Application of mitoxantrone in preparation of lymph tracer

Abstract

The invention provides an application of mitoxantrone and / or a pharmaceutical salt thereof in preparation of a lymphatic tracer, and the lymphatic tracer is used for lymph node tracing in gastric resection related diseases and has a good dyeing effect on gastric cancer lymph nodes.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical preparations, and particularly relates to the application of mitoxantrone in the manufacture of lymphatic tracers for diseases related to gastrectomy. Background Art

[0002] Gastric cancer is the fifth most common cancer globally and the third most common cause of cancer-induced death. There are approximately 1 million newly diagnosed gastric cancer patients each year, and more than 700,000 people die from it. In China, there are more than 400,000 newly diagnosed gastric cancer patients each year. In the treatment of advanced gastric cancer, with the development of chemotherapy and small molecule targeted therapy, the survival period of patients has been extended, but surgical resection remains the most effective way to cure cancer. The Japanese Gastric Cancer Treatment Guidelines 2010 define D2 dissection as the standard surgical procedure for gastric cancer treatment, which is also accepted by Eastern and Western scholars.

[0003] The Japanese Gastric Cancer Treatment Guidelines 2010 divide the lymph nodes involved in gastric cancer lymph node dissection into multiple groups, such as NO.1 - right gastric cardia lymph nodes, NO.2 - left gastric cardia lymph nodes, NO.3 - lesser curvature lymph nodes, NO.4 - greater curvature lymph nodes, etc. Currently, there are five standards for gastric cancer surgical lymph node dissection: the first level is radical resection of grade 0 (D0 surgery), the second level is radical resection of grade 1 (D1 surgery), the third level is radical resection of grade 2 (D2 surgery), the fourth level is radical resection of grade 3 (D3 surgery), and the fifth level is radical resection of grade 4 (D4 surgery). D0 surgery has a very low radicality and is basically not used by surgeons except for extremely early gastric cancer. D1 surgery performs the first-station lymph node dissection around the stomach, which has a certain radicality and high safety. D2 surgery performs the first-station and second-station lymph node dissections around the stomach, which has a higher radicality, but the surgical risk is relatively large, especially in non-large tumor centers, postoperative complications are likely to occur. Due to the issues of indications and surgical difficulty, D3 and D4 surgeries are usually only performed on patients with special medical conditions in large tumor centers, and the surgical risks of such surgeries are several times higher than that of D2 surgery. There are two benefits to removing as many lymph nodes as possible: 1. The surgery is thorough and the tumor residue is minimized to the greatest extent; 2. It provides a more accurate postoperative pathological stage, which is beneficial for the formulation of subsequent treatment (chemotherapy) plans. That is to say, the most important and fundamental point of the quality of gastric cancer surgery is whether the lymph nodes are completely removed.

[0004] The lymph nodes related to the stomach have the characteristics of a large number and a wide distribution area, and the lymph nodes are often hidden in the surrounding tissues, making it difficult to distinguish them with the naked eye. It is often very difficult to remove as many lymph nodes as possible. If a lymphatic tracer can be used during the surgery to stain and trace the living lymph nodes to help doctors completely remove the lymph nodes, it is the key to improving the treatment effect of malignant tumors.

[0005] Therefore, developing safe and effective lymphatic tracers and effectively performing lymph node dissection in the tumor-draining area are important means to improve the quality of life and prolong the lifespan of patients. Lymphatic tracers currently play a crucial role in the clinical treatment of malignant tumors at home and abroad. Injecting lymphatic tracers near the primary lesion can quickly stain the lymph nodes near the malignant tumor.

[0006] Currently known lymphatic tracers include nano-carbon, methylene blue, etc. However, nano-carbon and others have the following problems: Nano-carbon and the like are often used for intraoperative sentinel lymph node tracing in breast cancer and thyroid cancer. However, nano-carbon accumulates, is not metabolized in the body, and there is a risk of blocking capillaries when entering the blood and lymphatic circulation. Moreover, for cancer tissues with a hard and brittle texture, direct intratumoral injection can cause tumor tissue necrosis, shedding, and bleeding. Furthermore, nano-carbon is not easily taken up by small lymph nodes, so the tracing effect on sentinel lymph nodes in gastric cancer is poor, resulting in the continued proliferation and metastasis of tumor cells remaining in small lymph nodes after gastric cancer surgery, ultimately leading to poor prognosis. In addition, there are significant differences in the requirements for the size of lymph nodes that can be dissected by tracers between gastric cancer resection and breast cancer resection. Breast cancer tracing mainly focuses on sentinel lymph node tracing. If the sentinel lymph node has no metastasis, the possibility of metastasis in other lymph nodes in this area is very small. However, the tumor metastasis pathways of gastric cancer and breast cancer are different. Gastric cancer resection requires tracing as many lymph nodes as possible to the smallest size (≤1mm).

[0007] Therefore, seeking to develop a safe and effective lymphatic tracer for intraoperative lymph node tracing in gastric cancer to predict whether the tumor has metastasis is an important means to improve the quality of life and delay the lifespan of gastric cancer patients. Summary of the Invention

[0008] The present invention aims to provide the use of mitoxantrone and / or its pharmaceutically acceptable salts in the preparation of a lymphatic tracer for lymph node tracing in diseases related to gastric resection.

[0009] In the present invention, unless otherwise specified, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Moreover, the protein and nucleic acid chemistry, molecular biology, cell and tissue culture, microbiology, immunology-related terms and laboratory operation procedures used herein are all widely used terms and conventional procedures in the corresponding fields. At the same time, for a better understanding of the present invention, the following provides definitions and explanations of relevant terms.

[0010] It should also be understood that the terms used herein are only for the purpose of describing specific embodiments and are not intended to be restrictive.

[0011] As used herein, the terms "patient", "individual", and "subject" are used interchangeably and refer to any single animal desiring treatment, more preferably a mammal (including non-human animals such as cats, dogs, horses, rabbits, zoo animals, cows, pigs, sheep, and non-human primates). In certain embodiments, the patient herein is a human. The patient may have, be suspected of having, or be at risk of having a gastric tumor. As used herein, "disorder" refers to any condition that would benefit from treatment, including but not limited to chronic and acute disorders or diseases, including those pathological conditions that render a mammal susceptible to the disorder in question.

[0012] As used herein, "pharmaceutical formulation" refers to a formulation in a form that allows the bioactivity of the active ingredient contained therein to be effective and that does not contain other components that are unacceptably toxic to the subject to which the formulation is administered.

[0013] As used herein, "pH adjuster" refers to a compound or mixture of compounds that can be used to ensure that the pH of a reconstitution kit is within an acceptable range (pH of about 4.0 - 10.5) for human or mammalian administration. Suitable pH adjusters include pharmaceutical buffers such as tricine, phosphate, or TRIS (i.e., tris(hydroxymethyl)aminomethane); pharmaceutical acids such as pharmaceutical organic acids (e.g., formic acid, acetic acid) or mixtures thereof or inorganic acids (e.g., hydrochloric acid, phosphoric acid) or mixtures thereof, and pharmaceutical bases such as sodium carbonate, sodium bicarbonate, or mixtures thereof. When the conjugate used is in the form of an acid salt, the pH adjuster may optionally be provided in a separate vial or container so that the kit user can adjust the pH as part of a multi-step operation.

