TW200824699A - Arsenic compounds for the treatment of the arsenic-sensitive blast-cell related diseases - Google Patents

Abstract

The present invention is related to arsenic compounds for the arsenic-sensitive blast-cell related diseases, such as hypersensitivity diseases, immunologic diseases, fibroblastic diseases, inflammatory diseases and infectious diseases, more particularly, the present invention is related to the treatment for malaria, Trypanosoma, bronchial asthma and systemic lupus erythematosus.

Description

200824699 IX. INSTRUCTIONS: TECHNICAL FIELD OF THE INVENTION The present invention relates to a method of treating blast cell disease using an arsenide such as arsenic trioxide. [Prior Art] The basic cause of asthma in recent years is still the main cause of chronic inflammation (Chronic inflammation). Due to the dual effects of genetic inheritance and the external environment, allergic inflammatory reactions occur, affecting inflammatory cells, including dendritic cells, epithelial cells, T cells, beta cells, obese cells, granulosa cells, and eosinophils. In the war, how to detect early, prevent early or treat asthma early is the goal of hard work. The cause of asthma may be caused by various stimuli (inducing) to induce tracheal inflammation (ΑΙ) caused by abnormal control of immune response of sputum cells, which induce or aggravate tracheal hyperresponsiveness and cause bronchoconstriction' mucosal congestion and edema, hypersecretion, micro blood Leakage and tracheal remodeling in the end, resulting in tracheal stenosis, airflow limitation, clinical symptoms. Systemic lupus erythematosus (SLE) is a progressive connective tissue disease characterized by the production of a variety of autoantibodies in the body that can cause damage to many of its own constituent components. The disease is a multi-system autoimmune disease, the main clinical manifestations are: arthritis, glomerulonephritis, epileptic seizures, anemia or thrombocytopenia. The disease often has an episode and an alternating phase of remission. In the past, it was considered that SLE was difficult to treat and the prognosis was very poor. Recently, due to the advancement of medical science, SLE has a deeper understanding. In the case of early diagnosis and timely treatment, the damage of vital organs is reduced, and the survival rate is improved. The prognosis of 丨E is greatly improved. Due to the development of immunopathology in the past 30 years, it has been confirmed that the humoral immunity and cellular immunity of SLE patients have abnormal changes in different degrees. However, the exact cause of SLE has not yet been fully elucidated. 5 200824699 may be related to some of the following factors: genetic elements, sex hormones, viral infections. Malaria is called hernia in ancient China. It is called malaria in foreign countries. It is a synonym for Mala mala (bad) and aria (air). The disease is caused by the female malaria mosquito biting the parasitic malaria parasite into the human body when biting the human body. Clinical manifestations include intermittent, timed, episodic chills, high fever and sweating, as well as anemia and splenomegaly. There are frequent recurrences between vivax malaria and three-day malaria, and irregular malignant sores are characterized by a malfunction of the central nervous system and a dangerous episode. There are four types of human malaria caused by four different malaria parasites, namely vivax malaria (benign tertian), the pathogen is Plasmodium vivax, and the third disease (quartan φ malaria, malariae malaria), pathogen It is Plasmodium malariae; ovale malariae, the pathogen is Ovary ovale; falciparum malaria (malignant tertian), the pathogen is Plasmodium falciparum (Ρ· Falciparum). There are about 20 species of trypanosomes, and only two species are pathogenic to humans. Trypanosomiasis is a disease caused by parasitic parasites, including American trypanosomiasis and African trypanosomiasis. The former is also known as chagas disease, which is also known as African sleeping sickness. The pathogen of African trypanosomiasis is Trypanosoma brucei, and its three subspecies are pathogenic to humans. T. b. rhodesiense causes Rhodesia trypanosomiasis (eastern sleep disease); Trypanosoma chinensis (T·b, gambiense) caused by Gambia trypanosomiasis (Western non-sleep disease); cases caused by T. b. brucei, although clinically reported, but less It mainly causes the onset of cattle. African trypanosomiasis, also known as sleeping sickness, is one of the serious diseases of African zoonosis. The disease is caused by a parasitic protozoan that is associated with the flagella of the T. brucei Complex, which is the vector of transmission. Early manifestations of the disease are irregular fever, lymphadenitis, etc., and later manifested as damage to the central nervous system, severe headache, unresponsiveness, lethargy until drowsiness, and eventually death. 200824699 Traditional Chinese medicine is naturally natural, and many natural medicines are used in humans, which can cause side effects such as _ sex. Traditional Chinese medicine processing technology has long been one of the main methods to reduce the side effects of traditional Chinese medicine. The arsenic is a representative of the toxic drugs. Although the traditional Chinese medicine processing technology can reduce its toxicity to a certain extent, it has been a bad impression for a long time, and it has become a fascinating change, thus limiting the development of arsenic. There are many kinds of chemical components contained in arsenic, but its main component is arsenic trioxide. In the current known literature, the application of arsenic is mainly based on its main component, arsenic trioxide. Since Chinese herbal medicines have been highly valued by countries all over the world, the understanding and application of arsenic have increased, so the use of arsenic trioxide has also been paid attention to. For example, the use of arsenic trioxide for the treatment of cancers such as cervical cancer, skin cancer, and esophageal cancer. Or skin diseases such as corns. However, other applications of arsenic trioxide have yet to be further developed and studied by pharmaceutical researchers. SUMMARY OF THE INVENTION The present invention relates to a method of treating blast cell disease using an arsenide such as arsenic trioxide. The present invention relates to the use of arsenic trioxide for the preparation of a medicament for the treatment of allergic diseases, immune diseases, fibroblast diseases, inflammatory diseases and infectious diseases, particularly malaria, African trypanosomiasis, asthma and systemic lupus erythematosus. DETAILED DESCRIPTION OF THE INVENTION The technical solution employed by the present invention to address its technical aspects is as follows: The present invention relates to the use of arsenic trioxide for the preparation of a medicament for the treatment of arsenic-sensitive blast cell diseases. Preferably, the drug is used by injection. Preferably, the arsenic-sensitive blast cell disease is selected from the group consisting of an allergic disease, an immunological disease, a fibroblast disease, an inflammatory disease, an infectious disease, and a caterpillar disease. 7 200824699 More preferably, the immune disease is selected from the following: connective tissue disease, autoimmune thyroid disease, neuromuscular autoimmune disease, gastrointestinal autoimmune disease, autoimmune hepatitis, primary hepatobiliary sclerosis, Primary sclerosing cholangitis, autoimmune carditis and arteritis. Most preferably, the immune disease is selected from the group consisting of systemic lupus erythematosus, rheumatoid arthritis, sclerosis, Gracies disease, myasthenia gravis, multiple sclerosis, ulcerative colitis, and Crohn's disease. Preferably, the allergic disease is selected from the group consisting of asthma, hypersensitivity pneumonitis, diffuse interstitial fibrosis, allergic rhinitis, eosinophilic rhinitis, spring conjunctivitis, and succulent measles. Preferably, the inflammatory disease is selected from the group consisting of pneumonia, osteomyelitis, leprosy, syphilis, tuberculosis, hepatitis, tumor formation, and chronic obstructive pulmonary disease. Preferably, the fibroblastic disease is selected from the group consisting of liver fibrosis, pulmonary fibrosis, skin fibrosis, systemic fibrosis, pneumoconiosis, tuberculosis, atypical pneumonia (S AR S), and adult respiratory distress syndrome. Chronic obstructive pulmonary disease, scarring, swelling, dryness, invasive cystic fibrosis and neurofibromatosis. • Preferably, the infectious disease is a parasitic disease. The younger one is that the parasitic disease is malaria. Preferably, the arsenic-sensitive blast cells are selected from the group consisting of white blood cells, peripheral blood mononuclear cells, and fibroblasts. Preferably, the drug comprises arsenic trioxide from about 1 μM to about 20 μM. More specifically, the drug comprises arsenic trioxide from about 1 μΜ to about 15 μΜ. Most preferably, the drug comprises from about 0.1 μΜ to about 10 μΜ of arsenic trioxide. 200824699 The proper term "arsenic-sensitive blast cells" as used herein means mononuclear spheroid cells from white blood cells and peripheral blood. The term "mammal" as used in vomiting means mammals containing humans. The present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Example 1: Animal test for treating bronchial asthma with arsenic trioxide 1.1 Materials Test animals: commonly used Balb/c female rats, 6-β weeks old, Feed for six weeks, 15 in each group. BALF/c female rats of SPF six weeks old were purchased from National Taiwan University Animal Center, and each mouse was raised separately. • Room temperature was controlled at 22±2t: and light and dark cycle time was 12 hours each (6 in the morning, 6 in the afternoon) Point dark), free access to drinking water and AIN_76 formula powder feed, weekly food intake and body weight twice, "week blood collection once every other week; mice eight weeks old are randomly divided into four groups, sensitization steps and treatment The method is as follows. Drug: arsenic trioxide injection (Asadin®, 10mg/10ml/vial), diluted with PBS solution Reagent: Balanced salt solution (HBSS) lavage solution, Trypan blue, Hematoxylin-eosin, carbonate coating buffer, 1X PBS buffer containing 0.05% Tween 20, blocking solution (1X PBS buffer containing 1% BSA-0.05% Tween 20) Liquid), [3H] TdR (lf confirmed) Other: oval albumin (ovalbumin), wax film (paraffin), eotaxin EUSA kit (old L, Hamburg, Germany), anti-mouse IgE antibody. Main instrument: nebulizer (DeVilbiss, Somerset, PA), centrifuge, hemocytometer, microscope, eotaxln ELISA reader, microplate reader, liquid 200824699 scintillation counter, Plethymography. '1·2 test method 1·2·1 establishes respiratory tract inflammation The animal model was selected from the commonly used BALB/C female rats, 6-8 weeks old. The mouse sensitization step was first intraperitoneally injected into the mice with ovalbumin, and the same dose of ovalbumin was obtained on the second, 14, 21, and 28 days, respectively. The mice were repeatedly injected. Six to eight mice were simultaneously placed in a closed container, and the OVA antigen was made into an inhalable particle type by a nebulizer, and the mice were inhaled in the closed container for more than 20 minutes. The device will send 0.5-5m (7) of antigen particle molecules at a rate of ml3ml/min. 1_2·2 Determination of ovalbumin (ovab_in, _specific antibody EUSA measurement " 〇.5mg antigen dissolved in 100ml carbonate coverage buffer After that, the microplate was added to the mixture overnight. On the second day, the microplate was washed with 1×PBS buffer containing a〇5% Tween 20. Then the microplate was blocked with blocking solution and washed three times. Each well was added with 100 μΙ of the sample to be tested - diluted with serum-like blocking solution), and washed for β times after the reaction. Add 100 m 丨 anti-aging mouse 丨 g [antibody for reaction and wash 6 times. The above reactions are all at 37. (: For 45 minutes, transfer to _ to room temperature for 15 minutes. Finally, add 100 μ丨 ABTS to each well (2.2, -Azino-bis-3-

Ethylbenzthiazoline each Sulfonic Acid) was subjected to a coloring solution at room temperature for about 30 minutes, and the reaction was terminated with 100 μΙ of 5% SDS per micropore and the absorbance was read with a plate reader. 1·2·3 Calculation of the number of bronchoalveolar fluids and giant sun cells, lymphocytes, neutrophils and eosinophils White blood cells 24 hours after the last inhalation of antigen, all the mouse eye sockets were sacrificed after blood collection. Bronchial intubation was then performed immediately, and the lungs of the mice were lavaged four times with 1 ml of HBSS containing no calcium and magnesium ions, and the rinse was collected. The collected alveolar lavage fluid was centrifuged at 400 x g for 1 Torr at 4 °C. The cells obtained by centrifugation were suspended in 1 ml of HBSS, and then the total number of cells was counted as 200824699 by the blood cell counting agent. The cells were then fixed on a glass slide by Cytocentrifuged preparations, and the number of cells was counted by Liu's stain. The number of cells in each cell is calculated as the number of squamous cells, lymphocytes, neutrophils, and eosinophils in 300 cells, and is classified according to their standard type. 1 ·2·4 Determination of eotaxin content in bronchoalveolar lavage fluid The content of eotax丨η in bronchoalveolar lavage fluid (BAL) was determined by eotaxin ELISA kit, and the procedure was as directed by the ELISA kit manufacturer. 1.2.5 Respiratory resistance test in mice φ The mice were stimulated by increasing the concentration of methacholine (Mch.) (6·25 mg/m Bu 12·5 mg/m b 25 mg/mh 50 mg/ml, respectively). The plethymography method was used to determine the airway enhanced pause (PenH) values of the mice to see if the airway contraction of the mice was improved. 1.3 Experimental group The mice were randomly divided into four groups at the age of eight weeks, and each group of 15 patients received treatment in four groups as described later. Group A mice were sensitized and treated with only PBS. Group B mice were sensitized with ovalbumin but treated with PBS only. Group C mice were sensitized with ovalbumin and treated with arsenic trioxide at a dose of 2.5 mg/kg. Group D mice were sensitized with ovalbumin and treated with arsenic trioxide at a dose of 5.0 mg/kg. Table 1-1 sensitizing antigen treatment Α group PBS PBS group B OVA PBS c group 1 OVA ΑΤΟ 2.5 mg / kg D group OVA ΑΤΟ 5 mg / kg π 200824699 1 · 4 test steps 8 weeks old BALB / c female rats The method of feeding and grouping the mice is as described above. Methods and Routes for Sensitization in Old Rats As described above, the titer of antigen-specific antibodies was determined weekly by eye socket blood sampling after sensitization in mice. 1.5 treatment method was first intraperitoneal injection of ovalbumin (OVA) in mice, respectively, on the third, 14, 21 and 28 days, the same dose of ovalbumin repeated injection of mice to complete the sensitization step, the 38th day with PBS And the dose of 2.5mg / kg and 5.0mg / kg of arsenic trioxide in the peritoneal cavity continuous injection φ shot seven days of treatment 4 times. Mice were stimulated with inhaled ovalbumin antigen on days 42, 43 and 44. The airway resistance test of the mice was carried out on the 45th day. The results of the airway resistance test are shown in Table 1-2. Table 1.2: Call surface resistance (tracheal reaction) of the test. Airway resistance test Mch (mg/ml) 6.25 mg/ml 12.5 mg/ml 25 rng/rnl 50 mg/ml Group A 2.20 ± 0.28# 8.22 ± 1.22 11.99 ± 1.50 14_21 ± 1,06 Group B 2.50 ±0.40 8.00 ±0.93 14.52 ±1_42 19·15±3·35 Group C 3.62±0·47 7·83± 1.13 10.15 ±0·841 9·83±1·171 D Group 2.55 ± 0.35 8.82 ± 0.88 11.57 ± 1.52 12.51 ± 1.90

