MOFCu@WZ35 nanoparticles, a preparation method and application thereof

Abstract

The application discloses a kind of MOF Cu @WZ35 nanoparticles and its preparation method and application, WZ35 is encapsulated by Cu-based metal organic framework, and the MOF Cu @WZ35 can more effectively inhibit the proliferation of PASMCs, promote the death of PASMCs, so as to achieve the purpose of relieving HPH, and the development of HPH therapeutic agent can provide a promising treatment option for HPH patients.

Description

Technical Field

[0001] This invention belongs to the field of drug preparation, specifically relating to a MOF Cu @WZ35 nanoparticles, their preparation methods, and applications. Background Technology

[0002] Pulmonary hypertension (PH) is a rare but serious condition characterized by elevated pressure in the blood vessels of the lungs, which can lead to heart failure and death. PH can be caused by a variety of underlying conditions, such as lung disease, heart disease, or genetic factors. Hypoxic pulmonary hypertension (HPH) is a subtype of PH characterized by increased resistance in the pulmonary arterioles. HPH is caused by prolonged exposure to low oxygen levels, which leads to pulmonary artery stenosis and increased pulmonary vascular resistance. This chronic hypoxia triggers a series of molecular and cellular events that result in pulmonary vascular remodeling and right ventricular hypertrophy.

[0003] Pulmonary artery smooth muscle cells (PASMCs) play a crucial role in the pathogenesis of HPH. In response to chronic hypoxia, these cells undergo changes in gene expression, becoming more proliferative and contractile, leading to thickening of the arterial wall and narrowing of the vascular lumen. Excessive proliferation and migration of PASMCs also contribute to the development of pulmonary vascular resistance. Currently, there is no cure for HPH; existing treatments focus on improving symptoms and slowing disease progression. However, these treatments have limited efficacy and can cause significant side effects. Curcumin and its analogues have recently emerged as promising drugs for the treatment of HPH. Curcumin, a natural compound found in turmeric flavoring, has been shown to possess anti-inflammatory, antioxidant, and anti-proliferative properties. In animal models of HPH, curcumin and its analogues have been shown to inhibit the proliferation and migration of PASMCs and reduce pulmonary vascular resistance. These compounds also exhibit good safety and low toxicity, making them potential alternatives to or adjuvant therapies for current HPH treatments. Curcumin analogue WZ35, a derivative of curcumin, possesses superior biological function and has significantly greater cellular effects than curcumin alone. Analyses of various cell types showed that WZ35 was superior to curcumin at lethal doses. However, its low bioavailability and potential toxic side effects limit its application in clinical treatment. Summary of the Invention

[0004] This invention provides a MOF Cu @WZ35 nanoparticles, their preparation methods, and applications; this MOF Cu @WZ35 nanoparticles encapsulate WZ35 through a Cu-based metal-organic framework, which enhances the efficacy of WZ35 in treating HPH and could serve as a potential drug for treating HPH.

[0005] A MOF Cu The preparation methods of @WZ35 nanoparticles include:

[0006] (1) Dimethylimidazole and copper acetate were dissolved in methanol to obtain dimethylimidazole solution and copper acetate solution, respectively;

[0007] (2) Copper acetate solution was added dropwise to dimethylimidazole solution, and compound WZ35 was added simultaneously. The mixture was stirred at 30-40°C for 20-40 minutes, and then centrifuged and washed to obtain the MOF. Cu @WZ35 nanoparticles;

[0008] The structural formula of the compound WZ35 is as follows:

[0009]

[0010] Experimental results show that MOF Cu @WZ35 nanoparticles can inhibit the proliferation of rat PASMCs, promote apoptosis, and have the potential to reduce rat HPH. MOF Cu The development of @WZ35 nanoparticles as a therapeutic agent for HPH may provide a promising treatment option for HPH patients.

[0011] Preferably, in step (1), the ratio of dimethylimidazole to methanol is 1 mg: 2-3 mL;

[0012] The ratio of copper acetate to methanol is 1 mg: 5-6 mL.

[0013] The present invention also provides a MOF obtained by the preparation method described above. Cu @WZ35 nanoparticles.

[0014] The present invention also provides the aforementioned MOF Cu Application of @WZ35 nanoparticles in the preparation of drugs for pulmonary hypertension.

