A dual-cell-targeting luminescent near-infrared fluorescent probe and its preparation method and application

Abstract

The present invention provides a dual-cell-targeted luminescent near-infrared fluorescent probe (E)-4-(4-(bis(4-(2-morpholinoethoxy)phenyl)amino)styryl)-1-(2-hydroxyethyl)quinolin-1-ammonium hexafluorophosphate (V), namely the fluorescent probe MoTQu. This dual-cell-targeted fluorescent probe contains a quinolinium cation moiety and targets mitochondria through electrostatic interaction with the electronegative mitochondrial membrane; the introduction of a weakly basic morpholine ring in the triphenylamine tail chain of this fluorescent probe targets lysosomes. At the same time, the preparation method and application of this probe are disclosed. The fluorescent probe of the present invention can not only light up different organelles in response to time, but also effectively improve the killing efficiency of cancer cells through the synergy of mitochondrial- and lysosome-targeted photodynamic therapy, and thus has a powerful effect on the ablation of photodynamic cancer cells.

Description

Technical Field

[0001] The present invention belongs to the field of organic fluorescent probe molecules, and particularly relates to a dual-organelle-targeted and dual-cell-targeted luminescent near-infrared fluorescent probe, a preparation method thereof, and an application thereof. Background Art

[0002] Most traditional photosensitizers such as phthalocyanine, porphyrin, etc., the defects of their rigid structures and hydrophobicity often lead to π-π stacking of molecules, resulting in the phenomenon of aggregation-caused quenching (ACQ). This not only leads to weak emission in bioimaging, but also greatly reduces the therapeutic effect of photodynamic therapy (PDT) in cancer treatment. Encouragingly, the concept of aggregation-induced emission (AIE) provides a possibility to solve this problem. Materials with AIE phenomenon generally have a propeller-shaped structure. They mainly exist in the form of single molecules in good solvents, while the intramolecular motion of AIE molecules is restricted in the aggregated state, showing enhanced emission. In addition, AIE photosensitizers can increase their ability to generate reactive oxygen species (ROS) by enhancing intersystem crossing (ISC) or narrowing the energy gap between the singlet (S1) and triplet excited states (T1), and also show great potential in the PDT process.

[0003] As one of the most important organelles, mitochondria are closely related to signal transduction, calcium homeostasis, cell differentiation, apoptosis, etc. in cells. In addition, mitochondria are the sites where ROS are generated, and changing the intracellular redox state can effectively promote programmed cell death of cells. Lysosomes are sac-like organelles containing various acidic hydrolases and are the main sites for hydrolyzing biological macromolecules such as nucleic acids and proteins, and are called the "stomach" of cells. At the same time, lysosome damage can induce the permeability of the lysosomal membrane, allowing the release of cathepsin B / D to trigger programmed cell death. In addition, studies have found that the content of cathepsin in tumor cells is significantly higher than that in normal cells. Therefore, tumor cells have lower resistance to lysosome-dependent cell death. Considering the high spatio-temporal resolution of light-controlled imaging and the good therapeutic effect of dual-organelle-targeted therapy, a dual-organelle-targeted photoactivatable fluorescent probe with the ability to in-situ induce lysosome damage and destroy mitochondrial structure is a direction worthy of research and development. Summary of the Invention

[0004] Object of the Invention: Aiming at the deficiencies and defects of the prior art, the present invention provides a dual-cell-targeted luminescent near-infrared fluorescent probe, a preparation method thereof, and an application thereof, which can not only light up different organelles in response to time, but also effectively improve the killing efficiency of cancer cells through the synergy of mitochondrial and lysosome-targeted photodynamic therapy, and thus has a powerful effect on the ablation of photodynamic cancer cells.

