Synthesis method of novel inhalable lipidosome and application of novel inhalable lipidosome in ARDS treatment

Abstract

The invention provides a synthesis method of a novel inhalable liposome and an application of the novel inhalable liposome in ARDS (arterial thrombosis syndrome) treatment, and the synthesis method comprises the following steps: firstly, mixing DSPE-PEG2K-TK-NHS and tocilizumab (TCZ: Tocilizumab) to form a lead compound DSPE-PEG2K-TK-TCZ; the preparation method comprises the following steps: dissolving DPPC, cholesterol, DSPE-TK-PEG2k and FG4592 in 5mL of chloroform, carrying out reduced pressure evaporation in a sample bottle to form a film, and adding 3mL of deionized water to prepare Lip-coated RF; and then stirring and incubating the DSPE-PEG2K-TK-TCZ and the Lip (at) RF, so as to form the Lip (at) RFT. As a functional nano-material with ultra-small size, good biocompatibility, excellent ROS removal capability and ROS responsive drug release, the nano-material is used for treating ARDS. The invention has the advantages that: the Lip-coated RFT relieves lung oxidative stress, inflammatory response, protein permeability and histological severity in the LPS-induced mouse ALI model. Therefore, Lip at RFT can be used as a new way for treating ARDS. Aerosol inhalation of Lip-coated RFT can effectively improve the severity of ARDS.

Description

Technical Field

[0001] The invention relates to the technical field of biomedicine and molecular biology, and in particular to a synthesis method of a novel inhalable liposome and its application in the treatment of ARDS. Background Art

[0002] Acute Respiratory Distress Syndrome (ARDS) is a serious lung disease characterized by acute onset of diffuse alveolar damage and inflammatory response, leading to severe hypoxemia. ARDS usually occurs after other serious diseases or trauma, such as pneumonia, sepsis, inhalation of harmful substances, severe trauma, etc.

[0003] ARDS is estimated to occur in about 0.5% to 1% of all hospitalized patients and in a higher rate of about 10% of those admitted to intensive care units, meaning that about 5 to 7 people per 100,000 will develop ARDS each year.

[0004] Excessive inflammatory response is one of the fundamental disease mechanisms of ARDS. Various stimuli may lead to endothelial and epithelial damage, which will trigger the recruitment of leukocytes to the lungs. On the one hand, activated inflammatory cells can release toxic mediators, including reactive oxygen species (ROS) and cytokines, leading to tissue damage and dysfunction of the endothelial and epithelial barriers. On the other hand, increased barrier permeability promotes the influx of leukocytes, which will further aggravate inflammation and oxidative stress. Therefore, clearing ROS and controlling excessive inflammation are necessary steps to break this vicious cycle and restore organ function.

[0005] Currently, the clinical treatment of ARDS is mainly based on supportive care, including protective mechanical ventilation, prone ventilation, fluid management strategies, and extracorporeal membrane oxygenation (ECMO). Unfortunately, the morbidity and mortality of ARDS remain high. There is still a lack of specific and effective treatments. Therefore, specific and effective treatments for ARDS are urgently needed. Summary of the invention

[0006] The purpose of the present invention is to overcome the above problems and particularly provide a method for synthesizing a novel inhalable liposome and its application in the treatment of ARDS.

[0007] Liposomes are nanostructures composed of phospholipid bilayers. Their structure is extremely similar to that of cell membranes, so they are highly biocompatible and biodegradable and are commonly used in drug delivery systems.

[0008] Nebulizer inhalation therapy is a treatment method that converts liquid medicine into tiny particles through a nebulizer, allowing patients to inhale the medicine directly into the respiratory tract and lungs through breathing. It is an important treatment for respiratory diseases.

[0009] The use of reactive oxygen species (ROS)-responsive drug delivery systems is an emerging strategy that aims to improve the release efficiency and targeting of drugs in specific pathological environments. Such systems are usually designed to respond and release drugs in high ROS environments, thereby achieving precise treatment of diseased tissues.

[0010] Tocilizumab is a recombinant humanized monoclonal antibody that targets the interleukin-6 receptor (IL-6R) and is used to block the IL-6 signaling pathway. During the COVID-19 pandemic, Tocilizumab has been studied and used to treat severe COVID-19 patients due to its potential anti-inflammatory effects. Currently, it is usually administered by intravenous injection, and aerosol inhalation has not yet been clinically used.

[0011] FG-4592, also known as Roxadustat, is an oral, small molecule hypoxia-inducible factor prolyl hydroxylase inhibitor (HIF-PHI). It stabilizes hypoxia-inducible factor (HIF) by inhibiting HIF prolyl hydroxylase, thereby promoting the production of erythropoietin (EPO) and other gene expressions related to erythropoiesis, thereby being used to treat anemia in patients with chronic kidney disease (CKD). It has not yet been used to treat ARDS.

[0012] The present invention provides a method for synthesizing a novel inhalable liposome and its application in treating ARDS, specifically a method for synthesizing inhalable liposomes loaded with FG4592 and tocilizumab and having ROS response, and its application in treating acute respiratory distress syndrome.

[0013] Synthesis method:

[0014] First, DSPE-PEG2K-TK-NHS was mixed with Tocilizumab (TCZ) to form the lead compound DSPE-PEG2K-TK-TCZ. DPPC, cholesterol, DSPE-TK-PEG2k, and FG4592 were dissolved in 5 mL of chloroform and evaporated under reduced pressure in a sample bottle to form a film, and 3 mL of deionized water was added to prepare Lip@RF. DSPE-PEG2K-TK-TCZ and Lip@RF were then incubated with continuous stirring at 4°C for 12 hours to finally form Lip@RFT.