[0014] As used herein, "pharmaceutical excipient" refers to a non-toxic component of a pharmaceutical formulation other than the active ingredient. Pharmaceutical excipients include but are not limited to buffers, carriers, stabilizers, or preservatives.

[0015] As used herein, "pharmaceutical salt" denotes a salt that is not biologically or otherwise undesirable. Pharmaceutical salts include acid and base addition salts. The phrase "pharmaceutical" indicates that the substance or composition must be chemically and / or toxicologically compatible with the other components of the formulation and / or the mammal being treated therewith.

[0016] As used herein, "pharmaceutically acceptable acid addition salts" refers to those salts formed with inorganic acids and organic acids, said inorganic acids being selected from hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, carbonic acid, phosphoric acid, and said organic acids being selected from aliphatic, alicyclic, aromatic, aralkyl, heterocyclic, carboxylic and sulfonic organic acids, such as formic acid, acetic acid, propionic acid, glycolic acid, gluconic acid, lactic acid, pyruvic acid, oxalic acid, malic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, aspartic acid, ascorbic acid, glutamic acid, anthranilic acid, benzoic acid, cinnamic acid, mandelic acid, pamoic acid, phenylacetic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid.

[0017] The term "pharmaceutically acceptable base addition salts" denotes those salts formed with organic or inorganic bases. Examples of acceptable inorganic bases include sodium, potassium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Salts derived from pharmaceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, substituted amines (including naturally occurring substituted amines), salts of cyclic amines, and salts of basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-diethylaminoethanol, trimethylamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucosamine, theobromine, purine, piperazine, piperidine, N-ethylpiperidine, and salts of polyamine resins.

[0018] As used herein, "treatment" refers to a clinical intervention that attempts to alter the natural course of the individual being treated and can be used to effect prevention or be applied during the course of a clinical pathology. Desirable therapeutic outcomes include, but are not limited to, preventing the occurrence or recurrence of disease, alleviating symptoms, reducing any direct or indirect pathological consequences of the disorder, preventing metastasis, reducing the rate of disease progression, remission or alleviation of the disease state, and remission or improvement of the prognosis.

[0019] As used herein, "administration" refers to the method of giving a subject (e.g., a patient) a dose of a compound (e.g., mitoxantrone hydrochloride injection) or a pharmaceutical composition (e.g., a pharmaceutical composition containing an inhibitor or antagonist). Administration can be effected by any suitable means, including parenteral, intranasal, and inhalational administration, and, if desired for local treatment, by intralesional administration. Parenteral infusion includes, for example, intramuscular, intravenous, intraarterial, intraperitoneal, or subcutaneous administration. Administration can be effected by any suitable route, for example, by injection, such as intravenous or subcutaneous injection, partly depending on whether the administration is to be brief or long-term. A variety of dosing regimens are contemplated herein, including, but not limited to, single or multiple administrations at different time points, bolus administration, and pulse infusion.

[0020] As used herein, the Full Analysis Set (FAS): a set of subjects in accordance with the Intention To Treat (ITT) principle. It refers to the data set composed of all subjects who participated in the trial, received treatment, and had a baseline efficacy evaluation.

[0021] As used herein, the Per Protocol Set (PPS): refers to a subgroup of the treatment population that has completed the trial and excluded serious protocol violations (referring to the study subjects violating the inclusion criteria or exclusion criteria), which is a set of cases that meet the inclusion criteria, do not meet the exclusion criteria, and have completed the treatment protocol.

[0022] As used herein, "lymph node metastasis" means that gastric cancer cells metastasize to lymph nodes somewhere along the lymphatic tract, which is an important way for gastric cancer to metastasize. Generally speaking, lymph node metastasis of gastric cancer is from near to far. As the cancer tumor extends deeper, the chance of metastasis increases. According to the order of metastasis, it is divided into 3 stations: The first station is the superficial group of lymph nodes closest to the cancer body and attached to the gastric wall, such as the lymph nodes in the greater curvature of the stomach, lesser curvature of the stomach, above and below the pylorus, and beside the cardia (hereinafter, sometimes the lymph nodes of the first station are called "N1 lymph nodes", and the stained lymph nodes of the first station are called "N1 stained lymph nodes"); The second station is the deep group of lymph nodes that drain the superficial group of lymph nodes, such as the lymph nodes at the splenic hilum, common hepatic artery, left gastric artery trunk, and pancreaticoduodenal artery (hereinafter, sometimes the lymph nodes of the second station are called "N2 lymph nodes", and the stained lymph nodes of the second station are called "N2 stained lymph nodes"); The third station includes the lymph nodes around the celiac artery, para-aortic, porta hepatis, root of the mesentery, and around the middle colic artery, and sometimes metastasizes to the left supraclavicular lymph nodes (hereinafter, sometimes the lymph nodes of the third station are called "N3 lymph nodes", and the stained lymph nodes of the third station are called "N3 stained lymph nodes").

[0023] As used herein, "TNM tumor staging" refers to the AJCC / UICC TNM staging of gastric cancer. For the specific staging criteria, please refer to the Guidelines for the Diagnosis and Treatment of Gastric Cancer (2022).

[0024] The present invention provides the use of mitoxantrone and / or its pharmaceutically acceptable salts in the preparation of a lymphatic tracer for lymph node tracing in diseases related to gastrectomy.

[0025] In some specific embodiments, the lymph nodes are lymph nodes with a short diameter less than or equal to 2 mm, further may be lymph nodes with a short diameter less than or equal to 1.5 mm, and preferably are lymph nodes with a short diameter less than or equal to 1 mm.

[0026] The inventors of the present invention have found through research that mitoxantrone and / or its pharmaceutically acceptable salts described in the present invention can be used to prepare a lymphatic tracer for lymph node tracing during gastrectomy, which can stain lymph nodes with a short diameter less than or equal to 1 mm, and has a good lymph node staining effect during resection surgeries of various parts and orientations of the stomach and the perigastric region.

[0027] In some specific embodiments, the detection rate of the tracer for lymph nodes with a short diameter less than or equal to 1 mm is greater than 70%, and further greater than 75%.

[0028] In the present invention, the color development time of the tracer is less than 25 min, and further less than 20 min.

[0029] In some specific embodiments, the fading time of the tracer is greater than 180 minutes.

[0030] In some specific embodiments, the lymph node staining rate of the tracer during gastrectomy is greater than 50%, and further greater than 55%.

[0031] In some specific embodiments, the diseases related to gastrectomy are gastric polyps or gastric tumors.

[0032] In some specific embodiments, the gastric tumors include gastric benign tumors and gastric malignant tumors.

[0033] In some specific embodiments, the gastric benign tumors and gastric malignant tumors are selected from gastric cardia cancer, gastric body cancer, and gastric antrum cancer.