#数値 is the average ± SD Μ 1 Compared with the Β group, ρ < 0·05. After increasing the concentration of methacholine (Mch·) (6·25 mg/m b 12.5 mg/m b 25 mg/mh 50 mg/ml) in mice shown in Table 1. 2, the respiratory resistance of the mice was determined by plethymography. The tracheal allergic reaction-PenH is indicated by AHR; airway 200824699 hyper-reactivity-PenH (airway enhanced pause). Compared with group B mice not treated with arsenic trioxide, treatment with 2.5 mg/kg arsenic trioxide successfully blocked tracheal hyperimmune response in group C mice and reduced respiratory resistance in mice. 48 hours after the last inhalation of ovalbumin antigen stimulation All the mouse eye sockets were sacrificed after blood collection. Immediately after the bronchial intubation, the lungs of the rats were lavaged four times with 1 ml of HBSS containing no strontium ions, and the alveolar lavage fluid was collected. The collected alveolar lavage fluid was subjected to histopathological investigation, the number of various cells (macrophages, lymphocytes, neutrophils and eosinophils) was measured, and the content of eotaxin in the bronchoalveolar lavage fluid was determined. As shown in Table 1 · 3 and Table 1 · 4. • Table 1.3 Bronchoalveolar lavage fluid macrophage, lymphocytes, neutrophils and eosinophilic white blood cell count BALF mouse white blood cell count (χ1000 cells/ml) Macrophage eosinophilic white blood cell neutrophil lymph Ball * Group A 135.6 ± 54.5 # 3 · 2 ± 2 · 1 8.4 ± 4.9 12.9 ± 8.2 Group B 243.8 ± 69.3 105.3 ± 60 · 7 77.1 ± 32.8 Ί 37.8 ± 16.3 Group C 621.0 ± 199.6 28.6 ± 13.0 200.7 ± 75.2 37.0 ±9.4 D group 499.1±131.6 33.2±12.5 244.0±72.6 28.7±13.6 * The number of basophilic white blood cells is sparse and is not indicated here. Table 1.4 shows the average concentration and standard deviation of eotaxin content in mouse bronchoalveolar lavage fluid. Compared with group B sputum that was not treated with arsenic trioxide, mice in groups C and D received arsenic trioxide treatment, which reduced the amount of eotaxin in bronchoalveolar lavage fluid, especially in group C mice treated with 2.5 mg/kg arsenic trioxide. There was a significant difference in the arsenic trioxide-treated pot mice}3 200824699 (Ρ <〇·〇5).

Table 1.4 Determination of eotaxin in bronchoalveolar lavage fluid

Eotaxin (pg/ml) Average SD A group 14.42 8.39 Group B 20.90 4.97 Group C 12.38* 3.92 Group D 16.00 4.06 *Compared with Group B, ρ<0·05 was taken after /J, mouse received egg albumin antigen stimulation Rat blood and serum levels of ovalbumin-specific IgE values were measured as evidence of sensitization. Table 1.5 shows that the serum albumin-specific antibody content in the B, C, and D groups was significantly higher than that in the control group A mice, indicating that the mice in groups B, C, and D were successfully sensitized by ovalbumin antigen stimulation. Table 1.5 Ovalbumin 朿 [I-Amazing Index (OVAstimulatiorHndex) 0. Specific IgE Average SD Antibody (S1) Group A 0.30 0.09 Group B 2.06 0.81 Group C 2.20 0.70 Group D 1.96 0.31 Example 2: Treatment of systemic lupus erythematosus with arsenic trioxide Test

u 200824699 2·1 test material 2·1_1 test animal ISIZBxNZW F1 female mouse, about 7 months old; NZB (H-2d) female mouse; NZW (H-2Z) male mouse 2. 1.2 drug arsenic trioxide injection (Asadin®, 10 mg/10 ml/vial), diluted in PBS. 2·1·3 reagents (1) methylated bovine Serum AIbumin working solution) (2) calf thymus DNA working solution (3) blocking Liquid (Geletin-PBS solution) (4) Serum and positive control group: Serum was obtained by centrifugation (5000 rpm, 10 minutes, 4) of blood after blood collection. Serum was diluted with blocking solution at the time of use. The positive control group was anti-DNA IgG or IgM secreted by the fusionoma cells, and the antibody was taken from each culture plate and diluted to a 250-fold 500 μM solution, followed by serial dilution. (5) Level 2 antibody (2Ab; Horserad丨sh peroxldase (HRP)-conjugated anti-mice mouse Y chain or μ chain antibody; Horserapish peroxidase (HRP)-conjugated anti-mouse γ chain or μ chain Ab): (6) 2 , 2,-azino-bisC3-ethylbenzthiazoline-6-suIphonic acid (ABTS) solution (7) 5X citrate buffer (8) SDS solution 2_1 _4 main instrument: ELISA reader for measuring hungry body 200824699 2·2 test method 2·2 ·1 Establishing an animal model of human systemic erythema field. About 7 months old MZBxNZW F1 female mice and 8 焖^ breeding mice naturally develop an autoimmune syndrome similar to human systemic IBS surface (please confirm ). The autoimmune syndrome of NZB/NZW-matched mice produces antinuclear antibodies (丨gG) at four months of age, and glomerulonephritis and proteinuria occur at nine months of age, kidney failure Inducing uremia is the leading cause of death. _ 2.2.2 Determination of anti-dsDNA and anti-ssDNA antibodies ELISA S!

The anti-DNA-antibody content of anti-dsDNA and ssDNA in serum was determined by standard ELISA method. The steps were as follows: (1) Covering with mBSA: covering 100 μM of mBSA per microwell (10 Mg/mL) ), allowed to stand overnight at 4 ° C, and then washed twice with PBS. (2) dsDNA and ssDNA coverage: 100 μM of dsDNA and ssDNA (7.5 ijg/mL, respectively) were placed in each well, and allowed to stand overnight at 4 ° C, followed by washing twice with PBS. • (3 blocking blockage: cover 2 μl of blocking solution per microwell, and stand overnight in 4th generation (or room temperature for 2 hours or 37 ° C for 45 minutes) 15 minutes), wash three times with PBS-tween 20. (4) Add serum or positive control group or negative control group: Add 1 μμΙ serum or positive control group or negative control group to each well, at 37. After 45 minutes, room temperature for 15 minutes (or room temperature for 2 hours), washed six times with PBS-tween 20. (5) Add the above-mentioned grade 2 antibody: add 100 μΙ of secondary antibody per microwell, and let stand at 37 ° C /6 200824699 After 45 minutes at room temperature for 15 minutes, wash with PBS-tween 20 six times. (9) Add coloring agent: Add 10 μμ of coloring agent (ABTS solution) to each micropore, avoiding light reaction for 5-10 minutes. The stop solution (5% SDS solution) was added and the 0.D measurement was performed under the absorbance 405. (7) The amount of anti-DNA IgG is expressed in units of ELISA assay (Eli/ml) compared with mAb 10F10, 1 〇 “1〇 antibody produced at 74叩 plus concentration is defined as 1 EU/ml anti-DNA丨gG. Anti-ribosomal IgG concentration is measured in EU according to 10F10 antibody concentration. 10F10 from 231ng/ml The absorption enthalpy produced by the body is defined as 1 φ EU/ml anti-ribosomal 丨gG. 2.2.3 Detection of glomerulonephritis in mice. Evaluation of glomerulonephritis in mice, urine test with urine Tablet (Multistix®, Bayer) colorimetric Measure urine protein content. Colorimetric determination of urine protein content greater than 1g / L (2+) is defined as severe glomerulonephritis. When mice are 37th week old and 40th week old 2.2.4 Survival rate of mice The evaluation of mouse survival rate of each group of 6 mice was recorded to the 43th week. _ 2·3 test steps 2.3.1 test group mice were randomly divided into three at 24 weeks of age. Group, 6 rats in each group: (1) sputum group: treated with PBS only (1 〇 ml/kg); (2) group B: treated with arsenic trioxide at a dose of 2.5 mg/kg; (3) group C: dose Treatment with 5.0 mg/kg of arsenic trioxide. 2·3·2 test procedure mice were randomly divided into three groups at the age of 24 weeks, each group of 6 patients, each method of treatment / 7 200824699 one, three, five Each was intraperitoneally injected once for 1 month. Before treatment and after treatment was stopped, anti-SSDNA and anti-dsDMA antibodies were surface-measured every four weeks for 3 months. 2,3.3 EUSA was used to determine the content of anti-dsDNA and anti-ssDNA antibodies in mouse serum: the anti-dsDNA measurement results are shown in Table Z1. Table 2.1 丨gG anti-dsDNA titer (ELISA unit) Test time (week) test The first 4 8 12 PBS (Group A) 0-280 0.623 0.679 0.381 2.5mg/kg ΑΤΟ (Β group) 0.262 0.572 0.464 0.442 5mg/kg ΑΤΟ (Group C) 0.362 0.285 0.357 0.702 Test results show that B, treated with arsenic trioxide Group C mice had lower anti-DNA antibody levels in the serum of Group A mice that were not treated with arsenic and arsenic; and higher doses of arsenic trioxide were more potent. Because the anti-DNA antibody content is strongly correlated with SLE disease activity, the results of this trial show that arsenic trioxide may become a new therapeutic agent for SLE. Glomerulonephritis test results in mice in 2·3·4 The evaluation of glomerulonephritis in mice was performed by urine colorimetry using urine test. Colorimetric determination of urinary protein content greater than 1 g/L (2+) is defined as severe glomerulonephritis. Table 2. 2 The proportion of mice with severe glomerulonephritis 18 200824699 Time (week) 37 40 PBS (Group A) 25 too 2.5 mg/kg ATO (Group B) 16.67 40 5 mg/kg ΑΤΌ (C _ 25 25 Table 2.2 The trial data showed that treatment with arsenic trioxide significantly delayed the onset of severe kidney/J nephritis in mice in the β and C groups. Some mice that did not have severe renal nephritis did not even have normal renal function. The phenomenon of proteinuria. In group A mice that were not treated with arsenic trioxide, all of them developed severe glomerulonephritis. Compared with group A mice that were not treated with arsenic trioxide, arsenic trioxide treatment significantly reduced mice in group B and C. Glomerulone-nephritis occurrence and progression. 2.3_5 Mouse survival rate evaluation Table Z3 mouse survival rate evaluation (%) Time (4) 22 31 38 39 40 I 41 42 43 PBS (Group A) 100 100 66.7 50 50 16J 0 0 2.5mg/k gATO (B-touch 100 100 100 83.3 66.7 66.7 66.7 50 5mg/kg ΑΤΟ (Group C) 100 100 66.7 66.7 66.7 66.7 66.7 66.7 Table 2.6 shows groups B and C treated with arsenic trioxide, 43-week survival rate is 50 %, the caries in the sputum group that did not receive arsenic trioxide died. Compared with the group A mice that did not receive the oxidized/9 200824699 arsenic treatment, the arsenic trioxide treatment significantly prolonged the survival of the mice in group B. Example 3: In vitro test of Plasmodium falciparum poisoning by arsenic trioxide 3.1 Test material 3-11 Drug: Arsenic trioxide injection (Asadin®, 10mg/10ml/vial) 〇trioxide arsenide, molecular weight 197.64, test drug dosage form concentration 1mg/ Ml (5054·6μΜ), dilute it to 160_ before the test, and then semi-dilution, the final concentration is calculated as ng/ml 3.1 _2 Protozoa Plasmodium falciparum FCC1/_ cited from Hainan, laboratory continuous culture or cold storage 5 years to -3. 3.1.3 • Culture medium: The medium (liquid) used for continuous culture of P. falciparum in vitro, the main components are RPMM640 (GBCO), HEPES (GBCO), NaHC03 and AB serum, etc. 3- 1.4 Other materials Serum, red blood cells: supplied by Guangzhou Blood Center. Red blood cells (RBC) are ACD anticoagulated type B whole blood, stored at 4 ° C, no more than one month, washed 2-3 times with serum-free medium before use. 2 test methods Continuously culture FCC1/HN. Refer to Tube Weibin [73], Chen Lin [74] and other methods for micro-measurement of anti-malarial drug in vitro, add 20 μΙ per well to the culture plate, and add about 1% infection rate. Insect blood 180μΙ, capped, $ stem lightly shake for a few seconds, at 37 ° C, 5% CO 2 , cultured for 48 hours, go to the supernatant, take RBC coated with a thin film of blood, Gyr stain and microscopic examination, calculate the protozoa The infection rate was determined as 100 in the control group, and the worm reduction rate of each dose group was determined [worm reduction rate = (control group infection rate - test group infection rate)