[0005] Technical Solution: A dual-cell-targeted luminescent near-infrared fluorescent probe of the present invention, the chemical structural formula of the dual-cell-targeted probe is:

[0006]

[0007] Among them, the dual-cell-targeted fluorescent probe contains a quinoline cation moiety and targets mitochondria through electrostatic interaction with the electronegative mitochondrial membrane; the introduction of a weakly basic morpholine ring in the triphenylamine tail chain of the fluorescent probe targets lysosomes.

[0008] Preparation method of the dual-cell-targeted luminescent near-infrared fluorescent probe of the present invention: Place 4-(bis(4-(2-morpholinoethoxy)phenyl)amino)benzaldehyde (MPAA), piperidine, and 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate in a reaction flask, add ethanol to the reaction flask, heat the mixed materials to no less than 85 °C under nitrogen protection, cool the reaction solution to room temperature after the reaction ends, extract and combine the organic phases, perform drying and concentration under reduced pressure to obtain a crude product, and use dichloromethane / methanol as the eluent to purify the crude product by column chromatography to obtain the fluorescent probe (E)-4-(4-(bis(4-(2-morpholinoethoxy)phenyl)amino)styryl)-1-(2-hydroxyethyl)quinolin-1-ium hexafluorophosphate (V), that is, the fluorescent probe MoTQu.

[0009] Among them, the mixed molar ratio of the described 4-(bis(4-(2-morpholinoethoxy)phenyl)amino)benzaldehyde (MPAA) and 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate is 1:0.5 to 1:5.

[0010] Among them, the reaction time is 8 h to 16 h.

[0011] The fluorescent probe of the present invention contains a quinoline cation moiety and targets mitochondria through electrostatic interaction with the electronegative mitochondrial membrane; in addition, the introduction of a weakly basic morpholine ring in the triphenylamine tail chain of the fluorescent probe increases its ability to target lysosomes. Once it enters the lysosome, the morpholine ring will be protonated by the high acidity in the lysosome, which makes MoTQu have high hydrophilicity and lysosome retention rate. At this time, the fluorescence of MoTQu can only be observed in the lysosome region. The fluorescent probe of the present invention can not only light up different organelles in response to time, but also effectively improve the killing efficiency of cancer cells through the synergy of mitochondrial targeting and lysosome-targeted photodynamic therapy, so it has a powerful effect on the ablation of photodynamic cancer cells.

[0012] Application of the dual-cell-targeted luminescent near-infrared fluorescent probe of the present invention in detection.

[0013] Among them, for in vitro cells, the fluorescent probe was dissolved in DMEM culture medium at a concentration of 10 μM; incubating the live HepG2 cells with the culture medium containing the fluorescent probe for different times would light up different organelles as the evaluation index for the dual-organelle targeting of MoTQu. The detection process of the fluorescent probe for the endoplasmic reticulum localization in cells was as follows: the fluorescent probe was dissolved in DMEM culture medium at a concentration of 10 μM; the live HepG2 cells were co-incubated with the culture medium containing the fluorescent probe and a commercial mitochondrial localization probe (Mito-Tracker Green) or a commercial lysosome localization probe (Lyso-Tracker Green) for 20 or 60 min, and the co-localization effect was observed under confocal microscopy as the evaluation index for the localization of the fluorescent probe to mitochondria or lysosomes.

[0014] Among them, the detection process of the photostability of the fluorescent probe in live HepG2 cells was as follows: the HepG2 cells were incubated with the culture medium containing the fluorescent probe at room temperature, the concentration of the fluorescent probe in the culture medium was 10 μM, and the degree of fluorescence attenuation of the fluorescent probe in the HepG2 cells under confocal laser scanning was measured 30 min later as the evaluation index for the photostability of MoTQu staining in HepG2 cells.