[0034] In some specific embodiments, the diseases related to gastrectomy are selected from one or more of the tumors located in the following parts: cardia, cardia mucous flexure side, cardia posterior wall, cardia lesser curvature, cardia gastric body lesser curvature side, cardia gastric fundus lesser curvature anterior wall, gastric fundus, gastric fundus lesser curvature, gastric antrum anterior wall, greater curvature of the stomach, gastric antrum junction, gastric antrum lesser curvature, gastric body upper posterior wall, gastric body lower posterior wall, gastric fundus gastric body greater curvature side, gastric lesser curvature posterior wall, anterior and posterior walls of the lesser curvature, gastric fundus, gastric body lesser curvature posterior wall.

[0035] In some specific embodiments, the tumors are tumors in TNM tumor stage I, II, III, or IV.

[0036] In some specific embodiments, the gastrectomy is selected from total gastrectomy or subtotal gastrectomy.

[0037] In some specific embodiments, the lymphatic tracer is used for lymph node tracing in gastric tumors.

[0038] In some specific embodiments, the lymphatic tracer contains mitoxantrone and / or its pharmaceutically acceptable salts and a pharmaceutical excipient.

[0039] In some specific embodiments, the pharmaceutical excipients include, but are not limited to, one or more of an osmotic pressure regulator, an antioxidant, an adsorbent, a filler, a buffer, a carrier, a stabilizer, or a preservative.

[0040] In some specific embodiments, the lymphatic tracer is an injection.

[0041] In some specific embodiments, the dosage form of the injection is a solution, a lyophilized powder, an emulsion, a liposome, a nanoparticle, a nanocrystal, a microcrystal, a microsphere, or a gel.

[0042] In some preferred embodiments, the solution is sodium chloride injection or glucose injection.

[0043] In some specific embodiments, the injection is formulated as a dosage form for use at a concentration of 2 - 10 mg / mL, and the single-use dose of the drug is 0.5 - 3 mL, for example, it can be 0.5 mL, 1 mL, 1.5 mL, 2.5 mL, or 3 mL, preferably 2 - 3 mL.

[0044] In some preferred embodiments, the pharmaceutical salts are selected from one or several of mitoxantrone hydrochloride, mitoxantrone oxalate, mitoxantrone sulfate, mitoxantrone phosphate, mitoxantrone acetate, and mitoxantrone citrate.

[0045] Preferably, the pharmaceutical salt is mitoxantrone hydrochloride.

[0046] In some specific embodiments, the tracer is an injection, and its components contain mitoxantrone or mitoxantrone salts: 0.05% - 5% by mass / volume ratio (g / mL), and an osmotic pressure regulator: 0.1 - 10%.

[0047] In some specific embodiments, the tracer further contains a buffer: 0.01 - 0.1%, an antioxidant: 0.01 - 0.1%, an adsorbent: 0.05 - 1%, and a filler: 0 - 20%.

[0048] In some specific embodiments, the osmotic pressure regulator is a mixture of one or several substances selected from sodium chloride, glucose, sorbitol, mannitol, glycerol, phosphate, and citrate.

[0049] In some specific embodiments, the buffer is one or several of acetic acid, sodium acetate, citric acid, and sodium citrate.

[0050] In some specific embodiments, the antioxidant is one or several of sodium sulfite, sodium bisulfite, sodium metabisulfite, sodium thiosulfate, and disodium edetate. Polyethylene glycol is unstable at high temperatures, and antioxidants (such as sodium metabisulfite) help inhibit the oxidation to produce impurities.

[0051] In some specific embodiments, the filler is glucose, fructose, galactose, ribose or deoxyribose of monosaccharides, or sucrose, trehalose, maltose, lactose of disaccharides, or mannitol, sorbitol, lactitol, xylitol, maltitol, erythritol of polysaccharides, or one or more of them.

[0052] Preferably, the adsorbent is activated carbon.

[0053] In some specific embodiments, the lymph tracer contains a pH regulator.

[0054] In some preferred embodiments, the pH regulator is selected from one or more of hydrochloric acid, phosphoric acid, sulfuric acid, oxalic acid, acetic acid and citric acid.

[0055] In some specific embodiments, the pH value of the injection ranges between 2.8 and 4.3.

[0056] In some specific embodiments, the tracer further contains polyethylene glycol, and its mass concentration in the injection solution accounts for 0.01%-0.2%, which is beneficial to increasing the water solubility of mitoxantrone hydrochloride, improving its stability (for example, helping it not to decompose easily when encountering water), and is used for the preparation of nanoparticle drug carriers and controlled release systems, controlling drug delivery, and promoting lymphatic transport. Specifically, adding polyethylene glycol helps the drug to form a depot locally after injection, thereby prolonging the action time of the drug.

[0057] In one embodiment, the molecular weight of the polyethylene glycol is less than 2000, preferably less than 1000, further preferably 200, 400 or 600, and more preferably 400 or 600, which is more beneficial to the staining of mitoxantrone hydrochloride.

[0058] In some specific embodiments, the osmotic pressure is selected in the range of 285 to 2317 mmol / kg.

[0059] In some preferred embodiments, the osmotic pressure is selected in the range of 600-1200 mmol / kg. When the osmotic pressure is too small, the lymphatic drainage power is insufficient. When the osmotic pressure is too large, the concentration of the osmotic pressure regulator in the injection solution is too high, resulting in irritation at the injection site. Maintaining the osmotic pressure at 600-1200 mmol / kg is convenient for better staining of lymph nodes.

[0060] In some specific embodiments, the tracer has a nano-particle size of 20-100 nm under physiological conditions, can pass through the gaps between lymphatic endothelial cells, and produce a good lymphatic targeting effect. Preferably 30-60 nm, and further can be 40-60 nm.

[0061] In some preferred embodiments, the content of mitoxantrone in the mitoxantrone or its salt is 1 - 15 mg / ml by weight / volume; preferably 2 - 10 mg / ml; more preferably 2 mg / ml, 5 mg / ml or 10 mg / ml.

[0062] In some preferred embodiments, the content of sodium chloride is 0.3 - 1.8 mg / ml by weight / volume; preferably 0.4 - 1.6 mg / ml; more preferably 0.4 mg / ml, 0.8 mg / ml or 1.6 mg / ml.

[0063] In some preferred embodiments, the content of acetic acid is 0.015 - 1 mg / ml by weight / volume; preferably 0.023 - 0.092 mg / ml; more preferably 0.023 mg / ml, 0.046 mg / ml or 0.092 mg / ml.

[0064] In some preferred embodiments, the content of sodium acetate is 0.003 - 0.015 mg / ml by weight / volume; preferably 0.005 - 0.01 mg / ml; more preferably 0.005 mg / ml or 0.01 mg / ml.

[0065] In some preferred embodiments, the content of the antioxidant is 0.005 - 0.03 mg / ml by weight / volume; preferably 0.08 - 0.012 mg / ml; more preferably 0.01 mg / ml, 0.02 mg / ml or 0.03 mg / ml.

[0066] In some preferred embodiments, the content of sodium sulfate is 0.005 - 0.06 mg / ml by weight / volume; preferably 0.015 - 0.045 mg / ml; more preferably 0.015 mg / ml, 0.03 mg / ml or 0.045 mg / ml.

[0067] In some specific embodiments, the lymphatic tracer comprises:

[0068]

[0069] In some specific embodiments, the solvent is selected from: water, sodium chloride solution or glucose solution.