2Q 200824699 / control group infection rate x 100%], with the logarithm of the dose as the abscissa, the probability unit of the worm reduction rate as the ordinate as the regression equation, calculate the drug that reduces the proliferation of protozoa by 50% and 90% compared with the control group. the amount. 3. 3 Test Procedures and Results (1) 3_3_1 Drugs were prepared and sterilized with 8 penicillin bottles. Add RPMM640 complete medium and arsenic trioxide (ΑΤΟ) according to Table 3-1. Table 3.1 Concentration of arsenic trioxide in culture solution Ι (μΙ) 0 0.5 10 15 2.0 2.5 3.0 0 RPMI-1 640 medium (ml) 1 1 1 1 1 1 1 1 Concentration (uM) 0 2.5 5 7.5 10 12.5 15 0 3.3.2 Wet blood dilution Take continuous culture of FCC1/HN, smear, stain The infection rate was calculated, and the infection rate was adjusted to about 1% by sputum type RBC. 3·3·3 test According to the method of Chen Lin et al., the liquid medicine 2_ was added to each micropore on the 3599 culture plate, and then the blood containing 180 μΙ was added, and the test was performed for 4 lines. Then cover and shake for a few seconds, set 37. (: 5% in 〇2 boxes for 48 hours, and the number of RBCs infected with Plasmodium in 5000 RBCs was calculated by thin-staining. The protozoal infection rate was 0.98% when the results were started. Table 3.2 For the number of protozoa in the 200824699 ball per 5000 red blood in the table, the number of protozoa per 5000 red blood cells after cultivation 3.2 (control) BCDEFG sun exposure) 剂量 dose (βΜ) 0 2.5 5 7.5 10 12.5 15 0 1 172 82 79 81 60 53 39 199 2 306 163 48 33 47 49 33 294 3 251 125 36 29 46 32 17 213 4 198 74 57 55 53 48 26 208 As can be seen from the above table, the highest dose (15 μΜ) <3# not Can completely inhibit the development of Plasmodium. 3. 4 Test Procedures and Results (2) 3.4.1 Preparation of Drugs One sterile penicillin bottle is added, and RPiyn-1640 complete medium and arsenic trioxide (ΑΤΟ) are added separately according to Table 3. 3 below. Table 3.3 Concentration of arsenic trioxide in culture solution AJO (10) 0 1 2 3 4 5 6 7 8 RPM1- 1640 (ml) 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 0.5 Concentration 0 10 20 30 40 50 60 70 80 (μΜ) 3.4. 2 insects_dilution

Continuously cultured FCC1/HN, smear, stain, calculate the infection rate, and adjust the infection rate to about 1% with sputum type RBC. 22 200824699 3.4.3 mm Refer to the method of Weibin, Chen Lin, etc., on the 40-well culture plate, add 20 μM of the solution to each micropore, and then add 180 μ of the blood containing bacteria to do 4 lines. After shaking for a few seconds, set 37t:, 5% C02 box for 48 hours, remove the supernatant, take RBC smears, stain and microscopic examination, calculate the number of red blood cells infected with Plasmodium every 5000 RBC. 3.4_4 The protozoal infection rate was 1.6% when the culture started, and the number of protozoa in every 5000 red blood cells φ after culture in Table 3.4. Table 3.4 The number of protozoa per 5000 red blood cells after culture was observed) 2 3 4 5 6 7 8 9 /VTO III | ΜβΜ 0 10 20 30 40 50 60 70 80 A 227 96 49 46 45 45 50 40 25 B ' 209 50 68 63 64 60 70 56 19 C 188 70 79 90 61 87 67 "57 |23 D 229 71 77 77 67 74 69 66 30 As can be seen from the above table, the density of 80 μΜ# protozoa has decreased, but its development has not been completely inhibited. 3.5 Test procedure and results (3) 3.5.1 Drug preparation The stock solution was 20 μM in complete medium and half-diluted in a penicillin bottle for a total of 11 dose groups: 2522, 1216, 633, 316, 158, 79, 40, 20, 10, 5, 2.5, ΟμΜ. Insect blood dilution ίο 200824699 Take continuous culture of FCC1/HN, smear and stain, calculate the infection rate, and adjust the infection rate to about 1% with B-type RBC. 3_5·3 test: Refer to the methods of Guan Weibin, Chen Lin, etc. Add 20 μΙ of the drug solution to each micropore on the 3599 culture plate, and add 180 μM containing the blood of the insects to make 4 rows. Then cover and shake for a few seconds and set 37. : and 5% G02 box culture for 48 hours, remove the supernatant, take RBC smear, stain and microscopic examination, calculate the number of RBC infected with Plasmodium in 5000 RBC. • 3. 5 · 4 results when starting culture Protozoa infection rate was 1.8%. Table 3.5 shows the number of protozoa per 5000 red blood cells after culture. Table 3.5 Number of protozoa per 5000 red blood cells after culture 1 2 3 4 5 6 ΑΤΟ Measured / / 2522 1216 633 316 158 79 A 0 0 0 0 0 38 B 0 0 0 0 0 28 C 0 0 0 0 0 33 D 0 0 0 0 0 25 Four lines of mixing 0 0 0 0 0 32 Performance table) 7 8 9 10 11 12 ΑΤΟMeasuring Μ μ 40 20 10 5 2.5 〇Control A 63 75 88 108 206 503 24 200824699 Β 63 75 88 108 206 503 C 52 69 81 160 236 450 D 43 60 77 108 186 392 Four lines mixed 44 65 92 116 200 473 Even on the table, 158uM can completely inhibit FCC1 development. 3·5·5 Results Statistics: The results of the inhibition of FCC1 by arsenic trioxide (ΑΤΟ) are shown in Table 3.6 Table 3.6 Arsenic Trioxide (ΑΤΟ) on FCC1 inhibition results Statistical Table Dose-dose log-infection rate worm-reducing rate probability (uM) (X (%) (%) (Y) 0.25 0.6021 42.3 57.7 5.1942 0.5 -0.3010 24.5 75.5 5.6903 1.0 0 19.5 80.5 5.8596 20 0.3010 13.7 86.3 6.0939 4.0 0.6021 9.3 90.7 6.3225 8.0 0.9031 6.8 93.2 6.4909 3·6 Test procedure and results (4) 3.6 .1 Preparation of the drug: Take the stock solution 100μΙ, add the complete medium 3059μΙ, mix, this is the highest concentration liquid (160μΜ), take the highest concentration of the solution 200μΙ, and take 200μΙ medium in the penicillin bottle to mix, half-dilution, A series of solutions are available: I, 160, 80, 40, 20, 10, 5, 2.5, 1.25, 0.625, 0.3125, ΟμΜ. (Total 4 rows, 11 columns for the highest dose hole 160UM). 3.6.2 Dilution of blood:

Continuous culture of FCC1/HN, smear, staining, calculation of infection rate, sputum type RBC 25 200824699 regulation of infection rate of about 13⁄4 〇3·6·3 test: according to the method of tube Weibin, Chen Lin, etc., cultured in 3599 Add the drug solution 20|jh to each micropore and add 180 μM containing insect blood to make 4 rows. Then cover, shake the side for a few seconds, set 37t: incubated in 5% C02 box for 48 hours, remove the supernatant, take RBC smear and stain, microscopic examination, calculate the number of RBC infected with Plasmodium per 5000 RBC (Only nuclear cells without cytoplasmic dead insects are not counted). 3_6.4 Results: The protozoal infection rate was 1.5% at the start of culture, and the number of protozoa per 5000 red blood cells after culture in Table 37.

Table 3.7 Number of protozoa per 5000 red blood cells after culture 1 2 3 4 5 6 Tanning agent|--t Control 0.3125 0.625 1.25 2.5 5 .*—*-L· 里Μμ Ε 394 384 371 328 279 71 F 410 407 401 361 302 72 G 457 414 364 276 240 67 H 454 385 339 308 235 98 Mix 4 448 358 330 306 246 99 Row Stuffed) 7 8 9 10 11 Ingredients MM 10 20 40 80 160 Ε 70 50 24 19 0 200824699 F 45 ^22 34 25 0 G 55 36 28 24 0 Η 69 47 18 0 Mixed 4 rows 80 58 50 23 0 3.6.5 Statistical results The results of inhibition of FCC1 by arsenic trioxide (ΑΤΟ) are shown in Table 3.8. Table 3.8 Results of inhibition of FCC1 by arsenic trioxide (ΑΤΟ) Statistics ΑΤΟ dose dose logarithmic infection rate worm reduction rate probability (uM) (X) (%) (%) (Υ) 0.031 -1.509 79.9 20.1 4.1619 0.063 -1.201 73.7 26.3 4.3659 0.125 -0.903 68.3 317 4.5239 0.25 -0.6021 54.9 45.1 4.8743 0.5 -0.3010 44.4 55.6 5.1408 1.0 0 17.9 82.1 5.9192 2.0 0.3010 12.9 87.1 6.1311 4.0 0.6021 11.2 88.8 6.2160 8.0 0.9031 5.1 94.9 6.6352 Y=10904x+5.6582 Γ2=0.9706 IC50=0.25uM =49.28ng/ml 95% confidence limit (28.00-86.73ng/ml) IC90=3.73uM=737.94ng/ml 95% confidence limit (419.28-1298.77ng/ml) 3·7 test Steps and results (5) 3·7·1 drug preparation: Take 32μΙ of the original solution, add complete medium 0,5m丨, mix, this is the highest concentration of liquid (320μΜ), half-dilute in the marrow bottle, get a series The concentration of the drug solution is: 〇, 320, 160, 80, 40, 20, 10, 5 μΜ. (Total 12 rows, 11 歹[| is the highest dose hole 32〇um) 27 200824699 3.7.2 Worm blood _ : Take continuous culture of FCC1/HN, smear, stain, calculate infection rate, adjust infection with type B RBC The rate is around 1%. 3.7.3 Test: According to the method of Guan Weibin, Chen Lin, etc., add 20 μΙ of the drug solution per micropore on the 3599 culture plate, and then add 180 μM containing insect blood to make 12 rows. Then cover, shake for a few seconds, set 37. (: and cultured in 5% C02 box for 48 hours, remove the supernatant, mix, take RBC 5μΙ, mix, smear and stain φ color microscopy, calculate RBC infected with Plasmodium every 5000 RBC Number (only dead cells with no cytoplasm in the nucleus are not counted) • 3.7.4 Results

The protozoal infection rate at the start of culture was 1.1%. Table 3. The number of protozoa per 5000 RBCs after culture. Table 3.9 Number of protozoa per 5000 RBCs after culture [ IABCDEFG Η ΑΤΟ ft (μΜ) 0 5 10 20 40 80 160 320 Mix 256 75 34 21 9 5 0 0 Mix 247 72 39 25 10 4 0 0 Mix 231 69 28 20 9 6 0 0 Mix 258 57 26 17 8 4 0 0 28 200824699 3.7.5 Inhibition of FCC1 by arsenic trioxide (ΑΤΟ): Table 3.10 Inhibition of FCC1 by arsenic trioxide (ΑΤΟ) results Statistical data ΑΤΟ dose dose logarithmic infection Rate of worm reduction rate unit (uM) (X) (%) (%) (Y) 0.5 -0.3010 1.0 0 2.0 0.3010 4.0 0.6021 8.0 0.9031 5.6.0 8.32. 4 9 7· 2· 2 1 72.6 5.6008 87.1 5.1311 91.5 6.3722 96.3 6.7866 98.0 7.0537 Y=1.1832x+6.0328 r2=0.9849 _ IC50=0.13uM=25.72ng/ml 95% confidence limit (18.91-34.9ng/ml) - IC90=0.96uM=189.93ng/ml 95% confidence limit (139.65-258.3ng/mi) 3.8 Test procedure and results (6) 3·8·1 drug preparation: Take the stock solution 32μΙ, add complete medium 0.5ml, mix, this is the highest concentration drug Liquid _ (320μΜ), half-dilution in a penicillin bottle, to obtain a system of 歹{] concentration of liquid medicine: 〇, 320, 1 60, 80, 40, 20, 10, 5μΜ 1-6 1-6 columns, Η behavior highest dose hole 32〇uM). 3_8·2 dilution of worm blood: Take continuous culture of FCC1/HN, smear, stain, calculate the infection rate, adjust the infection rate to about 1% with sputum type RBC. 3-8.3 mm · According to the method of tube Weibin, Chen Lin, etc., add 20 μΙ per micropore on the 3599 culture plate, 29 200824699 and then add 180 μΙ containing insect blood, do 12 lines (1-6 is ΑΤΟ, 7- 12 is ATS). Then, cover and shake for several seconds, set at 37 ° C and incubate in 5% CO 2 box for 48 hours, remove the supernatant, mix, take RBC 5μ each, mix, smear and stain microscopy, calculate every 5000 RBC The number of RBCs infected with Plasmodium (only dead cells with no cytoplasm in the nucleus are not counted). 3·8·4 Results: The protozoan infection rate was 1.39% at the start of culture, and Table 3·11 was the number of protozoa per 5000 RBCs after culture.