[0015] Preferably, the MOF Cu @WZ35 nanoparticles are used to treat or alleviate hypoxic pulmonary hypertension.

[0016] Preferably, the MOF Cu @WZ35 nanoparticles are used to inhibit the proliferation of pulmonary artery smooth muscle cells.

[0017] Compared with the prior art, the beneficial effects of the present invention are reflected in:

[0018] This invention utilizes a Cu-based metal-organic framework (MOF) to encapsulate WZ35, which, compared to free WZ35, provides higher performance. Cu@WZ35 can more effectively inhibit the proliferation of PASMCs and promote their death, thereby alleviating HPH. Its development as a therapeutic agent for HPH may provide a promising treatment option for HPH patients. Attached Figure Description

[0019] Figure 1 Figure showing the diameter results of two types of particles, MOFCu and MOFCu@WZ35, measured using Zetasizer Nano ZS;

[0020] Figure 2 Images of MOFCu@WZ35 detected by transmission electron microscopy;

[0021] Figure 3 To detect cellular uptake of MOFCu-RhoB and RhoB using laser scanning confocal microscopy;

[0022] Figure 4 MOF Cu @WZ35 triggers cell viability in PASMCs. Detailed Implementation

[0023] Materials and Cells

[0024] Dimethylimidazole, methanol, and copper acetate were purchased from JingQiao Bio.Company. DAPI and DCFH-DA antibodies were from Dakewei Company. PASMCs were obtained from rats. PASMCs were cultured in DMEM medium containing 10% FBS under hypoxic pretreatment. All solutions and materials were used according to the manufacturer's instructions.

[0025] Example 1: Preparation and Characterization of Two Types of Nanoparticles

[0026] (1) 5 mg of dimethylimidazole was dissolved in 10 mL of methanol. 1 mg of copper acetate was dissolved in 5 mL of methanol. 2.5 mL of copper acetate solution was added dropwise to 5 mL of dimethylimidazole solution, and the mixture was stirred at 37 °C for 30 min. Finally, the prepared MOF... Cu Centrifuge at 8000 rpm for 20 min. Wash the collected nanoparticles three times with methanol. Store the purified nanoparticles at 4℃ for later use. Determine the particle size using dynamic light scattering.

[0027] (2) 5 mg of dimethylimidazole was dissolved in 10 mL of methanol. 1 mg of copper acetate was dissolved in 5 mL of methanol. 2.5 mL of copper acetate solution was added dropwise to 5 mL of dimethylimidazole solution. 5 mg of WZ35 was added to the mixed solution, and the mixture was incubated at 37 °C for 30 min. The prepared MOF... CuThe nanoparticles were centrifuged at 8000 rpm for 20 min using a WZ35 micrometer. The collected nanoparticles were washed three times with methanol. The purified nanoparticles were stored at 4 °C for later use. The MOF was measured by dynamic light scattering. Cu The diameter of the WZ35 nanoparticles was determined. Simultaneously, we used transmission electron microscopy to study the MOF... Cu The morphology of @WZ35 was measured.

[0028] The sizes of two types of nanoparticles were measured using Zetasizer Nano ZS, and the results are shown in [Figure number missing]. Figure 1 ; . Figure 1 The results show that MOF Cu Its diameter is approximately 116 nm. Figure 1 a), while MOF Cu The diameter of @WZ35 is approximately 119nm. Figure 1 b).

[0029] After characterization, the prepared MOFs were... Cu @WZ35 was used for further morphology analysis. The prepared nanoparticles were prepared at a concentration of 1 mg / mL and dispersed by ultrasound to ensure homogeneity. Then the prepared MOF was... Cu @WZ35 nanoparticles were dropped onto a copper grid. Excess liquid was removed with absorbent paper, and the sample was dried in an oven. Then, they were examined using a transmission electron microscope. It was clearly visible that the nanoparticles exhibited a spherical structure. Figure 2 Measurements show that the diameter of the nanoparticles is around 100 nm. Furthermore, the images also show some aggregates within the nanoparticles. These black dots are present in all the nanoparticles. Therefore, we hypothesize that MOF... Cu It may contain the drug WZ35.