[0015] Among them, for in vitro cells, the fluorescent probe was dissolved in DMEM culture medium at a concentration of 10 μM; the live HepG2 cells were incubated with the culture medium containing the fluorescent probe, and the amount of ROS generated by the fluorescent probe under light excitation in the cells was measured 30 min after light exposure as the evaluation index for the phototoxicity of MoTQu. The detection process of the fluorescent probe for ROS generation in cells was as follows: a culture medium containing the fluorescent probe (10 μM) and DCFH-DA solution (10 μM) was prepared, the live HepG2 cells were incubated with the culture medium containing the fluorescent probe for 30 min, and the degree of increase in green fluorescence under confocal laser scanning was used as the evaluation index for the generation of ROS by the fluorescent probe in cells.

[0016] Among them, for in vitro cells, the fluorescent probe was dissolved in DMEM culture medium at a concentration of 10 μM; the live HepG2 cells were incubated with the culture medium containing the fluorescent probe, and the degree of loss of mitochondrial membrane potential in the cells was measured 30 min after light exposure as the evaluation index for MoTQu targeting and damaging mitochondria.

[0017] Beneficial effects: Compared with the prior art, the present invention has the following remarkable advantages: By modifying the molecular structure, the fluorescent probe of the present invention enables the molecular structure to specifically target mitochondria and lysosomes in living cells over the incubation time. The fluorescence wavelength emitted by the fluorescent group in the fluorescent probe of the present invention is in the near-infrared region, so it can avoid interference from the autofluorescence of organisms during fluorescence detection. The fluorescent probe of the present invention has good biocompatibility. When cells are incubated with a culture medium containing MoTQu (80 μM) for 24 hours, the cell viability is still greater than 80%. The fluorescent probe of the present invention has excellent phototoxicity. When cells are incubated with a culture medium containing MoTQu (5 μM), irradiated with light for 30 minutes, and then cultured in an incubator for 24 hours, the cell viability is lower than 40%. The fluorescent probe of the present invention can generate a large amount of ROS under light excitation, promote functional damage of lysosomes, release a large number of apoptotic bodies, and induce apoptosis of cancer cells. After causing lysosomal dysfunction, the fluorescent probe of the present invention further targets and disrupts the mitochondrial membrane potential through cationic groups, and the combined action of the two accelerates the irreversible damage of cancer cells. Description of the Drawings

[0018] Figure 1 is the synthetic route diagram of the fluorescent probe of the present invention;

[0019] Figure 2 is the ultraviolet absorption diagram of the fluorescent probe of the present invention in dimethyl sulfoxide;

[0020] Figure 3 is the fluorescence emission spectrum diagram of the fluorescent probe of the present invention in the solid state;

[0021] Figure 4 is the emission spectrum diagram of the maximum fluorescence intensity of the fluorescent probe of the present invention in a mixed solvent of dimethyl sulfoxide / toluene with different ratios;

[0022] Figure 5 is the ratio spectrum diagram of the maximum fluorescence intensity of the fluorescent probe of the present invention in a mixed solvent of dimethyl sulfoxide / toluene with different ratios;

[0023] Figure 6 is the comparison diagram of the ultraviolet absorption and fluorescence emission spectra of the fluorescent probe of the present invention in dimethyl sulfoxide solvent;

[0024] Figure 7 is the confocal fluorescence imaging diagram after the fluorescent probe of the present invention and the commercial mitochondrial localization probe (Mito-Tracker Green) are co-incubated in HepG2 cells for 20 minutes; Figure A is the co-localization diagram between MoTQu and Mito-Traker Green; Figure B is the overlap coefficient and Pearson co-localization coefficient diagram between MoTQu and Mito-Traker Green; Figure C is the intensity scatter plot;

[0025] Figure 8 Confocal fluorescence imaging map after co-incubating the fluorescent probe of the present invention and the commercial lysosome localization probe (Lyso-Tracker Green) in HepG2 cells for 60 minutes; Figure A is the co-localization map between MoTQu and Lyso-Traker Green; Figure B is the overlap coefficient and Pearson co-localization coefficient map between MoTQu and Lyso-Traker Green; Figure C is the intensity scatter plot;

[0026] Figure 9 Time-dependent fluorescence imaging map of the fluorescent probe of the present invention in HepG2 cells;

