Application of 3-indole acetonitrile in preparing medicine for inhibiting novel coronavirus SARS-CoV-2
阅读说明：本技术 3-吲哚乙腈在制备抑制新型冠状病毒SARS-CoV-2药物中的应用 (Application of 3-indole acetonitrile in preparing medicine for inhibiting novel coronavirus SARS-CoV-2 ) 是由 金梅林 回显锋 杨丽 黄坤 张宇飞 赵雪锦 盛锋 张强 孙小美 钟鸣 杨欢欢 于 2021-03-22 设计创作，主要内容包括：本发明属于生物制剂领域,具体涉及3-吲哚乙腈在制备抑制新型冠状病毒SARS-CoV-2药物中的应用。本发明首次发现3-吲哚乙腈在细胞和动物水平均能对新型冠状病毒起抑制作用,并且能缓解机体由于新型冠状病毒感染导致的不良症状,3-吲哚乙腈在小鼠SARS-CoV-2感染模型中有效果,可以显著降低小鼠肺组织中的病毒含量,降低小鼠体重下降程度,提高小鼠的存活率,这使得3-吲哚乙腈具有非常大的临床治疗潜力。(The invention belongs to the field of biological preparations, and particularly relates to application of 3-indole acetonitrile in preparation of a medicine for inhibiting novel coronavirus SARS-CoV-2. The invention discovers for the first time that 3-indole acetonitrile can inhibit novel coronavirus at cell and animal levels and relieve adverse symptoms of organisms caused by infection of the novel coronavirus, and the 3-indole acetonitrile has an effect in a mouse SARS-CoV-2infection model, so that the virus content in lung tissues of mice can be obviously reduced, the weight reduction degree of the mice is reduced, the survival rate of the mice is improved, and the 3-indole acetonitrile has great clinical treatment potential.)
1.3-indole acetonitrile in preparing medicine for inhibiting SARS-CoV-2.
2.3-indole acetonitrile in the preparation of drugs for treating or preventing SARS-CoV-2 infection.
3.3-indole acetonitrile in the preparation of relieving SARS-CoV-2infection after body weight reduction drug application.
4.3-indole acetonitrile in preparing medicine for improving organism survival rate after SARS-CoV-2 infection.
5.3-indole acetonitrile in the preparation of reducing SARS-CoV-2infection in the lung virus content of the drug application.
6.3-indole acetonitrile in the preparation of treatment or prevention of mice SARS-CoV-2infection drug application.
Application of 3-indole acetonitrile in preparing medicine for improving survival rate of mouse SARS-CoV-2infection is provided.
8.3-indole acetonitrile in the preparation of relieving mice SARS-CoV-2infection after body weight reduction drug application.
9.3-indole acetonitrile in the preparation of reducing mouse SARS-CoV-2infection in the organism lung virus content in the drug application.
10. The use according to claim 6 or 7 or 8 or 9, wherein SARS-CoV-2 infected by the mouse is a novel coronavirus SARS-CoV-2/WBP-1 with a collection number of CCTCC NO: v202031.
Technical Field
The invention belongs to the field of biological preparations, and particularly relates to application of 3-indole acetonitrile in preparation of a medicine for inhibiting novel coronavirus SARS-CoV-2.
Background
The novel coronavirus pneumonia (CoVID-19) caused by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) seriously jeopardizes human health and public health safety, and brings huge burden to the world society and economy. Many foreign and domestic scholars speculate that new coronavirus or similar influenza virus will coexist with human for a long time. However, to date, there has been no widely recognized effective new method for treating coronary pneumonia or anti-SARS-CoV-2 virus, and therefore, it is of great importance to research and develop new anti-SARS-CoV-2 preparations or drugs.