[0070] Preferably, the water is water for injection.

[0071] In a preferred embodiment, the content of impurity I in the mitoxantrone preparation is between 0 and 1.5%, and the structural formula of impurity I is shown in formula (II):

[0072]

[0073] In a preferred embodiment, the content of impurity II in the mitoxantrone preparation is between 0 and 1.5%, and the structural formula of the impurity II is shown in formula (III):

[0074]

[0075] In a preferred embodiment, the content of impurity III in the mitoxantrone preparation is between 0 and 1.5%, and the structural formula of the impurity III is shown in formula (IV):

[0076]

[0077] In a preferred embodiment, the content of impurity IV in the mitoxantrone preparation is between 0 and 1.5%, and the structural formula of the impurity IV is shown in formula (V):

[0078]

[0079] In some specific embodiments, the lymphatic tracer comprises: 2.91 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.9 g of sodium sulfate, and water for injection is added to 2000 ml;

[0080] or, 5.82 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.9 g of sodium sulfate, and water for injection is added to 2000 ml;

[0081] or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.9 g of sodium sulfate, and water for injection is added to 2000 ml;

[0082] or, 23.28 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.9 g of sodium sulfate, and water for injection is added to 2000 ml;

[0083] or, 11.64 g of mitoxantrone, 32 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.9 g of sodium sulfate, and water for injection is added to 2000 ml;

[0084] or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.2 g of sodium metabisulfite, 0.3 g of sodium sulfate, and water for injection is added to 2000 ml;

[0085] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 0.3 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0086] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 1.84 g of acetic acid, 0.2 g of sodium acetate, 0.2 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0087] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 1.84 g of acetic acid, 0.2 g of sodium acetate, 0.4 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0088] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 1.84 g of acetic acid, 0.2 g of sodium acetate, 0.6 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0089] Or, 11.64 g of mitoxantrone, 32 g of sodium chloride, 1.84 g of acetic acid, 0.2 g of sodium acetate, 0.3 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0090] Or, 11.64 g of mitoxantrone, 32 g of sodium chloride, 1.84 g of acetic acid, 0.2 g of sodium acetate, 0.3 g of sodium metabisulfite, 0.6 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0091] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 40 g of polyethylene glycol 400, 0.2 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection;

[0092] Or, 11.64 g of mitoxantrone, 16 g of sodium chloride, 0.92 g of acetic acid, 0.1 g of sodium acetate, 20 g of polyethylene glycol 1000, 0.2 g of sodium metabisulfite, 0.3 g of sodium sulfate, q.s. to 2000 ml with water for injection

[0093] In some specific embodiments, the lymphatic tracer is an injection, which is prepared by a method comprising the following steps:

[0094] Weigh the excipients in the prescribed amounts, add them to the prescribed amount of water for injection, stir to dissolve, and after dissolution, add the prescribed amount of mitoxantrone hydrochloride and / or its pharmaceutically acceptable salts.

[0095] It can be understood that the aforementioned osmotic pressure parameters can be adjusted by controlling the excipient formulation and content, etc., specifically based on the detection meeting the standards.

[0096] In some specific embodiments, the injection is prepared by the following method:

[0097] (1) Weigh the prescribed amounts of acetic acid, sodium acetate, sodium chloride, and disodium edetate; or weigh the prescribed amounts of acetic acid, sodium acetate, sodium chloride, and sodium metabisulfite; or weigh the prescribed amounts of acetic acid, sodium acetate, sodium chloride, sodium metabisulfite, and sodium sulfate, add them to the prescribed amount of injection water, and stir to dissolve to obtain an excipient mixture.

[0098] (2) Add the prescribed amount of mitoxantrone hydrochloride to the excipient mixture obtained in step (1), and stir to dissolve to obtain mitoxantrone hydrochloride injection; preferably, stir for 10 - 30 min to dissolve.

[0099] In some specific embodiments, the method further includes:

[0100] (3) Filter; preferably, filter through 0.45 μm and 0.22 μm filter membranes.

[0101] In some specific embodiments, the method further includes:

[0102] (4) Fill with nitrogen, 2 ml per vial, crimp the cap, and sterilize at 121 °C for 15 min; the pH value ranges between 2.8 - 4.3.

[0103] In some specific embodiments, step (1) further includes: weighing the prescribed amounts of polyethylene glycol, acetic acid, sodium acetate, sodium chloride, and disodium edetate; or weighing the prescribed amounts of polyethylene glycol, acetic acid, sodium acetate, sodium chloride, and sodium metabisulfite; or weighing the prescribed amounts of polyethylene glycol, acetic acid, sodium acetate, sodium chloride, sodium metabisulfite, and sodium sulfate, add them to the prescribed amount of injection water, and stir to dissolve to obtain an excipient mixture.

[0104] In some preferred embodiments, the injection is formulated into specifications of 2 ml:10 mg, 1 ml:5 mg, or 0.5 ml:2.5 mg.

[0105] In some specific embodiments, the administration of the injection is peritumoral injection, or injection at multiple points on the greater curvature and lesser curvature of the stomach.

[0106] In some preferred embodiments, the administration of the injection is at 4 - 6 injection points on the normal subcutaneous tissue of the greater curvature and lesser curvature of the stomach.

[0107] In another aspect of the present invention, there is provided a method for lymph node tracing in gastric resection - related diseases, which includes the following main steps;

[0108] S1: Prepare the pathological information of the experimental patients;

[0109] S2: Use mitoxantrone and / or its pharmaceutically acceptable salts to conduct a tracing experiment on eligible patients in the preparation of a lymph node tracer;

[0110] S3: Observe the indicators of the patient after using mitoxantrone and / or its pharmaceutically acceptable salts in the preparation of lymphatic tracers;

[0111] S4: Conduct statistical analysis on the obtained indicator data.

[0112] In the said method, the usage, dosage, composition, preparation method, etc. of the tracer are adapted to those in the foregoing use part.

[0113] By adopting the above technical solution; due to the lymphatic system tropism of mitoxantrone itself, its own color (showing blue) stains the lymph nodes near the tumor. When locally injected during gastric cancer tumor surgery, it stains the lymph nodes near the tumor, helping clinical lymph node localization and dissection. Through in-depth research on the preclinical pharmacodynamics of mitoxantrone hydrochloride, it is found that this product has a high affinity for lymph nodes when subcutaneously injected, can blue-stain lymph nodes, and can be used as a lymphatic tracer. Using it for lymph node tracing in gastric resection surgery can increase the number of detected lymph nodes, thereby reducing staging bias and improving the accuracy of gastric cancer staging.

[0114] By adopting the above technical solution, the lymph nodes and the first-station lymph nodes can be significantly blue-stained, clearly visible during the operation, the blue-staining rate of lymph nodes is improved, and thus the total number of detected lymph nodes is significantly increased. It has the characteristics of being convenient and feasible, fast staining, long staining time, and high lymph node blue-staining rate. Compared with the traditional nano-activated carbon tracing method, it can further increase the number of detected lymph nodes, thereby reducing staging bias and improving the accuracy of gastric cancer staging.