[0027] Figure 10 Fluorescent imaging map of the fluorescent probe of the present invention generating ROS in HepG2 cells;

[0028] Figure 11 JC-1 flow cytometry map of the fluorescent probe of the present invention causing mitochondrial membrane potential damage in HepG2 cells;

[0029] Figure 12 Cell dark toxicity map of the fluorescent probe of the present invention on HepG2 cells;

[0030] Figure 13 Cell phototoxicity map of the fluorescent probe of the present invention on HepG2 cells;

[0031] Figure 14 Confocal fluorescence imaging map of live and dead cells stained with CalceinAM / PI medium after the fluorescent probe of the present invention was incubated in HepG2 cells for 60 minutes and irradiated for 30 minutes. Detailed implementation manners

[0032] The technical solution of the present invention will be further described below in conjunction with the accompanying drawings and specific implementation manners.

[0033] Figure 1This is the synthetic route diagram of the fluorescent probe MoTQu of the present invention, from which the fluorescent probe (E)-4-(4-(bis(4-(2-morpholinoethoxy)phenyl)amino)styryl)-1-(2-hydroxyethyl)quinolin-1-ammonium hexafluorophosphate(V) of the present invention, namely the fluorescent probe MoTQu, is obtained. This fluorescent probe MoTQu uses triphenylamine quinoline as the backbone to construct a near-infrared fluorescent probe with a dual-cell targeting luminescence effect. MoTQu contains a quinoline cation moiety and targets mitochondria through electrostatic interaction with the electronegative mitochondrial membrane; the introduction of a weakly basic morpholine ring in the triphenylamine tail chain increases the ability to target lysosomes. This synthetic route is simple and has a high yield; it has a large Stokes shift, good anti-photobleaching property, and its emission is in the near-infrared region, which can effectively resist background interference; the fluorescent probe of the present invention can not only light up different organelles in response to time, but also effectively improve the killing efficiency of cancer cells through the synergy of mitochondrial and lysosome-targeted photodynamic therapy, so it has a powerful effect on the ablation of photodynamic cancer cells.

[0034] Specific steps: Place 1 mmol of MPAA (MPAA of the present invention and 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate are prepared by the synthesis method disclosed in the following literature: (ACS Sens. 2020, 5, 225-233)), 200 μL of piperidine and 2 mmol of 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate in a 25 mL two-necked flask, then add 20 mL of anhydrous ethanol to the two-necked flask, heat the reaction materials to 85 °C under nitrogen protection, and stir the reaction for 8 hours; after the reaction is completed, cool the reaction solution to room temperature, extract and combine the organic phases, perform drying and concentration under reduced pressure to obtain a crude product, and use dichloromethane / methanol as the eluent to purify the crude product by column chromatography. (The developing agent is: dichloromethane:methanol = 5:1, v:v). A dark purple solid is obtained (yield 69.8%). 1HNMR(600MHz, DMSO) δ 9.08 (d, J = 6.7 Hz, 1H), 8.99 (d, J = 8.5 Hz, 1H), 8.50 (d, J = 9.0 Hz, 1H), 8.40 (d, J = 6.7 Hz, 1H), 8.22–8.17 (m, 1H), 8.11 (dd, J = 37.2, 15.7 Hz, 2H), 8.01–7.97 (m, 1H), 7.81 (d, J = 8.8 Hz, 2H), 7.14 (d, J = 8.8 Hz, 4H), 7.00 (d, J = 8.9 Hz, 4H), 6.77 (d, J = 8.8 Hz, 2H), 5.14 (t, J = 5.6 Hz, 1H), 5.01–4.97 (m, 2H), 4.10 (s, 4H), 3.91 (dd, J = 9.8, 5.1 Hz, 2H), 3.59 (s, 8H), 2.71 (s, 4H). 13 C NMR(151MHz, DMSO) δ 156.11 (s), 153.28 (s), 151.14 (s), 147.63 (s), 143.74 (s), 138.84 (s), 138.26 (s), 134.77 (s), 130.87 (s), 128.82 (s), 127.92 (s), 126.58 (s), 119.26 (s), 117.33 (s), 115.90 (s), 115.61 (s), 114.68 (s), 66.22 (s), 65.59 (s), 59.08 (s), 57.11 (s), 53.70 (s), 29.09 (s), 22.36 (s). HR-MS: m / z calcd for C 43 H 49 N4O5 701.3697; found, 701.37062 [M] + 。