Table 3.11 Number of protozoa per 5000 RBCs after culture ABCDEFG Η 计量 Metering (uM) 0 5 10 20 40 80 160 320 Mixing 420 103 66 38 17 8 0 0 3_8_5 Result statistic: Arsenic trioxide (/VFO) vs. FCC1 The statistical table of inhibition results is shown in Table 3.12. Table 3.12 Results of inhibition of FCC1 by arsenic trioxide (ΑΤΟ) 剂量 dose (uM) dose log (X) infection rate (%) worm reduction rate (%) probability unit (Y) 0.5 -0.3010 24.5 75.5 5.6903 1.0 0 15.7 84.3 6.0027 2.0 0.3010 9.0 91.0 6.3408 4.0 0.6021 4.0 96.0 6.7507 8.0 0.9031 1.9 98.1 7.0749 200824699 Y=1.5032X+5.8726 r2=0.9495 lC50=0.32uM=63.31ng/mI 95% confidence limit (40.32-99.4ng/mI) IC90= 2.09uM=413.48ng/ml 95% confidence limit (263.06-258.30ng/ml) 3·9· Through several experiments, it was found that 丨C50 is between 19.78-63.31ng/m丨, with an average of 35.57. The ng/m Bu IC90 is between 185.97-737.94 ng/ml, and the average dose is 451O7 ng/m. This dose is about 10 times larger than the existing antimalarial drug chloroquine, which is equivalent to the dose of pigoquine. Example 4: In vitro test of arsenic trioxide poisoning African trypanosomiasis 4.1 Test material 4.11 Drug: Arsenic trioxide injection (eight 33 < ^ 10 19/10111 pair 31). 4.1.2 Protozoa:

Trypanosoma brucei is labeled "427". The protozoan strain was obtained from Dr. Li Mei from New York University (Dr_ Mary Lee Bu 41.3 medium: SDM79 (JRH 57453) medium containing fetal bovine serum. 4_2 Test method The number and type of living cells were observed by microscope. 3 test steps

V 200824699 was treated with arsenic trioxide aimg/ml/day for 3 days, and the poisoning effect on the 7th was observed. 4.4 Test results Cytology found that the death of the worms was linear rather than a generally small circle. A temporary observation showed that the toxic effect of arsenic trioxide on the cultured Trypanosoma brucei was comparable to that of Melarsoprol. Example 5: In vitro test of arsenic trioxide on apoptosis of human peripheral blood mononuclear leukocytes. 5.1 Test materials 5.1·1 cell line: Fresh human peripheral blood mononuclear leukocytes isolated by self. 5.1 · 2 drugs: arsenic trioxide injection (Asadin®, 10mg/10ml/vial). 5.13 Reagent: White blood cell separation reagent. 5_2 Test methods and procedures 5·2_1 cell line isolation: Human centrifugal mononuclear leukocytes were isolated from human blood, and the Ficoll Plaque method was used to separate the other blood cells from the peripheral blood mononuclear leukocytes. 5.2.2 Cell culture: The isolated human peripheral blood mononuclear leukocytes were cultured in RPMI 1640 and added with 10% fetal bovine serum at 5% CO 2 and 95% air, 37. (: Next culture. 5.2.3 Apoptosis detection: After staining by P丨 staining, it is detected by flowcytometry. 32 200824699 5_2·4 Apoptosis test of human peripheral blood mononuclear leukocytes: combination After single-body antibody-mum-fluorescein tandem staining and Phiphilux treatment, the classification was detected by flow cytometry. 5.3 Experimental results 5.3.1 Arsenic trioxide induced apoptosis of human leukocyte cells in Zhouchi liquid It has been shown in experiments that it is found that arsenic trioxide induces apoptosis of human peripheral blood mononuclear leukocytes. • When arsenic trioxide (different 〇, 〇·5, 2.5, 5, or 10 μΜ, respectively) with different concentrations, all human peripheral blood singles The nucleated white blood cells were treated for 24 hours and 48 hours respectively, and the peripheral blood mononuclear white blood cells were stained with sputum staining, and the percentage of apoptosis of all human peripheral blood mononuclear white blood cells was detected by flow cytometry. It was shown that in the two groups treated for 24 hours and 48 hours, the arsenic trioxide was between 2.5 and 10 μΜ, and the arsenic trioxide was induced. There is no dose-dependent relationship between the apoptosis of human peripheral blood mononuclear leukocytes; there is a slightly higher rate of death than φ in the 48-hour group, and perhaps some specific sputum peripheral blood mononuclear leukocytes to arsenic trioxide ratio Other cells are sensitive. 5.3·2 Use caspase_3 activity to distinguish arsenic trioxide-sensitive white blood cell types with special cell staining patterns to distinguish human peripheral blood mononuclear leukocytes from the arsenic trioxide-induced apoptosis. The results are shown in Table 5.2. Using the caspase-3 activity test, combined with a special type of cell staining mode, the surface markers of human peripheral blood mononuclear white blood cells are identified as "CD3+" by the T lymphocytes, and the B lymphocytes are identified as "20082008". From the test results in Table 5.2, after treatment with arsenic trioxide at a concentration of 5.0 and 1/1 for 48 hours, 96% of T lymphoids died. Therefore, in the arsenic trioxide-induced apoptosis of specific human peripheral blood mononuclear leukocytes, the axillary lymphocytes have certain sensitivity. Table 5.2 After treatment with arsenic trioxide for 48 hours, the apoptosis of mononuclear leukocytes induced by specific human peripheral blood was controlled by 34.5 (%) 1 μΜ 36.1 (%) 5 μΜ 96.2 (%) Example 6: cells of arsenic trioxide against human fibroblasts Apoptosis and in vitro assays affecting the cell cycle' 6.1 Test Materials. 6.1 Cell Line: Human fibroblasts purchased from ATCC, R-90 [丨MR90]. . 6.1.2 Drugs: Arsenic Trioxide Injection (Asadin®, 10 mg/10 ml/vial). # 6·1·3 Reagents:

Triton Χ-100, sodium chloride, EGTA, ΝΡ-40, sodium fluoride, Na3V04, aprotinin, leupeptln, PBS, phenylmethyl-sulfonylfluoridepMSF). 6.2 Test method 6.11 sensitization of arsenic trioxide inhibiting the proliferation of fibroblastic cell lines 6.2·1·1 cell lysing test τ4 200824699 Different concentrations of arsenic trioxide ((, 1.0, 2·5, respectively) 5, or 10 μΜ), after treatment with human fibroblastic cell lines for 24 hours, staining with iodide staining (propidium iocHde staining), cell cycle arrest was measured by flow cytometry. 6-2, 1_2 Measurement of cell viability by MTT colorimetry In a 96-well microplate, 5 x 103 cells were added to each well. After the cells were cultured at 37 ° C, 30 μM of MTT solution was added, followed by incubation for 4 hours in the dark. After the Formazan grain was dissolved in DMSO, the absorbance enthalpy was measured at 570 nm using an EL]SA machine. Observation of apoptotic bodies by β_2.1.3 The type of apoptotic bodies was observed by Hoechst staining. The culture solution was removed and fixed by a 4% formaldehyde chamber in PBS for 5 minutes. Then, Hoechst dye 33258 5pg/mMn PBS was added for 5 minutes, and after washing, a reagent (PBS: glycerol = 3:1) was added. The nuclei stained with fluorescence were observed using a fluorescence microscope. Hoechst 33258 staining was used to observe the morphology of apoptosis. 6·2·2 Experimental results 6_2.2.1 Arsenic trioxide induced cell cycle arrest in human fibroblasts with different concentrations of arsenic trioxide (0, 1·0, 2.5, 5, 10 μΜ, respectively) for human fibroblastic cell lines After 24 hours of treatment, the cell cycle was analyzed by iodide staining and flow cytometry. It was found that the cell cycle of human fibroblasts arrested by arsenic trioxide was arrested at G2/M 〇6·2·2·2 arsenic trioxide induced human fibroblasts. The cell death of the mother cells was 0, 1·0, 2.5, 5, 10 μΜ with different concentrations of arsenic trioxide. After treatment with human fibroblastic cell lines for 24 hours, the cell activity was measured by 11 colorimetry. Arsenic trioxide is cytotoxic to human fibroblasts. 55* 200824699 At the same time, human fibroblastic cell lines were treated with different concentrations of arsenic trioxide (〇, 1·〇, 2·5, 5, 10 μΜ, respectively) for 24 hours, and the apoptosis was small by DAPI staining. body. The results showed that there was no apoptosis in the control group, 1·〇 and 2·5 μΜ arsenic trioxide induced cell bodies; 5.0 and 10 μΜ showed obvious apoptotic bodies, and apoptosis of human fibroblasts occurred. Necrosis. 6.Ζ2.3 Analytical method The experiment uses the variance analysis of the two elemental factorial design data. The data is done using the SPSS suite of software. According to the present invention, various modifications and changes may be made without departing from the scope and spirit of the invention, and although the invention has been specific to the key, it is undoubted that the present invention is not Unduly limited to these specific touches. Specialized in the application of the present invention, for the familiarity of the item 113⁄4

It is intended to be within the following claims. Different amendments that are obvious to the user: [Embodiment] [Simple description of the diagram] [Explanation of main component symbols] 36

Claims (1)