[0115] According to the above technical solution, the invention of this application can completely dissect the lymph nodes (especially the smallest lymph nodes) during the metastasis process of gastric cancer, can prevent the recurrence and secondary metastasis of gastric cancer, improve the quality of life of gastric cancer patients, and can delay the lifespan of patients. Detailed Embodiments

[0116] For the purpose of clear and concise description, features are described herein as part of the same or separate embodiments. However, it is to be understood that the scope of the present invention may include some embodiments having combinations of all or some of the described features.

[0117] Preparation of Injection Formula 1 of Mitoxantrone Hydrochloride in Example 1

[0118] Table 1: Prescription of Example 1

[0119]

[0120] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, and disodium edetate, add them to the prescribed amount of water for injection, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.5.

[0121] Preparation of Mitoxantrone Hydrochloride Injection Formula 2 in Example 2

[0122] Table 2: Prescription of Example 2

[0123]

[0124] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, sodium metabisulfite, and sodium sulfate, add them to the prescribed amount of water for injection, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.4.

[0125] Preparation of Mitoxantrone Hydrochloride Injection Formula 3 in Example 3

[0126] Table 3: Prescription of Example 3

[0127]

[0128] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, and disodium edetate, add them to the prescribed amount of water for injection, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.6.

[0129] Preparation of Mitoxantrone Hydrochloride Injection Formula 4 in Example 4

[0130] Table 4: Prescription of Example 4

[0131]

[0132] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, sodium metabisulfite, and sodium sulfate, add them to the prescribed amount of water for injection, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.7.

[0133] Preparation of Mitoxantrone Hydrochloride Injection Formula 5 in Example 5

[0134] Table 5: Prescription of Example 5

[0135]

[0136]

[0137] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, sodium metabisulfite and sodium sulfate, add them to the prescribed amount of injection water, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 min to dissolve, filter through 0.45 μm and 0.22 μm filter membranes for fine filtration, fill with nitrogen, seal the cap, sterilize at 121 °C for 15 min, and that's it. Measure the pH value to be 3.6.

[0138] Preparation of Mitoxantrone Hydrochloride Injection Formula 6 in Example 6

[0139] Table 6: Prescription of Example 6

[0140]

[0141] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate and disodium edetate, add them to the prescribed amount of injection water, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 min to dissolve, filter through 0.45 μm and 0.22 μm filter membranes for fine filtration, fill with nitrogen, seal the cap, sterilize at 121 °C for 15 min, and that's it. Measure the pH value to be 3.7.

[0142] Preparation of Mitoxantrone Hydrochloride Injection Formula 7 in Example 7

[0143] Table 7: Prescription of Example 7

[0144]

[0145] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate and sodium metabisulfite, add them to the prescribed amount of injection water, stir to dissolve. After dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 min to dissolve, filter through 0.45 μm and 0.22 μm filter membranes for fine filtration, fill with nitrogen, seal the cap, sterilize at 121 °C for 15 min, and that's it. Measure the pH value to be 3.9.

[0146] Preparation of Mitoxantrone Hydrochloride Injection Formula 8 in Example 8

[0147] Table 8: Prescription of Example 8

[0148]

[0149] Weigh the prescribed amounts of sodium chloride, acetic acid, sodium acetate, sodium metabisulfite and sodium sulfate, add them to the prescribed amount of water for injection, stir to dissolve, and after dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, seal the cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.5.

[0150] Preparation of Mitoxantrone Hydrochloride Injection Formula 9 in Example 9

[0151] Table 9: Prescription of Example 9

[0152]

[0153] Weigh the prescribed amounts of sodium chloride, polyethylene glycol 400, sodium chloride, acetic acid, sodium acetate, disodium edetate, add them to the prescribed amount of water for injection, stir to dissolve, and after dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, seal the cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.5.

[0154] Preparation of Mitoxantrone Hydrochloride Injection Formula 10 in Example 10

[0155] Table 10: Prescription of Example 10

[0156]

[0157]

[0158] Weigh the prescribed amounts of sodium chloride, polyethylene glycol 1000, sodium chloride, acetic acid, sodium acetate, disodium edetate, add them to the prescribed amount of water for injection, stir to dissolve, and after dissolution, add the prescribed amount of mitoxantrone hydrochloride, stir for 30 minutes to dissolve, filter through 0.45μm and 0.22μm membranes, fill with nitrogen, seal the cap, sterilize at 121°C for 15 minutes, and that's it. Measure the pH value to be 3.5.

[0159] Osmotic Pressure Investigation in Example 11

[0160] 11.1 Influence of Osmotic Pressure on Irritation at the Injection Site

[0161] Using mitoxantrone hydrochloride as a model drug, the osmotic pressure values of the injection solution (Example 4) were adjusted to 196, 285, 307, 600, 900, 1200, 1503, 2317, and 3011 mmol / kg respectively. The interstitial injection route was selected and the injection was administered subcutaneously to the soles of the feet of Kunming mice. At the preset time points, the mice were sacrificed by cervical dislocation, and the injection sites of the mice were excised for H&E staining to evaluate local irritation. When the osmotic pressure of the injection solution was 3011 mmol / kg, redness and swelling occurred, and the H&E staining results showed local irritation. No obvious local irritation was shown at other osmotic pressure values.

[0162] 11.2 Influence of Osmotic Pressure on Drug Targeting Ability

[0163] Using mitoxantrone hydrochloride as a model drug, the osmotic pressure values of the injection solution (Example 4) were adjusted to 196, 285, 307, 600, 900, 1200, 1503, 2317, and 3011 mmol / kg respectively. The interstitial injection route was selected and the injection was administered subcutaneously to the soles of the feet of Kunming mice. At the preset time points, the mice were sacrificed by cervical dislocation, and the first, second, and third-level lymph nodes were excised to observe their staining conditions. When the osmotic pressure value was less than 285 mmol / kg, only the first-level lymph nodes could be stained. When the osmotic pressure value was between 285 - 2317 mmol / kg, the third-level lymph nodes could all be stained. However, when the osmotic pressure values were 600, 900, and 1200 mmol / kg, the blue color of the third-level lymph nodes was dark blue, and when the osmotic pressure values were 285, 307, and 1503 mmol / kg, the color of the third-level lymph nodes was normal blue.

[0164] Osmotic Pressure Investigation in Example 12

[0165] 12.1 Influence of Osmotic Pressure on Irritation at the Injection Site

[0166] Using mitoxantrone hydrochloride as a model drug, two groups of mice, group A and group B, were taken. The osmotic pressure values of the injection solutions of Example 9 and Example 1 were adjusted to 196, 285, 307, 600, 900, 1200, 1503, 2317, and 3011 mmol / kg respectively. The interstitial injection route was selected and the injection was administered subcutaneously to the soles of the feet of Kunming mice in group A and group B respectively. At the preset time points, the mice were sacrificed by cervical dislocation, and the injection sites of the mice were excised for H&E staining to evaluate local irritation. When the osmotic pressure of the injection solution was 3011 mmol / kg, redness and swelling occurred in both cases, and the H&E staining results showed local irritation. No obvious local irritation was shown at other osmotic pressure values.