[0035] Prepare a dimethyl sulfoxide solution of the fluorescent probe MoTQu at 10 μM and measure it on an ultraviolet spectrophotometer. The results are as Figure 2 。Prepare the fluorescence emission spectrum of the fluorescent probe MoTQu in the solid state at 10 μM, as Figure 3 。From Figures 2 to 3 It can be seen that the maximum absorption peak of the fluorescent probe molecule MoTQu prepared in the present invention is around 540 nm, and the maximum emission peaks in the solid state all fall in the near-infrared region I around 800 nm, showing significant near-infrared fluorescence emission performance.

[0036] Prepare a mixed solution of the fluorescent probe MoTQu at 10 μM in different ratios of dimethyl sulfoxide / toluene and measure it on a fluorescence visible spectrophotometer. The results are as shown in the appendix Figure 4 。From Figure 4It can be seen that as the proportion of the poor solvent increases, the fluorescence emission intensity in the mixed solution of MoTQu increases significantly.

[0037] The graph of the maximum fluorescence intensity ratio (I / I0) of the 10 μM fluorescent probe MoTQu in the mixed solution of different ratios of dimethyl sulfoxide / toluene is prepared, and the results are as follows Figure 5 . From Figure 5 It can be seen that the fluorescent probe MoTQu has obvious AIE performance, that is, the fluorescence intensity gradually increases with the increase of the degree of molecular aggregation.

[0038] The comparison graph of the ultraviolet absorption and fluorescence emission of the 10 μM fluorescent probe MoTQu in dimethyl sulfoxide solution is prepared. By Figure 6 It can be known that the fluorescent probe MoTQu of the present invention has a large Stokes shift (240 nm), which is more conducive to anti-interference of fluorescence molecular imaging.

[0039] The confocal fluorescence imaging graph of the 10 μM fluorescent probe MoTQu and the commercial mitochondrial probe (Mito-Tracker Green) after co-incubation in HepG2 cells for 20 minutes is prepared. From Figure 7 It can be seen that the Pearson coefficient of co-localization of the fluorescent probe MoTQu and the commercial mitochondrial fluorescent probe is 0.867, indicating that the fluorescent probe MoTQu has a very high mitochondrial localization ability when co-incubated with cells for 20 minutes.

[0040] The confocal fluorescence imaging graph of the 10 μM fluorescent probe MoTQu and the commercial endoplasmic reticulum probe (Lyso-Tracker Green) after co-incubation in HepG2 cells for 60 minutes is prepared. From Figure 8 It can be seen that the Pearson coefficient of co-localization of the fluorescent probe MoTQu and the commercial endoplasmic reticulum fluorescent probe is 0.911, indicating that the fluorescent probe MoTQu has a very high mitochondrial localization ability when co-incubated with cells for 60 minutes. Once the fluorescent probe enters the lysosome, the morpholine ring will be protonated by the high acidity in the lysosome, which makes MoTQu have high hydrophilicity and lysosome retention rate. At this time, the fluorescence of MoTQu can only be observed in the lysosome region.