200824699 X. Patent application: The therapeutic application of an arsenic-containing blast-sensitive germ cell-related disease, the invention relates to the preparation of arsenic trioxide in the treatment of allergic diseases, immune diseases, fibroblast diseases, inflammation Use in diseases and infectious diseases, especially in malaria, African trypanosomiasis, asthma and systemic lupus erythematosus. It is apparent to those skilled in the art that various modifications and variations can be made without departing from the scope and spirit of the invention. Although the present invention has been described in terms of specific preferred embodiments, it should be understood that the invention should not be In fact, the various modifications that are obvious to those skilled in the art are also encompassed by the following claims. "1. Application of arsenic trioxide in the preparation of a medicament for treating arsenic-sensitive blast cell diseases. 2. The use according to claim 1, wherein the medicament is used by injection. 3. The claim 1 or 2 The use thereof, wherein the arsenic-sensitive blast cell disease is selected from the group consisting of an allergic disease, an immune disease, a fibroblast disease, an inflammatory disease, and a host disease. 4. The method according to claim 3. Application, wherein the immune disease is selected from the following: connective tissue disease, autoimmune thyroid disease, neuromuscular autoimmune disease, gastrointestinal autoimmune disease, autoimmune hepatitis, primary hepatobiliary sclerosis, primary sclerosis Spontaneous vasculitis, autoimmune carditis, and arteritis. 5. The use according to claim 4, wherein the immune disease is selected from the group consisting of systemic lupus erythematosus, rheumatoid arthritis, sclerosis, Gracies disease, Myasthenia gravis, multiple sclerosis, 200824699 ulcerative colitis and Crohn's d丨sease. 6. The use according to claim 3, wherein allergies The sexual disease is selected from the group consisting of asthma, over-related rhinitis, spring conjunctivitis and urticaria. 7. The use according to claim 3, wherein the inflammatory disease is selected from the group consisting of pneumonia, osteomyelitis, leprosy, syphilis, tuberculosis, Hepatitis, tumor formation and chronic obstructive pulmonary disease. 8. The use according to claim 3, wherein the fibroblastic disease is selected from the group consisting of: liver fibrosis, retinalization, pulmonary fibrosis, skin fibrosis, systemic fibrosis , pneumoconiosis, tuberculosis, atypical pneumonia (SARS), adult respiratory distress syndrome, chronic obstructive pulmonary disease, scarring, edema, dryness, invasive cystic fibrosis, and neurofibromatosis. The application according to claim 3, wherein the parasitic disease is malaria or trypanosomiasis. 10. The use according to claim 1, wherein the arsenic-sensitive blast cells are selected from the group consisting of white blood cells and Zhouchi liquid mononuclear cells. And a fibroblast. The use of the medicament according to claim 1, wherein the medicament comprises arsenic trioxide of from about 0.001 μΜ to about 20 μΜ·1 2 as claimed in claim 1 The use, wherein the medicament comprises arsenic trioxide of from about 0.1 μm to about 15 μΜ. The use of the medicament according to claim 12, wherein the medicament comprises from about 0.1 μΜ to about 10 μΜ of arsenic trioxide. 3S 200824699 VII. Designation: ( a) The representative representative of the case is: ( ). (2) The symbol of the symbol of the representative figure is simple: 8. If there is a chemical formula in this case, please disclose the chemical formula that best shows the characteristics of the invention. 200824699 |i Correction 1 Supplement to the invention patent specification (Do not change the format, order, and bold text of this manual. Please do not fill in the ※ part of the symbol.) ※Application number·· 0,^200βΜ) ※Application date·· "9 classification: life up (_6 plus one, invention name · (Chinese / English) treatment of estrogen-sensitive disease-related disease with steroids. ARSENIC COMPOUNDS FOR THE TREATMENT OF THE ARSENIC-SENSITIVE BLAST- CELL RELATED DISEASES 馨二,本人·(一人) Name: (Chinese/English) ^ 辛绍祺HSIN, SHAO-CHI ID: E121185156 Representative: (Chinese / English) Xin Shaozhen HSIN, SHA0-CHI Residence or Business Address: (Chinese / English) 242 1st Floor, No. 1-11, Lane 377, Zhongping Road, Xinzhuang City, Taipei County 242 Nationality: (Chinese / English) Republic of China REPUBLIC OF CHINA ® III, Inventor · (Total People) Name :(Chinese / English) Xin Shaozhen HSIN, SHA0-CHI ID: E121185156 Nationality: (Chinese / English) Republic of China REPUBLIC OF CHINA 200824699 IV. Declaration: □ Lord The facts of the second paragraph or the second paragraph of Article 22 of the Patent Law are as follows: the period of the year is: Years and months. □ Before applying, the patent has been applied to the following countries (regions): Please follow the following: order of acceptance of the country (region), application date, and application number] □ There is a claim for patent law Article 27, the first international priority: G no claim patent law, Article 27, first international priority right: Advocate the first domestic priority of Article 29 of the Patent Law: [Please follow the format of application date and application number]. Article 30 of the Patent Law: Biomaterials: _Guide to store biological materials · Domestic organisms Material [format: Please note according to the order of the depository, date, and number] Foreign biomaterials [format: Please note according to the order of the country, organization, date, and number of the deposit] I i do not need to deposit biomaterials: It is usually in the technical field. When the knowledge is easy to obtain, there is no need to register. 200824699 IX. INSTRUCTIONS: TECHNICAL FIELD OF THE INVENTION The present invention relates to a method of treating blast cell disease using an arsenide such as arsenic trioxide. [Prior Art] The basic cause of asthma in recent years is still the main cause of chronic inflammation (Chronic inflammation). Due to the dual effects of genetic inheritance and the external environment, allergic inflammatory reactions occur, and the affected inflammatory cells, including dendritic cells, epithelial cells, T cells, B cells, obese cells, granulosa cells, and eosinophils are involved. In this war, early detection, early prevention, or early treatment of asthma is the goal of the effort. The cause of asthma may be caused by various stimuli (inducing) to induce tracheal inflammation (AI) caused by abnormal control of T cell immune response, which induces or aggravates tracheal hyperresponsiveness and causes bronchoconstriction, mucosal congestion and edema, hypersecretion, micro blood Leakage and gradually remodeling of the trachea eventually lead to tracheal stenosis, restricted airflow, and clinical symptoms. Systemic lupus erythematosus (SLE) is a progressive connective tissue disease characterized by the production of a variety of autoantibodies in the body that can cause damage to many of its own constituent components. The disease is a multi-system autoimmune disease, the main clinical manifestations are: arthritis, glomerulonephritis, epileptic seizures, anemia or thrombocytopenia. The disease often has an episode and an alternating phase of remission. In the past, it was considered that SLE was difficult to treat and the prognosis was very poor. Recently, due to the advancement of medical science, SLE has a deeper understanding. In the case of early diagnosis and timely treatment, the damage of vital organs is reduced, and the survival rate is improved. The old prognosis has changed a lot. Due to the development of immunopathology in the past 30 years, it has been confirmed that the humoral immunity and cellular diseases of SLE patients have abnormal changes in different degrees. However, the exact cause of SLE has not yet been fully elucidated. 200824699 may be related to some of the following factors: genetic elements, sex hormones, viral infections, drugs. Malaria is called hernia in ancient China. It is called malaria in foreign countries. It is mala mala (bad) and aria (air two-synthesis. The disease is a parasite that parasitizes the human body when it is bitten by a female malaria mosquito. Caused by introduction into the human body. Clinical manifestations of intermittent, timed, episodic chills, high fever and sweating, as well as anemia and splenomegaly. Frescoes and vaginal malaria often recur, falciparum malaria fever irregular, patients It is characterized by dysfunction of the central nervous system and presents a dangerous attack. There are 4 kinds of human malaria caused by 4 different malaria parasites, namely vivax malaria (benign tertian), the pathogen is Plasmodium vivax; Qurtan malaria (malaiiae malaria), the pathogen is P. malariae; Oval malariae, the pathogen is Plasmodium falciparum (卩.〇7316); falciparum malaria 1_3"1^ 1113丨3113, malignant tertian), the pathogen is Plasmodium falciparum. There are more than 20 kinds of trypanosomes, only two species of human pathogens. Trypanosomiasis is a disease caused by trypanosomes, including the Americas. Trypanosomiasis (Ameiican trypanos Omisis) and African trypanosomiasis, the former also known as chagas disease, the latter also known as African sleeping sickness ° The African trypanosomiasis pathogen is Trypanosoma brucei, and its three subspecies are Disease. T. b. rhodesiense causes Rhodosia trypanosomiasis frozen non-sleep disease); T. b. gambiense causes Gambia trypanosomiasis ( Cases caused by T. b. brucei, although clinically reported, but less, mainly caused by the occurrence of cattle. African trypanosomiasis is also called drowsiness. Sleeping sickness is one of the serious diseases of African zoonotic diseases. It is a disease caused by the T. brucei Complex parasitic protozoa. Tsetse flies are the vector. Early manifestations of the disease include irregular fever, lymphadenitis, etc., and later manifestations of central nervous system damage, severe headache, unresponsiveness, lethargy, and lethargy, and eventually death. 200824699 Traditional Chinese medicine is used in natural habitats for many natural medicines. to The human body can not cause side effects such as toxicity. Chinese medicine processing technology has been one of the main methods to reduce the side effects of traditional Chinese medicine for a long time. The arsenic is a representative of the toxic drugs, although the traditional Chinese medicine processing technology can reduce its toxicity to a certain extent, but long-term It still gives people a bad impression and is fascinating, so it limits the development of arsenic. There are many kinds of chemical components contained in arsenic, but its main component is arsenic trioxide. In the current known literature, the application of arsenic is mainly based on its main component, arsenic trioxide. Since Chinese herbal medicines have been highly valued by countries all over the world, the understanding and application of base creams have increased the use of arsenic trioxide. For example, the use of arsenic trioxide for the treatment of cancers such as cervical cancer, skin cancer, and esophageal cancer. Or skin diseases such as corns. However, other applications of arsenic trioxide have yet to be further developed and studied by pharmaceutical researchers. SUMMARY OF THE INVENTION The present invention relates to a method of treating blast cell disease using an arsenide such as arsenic trioxide. The present invention relates to the preparation of arsenic trioxide in the treatment of allergic diseases, immune diseases, fibroblastic diseases, inflammatory diseases and infectious diseases, particularly malaria, African trypanosomiasis, asthma and systemic lupus erythematosus application. DETAILED DESCRIPTION OF THE INVENTION The present invention relates to the use of arsenic trioxide for the preparation of a medicament for the treatment of arsenic-sensitive blast cell diseases. Preferably, the drug is used by injection. Preferably, the arsenic-sensitive blast cell disease is selected from the group consisting of an allergic disease, an immunological disease, a fibroblast disease, an inflammatory disease, an infectious disease, and a parasitic disease. 7 200824699 More preferably, the immune disease is selected from the following: connective tissue disease, autoimmune thyroid disease, neuromuscular autoimmune disease, gastrointestinal autoimmune disease, autoimmune hepatitis, primary hepatobiliary sclerosis, Primary sclerosing cholangitis, autoimmune carditis and arteritis. Most preferably, the immune disease is selected from the group consisting of systemic lupus erythematosus, rheumatoid arthritis, sclerosis, Gracies disease, myasthenia gravis, multiple sclerosis, ulcerative colitis, and Crohn's disease. Preferably, the allergic disease is selected from the group consisting of asthma, hypersensitivity pneumonitis, diffuse pulmonary fibrosis, allergic rhinitis, eosinophilic rhinitis, spring conjunctivitis, and measles. Preferably, the inflammatory disease is selected from the group consisting of pneumonia, osteomyelitis, leprosy, syphilis, tuberculosis, hepatitis, tumor formation, and chronic obstructive pulmonary disease. Preferably, the fibroblastic disease is selected from the group consisting of liver fibrosis, pulmonary fibrosis, skin fibrosis, systemic fibrosis, pneumoconiosis, tuberculosis, atypical pneumonia (SARS), adult respiratory distress syndrome, chronic obstruction Sexual lung disease, scarring, swelling, dryness, invasive cystic fibrosis, and neurofibromatosis. • Young legs, Lin disease, worm disease. The car's father is that 'the worm disease is sore or trypanosomiasis. Preferably, the arsenic-sensitive blast cells are selected from the group consisting of white blood cells, peripheral blood mononuclear cells, and fibroblasts. &lt; Preferably, the medicament comprises from about 0.001 μΜ to about 20 μΜ of arsenic trioxide. ^ Further, the drug comprises about 1 μΜ to about 15 μΜ of arsenic trioxide. Most preferably, the drug comprises from about 1 μΜ to about 10 μΜ of arsenic trioxide. 