[0167] 12.2 Influence of Osmotic Pressure on Drug Targeting Ability

[0168] Using mitoxantrone hydrochloride as a model drug, two groups of mice, group C and group D, were taken. The osmotic pressure values of the injection solutions in Examples 9 and 10 were adjusted to 196, 285, 307, 600, 900, 1200, 1503, 2317, and 3011 mmol / kg respectively. The interstitial injection route of administration was selected, and the injection was given subcutaneously into the soles of the feet of Kunming mice in group C and group D. At the preset time points, the mice were sacrificed by cervical dislocation, and the first, second, and third-level lymph nodes were removed to observe their staining conditions. In the lymph nodes of the mice in group C (Example 9), when the osmotic pressure value was less than 285 mmol / kg, only the first-level lymph nodes could be stained. When the osmotic pressure value was between 285 and 2317 mmol / kg, the third-level lymph nodes could all be stained. However, when the osmotic pressure values were 600, 900, and 1200 mmol / kg, the color of the third-level lymph nodes was blackish blue, and when the osmotic pressure values were 285, 307, and 1503 mmol / kg, although the third-level lymph nodes could be stained, the color was dark blue. However, in the lymph nodes of the mice in group D (Example 10), when the osmotic pressure value was less than 285 mmol / kg, only the first-level lymph nodes could be stained. When the osmotic pressure value was between 285 and 2317 mmol / kg, the third-level lymph nodes could all be stained. However, when the osmotic pressure values were 600, 900, and 1200 mmol / kg, the color of the third-level lymph nodes was normal blue, and when the osmotic pressure values were 285, 307, and 1503 mmol / kg, although the third-level lymph nodes could be stained, the blue color was lighter. The above examples show that after injecting the above injection solution, mitoxantrone nanocrystals gradually precipitate from the solution. The nanocrystals can pass through the endothelial stroma, be phagocytosed by endothelial pinocytosis and enter the capillary lymphatics, and then accumulate in the local lymph nodes through lymphatic drainage. Mitoxantrone enters and temporarily stays in the lymph nodes, making them blue. And under the action of the optimized formulation components and osmotic pressure, its main active ingredient can penetrate the lymphatic vessels faster, with a stronger lymph node staining effect. Moreover, the size of the nanocrystals is larger than the capillary container. Therefore, they rarely enter the blood circulation and cause systemic toxicity.

[0169] Test Example 1 Pharmacokinetics and Pharmacodynamics Study of Mitoxantrone Hydrochloride Injection

[0170] In order to investigate the tracer safety and effectiveness of mitoxantrone hydrochloride injection for lymphatic tracing on the draining lymph nodes of cancer foci in gastric cancer patients, as well as the tolerance test and in vivo pharmacokinetic test of mitoxantrone hydrochloride injection for lymphatic tracing in gastric cancer subjects, and to determine the safe dose range, this example adopted a single-blind, single-center, parallel control, pharmacokinetic, and tolerance clinical trial design. According to the principle of dose escalation, the human tolerance, pharmacokinetics, and safety studies of mitoxantrone hydrochloride injection for lymphatic tracing in gastric cancer subjects were carried out group by group. After obtaining the safe dose range for use, the effectiveness study was carried out.

[0171] Specifically, 12 gastric cancer subjects were selected in this stage. Using the mitoxantrone hydrochloride injection (5 mg / ml) prepared in Example 4, 4 dose groups of 1.0 ml, 1.5 ml, 2.0 ml, and 3.0 ml were tested from low to high, with 3 cases in each group. Each subject only participated in one dose group test. Observe the safety of the test drug and explore the optimal dosage range of the test drug. This stage of the test consists of a screening period, intraoperative observation, and safety evaluation after medication.

[0172] Experimental group: used during surgery, mitoxantrone hydrochloride injection for tracer was directly injected into the greater and lesser curvatures of the stomach around the gastric tumor. The posterior wall and the stomach in the blind area of ​​the visual field were first freed, and the drug was injected after the tumor position was fully exposed. Specific method: After the tracer mitoxantrone hydrochloride injection was injected into the gastric serosa about 1.0 cm away from the edge of the tumor lesion, it was injected at multiple points, and the lymph nodes around the stomach were cleared after staining. The total doses for each subject were 1.0 ml, 1.5 ml, 2.0 ml, and 3.0 ml, respectively.

[0173] Blood sample collection: 4 ml of venous blood samples were collected before administration (within 60 minutes) and at 15±1min, 30±1min, 60±2min, 120±2min, 240±2min, and 360±2min after administration.

[0174] All subjects in this phase will be included in the safety and efficacy evaluation. Dose-limiting toxicity (DLT) will be observed until 14±2 days after administration. If no DLT occurs, the next dose group will be observed. Safety evaluation will be observed until 28±3 days after administration. Safety will be evaluated by comparing the results of our hospital examinations within one week before screening and postoperative laboratory test results, as well as by evaluating adverse events based on the "NCI CTCAE5.0 evaluation criteria".

[0175] Table 12: Pharmacokinetic data after injection of mitoxantrone hydrochloride injection

[0176]

[0177]

[0178] Table 13: Total number of lymph nodes detected after injection of mitoxantrone hydrochloride injection

[0179]

[0180] Table 14: Metastatic lymph node staining rate after injection of mitoxantrone hydrochloride injection

[0181]

[0182]

[0183] From the pharmacokinetic data in Table 12, it can be seen that after the tracer mitoxantrone hydrochloride injection is administered peritumorally to gastric cancer subjects, it is rapidly absorbed, the peak concentration and exposure are very low, and it is cleared quickly without causing toxic side effects.

[0184] After the subjects were injected with the test drug, the lymph node staining in the 1.0mL-2mL group generally showed a dose-dependent trend, and the total number of lymph nodes detected in the 2.0ml and 3.0ml dose groups was higher; and its components in the 3.0ml dose group could no longer be detected at 240min, indicating that it can be metabolized in the body. Therefore, the safe dose range of the test drug was set at 2.0-3.0ml.

[0185] Test Example 2 Application of Mitoxantrone Hydrochloride Injection in Lymphatic Tracing in Patients Undergoing Gastric Cancer Surgery

[0186] 1. Clinical trial design

[0187] The experiment adopts a single-center, positive, self-controlled experimental design, and intraoperative injection (injection of Example 4) is administered in groups for human tolerance test and pharmacokinetic test, and the effectiveness of the experimental drug is observed at the same time. This experiment intends to enroll 20 gastric cancer subjects, respectively, in the experimental group and the control group, and randomly assign them to each group, with 10 subjects in each group. Finally, 17 subjects completed the experiment, and the subjects had good compliance, including 9 subjects in the experimental group and 8 subjects in the control group.

[0188] The effectiveness and safety of the test drug were observed based on the dose range of 2.0-3 ml determined in Test Example 1. This phase of the trial consisted of a screening period, intraoperative observation, and postoperative safety evaluation.

[0189] Test group: used during surgery, mitoxantrone hydrochloride injection for tracer was directly injected into the greater and lesser curvatures of the stomach around the gastric tumor. The posterior wall and the stomach in the blind area of ​​the visual field were first freed, and the drug was injected after the tumor position was completely exposed. Specific method: The mitoxantrone hydrochloride injection for tracer was injected into the gastric serosa about 1.0 cm away from the edge of the tumor lesion, and then injected at multiple points. The lymph nodes around the stomach were cleared after staining. The total dose for each subject was 3.0 ml.