[0030] Example 2 In vitro cell uptake assay

[0031] To verify MOF Cu To assess the targeting ability of PASMCs, we labeled MOFs with Rhobmine B. Cu : Mix 1 mg of Rhobmine B with 1 mL of prepared MOF Cu The nanoparticles were mixed overnight at 5000 rpm at room temperature, allowing them to be indicated by red fluorescence and visualized in cells. Rhobmine B was used as a control group for comparison. The fluorescently labeled nanoparticles and free dye were incubated with pasmc cells (cultured in hypoxic pretreatment) for 8 hours. Cells were then washed three times with PBS. Subsequently, the cytoskeleton was stained with green phthalide. The cell nuclei were simultaneously stained with 0.1% DAPI. The sections were then mounted on glass with an anti-fluorescence quencher. Finally, they were observed using a laser scanning confocal microscope.

[0032] To visualize using MOF Cu To assess the drug delivery effect, we decided to use Rhomine B fluorescent dye to label the nanoparticles. Then, we applied MOF... Cu RhoB red nanoparticles were incubated with pasmc. Cells were collected after incubation and stained with DAPI and phalloidine. They were then examined using laser scanning confocal microscopy. We discovered MOFs. Cu -The red fluorescence of the RhoB group was much stronger than that of the RhoB group. Figure 3 The results show that MOF Cu It can deliver more payload to cells.

[0033] Example 3: In vitro cell viability assay

[0034] This embodiment uses MOFs of different concentrations. Cu Cells were incubated with WZ35, and MOFs were added at concentrations of 2.5 μg / mL (based on WZ35), 5 μg / mL, and 10 μg / mL, respectively. Cu @WZ35 particles and 10,000 PASMCs were co-incubated at 37°C for 24 hours to determine the effect of inhibition on cell proliferation. PASMC viability was assessed using a CCK8 assay kit. Cell viability was calculated compared to the control group. Results showed that MOF... Cu @WZ35 treatment can affect the proliferation of PASMCs.

[0035] Using free WZ35 10ug / mL and different concentrations of MOF Cu @WZ35 cultured PASMCs in a hypoxic microenvironment. Since HPH is caused by malignant cell proliferation under hypoxic conditions, we also selected PASMCs cultured under normoxic conditions as a control. The remaining cells were cultured in a hypoxic microenvironment to maintain their HPH disease potential. Results showed MOF... Cu @WZ35 cell viability was lower than that of the control group and the free WZ35 group. Furthermore, we found that MOF... Cu Cell viability in the WZ35 group decreased with increasing WZ35 content. Figure 4 Therefore, the results indicate that nanoparticles can facilitate drug delivery to target cells. These results confirm that this nano-drug delivery system can effectively inhibit the abnormal proliferation of PASMCs under hypoxic conditions and has the potential for application in the treatment of HPH.

Claims

1. A MOF Cu The method for preparing @WZ35 nanoparticles is characterized by, include: (1) Dimethylimidazole and copper acetate were dissolved in methanol to obtain dimethylimidazole solution and copper acetate solution, respectively; (2) Copper acetate solution was added dropwise to dimethylimidazole solution, and compound WZ35 was added simultaneously. The mixture was stirred at 30-40°C for 20-40 minutes, and then centrifuged and washed to obtain the MOF. Cu @WZ35 nanoparticles; The structural formula of the compound WZ35 is as follows:

2. The MOF according to claim 1 Cu The method for preparing @WZ35 nanoparticles is characterized by, In step (1), the ratio of dimethylimidazole to methanol is 1 mg: 2-3 mL; The ratio of copper acetate to methanol is 1 mg: 5-6 mL.

3. A MOF obtained by the preparation method according to claim 1 or 2 Cu @WZ35 nanoparticles.

4. A MOF according to claim 3 Cu Application of @WZ35 nanoparticles in the preparation of drugs for pulmonary hypertension.

5. The MOF according to claim 4 Cu The application of @WZ35 nanoparticles in the preparation of drugs for treating pulmonary hypertension is characterized by... The MOF Cu @WZ35 nanoparticles are used to treat or alleviate hypoxic pulmonary hypertension.

6. The MOF according to claim 4 Cu The application of @WZ35 nanoparticles in the preparation of drugs for treating pulmonary hypertension is characterized by... The MOF Cu @WZ35 nanoparticles are used to inhibit the proliferation of pulmonary artery smooth muscle cells.

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