2008 2008699 The term "arsenic-sensitive blast cells" as used herein means white blood cells, peripheral blood mononuclear cells and fibrils. cell. The term "mammal" as used in vomiting includes human milky animals i. [Embodiment] The present invention will be further described in detail below with reference to the accompanying drawings and embodiments: Example 1: Treatment of bronchial asthma with arsenic trioxide Animal test φ 1·1 material test animals: commonly used Balb/C female rats, 6-8 weeks old, fed for six weeks, 15 in each group. BALF/c female rats of SPF six-week-old were purchased from the National Taiwan University Animal Center. Each mouse was housed at room temperature, controlled at 22±2t at room temperature, and 12 hours in light and dark cycle time (6 in the morning, 6 in the afternoon) Dark), free access to drinking water and AIN-76 formula powder feed, weekly food intake and body weight were recorded twice a week, blood was collected once every other day; mice were randomly divided into four groups at eight weeks of age, sensitization steps and treatment The method is as follows. Drug: arsenic trioxide injection (Asadin®, 10 mg/10 ml/vial) diluted with PBS_solution. Reagents: Balanced salt solution (HBSS) lavage fluid, Trypan blue, hematoxylin-eosin, carbonate coating buffer, containing 〇.〇5% Tween 20% PBS buffer, blocking solution (blocking solution; 1X PBS buffer containing 1% BSA-0.05% Tween 20), [3H] TdR (please confirm) Other: oval albumin (ovalbumin), wax film ( Paraffin), eotaxin ELISA kit (old L, Hamburg, Germany), anti-mouse 丨gE antibody. 9 200824699 Main instruments: nebulizer (DeVilbiss, Somerset, PA), centrifuge, hemocytometer, microscope, eotaxin ELISA read Machine, microplate reader, liquid scintillation counter, Plethymography 〇1.2 test method 1.2.1 Establish respiratory tract inflammatory animal model Use commonly used BALB/c female mice, 6-8 weeks old, mouse sensitization step first Ovalbumin was intraperitoneally injected into mice, and mice were repeatedly injected with the same dose of ovalbumin on Days 14, 14, 21, and 28, respectively. Six to eight mice were placed in a closed container at the same time. After the OVA antigen is made into a respirable particle type by a nebulizer, the mouse is inhaled in the closed container for more than 20 minutes. The nebulizer will deliver 0.5-5 m (7) of the antigen at a rate of ml3 ml/min. Particle molecules. 1·2·2 ELISA assay for determination of ovalbumin (OVA)-specific antibody (J method) Dissolve 0.5 mg of antigen in 100 ml of carbonation-covering buffer, and add a micro-culture plate to stand at 4 overnight. The microplate was washed with IX PBS buffer containing 0.05% Tween 20 for 2 days, then the microplate was blocked with blocking solution and washed 3 times. Then 100 μmol of the sample to be tested was added to each well. The reaction was diluted with liquid, and washed 6 times after the reaction. 100 ml of anti-mouse IgE antibody was added to carry out the reaction and washed 6 times. The above reactions were all carried out at 37T for 45 minutes, and then moved to room temperature for 15 minutes. Finally, each microporous Then add 10 (^|6-8 butyl 3 (2.2 ^ 〇 〇 - fine - 3 Ethylbenzthiazoline -S-Sulfonic Acid) to the solution at room temperature for about 30 minutes, stop the reaction with 100 μΙ of 5% SDS per micropore And read the light absorption 以 with a culture disk reader. 12_3 Calculation of the number of bronchoalveolar lavage fluid and macrophages, lymphocytes, neutrophils and eosinophils White blood cells 24 hours after the last inhalation of antigen, all the mouse eye sockets were sacrificed after blood collection. Bronchial intubation was then performed immediately, and the lungs of the mice were lavaged four times with 1 ml of HBSS containing no calcium and magnesium ions, 200824699 and the rinse was collected. The collected alveolar lavage fluid was centrifuged at 400 x g for 1 Torr at 4 °C. The cells obtained by centrifugation were suspended in 1 ml of HBSS, and then the total number of cells was counted using a blood cell counter. The cells were then fixed on a glass slide by Cytocentrifuged preparations, and the number of cells was counted by Liu's stain. The number of cells in each cell is calculated as the number of macrophages, lymphocytes, neutrophils, and eosinophils in 300 cells, and is classified according to their standard type. 1.2.4 Determination of eotaxin content in bronchoalveolar lavage fluid Bronchoalveolar lavage fluid (BAL rushed eotaxin content, determined by eotaxin ELISA kit, the procedure is as directed by the ELISA kit manufacturer. 1 2 · 5 mice respiratory resistance test Mice were stimulated by increasing the concentration of methacholine (Mch.) (6.25 mg/m Bu 12-5 mg/m b 25 mg/m b 50 mg/ml, respectively), and the respiratory tract of the mice was determined by plethymography. Airway enhanced pause (PenH) values to see if the airway contraction in mice is improved. When the mice were eight weeks old, they were randomly divided into four groups of 15 mice each, and the four groups of mice were treated as described later. Group A mice were sensitized and treated with only PBS. Group B mice were sensitized with ovalbumin but treated with PBS only. Group C mice were sensitized with ovalbumin and treated with arsenic trioxide at a dose of 2.5 mg/kg. Group D mice were sensitized with ovalbumin and treated with arsenic trioxide at a dose of 5.0 mg/kg. Table 1.1 Sensitization antigen treatment Group A PBS PBS Group B OVA PBS Group C OVA ΑΤΟ 2.5 mg / kg Group D OVA ΑΤΟ 5 mg / kg 1_4 Test Step 11 200824699 8 weeks old BALB / c female mice, mouse feeding methods and The grouping is as described above. Methods and Routes for Sensitization in Old Rats As described above, the titer of antigen-specific antibodies was determined weekly by eye socket blood sampling after sensitization in mice. 1.5 The treatment method was first intraperitoneally injected with ovalbumin (〇VA) into mice, and the mice were repeatedly injected with the same dose of ovalbumin on the third, 14th, 21st and 28th day to complete the sensitization step. The mice were treated with PBS and a dose of 2.5 mg/kg and 5.0 mg/kg of arsenic trioxide intraperitoneally for 7 days. Mice were stimulated with inhaled ovalbumin antigen on days 42, 43 and 44. The airway resistance test of the mice was carried out on the 45th day. The respiratory resistance test results are shown in Table 1.2. Table 12 Respiratory resistance (tracheal reaction) of the test. Respiratory resistance test Mch (mg/ml) 6.25 mg/ml 12.5 mg/ml 25 mg/ml 50 mg/ml Group A 2 ·20 ±0.28# 8.22 ±1.22 11.99 ±1·50 14.21 ± 1 〇6 —--- Group B 2.50 ±0.40 8.00 ±0.93 14.52 ±1.42 19.15 ±3.35 ----- Group C 3.62 ±0.47 7.83 ±1.13 10.15 ±0.84* 9.83 ±1.17* ------ Group D 2.55 ± 0.35 8.82 ±0.88 11.57 ±1.52 12.51 ± 190 --1 -1 #数値 is the average soil SD * compared with group B, p &lt; 0.05. After stimulation of mice by increasing the concentration of methacholine (Mch·) (6.25 mg/m b 12.5 mg/ml, 25 mg/mh 50 mg/ml), the respiratory tract resistance of mice was determined by plethymography, and the tracheal allergic reaction-PenH (AHR; airway hyper-reactivity-PenH (airway enhanced pause)). Compared with group B mice that were not treated with arsenic dioxide, the dose of 2.5 mg/kg arsenic trioxide successfully blocked the tracheal high immunoreactivity of mice in group C and reduced the respiratory resistance of mice. 12 200824699 Last inhalation of ovalbumin After 48 hours of antigen stimulation, all mice were sacrificed after blood collection. Immediately, bronchial intubation was performed, and the lungs of the rats were lavaged four times with 1 ml of HBSS containing no calcium and magnesium ions, and alveolar lavage fluid was collected. The collected alveolar lavage fluid was subjected to histopathological investigation, the number of various cells (macrophages, lymphocytes, neutrophils and eosinophils) was measured, and the content of eotaxin in the bronchoalveolar lavage fluid was determined. As shown in Table 1.3 and Table 1-4. Table 1-3 Bronchoalveolar lavage fluid macrophage, lymphocyte, neutrophil and eosinophilic white blood cell count BALF mouse white blood cell count (x1000ce 丨ls/ml) macrophage eosinophilic white blood cell neutrophil Ball lymphocytes + A group 135.6±54.5# 3.2±2.1 8.4±4.9 12.9±8,2 Group B 243.8±69.3 105.3±60.7 77.1±32.8 37.8±16.3 Group C 621 _0±199.6 28.6±13.0 200.7±75.2 37_0±9.4 Group D 499.1±131.6 33.2±12.5 244.0±72.6 28.7±13.6 The number of white blood cells in the mouth is scarce and is not indicated here. Table 1.4 shows the average concentration of eotaxin in the bronchoalveolar lavage fluid of mice and the standard deviation. Compared with group B mice that were not treated with arsenic trioxide, mice in groups C and D received arsenic trioxide treatment to reduce eotaxin levels in bronchoalveolar lavage fluid, especially in group C mice treated with 2.5 mg/kg arsenic trioxide. There was a significant difference in the B group of mice treated with arsenic trioxide (P &lt; 〇.〇5). Table 1.4 Determination of eotaxin in bronchoalveolar lavage fluid Eotaxin (pg/ml) Average SD A group 14.42 8.39 Group B 20.90 4.97 Group C 12.38* 3.92 Group D 16.00 4.06 *Compared with Group B, p&lt;0.05 in mice receiving egg white After stimulation with protein antigens, mouse blood was taken and serum levels of ovalbumin-specific IgE values (〇VA-specific IgE values) were measured as evidence of sensitization. Table 1.5 shows that the serum albumin-specific antibody content in the B, .C, and D groups was significantly higher than that in the control group of 200824699, indicating that mice in groups B, C, and D received ovalbumin antigen stimulation. Sensitization was successful. Table 1.5 OVA stimulation index OVA-specific IgE antibody (SI.) Average SD A group 0.30 0.09 B group 2.06 0.81 C group 2.20 0.70 D group 1.96 0.31 Example 2 treatment of systemic lupus erythematosus with arsenic trioxide Test 2_1 Test material 2.1.1 Test animal NZBxNZW F1 female mouse, about 7 months old; NZB (H-2d) female mouse; NZW (H-2z) male mouse 2·1·2 drug arsenic trioxide injection ( Asadin ®, 10 mg/10 ml/vial), diluted 卩83. 2·13 try Qi (J (1) methylated Bovine Serum Albumin working solution) (2) calf thymus DNA working solution (3 ancestor Liquid (Geletin-PBS solution) (4 洫 clear and positive control group: serum was obtained by centrifugation (5000 rpm, 10 minutes, 4) of blood after blood collection. Serum was diluted with blocking solution at the time of use. The anti-DNA IgG or IgM secreted by the hybridoma is taken from each culture plate and diluted into 250-fold 500 μM solution for serial dilution. 14 200824699 (5) Level 2 antibody (2Ab; Horseradish pe "oxidase (HRP)_conjugatecl anti-mice mouse Y bond or μ bond antibody; Horserapish peroxidase (HRP)-conjugated anti-mouse γ chain or μ chain Ab): (6) 2,2'-azino-bisC3_ethylbenzthiazoline-6-sulphonic acid (ABTS Solution (7) 5X citrate buffer (8) SDS solution 2.1.4 Main instrument: Determination of antibody day 1 8 reading machine 2.2 Test method 2.2.1 Establishment of human systemic erythematosus animal model about 7 NRB NZBxNZW F1 female mice and NZB/NZW The mouse will naturally develop an autoimmune syndrome similar to human systemic lupus erythematosus (please confirm). The autoimmune syndrome of NZB/NZW-matched mice produces anti-nuclear antibodies at four months of age. (antinuclear antibodies (IgG)), glomerulonephritis and proteinuria occur at nine months of age, and uremia is the leading cause of death due to kidney failure. 2·2·2 ELISA for determination of anti-dsDNA and anti-ssDNA antibodies The method uses standard ELISA to determine the anti-DNA and anti-DNA antibody content of anti-dsDNA and ssDNA in serum. The steps are as follows: (1) mBSA-like coating: covering 100 μM of mBSA (10 pg/mL) per microwell. After standing overnight at 4 ° C, it was washed twice with PBS. (2) dsDNA and ssDNA coverage: 100 μΙ of dsDNA and ssDNA (7.5 Mg/mL, respectively) per microwell, and silenced at 4 ° C Set overnight, then rinse twice with PBS. 15 200824699 (3 blocking blockage: cover 200 μΙ of blocking solution per microwell, and place it on the 44 core overnight (or room temperature for 2 minutes or 37 ° C for 45 minutes and room temperature for 15 minutes) ), wash with PBS-tween 20 three times. (4) Add serum or positive control group or negative control group, add 1 μμ| serum or positive control group or negative control group, and let stand at 37 °C. After 45 minutes, room temperature was 15 minutes of iron or room temperature for 2 hours), and washed six times with PBS-tween 20. (5) The above-mentioned class 2 antibody was added: 1 μμ of the secondary antibody was added to each well, and the mixture was incubated at 37 ° C for 45 minutes, at room temperature for 15 minutes, and washed with PBS-tween 20 six times. (6) A coloring agent (ABTS solution) was added to the sea micropore of the coloring agent, and the reaction was carried out for 5-10 minutes in the dark, and a stop solution (5% SDS solution) was added and O.D. measurement was carried out at a light absorption 値405 nm. (7 The amount of filthy-DNA IgG is expressed in units of ELISA assay (EU/ml). Compared with mAb 10F10, the OD 产生 produced by the 10F10 antibody at a concentration of 74 ng/m 値 is defined as 1 EU/m丨. Anti-DNA IgG. Anti-ribosomal IgG concentration is measured in EU according to 10F10 antibody concentration. Absorption enthalpy produced by 231 ng/ml of 10F10 antibody is defined as 1 _ EU/ml leg IgG IgG. 2·2·3 Glomerulonephritis in mice was evaluated for glomerulonephritis in mice. Urine protein content was determined by urine staining (Multistix®, Bayer). Colorimetric determination of urine protein content was greater than 1 g/L (2+) It is defined as severe glomerulonephritis. It is detected in mice at the 37th week and the 40th week. 2.2.4 Survival rate of mice Evaluation of 6 mice per group, record the survival rate of mice to the 43rd week 16 200824699 2.3 Test procedure 2·3_1 The test group mice were randomly divided into three groups at 24 weeks of age, with 6 rats in each group: (1) sputum group: treated with PBS only (10 ml/kg); (2) group B: Treatment with arsenic trioxide at a dose of Z5 mg/kg; (3) Group c: treatment with arsenic trioxide at a dose of 5.0 mg/kg. 2·3_2 test procedure mice were randomly divided into three groups at 24 weeks of age. Each group of 6 patients received a treatment method. Each of the stars, one, three, and five each were injected intraperitoneally for one month. Before the treatment and after the treatment was stopped, the blood test was taken every four weeks by the orbital blood collection method. -ssDNA and anti-dsDNA antibody for 3 months. 