[0015] As a functional nanomaterial with ultra-small size, good biocompatibility, excellent ROS scavenging ability and ROS-responsive drug release, it is used to treat ARDS.

[0016] Administration by nebulized inhalation significantly improved lung oxidative stress, inflammatory response, protein permeability, and histological severity in ALI mice induced by various pathogens without causing any safety issues.

[0017] The ALI mouse model was established by administering LPS through endotracheal intubation to simulate the pathophysiological process of ARDS

[0018] Nebulized inhalation of Lip@RFT reduced pulmonary oxidative stress, inflammatory response, protein permeability, and histological severity without causing any safety issues.

[0019] The advantages of the present invention are:

[0020] The present invention uses LPS to create an ALI mouse model by endotracheal intubation to simulate the pathological and physiological process of ARDS. After aerosol inhalation of Lip@RFT, pathological staining of lung tissue shows reduced exudation of inflammatory cells in the lungs, weakened thickening of alveolar walls, and reduced appearance in alveoli. ELISA shows a decrease in the level of inflammatory factor IL-6. Lung dry-wet ratio and bronchoalveolar lavage fluid protein detection found a decrease in lung wet-dry ratio and a decrease in bronchoalveolar lavage fluid protein level. ROS detection found an increase in lung tissue GSH level and a decrease in MDA level.

[0021] The results showed that Lip@RFT reduced lung oxidative stress, inflammatory response, protein permeability and histological severity in the LPS-induced ALI model in mice. Therefore, Lip@RFT can be used as a new approach to treat ARDS. Inhalation of Lip@RFT can effectively improve the severity of ARDS. DETAILED DESCRIPTION

[0022] The present invention will be further explained below in conjunction with specific implementation schemes, but the present invention is not limited thereto. The structures, proportions, sizes, etc. illustrated in the specification are only used to match the contents disclosed in the specification for people familiar with this technology to understand and read, and are not used to limit the restrictive conditions under which the present invention can be implemented. Therefore, they have no substantive technical significance. Any structural modification, change in proportional relationship, or adjustment of size should still fall within the scope of the technical content disclosed by the present invention without affecting the effects and purposes that can be achieved by the present invention.

[0023] This embodiment provides a method for synthesizing a novel inhalable liposome and its application in the treatment of ARDS, namely, a method for synthesizing inhalable ROS-responsive liposomes loaded with FG4592 and tocilizumab and its application in the treatment of acute respiratory distress syndrome.

[0024] Synthesis method:

[0025] First, DSPE-PEG2K-TK-NHS was mixed with Tocilizumab (TCZ) to form the lead compound DSPE-PEG2K-TK-TCZ. DPPC, cholesterol, DSPE-TK-PEG2k, and FG4592 were dissolved in 5 mL of chloroform and evaporated under reduced pressure in a sample bottle to form a film, and 3 mL of deionized water was added to prepare Lip@RF. DSPE-PEG2K-TK-TCZ and Lip@RF were then incubated with continuous stirring at 4°C for 12 hours to finally form Lip@RFT.

[0026] As a functional nanomaterial with ultra-small size, good biocompatibility, excellent ROS scavenging ability and ROS-responsive drug release, it is used to treat ARDS.

[0027] Administration by nebulized inhalation significantly improved lung oxidative stress, inflammatory response, protein permeability, and histological severity in ALI mice induced by various pathogens without causing any safety issues.

[0028] The ALI mouse model was established by administering LPS through endotracheal intubation to simulate the pathophysiological process of ARDS

[0029] Nebulized inhalation of Lip@RFT reduced pulmonary oxidative stress, inflammatory response, protein permeability, and histological severity without causing any safety issues.

[0030] Matters not covered by the present invention are known technologies.

[0031] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A method for synthesizing a novel inhalable liposome, characterized in that: The synthesis method of the novel inhalable liposomes, specifically the synthesis method of inhalable liposomes loaded with FG4592 and tocilizumab with ROS response, comprises the following steps: First, DSPE-PEG2K-TK-NHS was mixed with Tocilizumab (TCZ) to form the lead compound DSPE-PEG2K-TK-TCZ; DPPC (dipalmitoylphosphatidylcholine), cholesterol, DSPE-TK-PEG2k (distearoylphosphatidylethanolamine-thioketal-polyethylene glycol) and FG4592 (roxadustat) were mixed in a molar ratio of 12:8:1:3 and dissolved in 5 mL of chloroform. The mixture was evaporated under reduced pressure in a sample bottle to form a film. 3 mL of deionized water was added to prepare Lip@RF (ROS-responsive FG4592-loaded liposomes). Then, DSPE-PEG2K-TK-TCZ and Lip@RF were incubated with continuous stirring at 2-8°C (it is best to give a range of temperature values) for 12 hours to finally form Lip@RFT (ROS-responsive liposomes loaded with FG4592 and tocilizumab).

2. An application of the synthesis method of the novel inhalable liposome as claimed in claim 1 in the treatment of ARDS, characterized in that: The novel inhalable liposomes, specifically inhalable ROS-responsive liposomes loaded with FG4592 and tocilizumab, are functional nanomaterials with ultra-small size, good biocompatibility, excellent ROS scavenging ability and ROS-responsive drug release, and are used to treat ARDS.