[0041] Time-dependent fluorescence imaging experiment of the fluorescent probe MoTQu (10 μM) in HepG2 cells. The HepG2 cells were seeded into cell culture dishes for confocal imaging, 1 mL of the corresponding medium (containing 10% fetal bovine serum) was added, and the cells were incubated in a constant temperature incubator (5% CO2, 37 °C) for 24 hours; then they were washed 3 times with PBS (1 mL), and then incubated with the medium containing MoTQu (10 μM) and the cells; finally, the changes in the intracellular fluorescence emission intensity at different time points were observed under a confocal microscope. The excitation wavelength was 540 nm, and the fluorescence collection range was 650 - 750 nm. Through Figure 9 it can be seen that the fluorescent probe MoTQu of the present invention has excellent photostability in HepG2 cells.

[0042] Time-dependent ROS generation experiment of the fluorescent probe MoTQu (10 μM) in HepG2 cells. The HepG2 cells were seeded into cell culture dishes for confocal imaging, 1 mL of the corresponding medium (containing 10% fetal bovine serum) was added, and the cells were incubated in a constant temperature incubator (5% CO2, 37 °C) for 24 hours; then they were washed 3 times with PBS (1 mL), and then incubated with the medium containing MoTQu (10 μM) and DCFH-DA (10 μM) (which can be oxidized by ROS to 2,7-dichlorodihydrofluorescein to emit bright green light, used to evaluate the generation of total reactive oxygen species ROS) and the cells; finally, the changes in the intracellular fluorescence emission intensity at different time points were observed under a confocal microscope. The excitation wavelength was 488 nm, and the fluorescence collection range was 500 - 600 nm. Through Figure 10 it can be seen that the fluorescent probe MoTQu of the present invention has the ability to generate a large amount of ROS in cells under light excitation.

[0043] JC-1 flow cytometry experiment on the damage of mitochondrial membrane potential caused by the fluorescent probe MoTQu (10 μM) in HepG2 cells. The HepG2 cells were seeded into cell culture dishes for confocal imaging, 1 mL of the corresponding medium (containing 10% fetal bovine serum) was added, and the cells were incubated in a constant temperature incubator (5% CO2, 37 °C) for 24 hours; then they were washed 3 times with PBS (1 mL), and then incubated with the medium containing MoTQu (10 μM) and the HepG2 cells for 60 minutes and then irradiated for 30 minutes, and then incubated with JC-1 (2 μM) (JC-1 emits red light when aggregated and green light when in monomer form) and the cells; finally, the damage condition of intracellular mitochondria was observed under a flow cytometer. Through Figure 11 it can be seen that after the fluorescent probe MoTQu of the present invention causes lysosomal dysfunction, it further targets and destroys the mitochondrial membrane potential through cationic groups.

[0044] Detection of the dark toxicity of the fluorescent probe MoTQu to cells. The cell viability was determined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method. First, HepG2 cells (human liver cancer cells) were seeded in 96-well culture plates, and medium (100 μL) containing 10% fetal bovine serum (FBS) was added, and then incubated at 37 °C in 5% CO2 for 24 hours. The medium of HepG2 cells was replaced with medium (100 μL) containing different concentrations of MoTQu (0, 5, 10, 20, 30, 40, 80 μM) and incubated for another 24 hours. The medium was aspirated and the cells were washed with PBS (100 μL). Then 100 μL of medium containing 0.05% MTT was added to each well and incubated for 4 hours. After the incubation, the medium was removed and 150 μL of dimethyl sulfoxide was added to each well, and the formazan formed in the wells was dissolved by gently shaking at low speed for 10 minutes. Finally, the absorbance of each well at 490 nm was measured with an enzyme-linked immunosorbent assay (ELISA) reader and compared with the absorbance of the control group (100% survival rate) to obtain the relative survival rate of cells at different concentrations of the fluorescent probe. By Figure 12 It can be seen that the fluorescent probe of the present invention has good biocompatibility.