2.3.3 Results The levels of anti-dsDNA and anti-ssDNA antibodies in mouse serum were determined by ELISA: the anti-dsDNA measurements are shown in Table 2.1. Table 2.1 IgG Anti-dsDNA titer (EUSA unit) Test time (week) Before the test 4 8 12 PBS (Group A) 0.280 0.623 0.679 0.381 2_5mg/kg ΑΤΟ (Group B) 0.262 0.572 0.464 0.442 5mg/kgAT〇 (Group C) 0.362 0.285 The results of 0.357 0.702 showed that the anti-DNA antibody levels in the B and C groups treated with arsenic trioxide were lower than those in the group A mice not treated with arsenic trioxide; and the higher dose of arsenic trioxide was stronger because of anti-DNA. The content of antibody has a strong correlation with the activity of SLE disease. The results of this test show that arsenic trioxide may become a new therapeutic agent for SLE. 2.3.4 Results of glomerulonephritis in mice17 200824699 Kidney of mice The glomerulonephritis assessment was performed using a urine test dipstick colorimetric assay for urine protein content. Colorimetric determination of urinary protein content greater than ig/L (2+) is defined as severe glomerulonephritis. Table 2.2 Proportion time of mice with severe glomerulonephritis (week) 37 40 PBS (group A) 25 100 2.5 mg/kg AT〇 (group B) 16.67 40 5 mg/kg ΑΤΟ (group C) 25 25 Table 2.2 The test data showed that the treatment with arsenic trioxide could significantly delay the occurrence of severe glomerulonephritis in the mice. Some mice with severe glomerulonephritis did not develop any of the kidneys. The phenomenon of proteinuria. In the sputum group that did not receive arsenic trioxide treatment, all patients developed severe glomerulonephritis. Compared with group A mice that were not treated with arsenic trioxide, arsenic trioxide treatment significantly reduced severe kidney in mice in group B and C. Occurrence and progression of glomerulonephritis 2.3.5 Evaluation of mouse survival rate Table 2.3 Evaluation of mouse survival rate (%) Time (week) 22 31 38 39 40 41 42 43 pbs (group a) 100 100 66.7 50 50 16.7 0 0 2.5mg/kg ΑΤΟ (Group B) 100 100 100 83.3 66.7 66.7 66.7 50 5mg/kg ΑΤΟ (Group C) 100 100 66.7 66.7 66.7 66.7 66.7 66.7 Table 2.3 shows Group B and C mice treated with arsenic trioxide, 43 Weekly survival rate For the 50%, group A mice that were not treated with arsenic trioxide had all deaths. Compared with group A mice that were not treated with arsenic trioxide, arsenic trioxide treatment significantly prolonged the survival of mice in groups B and C. Example 3: arsenic trioxide In vitro test for plague of Plasmodium falciparum malaria 18 200824699 3.1 Test material 3.1.1 Drug: Arsenic trioxide injection (Asadin®, 10mg/10ml/vial) 〇trioxide arsenide, molecular weight 197.84, test drug concentration is 1mg/ml ( 5054.6μΜ), dilute it to 160μΜ before the test, and then half-dilution, the final concentration is calculated as ng/ml. 3.1.2 Protozoa Plasmodium falciparum FCC1/HN from Hainan, laboratory continuous culture or refrigerated storage 5 3. 3. Culture medium: The medium (liquid) used for continuous culture of Plasmodium falciparum in vitro, the main components are RPMM640 (GBC〇), HEPES (GBC〇), NaHC〇3ft! AB serum, etc. 1.4 Other materials Serum, red blood cells: supplied by Guangzhou Blood Center. Red blood cells (RBC) are ACD anticoagulated type B whole blood, stored at 4 °C, no more than one month, no serum medium before use. -3 times. 3.2 Test method _ Continuously culture FCC1/HN. Referring to the methods of in vitro micro-measurement of anti-malarial drugs, such as Guan Weibin [73] and Chen Lin [74], the drug solution was added to the wells at a rate of 2 μμΙ per well, and then the infection rate was about 1%. Cap, gently shake for a few seconds, at 37 ° C, 5% C 〇 2, culture for 48 hours, remove the supernatant, take RBC a thin film of blood, Gyr stain and microscopic examination, calculate the protozoal infection rate, In the control group/100, the worm reduction rate of each dose group was determined [worm reduction rate = (control group infection rate, test group infection rate) / control group infection rate x, with dose logarithm as abscissa' The probability unit is the vertical seat, which is labeled as a regression equation, and the amount of drug that reduces protozoal proliferation by 50% and 90% compared with the control group is calculated. 3.3 Test procedure and results (1) 19 200824699 3.3.1 Drug preparation and sterilization of penicillin bottles 8 'In accordance with Table 3-1, respectively, add RPMI-1640 complete medium and arsenic trioxide (MO) Table 3.1 The concentration of arsenic trioxide in the culture solution ΑΤΟ ( μΙ) 0 0.5 1.0 1.5 2.0 2.5 3.0 0 RPMM 640 medium (10)) 1 1 1 1 1 1 1 1 concentration _) 0 2.5 5 7.5 10 12.5 15 0 3·3·2 Dilute the blood to take the continuously cultured FCC1/HN, Smear, staining, calculate the infection rate, and adjust the infection rate to about 1% with sputum type RBC. 3·3·3 test According to the method of Chen Lin et al., 20 μΙ of the drug solution was added to each micropore on the 3599 culture plate, and then 180 μΙ containing insect blood was added, and 4 lines were tested. After that, the cells were shaken for several seconds, cultured in a 5% CO 2 box at 37 ° C for 48 hours, and the number of RBCs infected with Plasmodium in 5000 RBCs was calculated by thin-film staining. 3.3.4 Results The protozoal infection rate was 0.98% at the start of culture. Table 3.2 shows the number of protozoa per 5000 red blood cells after culture. Table 3.2 Number of protozoa per 5000 red blood cells after culture (Control) BCDEFG Η ( Control) 剂量 dose (//Μ) 0 2.5 5 7.5 10 12.5 15 0 1 172 82 79 81 60 53 39 199 2 306 163 48 33 47 49 33 294 3 251 125 36 29 46 32 17 213 4 198 74 57 55 53 48 26 208 It can be seen from the above table that the highest dose (15 μΜ) G# failed to completely inhibit Plasmodium development. 20 200824699 3.4 Test procedure and results (2) 3.4.1 Drug preparation 9 sterile penicillin bottles, respectively, add RPMI-1640 complete medium and arsenic trioxide (ΑΤΟ) according to Table 3.3. Table 3.3 Concentration of arsenic trioxide in culture solutionΑΤΟ , (μΙ) 0 -----n 1 2 3 4 5 6 7 8 RPMI-1640(m!) 0.5 0.5~ 0.5 0.5 0.5 0.5 0.5 0.5 0.5 Concentration_) 0 10 —' 20 30 40 50 60 70 80 3.4.2 Dilute blood was taken from continuous cultured FCC1/HN, smeared, stained, and the infection rate was calculated. The infection rate was adjusted to about 1% by sputum type RBC. 3.4_3 Test According to the method of tube Weibin, Chen Lin, etc., 20 μM of the drug solution was added to each well of the 40-well culture plate, and then the blood containing 180 μ of the insect blood was added for 4 lines. Then, it was shaken for several seconds, placed in a 37 ° C, 5% C〇2 box for 48 hours, and the supernatant was removed. RBC coated sheets were taken, stained and microscopically examined to calculate the infection of Plasmodium in 5000 RBCs per φ. The number of red blood cells. The protozoal infection rate was 1.6% when the results were started, and the number of protozoa per 5000 red blood cells after the culture in Table 3.4. Table 3.4 Number of protozoa per 5000 red blood cells after culture 3.4 (control) 2 3 4 5 6 7 8 9 八丁〇量//|\/| 0 10 20 30 40 50 60 70 80 A 227 96 49 46 45 45 50 40 25 B 209 50 68 63 64 60 70 56 19 C 188 70 79 90 61 87 67 57 23 21 200824699 D 229 71 77 77 67 74 69 66 30 J As can be seen from the above table, the density of 80 μΜ# protozoa has decreased, but its development has not been completely inhibited. 3.5 Test procedure and results (3) 3.5.1 Preparation of the drug Take 20 μΙ of the original solution and complete the medium, and half-dilute a total of 11 dose groups in the penicillin bottle. They are: 2522, 1216, 633, 316, 158, 79, 40, 20, 10, 5, 2.5, ΟμΜ. 3.5.2 Dilution of worm blood The continuous culture of FCC1/HN, smear and staining, calculate the infection rate, and adjust the infection rate to about 1% with sputum type RBC. 3.5.3 Test: According to the method of Guan Weibin, Chen Lin, etc., add 20 μΙ of the drug solution per micropore on the 3599 culture plate and add 180 μM containing insect blood to make 4 rows. Then cover, shake for a few seconds, set at 37 ° C and 5% CO2 box for 48 hours, remove the supernatant, take RBC smear, stain and microscopic examination, calculate the number of RBC infected with Plasmodium per 5000 RBC . 3.5.4 Results The protozoal infection rate was 1.8% at the start of culture. Table 3.5 shows the number of protozoa per 5000 red blood cells after culture. Table 3. Number of protozoa per 5000 red blood cells after culture. 1 2 3 4 5 6 ΑΤΟ Metering μ Μ 2522 1216 633 316 158 79 A 0 0 0 0 0 38 Β 0 0 0 0 0 28 C 0 0 0 0 0 33 D 0 0 0 0 0 25 Four lines mixing 0 0 0 0 0 32 22 200824699 (Continued from the above table) 7 8 9 10 11 12 ΑΤΟMeasuring Μ μ 40 20 10 5 2.5 〇Control A 63 75 88 108 206 503 B 63 75 88 108 206 503 C 52 69 81 160 236 450 D 43 60 77 108 186 392 Four lines of mixing 44 65 92 116 200 473 As can be seen from the above table, 158 uM can completely inhibit FCC1 development. 3.5.5 Results Statistics: The results of the inhibition of FCC1 by arsenic trioxide (ΑΓΟ) are shown in Table 3.6 Table 3.6 Arsenic Trioxide (ΑΤΟ) inhibition of FCC1 Results Statistical Table Dose-dose Logarithmic Infection Rate Reduced Insect Rate Rate Unit (uM) (X) ( %) (%) (丫) 0.25 -0.6021 42.3 57.7 5.1942 0.5 -0.3010 24.5 75.5 5.6903 1.0 0 19.5 80.5 5.8596 2.0 0.3010 13.7 86.3 6.0939 4.0 0.6021 9.3 90.7 6.3225 8.0 0.9031 6.8 93.2 6.4909 3.6 Test procedure and results (4) 3.6. 1 Preparation of the drug: Take the stock solution 100μΙ, add the complete medium 3059μΙ, mix, this is the highest concentration liquid (160μΜ), take the highest concentration of the solution 200μΙ, and take 200μΙ medium in the penicillin bottle to mix, half-dilution, A series of concentrations of the drug solution are: 〇, 160, 80, 40, 20, 10, 5, 2_5, 1 · 25, 0.625, 0.3125, ΟμΜ. (Total 4 rows, 11 columns for the highest dose hole 160uM). 3·6·2 Insect blood dilution: 23 200824699 Continuous culture of FCC1/HN, smear, staining, calculation of infection rate, B-type RBC regulation of infection rate of about 1%. 3.6.3 fil test: According to the method of Guan Weibin, Chen Lin, etc., add 20 μΙ of the drug solution per micropore in the 3599 culture plate, and then add 180 pl of insect-containing blood to do 4 rows. Then cover, shake for a few seconds, set 37t: incubated in 5% C〇2 box for 48 hours, remove the supernatant, take RBC smear and stain, microscopic examination, calculate RBC infected with Plasmodium every 50 (five) RBC Number (only dead cells with no cytoplasmic nucleus are not counted). 3·6·4 Results: The protozoal infection rate was 1.5% at the start of culture, and the number of protozoa per 5000 red blood cells after culture in Table 3.7. Table 3.7 Number of protozoa per 5000 red blood cells after culture 1 2 3 4 5 6 AT〇Dose Μμ Control 0.3125 0.625 1.25 2.5 5 Ε 394 384 371 328 279 71 F 410 407 401 361 302 72 G 457 414 364 276 240 67 Η 454 385 339 308 235 98 Mixed 4 lines 448 358 330 306 246 99 (Continued Above table) 7 8 9 10 11 ΑΤΟDose Μ μ 10 20 40 80 160 Ε 70 50 24 19 0 F 45 22 34 25 0 G 55 36 28 24 0 H 69 53 47 18 0 Mixed 4 lines 80 58 50 23 0 3.6 .5 Statistical Results The results of inhibition of FCC1 by arsenic trioxide (ΑΤΟ) are shown in Table 3.8. 24 200824699 Table 3.8 Results of inhibition of FCC1 by arsenic trioxide (ΑΤΟ) Statistics ΑΤΟ dose dose logarithmic infection rate worm reduction rate unit (uM) (X) (%) (%) (丫) 0 0.031 0.063 0.125 0.25 5 0. ο ο ο ο 12.4.8. -1.509 79.9 20.1 4.1619 -1201 73.7 26.3 4.3659 •0.903 68.3 31.7 4.5239 -0.6021 54.9 45.1 4.8743 -0.3010 44.4 55.6 5.1408 0 17.9 82.1 5.9192 0.3010 12.9 87.1 6.1311 0.6021 11.2 88.8 6.2160 0.9031 5.1 94.9 6.6352 Y=1.0904 x+5.6582 "2=0.9706 IC50=O.25uM=49.28ng/ml 95% confidence limit (28.00-86.73ng/ml) IC90=3.73uM=737.94ng/ml 95% confidence limit (confidence limit) (419.28-1298.77ng/ml) 3.7 Test procedure and results (5) 3_ 7.1 Drug preparation: Take the stock solution 32μΙ, add the complete medium 〇_5ml, mix, this is the highest concentration of liquid (320μΜ) 'in the penicillin bottle For half-dilution, a series of concentrations of the drug solution are: 〇, 32〇, 160, 80, 40, 20, 10, 5μΜ. (Total 12 rows, 11 columns for the highest dose hole 32〇_) 3_7, 2 insects Blood dilution: Take continuous culture of FCC1/HN Smear, staining, calculate the infection rate, adjust the infection rate to about 1% with β-type RBC. 3.7.3 Test: 25 200824699 Referring to the method of Guan Weibin, Chen Lin, etc., add liquid per micropore on the 3599 culture plate. 2〇μ|, then add 180μΙ containing worm blood, do 12 lines. Then cover, shake for a few seconds, set at 37 ° C and incubate in 5% C02 box for 48 hours, remove the supernatant, mix, each take RBC 5μΙ , mix, smear and stain microscopy to calculate the number of RBCs infected with Plasmodium per 5000 RBCs (only dead cells with no cytoplasm in the nucleus are not counted) 3.7.4 Results The protozoal infection rate was 1 when the culture started. , 1%, Table 3.9 shows the number of protozoa in every 50 rbc • after culture. Table 3.9 Number of protozoa per 5000 RBCs after culture ABCD Γε FG Η ΑΤΟ Measurement _) 0 5 10 20 40 80 160 320 Mix 256 75 34 21 9 5 0 0 Mix 247 72 39 25 10 4 0 0 Mix 231 69 28 20 9 6 0 0 Mix 258 57 26 17 8 4 0 0 3.7_5 Arsenic trioxide (ΑΤΟ) inhibits FCC1: 3.10 arsenic trioxide (ΑΤΟ) inhibition of FCC1 results statistical table ΑΤΟ dose dose logarithmic infection rate worm reduction rate Rate unit (uM) (X) (%) (%) (丫) 72.6 5.6008 87.1 5.1311 91.5 6,3722 96.3 6.7866 98.0 7.0537 5 6 0 8.3.2. 4 9 72. 