[0190] Control group: used during surgery. After the surgical field is exposed, 1 ml (50 mg) of nanocarbon suspension injection is taken and injected subserously at 4-6 points around the tumor using a skin test needle, 0.1-0.3 ml is injected at each point, and the injection is pushed slowly for about 3 minutes.

[0191] The safety evaluation of this clinical study was up to 28 days ± 3 days after the operation. Safety was evaluated by comparing the examination results in our hospital within one week before screening and enrollment with the postoperative laboratory examination results, and according to the "CTCAE 5.0 evaluation criteria of NCI" for adverse event evaluation. The safety and efficacy data obtained from this trial were processed using statistical software and statistical methods recognized by the NMPA, and the conclusions were reliable.

[0192] Table 15: Lymph node staining rate after injection of mitoxantrone hydrochloride injection

[0193]

[0194]

[0195] Note: Statistical method for data: Chi-square test

[0196] *: P < 0.05, there is a significant difference with statistical significance

[0197] Lymph node staining rate = (total number of stained lymph nodes / total number of detected lymph nodes) × 100

[0198] N1 lymph node staining rate = (total number of N1 stained lymph nodes / total number of N1 detected lymph nodes) × 100

[0199] N2 lymph node staining rate = (total number of N2 stained lymph nodes / total number of N2 detected lymph nodes) × 100

[0200] (1) Comparison of lymph node staining rate between the experimental group and the control group

[0201] According to the data in Table 15, it can be seen that: the total number of detected lymph nodes in the experimental group was 445, the total number of stained lymph nodes was 265, and the staining rate was 59.6%; the total number of detected lymph nodes in the control group was 484, the total number of stained lymph nodes was 209, and the staining rate was 43.2%. Using the chi-square test, the P value was less than 0.0001. Therefore, according to the test level of bilateral α = 0.05, it can be determined that the difference in lymph node staining rate between the experimental group and the control group has statistical significance.

[0202] The total number of N1 detected lymph nodes in the experimental group was 341, the total number of N1 stained lymph nodes was 207, and the N1 lymph node staining rate was 60.7%; the total number of N1 detected lymph nodes in the control group was 366, the total number of N1 stained lymph nodes was 175, and the N1 lymph node staining rate was 47.8%. Using the chi-square test, the P value was 0.0006. Therefore, according to the test level of bilateral α = 0.05, it can be determined that the difference in N1 lymph node staining rate between the experimental group and the control group has statistical significance.

[0203] The total number of lymph nodes detected in the experimental group N2 was 104, the total number of lymph nodes stained with N2 was 58, and the N2 lymph node staining rate was 55.8%; the total number of lymph nodes detected in the control group N2 was 118, the total number of lymph nodes stained with N2 was 34, and the N2 lymph node staining rate was 28.8%. Using the chi-square test, the P value was less than 0.0001. Therefore, according to the test level of bilateral α = 0.05, it can be determined that the difference in the N2 lymph node staining rate between the experimental group and the control group is statistically significant.

[0204] (2) Comparison of the detection rates of lymph nodes with a short diameter ≤ 1 mm between the experimental group and the control group

[0205] The number of cases of lymph nodes with a short diameter ≤ 1 mm detected in the experimental group was 7, and the detection rate was 77.8%; the number of cases of lymph nodes with a short diameter ≤ 1 mm detected in the control group was 2, and the detection rate was 25.0%. Using the chi-square test, the P value was 0.0295. Therefore, according to the test level of bilateral α = 0.05, it can be determined that the difference in the detection rates of lymph nodes with a short diameter ≤ 1 mm between the experimental group and the control group is statistically significant.

[0206] (3) Comparison of the detection rates of lymph nodes with a short diameter ≤ 2 mm between the experimental group and the control group

[0207] The number of cases of lymph nodes with a short diameter ≤ 2 mm detected in the experimental group was 8, and the detection rate was 88.9%; the number of cases of lymph nodes with a short diameter ≤ 2 mm detected in the control group was 6, and the detection rate was 75.0%. Using the chi-square test, the P value was 0.4534. Therefore, according to the test level of bilateral α = 0.05, it can be determined that the difference in the detection rates of lymph nodes with a short diameter ≤ 2 mm between the experimental group and the control group is not statistically significant.

[0208] 2. Results of the effectiveness of the clinical trial

[0209] It can be seen from the above experimental data that: the differences in the lymph node staining rate, N1 lymph node staining rate, N2 lymph node staining rate, and the detection rate of lymph nodes with a short diameter ≤ 1 mm between the experimental group and the control group are statistically significant, and the experimental group is superior to the control group.

[0210] Test Example 3: Tracer effect of mitoxantrone injection on tumors in different parts

[0211] Referring to the method of Test Example 1, for the enrolled patients, intraoperative tracer was performed with the injection doses listed in Table 16 below, and the lymph node staining situation was statistically analyzed.

[0212] Table 16: Tracer effect of tumors in different parts

[0213]

[0214]

[0215] Table 17 Tracer effects of mitoxantrone and carbon nanoparticles suspension on N1 / N2 stage tumors

[0216]

[0217] In summary, the tracer mitoxantrone hydrochloride injection is rapidly absorbed after administration by injection into various parts of the stomach and perigastric area. The peak concentration is basically reached at 0.5 h after injection, and the drug is rapidly eliminated after entering the blood. The blood drug concentrations after administration in different dose groups generally show a dose-dependent trend. The highest detected concentration and maximum exposure are 37.40 ng / ml in the 2 ml dose group and 34.47 h*ng / mL in the 3 ml dose group, respectively, which are far lower than the peak concentration and exposure of mitoxantrone administered by intravenous bolus injection reported in the literature. The highest detected concentration is only 7% of the Cmax (510 ± 206 ng / m) of mitoxantrone administered by intravenous bolus injection (10 - 12 mg / m2 / d) as a chemotherapeutic drug in the literature, indicating that the exposure of the tracer mitoxantrone hydrochloride injection is very low after peritumoral injection in gastric cancer and will not cause toxic side effects.

[0218] Among the subjects in the full analysis set of the clinical study, a total of 13 adverse event records occurred in 4 subjects in different dose groups, and no subject withdrew from the trial due to adverse events; 2 subjects had 2 cases of serious adverse events; severity: 10 cases of grade 1, 1 case of grade 2, and 2 cases of grade 3; relationship with the study drug: "possibly unrelated" in 3 cases and "definitely unrelated" in 10 cases; outcome of adverse events: remission in 3 cases and disappearance in 10 cases. The occurrence of DLT was observed in 12 subjects up to 14 ± 2 days after administration. None of the 12 subjects showed dose-limiting toxicity (DLT), that is, no grade 3 or above toxicity reactions in blood and skin occurred. It is suggested that the test drug has good safety.