[0045] Detection of the phototoxicity of the fluorescent probe MoTQu to cells. The cell viability was determined by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method. First, HepG2 cells (human liver cancer cells) were seeded in 96-well culture plates, and medium (100 μL) containing 10% fetal bovine serum (FBS) was added, and then incubated at 37 °C in 5% CO2 for 24 hours. The medium of HepG2 cells was replaced with medium (100 μL) containing different concentrations of MoTQu (0, 5, 10, 20, 30, 40 μM) and irradiated with white light at 10 mW·cm -2 for 30 minutes, and then incubated for another 24 hours. The medium was aspirated and the cells were washed with PBS (100 μL). Then 100 μL of medium containing 0.05% MTT was added to each well and incubated for 4 hours. After the incubation, the medium was removed and 150 μL of dimethyl sulfoxide was added to each well, and the formazan formed in the wells was dissolved by gently shaking at low speed for 10 minutes. Finally, the absorbance of each well at 490 nm was measured with an enzyme-linked immunosorbent assay (ELISA) reader and compared with the absorbance of the control group (100% survival rate) to obtain the relative survival rate of cells at different concentrations of the fluorescent probe. By Figure 13 It can be seen that the fluorescent probe of the present invention has excellent ability to ablate cancer cells by photodynamic therapy.

[0046] Live / Dead Cell Experiment of Fluorescent Probe MoTQu in HepG2 Cells. HepG2 cells were seeded in a 35 mm glass-bottom culture dish at 37 °C, then incubated with MoTQu (10 μM) for 60 min and irradiated for 30 min. The culture plate was further incubated in the dark for 24 h. Subsequently, the cells were stained with 1 μM Calcein AM and 1 μg / mL PI medium for 10 min, and the cells were photographed by CLSM. The excitation of Calcein AM was 488 nm and the emission was 500 - 550 nm; the excitation wavelength of propidium iodide (PI) was 560 nm and the emission wavelength was 550 - 650 nm. Through Figure 14 It can be seen that the fluorescent probe MoTQu of the present invention can effectively improve the killing efficiency of cancer cells through the synergy of mitochondrial-targeted and lysosome-targeted photodynamic therapy, and thus has a powerful effect on the ablation of photodynamic cancer cells.

Claims

1. A dual-cell-targeted luminescent near-infrared fluorescent probe, characterized in that: The chemical structural formula of the probe is as follows:

2. The preparation method of the dual-cell-targeting luminescent near-infrared fluorescent probe according to claim 1, wherein: 4-(Bis(4-(2-morpholinoethoxy)phenyl)amino)benzaldehyde, piperidine, and 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate are placed in a reaction flask. Ethanol is added to the reaction flask, and the mixed materials are heated to no less than 85 °C under nitrogen protection. After the reaction is completed, the reaction solution is cooled to room temperature, extracted, and the organic phases are combined, dried, and concentrated under reduced pressure to obtain a crude product. Using dichloromethane / methanol as the eluent, the crude product is purified by column chromatography to obtain the fluorescent probe (E)-4-(4-(bis(4-(2-morpholinoethoxy)phenyl)amino)styryl)-1-(2-hydroxyethyl)quinolin-1-ium hexafluorophosphate (V), namely the fluorescent probe MoTQu.

3. The preparation method of the dual-cell-targeting luminescent near-infrared fluorescent probe according to claim 2, characterized in that: The mixed molar ratio of the described 4-(bis(4-(2-morpholinoethoxy)phenyl)amino)benzaldehyde and 1-(2-hydroxyethyl)-4-methylquinolin-1-ium hexafluorophosphate is 1:0.5 to 1:

5.

4. The preparation method of the dual-cell-targeted luminescent near-infrared fluorescent probe according to claim 2, wherein: The described reaction time is 8 h to 16 h.

5. Application of the dual-cell-targeting luminescent near-infrared fluorescent probe according to claim 1 in the detection of non-disease treatment and diagnosis uses. For in vitro cells, the fluorescent probe is dissolved in DMEM culture medium, and the concentration of the fluorescent probe is 10 μM; incubating the live cells of HepG2 with the culture medium containing the fluorescent probe for different times will light up different cell organelles as an evaluation index for the dual-organelle targeting of MoTQu.

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