2 1 5 0 0 0 0 O.12.4 .06 -0.3010 0 0.3010 0.6021 0.9031 26 200824699 Y=1.1832X+6.0328 "2=0.9849 IC50=0.13uM=25.72ng/ml 95% confidence limit (18.91-34.9ng/m!) lC90=0.96uM =189.93ng/ml 95% confidence limit (139.65_258.3ng/ml) 3.8 Test procedure and results (6) 3.8.1 Preparation of the drug: Take the stock solution 32μΙ, add the complete medium 0.5ml, mix, this is The highest concentration of the drug solution (320 μΜ) was half-diluted in a penicillin bottle to obtain a series of concentrations of the drug solution: 〇, 320, 160, 80, 40, 20, 10, 5 μΜ (doing 1-6 columns, Η behavior) The highest dose hole is 320uM). 3_8·2 dilution of worm blood: Take continuous culture of FCC1/HN, smear, stain, calculate the infection rate, adjust the infection rate to about 1% with sputum type RBC. 3.8.3 Test: According to the method of Guan Weibin, Chen Lin, etc., add 20 μΙ of liquid medicine per micropore on the 3599 culture plate, and then add 180 μΙ containing insect blood to make 12 rows (1-6 for ΑΤΟ, 7-12 for ATS). Then cover and shake for a few seconds, set at 37 ° C and incubate in 5% C 〇 2 box for 48 hours, remove the supernatant, mix, take RBC 5μΙ each, mix, smear and stain microscopic examination, calculate every 5000 The number of RBCs infected with Plasmodium in RBC (only dead cells with no cytoplasm in the nucleus are not counted). 3.8.4 Results: The protozoal infection rate was 1.39% at the start of culture, and Table 3.11 shows the number of protozoa per 5000 RBCs after culture. 27 200824699 Table 3.11 Number of protozoa per 5000 RBCs after culture ABCDEFG Η ΑΤΟ Measurement (uM) 0 5 10 20 40 80 160 320 Mix 420 103 66 38 17 8 0 0 3·8·5 Result statistic: Arsenic trioxide (ΑΤΟ The statistical table of the results of inhibition of FCC1 is shown in Table 3"2. Table 3.12 Arsenic Trioxide (Inhibition Results of FCC1) dose (uM) dose log (X) infection rate (%) worm reduction rate (%) probability unit (丫) 0.5 -0.3010 24.5 75.5 5.6903 1.0 0 15.7 84.3 6.0027 2.0 0.3010 9.0 91.0 6.3408 4.0 0.6021 4.0 96.0 6.7507 8.0 0.9031 1.9 98.1 7.0749 丫=1_5032χ+5·8726 "2=0.9495 IC50=0.32uM=63.31 ng/ml 95% confidence limit (40_32-99.4ng/ml) IC90=2.09uM=413.48ng/ml 95% confidence limit (263.06-258.30ng/ml) 3.9 Conclusion Through several experiments, the IC50 was found to be between 19.78-63.31ng/ml with an average of 35.57ng/ m Bu IC90 is between 185.97-737.94 ng/ml with an average of 451_07 ng/ml. This dose is about 10 times larger than the existing antimalarial drug chloroquine, which is comparable to the dose of piperaquine phosphate. Example 4: In vitro test of arsenic trioxide poisoning African trypanosomiasis 4.1 Test material 28 200824699 4.1_1 Drug: Arsenic trioxide injection (Asadin®, 10 mg/10 ml/vial). 4.1.2 Protozoa: Trypanosoma brucei is labeled "427". The protozoan strain was obtained from Dr. Mary Lee of New York University. 4.1.3 Culture medium: SDM79 (JRH 57453) culture medium containing fetal bovine serum. 4.2 Test method The number and type of living cells were observed by microscope. 4.3 Test procedure The arsenic trioxide was treated at 0.1 mg/ml/day for 3 days, and the poisoning effect on the 7th day was observed. 4.4 Test results Cytology found that the death of the worms was linear rather than a generally small circle. A temporary observation showed that the toxic effect of arsenic trioxide on the cultured Trypanosoma brucei was comparable to that of Melarsoprol. Example 5: In vitro test of arsenic trioxide on apoptosis of human peripheral blood mononuclear leukocytes. 5.1 Test material 5_11 cell line: fresh human peripheral blood mononuclear leukocytes isolated by self. 5_ 1.2 Drug: Arsenic trioxide injection (Asadin 10mg/10ml/vial). 29 200824699 5.1.3 Reagents: white blood cell separation reagent. 5.2 Test methods and procedures 5_2·1 cell line isolation: Human peripheral blood mononuclear leukocytes were isolated from human blood, and the Ficoll Plaque method was used to separate the blood cells from the peripheral blood mononuclear leukocytes. 5.2.2 Cell culture: The isolated human peripheral blood mononuclear leukocytes were cultured in RPM11640 and added with 10% fetal bovine serum, and cultured in 5% CO 2 and 95% air at 37 °C. 5.2_3 Apoptosis assay: After staining by PI staining, it was detected by flowcytometry. 5_2,4 Apoptosis test of human peripheral blood mononuclear leukocytes: combined with single-body antibody fluorescent tandem staining (mAb-fluorescein tandem staining) and Phiphilux treatment, and then classified by flow cytometry. 5. 3 Experimental Results ^ 5.3.1 Arsenic Trioxide Induces Apoptosis of Human Peripheral Blood Mononuclear Leukocytes In the experiment, it was found that arsenic trioxide induced the apoptosis of human peripheral blood mononuclear leukocytes. When different concentrations of arsenic trioxide (0, 0.5, 2.5, 5, or 10 μΜ, respectively) were applied to the peripheral blood mononuclear leukocytes of the whole human body for 24 hours and 48 hours, respectively, the human peripheral blood mononucleus was stained with ΡΙ staining. White blood cells were stained, and the percentage of apoptosis of all human nuclear leukocytes was measured by flow cytometry. 30 200824699 It was shown in the experiment that there was no dose-dependent relationship between arsenic trioxide-induced apoptosis of human peripheral blood mononuclear leukocytes in the two groups treated at 24 and 48 hours (dose-dependent). There is a slightly higher rate of death in the 48-hour group. Perhaps some specific types of human peripheral blood mononuclear white blood cells are more sensitive to arsenic trioxide than other cells. 5.3.2 Differences in arsenic trioxide-sensitive white blood cells using caspase-3 activity Combined with special cell staining patterns, the apoptotic response induced by arsenic trioxide is distinguished by the most significant human peripheral blood mononuclear leukocytes. The results are shown in Table 5.2. The caspase-3 activity test was combined with a special type of cell staining mode to identify the surface of human peripheral blood mononuclear white blood cells (for example, T lymphocytes were identified by "CD3+" and B lymphocytes were identified by "CD19+"). From the test results in Table 5.2, after treatment with arsenic trioxide at a concentration of 5.0 μΜ for 48 hours, the sputum lymphocytes reached 96% of death. Therefore, in the arsenic trioxide-induced apoptosis of specific human peripheral blood mononuclear leukocytes, the axillary lymphocytes have certain sensitivity. Table 5. 2 After treatment with arsenic trioxide for 48 hours, the apoptosis of mononuclear leukocytes induced by specific human peripheral blood was controlled. 34.5 (%) 1 μΜ 36.1 (%) 5 μΜ 96.2 (%) Example 6: Arsenic trioxide on human fibroblasts Apoptosis and in vitro assays affecting cell cycle 6_1 Test material 6.1.1 Cell line: Human fibroblast IMR-90 [IMR90] purchased from ATCC. . 200824699 6·1·2 drugs: arsenic trioxide injection (Asadin®, 10mg/10ml/vial). 6_1_3 reagent: Triton X-1 hydrazine, sodium chloride, EGTA, NP-40, sodium fluoride, Na3V04, aprotinin, leupeptin, PBS, phenylmethyl-sulfonylfluoride (PMSF). 6_2 Test method 6.2.1 Sensitivity of arsenic trioxide to inhibit proliferation of human fibroblastic cell lines 6.1.2.1.1 Cell cycle test with different concentrations of arsenic trioxide (〇, 1.0, 2.5, 5, or 10 μΜ, respectively) After 24 hours of treatment with the human fibroblastic cell line, cell cycle arrest was measured by flow cytometry after staining with propidium iodide staining. 6.2.12 Measurement of cell viability using MTT colorimetry In a 96-well microplate, 5 x 103 cells were added to each well. After the cells were cultured at 37 ° C, 30 μM of MTT solution was added, followed by incubation for 4 hours in the dark. After the Formazan grain was dissolved in DMSO, the absorbance was measured at 570 nm using an ELISA reader. 6.2.13 Observation of apoptotic bodies The pattern of apoptotic bodies was observed by Hoechst staining. The culture broth was removed and fixed for 5 minutes at room temperature in 4% formaldehyde in PBS. Then, Hoechst dye 33258 5Mg/ml in PBS was added for 5 minutes, and after washing off, a reagent (PBS: glycerol = 3:1) was added. The nuclei that were stained with fluorescence were observed using a perfocal microscope. Hoechst 33258 staining was used to observe the morphology of 32 200824699 cells. 6.2.2 Experimental results 6.2.2.1 Arsenic trioxide induced cell cycle arrest in human fibroblasts Treatment of human fibroblastic cell lines with different concentrations of arsenic trioxide (〇, 1·〇, 2.5, 5, 10 μΜ, respectively) After an hour, the cell cycle was analyzed by iodide staining and flow cytometry, and it was found that the cell cycle of human fibroblasts arrested by arsenic trioxide was arrested at G2/M. 6·2_2·2 Arsenic trioxide induced cell death in human fibroblasts with different concentrations of arsenic trioxide (〇, 〇, 2.5, 5, ΙΟμΜ, respectively), after treatment with human fibroblasts for 24 hours, The cell viability was measured by the ruthenium colorimetric method and it was found that arsenic trioxide was cytotoxic to human fibroblasts. At the same time, human fibroblastic cell lines were treated with different concentrations of arsenic trioxide (〇, 1.0, 2.5, 5, 10 μΜ, respectively) for 24 hours, and the cell attenuated bodies were tested by DAPI staining. The results showed that there was no apoptosis in the control group, 1.0 and 2.5 μΜ arsenic trioxide induced cell bodies; 5.0 and 10 μΜ showed obvious apoptotic bodies, and apoptosis and necrosis occurred in human fibroblasts. • 6_2·2.3 Analytical method The test uses the variance analysis of the two elemental factorial design data. The data is done using the SPSS suite of software. The various modifications and variations of the present invention are obvious to those skilled in the art without departing from the scope and spirit of the invention. While the present invention has been described in detail, it is to be understood that the present invention should not be unduly limited to the nature of the particulars. In fact, the various modifications that are obvious to those skilled in the art are also included in the following claims. 33 200824699 [Simplified description of the schema] 益 [Main component symbol description] 200824699 V. Abstract of Chinese invention: Therapeutic application of arsenic-containing arsenic-sensitive embryonic cell-related disease, the present invention relates to the preparation of therapeutic arsenic trioxide Use of allergic diseases, immune diseases 'fibroblast diseases, inflammatory diseases and infectious diseases, especially malaria, African trypanosomiasis 'gas sputum and 1 systemic lupus erythematosus. Sixth, English invention summary: The present invention is related to arsenic compounds for the arsenic-sensitive blast-cell related diseases, such as hypersensitivity diseases, immunologic diseases, fibroblastic diseases, inflammatory diseases and infectious diseases, more particularly, the present invention is related to the treatment for malaria, Trypanosoma, bronchial asthma and systemic lupus erythematosus. 200824699 X. Patent application scope: 丨 疗 含 含 含 含 含 含 含 ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ ^ The use of immunological diseases II, cytopathic, inflammatory and infectious diseases, in particular malaria, African trypanosomiasis, lupus erythematosus. Different modifications and changes are possible according to the invention. It is apparent that the scope and spirit of the invention are not departed from the scope of the invention. It is to be understood that the invention is not to be construed as being limited to the specific details. </ RTI> </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; </ RTI> <RTIgt; Use in a medicament for blast cell disease. 2. Use according to claim 1 wherein the medicament is used in a variety of ways that may be administered to the human or other organisms, including injection, smearing, oral, spraying, etc. φ 3. The use according to claim 1 or 2, wherein the arsenic-sensitive blast cell disease is selected from the group consisting of an allergic disease, an immune disease, a fibroblast disease, an inflammatory disease, and a parasitic disease. The use according to claim 3, wherein the immune disease is selected from the group consisting of connective tissue disease, autoimmune thyroid disease, neuromuscular autoimmune disease, gastrointestinal autoimmune disease, autoimmune hepatitis, and primary hepatobiliary sclerosis. Symptoms, primary sclerosing cholangitis, autoimmune carditis, and arteritis. 5. The use according to claim 4, wherein the immune disease is selected from the group consisting of As follows: Systemic erythema 35 200824699 Lupus, rheumatoid arthritis, sclerosis, Gracies, myasthenia gravis, multiple sclerosis, ulcerative colitis, and CrohrVs disease. The use according to claim 3, wherein the allergic disease is selected from the group consisting of asthma, allergic pneumonia, diffuse pulmonary fibrosis, allergic rhinitis, eosinophilic rhinitis, spring conjunctivitis and urticaria. The use according to claim 3, wherein the inflammatory disease is selected from the group consisting of pneumonia, osteomyelitis, leprosy, syphilis 'tuberculosis, hepatitis, tumor formation and chronic obstructive pulmonary disease. 8. The use according to claim 3, wherein the fibroblastic disease is selected from the group consisting of liver fibrosis, pulmonary fibrosis, skin fibrosis, systemic fibrosis, pneumoconiosis, tuberculosis, atypical pneumonia (SARS), and adults. Respiratory distress syndrome, chronic obstructive pulmonary disease, scarring, swelling, dryness, invasive cystic fibrosis, and neurofibromatosis. 9. The use of claim 3 wherein the parasitic disease is malaria or trypanosomiasis. The use according to claim 1, wherein the arsenic-sensitive blast cells are selected from the group consisting of white blood cells, peripheral blood mononuclear cells, and fibroblasts. An application according to claim 1 wherein the medicament comprises from about 0.001 μΜ to about 20 μΜ of arsenic trioxide. The use according to claim 11, wherein the drug comprises arsenic trioxide of from about 1 μΜ to about 15 μΜ. 13. The use according to claim 12 wherein the drug comprises from about 0.1 μΜ to about 10 μΜ of arsenic trioxide. XI. Schema: None 36