[0219] The efficacy analysis of the subjects showed that the total number of lymph nodes detected by the test drug in the 2.0 ml dose group was 60.3 ± 14.57 (pieces), the total number of N1 lymph nodes detected was 57.3 ± 13.58, and the total number of N2 lymph nodes detected was 3.0 ± 1.00 (pieces). The total number of lymph nodes detected by the test drug in the 3.0 ml dose group was 60.3 ± 14.57 (pieces), the total number of N1 lymph nodes detected was 57.3 ± 13.58, and the total number of N2 lymph nodes detected was 3.0 ± 1.00 (pieces). The lymph node staining situation generally shows a dose-dependent trend, and the total number of lymph nodes detected in the 2.0 ml and 3.0 ml dose groups is relatively high; and the metabolism of the 3.0 ml dose group is complete at 240 min. Therefore, the safe dose range of the test drug is 2.0 - 3.0 ml.

[0220] In this clinical study, although there was no statistically significant difference in the total number of lymph nodes detected, the total number of lymph nodes detected in the first station (N1), and the total number of lymph nodes detected in the second station (N2) between the experimental group and the control group, as well as in the detection rate of lymph nodes with a short diameter ≤2 mm; there were statistically significant differences in the lymph node staining rate, N1 lymph node staining rate, N2 lymph node staining rate, metastatic lymph node staining rate, N1 metastatic lymph node staining rate, and the detection rate of lymph nodes with a short diameter ≤1 mm between the experimental group and the control group. This indicates that the total number of lymph nodes detected by the test drug is non-inferior to the control drug, but the lymph node staining rate and the detection rate of lymph nodes with a short diameter ≤1 mm of the test drug are significantly better than those of the control drug. In addition, the control drug, nanocarbon, cannot be metabolized in the body and there is a problem of permanent residue; while mitoxantrone has specific lymphatic affinity, fast staining, and long-lasting action without accumulation. Among the subjects in the safety set of the clinical study, a total of 1 subject had 1 adverse event record of "hypertension", with a severity of grade 3, and the relationship with the study drug was "definitely unrelated", and the outcome of the adverse event was "resolved", indicating that the clinical trial of the 2-ml dose group was safe and effective without adverse reactions related to the test drug.

[0221] The above research shows that mitoxantrone hydrochloride injection for tracer use is stable, efficient, safe and effective for lymphatic tracer in gastric cancer patients.

[0222] The foregoing description of specific exemplary embodiments of the invention has been presented for purposes of illustration and example. These descriptions are not intended to limit the invention to the precise forms disclosed, and it is apparent that many modifications and variations are possible in light of the above teaching. The purpose of selecting and describing the exemplary embodiments is to explain the specific principles of the invention and its practical application, so that those skilled in the art can implement and utilize various different exemplary embodiments of the invention, as well as various different selections and modifications. The scope of the invention is intended to be defined by the claims and their equivalents.

Claims

1. Use of mitoxantrone and / or its pharmaceutically acceptable salts in the preparation of a lymphatic tracer for lymph node tracing in diseases related to gastrectomy.

2. The use according to claim 1, wherein, The lymph node is a lymph node with a short diameter less than or equal to 2 mm, preferably a lymph node with a short diameter less than or equal to 1 mm.

3. The use according to claim 1, wherein, The diseases related to gastrectomy are gastric polyps or gastric tumors; Preferably, the gastric tumors include gastric benign tumors and gastric malignant tumors; More preferably, the gastric benign tumors and gastric malignant tumors are selected from gastric cardia cancer, gastric body cancer, and gastric antrum cancer.

4. The use according to claim 1, wherein, The gastrectomy is selected from total gastrectomy, partial gastrectomy, hemigastrectomy, or gastric antrum resection.

5. The use according to claim 1, wherein the diseases related to gastrectomy are gastric polyps or gastric tumors, and the lymphatic tracer is used for lymph node tracing in the gastric polyps or the gastric tumors.

6. The use according to any one of claims 1-5, wherein, The lymphatic tracer contains mitoxantrone and / or its pharmaceutically acceptable salts and pharmaceutical excipients; Preferably, the pharmaceutical excipients include, but are not limited to, one or more of osmotic pressure regulators, antioxidants, adsorbents, fillers, buffers, carriers, stabilizers, or preservatives; Preferably, the lymphatic tracer is an injection; more preferably, the dosage form of the injection is a solution, freeze-dried powder, emulsion, liposome, nanoparticle, nanocrystal, microcrystal, microsphere, or gel.

7. The use according to claim 6, wherein, The tracer contains mitoxantrone or its pharmaceutically acceptable salt in a mass-to-volume ratio of 0.05%-5%, and an osmotic pressure regulator of 0.1-10%; Preferably, the tracer also contains a buffer of 0.01-0.1%, an antioxidant of 0.01-0.1%, an adsorbent of 0.05-1%, and a filler of 0-20%; Preferably, the osmotic pressure regulator is a mixture of one or several substances selected from sodium chloride, glucose, sorbitol, mannitol, glycerol, phosphate, and citrate; Preferably, the buffer is one or several selected from acetic acid, sodium acetate, citric acid, and sodium citrate; Preferably, the antioxidant is one or several selected from sodium sulfite, sodium bisulfite, sodium metabisulfite, sodium thiosulfate, and disodium edetate; Preferably, the filler is one or several selected from monosaccharides such as glucose, fructose, galactose, ribose, or deoxyribose, or disaccharides such as sucrose, trehalose, maltose, lactose, or polysaccharides such as mannitol, sorbitol, lactitol, xylitol, maltitol, or erythritol; 8. The use according to claim 6, wherein, The tracer also includes polyethylene glycol, and the molecular weight of the polyethylene glycol is less than 2000, more preferably less than 1000, and further preferably 200, 400, or 600; and / or, The injection is formulated into a dosage form for use at a concentration of 2-10 mg / mL, and the single-use dose of the drug is 0.5-3 mL, preferably 2-3 mL; and / or, The osmotic pressure of the injection is 285 to 2317 mmol / kg, preferably 600-1200 mmol / kg.

9. The use according to any one of claims 1, wherein, The lymphatic tracer contains:

10. The use according to any one of claims 1-5, wherein, The injection is administered by peritumoral injection or at multiple points on the greater curvature and lesser curvature of the stomach; preferably, the injection is administered at multiple points on the greater curvature and lesser curvature of the stomach; and / or, The pharmaceutically acceptable salts of mitoxantrone are one or several selected from mitoxantrone hydrochloride, mitoxantrone oxalate, mitoxantrone sulfate, mitoxantrone phosphate, mitoxantrone acetate, and mitoxantrone citrate.

11. The use according to claim 1, wherein, The lymphatic tracer is an injection, which is prepared by a method including the following steps: Weigh the excipients in the prescribed amounts, add them to the prescribed amount of water for injection, stir to dissolve, and after dissolution, add the prescribed amount of mitoxantrone hydrochloride and / or its pharmaceutically acceptable salts.

12. A method for lymph node tracing in diseases related to gastrectomy, which comprises the following main steps; S1: Prepare the pathological information of the experimental patients; S2: Use mitoxantrone and / or its pharmaceutically acceptable salts to prepare a lymphatic tracer for performing a tracer experiment on eligible patients; S3: Observe the indicators of the patients after using mitoxantrone and / or its pharmaceutically acceptable salts to prepare a lymphatic tracer; S4: Perform statistical analysis on the obtained indicator data.

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