thermal imager. Subsequently, the photothermal cycling stability of the nanoparticles was tested. The 100 μM Ir 3-NO NPs aqueous solution was irradiated with the same power laser, and when the solution temperature reached the highest temperature, the laser was turned off. When the solution naturally cooled to room temperature, it was irradiated again, and the cycle was repeated five times.

[0140] Figure 16 The temperature change curve of the aqueous solution of Ir 3-NO NPs with different concentrations under 730 nm laser irradiation over time (a) and the temperature change curve of the solution with a concentration of 100 μM during five heating-cooling cycles (b) in Example 10

[0141] Figure 16 It was shown that under 730 nm laser irradiation, pure water did not have a significant temperature change, while the solution of Ir 3-NO NPs showed a significant temperature rise, indicating that the nanoparticles had obvious photothermal properties. The reversible temperature rise after five heating-cooling cycles indicated that the nanoparticles had excellent photothermal stability.

[0142] Example 11

[0143] The toxicity of Ir 3-NO NPs (preparation method same as Example 8) on 4T1 cells under normoxic and hypoxic conditions, in the dark and under 730 nm laser irradiation was tested:

[0144] The cytotoxicity of different formulations on 4T1 cells was determined by cell counting kit (CCK8) method. Cells were seeded in 96-well plates (1 x 10 4 per well) and incubated for 24 h, 200 μL of medium containing different concentrations of Ir 3-NO NPs was added, incubated for 4 h, and part of it was treated with 730 nm laser (1 W-cm -2 ) for 5 min, and after treatment, incubated for 20 h, the medium was removed, and the cells were gently washed with fresh PBS. Then 100 μL of fresh DMEM and 10 μL of CCK8 solution were added to each well, and the cells were incubated for 30 min. The absorbance of CCK8 at 450 nm was determined by SpectraMax M5 microplate reader, and the cell viability was determined by the ratio of the absorbance of 4T1 cells incubated with Ir 3-NO NPs to the absorbance of PBS solution (control). The cell viability of 4T1 cells before and after 730 nm laser irradiation was observed, respectively. The formula for calculating the relative viability (CV) of cells is as follows:

[0145] Cell viability (%) = (OD 样品 - OD 背景 ) / (OD 空白 - OD 背景 ) x 100%;

[0146] Figure 17 The results of the toxicity test of iridium complex photosensitizer Ir 3-NO NPs on 4T1 cancer cells under normoxic (a) and hypoxic (b) conditions, in the dark and under 730 nm laser irradiation in Example 11.

[0147] Figure 17 It is shown that under dark conditions, the cytotoxicity of Ir 3-NO NPs on 4T1 cancer cells is also negligible at a high concentration of 100 μM, which means that it has low dark toxicity, reflecting its excellent biocompatibility. However, when irradiated with 730 nm laser (1 W-cm -2 ) for 5 min, Ir 3-NO NPs showed severe photo-cytotoxicity, which indicates that Ir 3-NO NPs have the potential for cancer phototherapy.

[0148] Figure 18 The activation mechanism of photosensitizer Ir 1, 2, 3.

[0149] The above described embodiments are only to illustrate the preferred modes of the present application, and are not intended to limit the scope of the present application. Any modification and improvement made by those skilled in the art to the technical solutions of the present application without departing from the design spirit of the present application shall fall within the protection scope of the present application.

Claims

1. A NO / acid dual-factor activated near-infrared light-excited iridium (III) complex photosensitizer, characterized in that: The structural formula is as follows: In the above formula, the C^N ligand is The X group is R in the X group is any one of R1 to R5:

2. A NO / acid dual-factor activated iridium (III) complex photosensitizer, characterized in that: The structural formula is as follows: In the above formula, the C^N ligand is The X group is R in the X group is any one of R1 to R5:

3. A method for preparing the NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer according to claim 1, characterized in that: The following steps are involved: Compound A, hydrazine hydrate, a solvent and a catalyst are mixed and reacted under an inert atmosphere to obtain the NO / acid dual-factor activated near-infrared light-excited iridium (III) complex photosensitizer; The structural formula of the compound A is as follows:

4. The method for preparing the NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer according to claim 3, characterized in that: The catalyst is a palladium-carbon catalyst.

5. The method for preparing the NO / acid dual-factor activated near-infrared light excited iridium (III) complex photosensitizer according to claim 3, characterized in that: The reaction temperature is 70-90° C. and the reaction time is 8-14 hours.

6. A method for preparing the NO / acid dual-factor activated iridium (III) complex photosensitizer according to claim 2, characterized in that: The following steps are involved: The iridium (III) complex photosensitizer according to claim 1, an NO donor and an acid solution are mixed and reacted under an inert atmosphere to obtain the NO / acid dual-factor activated iridium (III) complex photosensitizer.

7. The method for preparing the NO / acid dual-factor activated iridium (III) complex photosensitizer according to claim 6, characterized in that: The pH value of the acid solution ranges from 5.4 to 6.

4.

8. The method for preparing the NO / acid dual-factor activated iridium (III) complex photosensitizer according to claim 6, characterized in that: The reaction time is 2 to 4 hours.

9. Use of the NO / acid dual-factor activated near-infrared light-excited iridium (III) complex photosensitizer according to claim 1 in the preparation of a photosensitizer for photodynamic therapy.

10. Use of the NO / acid dual-factor activated iridium (III) complex photosensitizer according to claim 2 in the preparation of a photosensitizer for photodynamic therapy.

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