At present, a large number of drugs having an inhibitory effect on the new coronavirus at the cellular level have appeared, but no effective therapeutic drugs are still available clinically at present, and some drugs are used but are ineffective. Pauline Maisonasse et al (2020, Hydroxychloroquine use against SARS-CoV-2infection in non-human precursors, nature) published research shows that Hydroxychloroquine (HCQ) has a wide antiviral effect according to previous extensive research, so HCQ is included in many projects for screening anti-SARS-CoV-2 drugs, and when the antiviral effect of Hydroxychloroquine (HCQ) is evaluated in vitro, the effect of resisting SARS-CoV-2 is proved to be really achieved in vitro; in combination with the existing literature report that HCQ can be used as an immunomodulator, the HCQ used by lupus patients can reduce the level of inflammatory cytokines, and the problem of cytokine storm exists when COVID-19 patients are treated; based on these properties, HCQ is thought to be useful in treating COVID-19 and has been used globally for thousands of patients infected with SARS-CoV-2 resulting in COVID-19, but there is no clear data to date that HCQ is effective in treating COVID-19. Thus, Pauline Maisonasse et al established a model of SARS-CoV-2infection in non-human primate (NHP) animals, which model could be used for the analytical evaluation of the early stage of SARS-CoV-2infection in humans, and evaluated the antiviral and therapeutic effects of HCQ in vivo after cynomolgus monkey infection in non-human primate animal models was also evaluated against SARS-CoV-2in vitro, and the results showed that HCQ exhibited anti-SARS-CoV-2 virus activity in VERO E6 cells, but did not have anti-SARS-CoV-2 effect in the reconstituted human airway epithelial model; in a cynomolgus monkey animal model, compared with a placebo, after the cynomolgus monkey is infected with SARS-CoV-2, no matter in the early stage or the middle and late stage of infection, the HCQ alone or the azithromycin is used together, so that the virus load in tissues is not significantly influenced, and the clinical use effect is not obvious; furthermore, HCQ used as a prophylactic prior to SARS-CoV-2infection also failed to provide protection against SARS-CoV-2 infection.
In 2020, WHO announced that a global clinical trial named a conglomerate trial was initiated, and selected 4 drugs most likely to reduce the new coronary mortality at that time, namely reidesavir, hydroxychloroquine, lopinavir and interferon beta 1a, and through the last year of trial, more than 400 hospitals and more than 1 million patients in more than 30 countries worldwide participated in, but the trial results were not ideal because there was no evidence that hydroxychloroquine and lopinavir could reduce the mortality, and the world health organization stopped the trial of these 2 drugs in 6 and 8 months, respectively. Reidesciclovir (Remdesivir GS-5734) is considered as an effective drug against SARS-CoV-2in the early stage of a new coronary outbreak, but the effect of Reidesciclovir is still under debate at present after more than one year of verification. In 10-15 days in 2020, the world health organization declares that the Reidcisvir cannot obviously reduce the mortality rate of the new coronary virus and improve the treatment process of the patient, and recommends not to use the Reidcisvir to treat the hospitalized patient with the new coronary pneumonia, but in 22 days in 10 months, Reidcisvir research and development company, United states Gilidd science company announces that the Reidcisvir can shorten the hospitalization time of the hospitalized patient with the new coronary pneumonia by 5 days, and becomes the first new coronary treatment drug approved by FDA in the United states. Random double-blind multi-center clinical trials of Reid Wer were conducted in 10 hospitals including Nitrocan Hospital, in the most severe period of new coronary outbreaks in China, and as a result, Reid Wer was found to be indistinguishable in overall time and survival improvement of clinical course, and relevant data was published on famous journal lancets (Wang Yomingtet. Remdesier in adults with segment COVID-19: a randomised, double-blind, placebo-controlled, multicentre trial [ J ] Lancet,2020, 20200516, 39510236 (10236)). These results indicate that the pathogenesis and therapeutic strategy of the novel coronavirus, a new infectious agent, are not well defined at present, and it is not easy to find an antiviral effective drug.
3-Indoleacetonitrile (3-Indoleacetonitrile) is an indole compound isolated from cabbage leaves in E.R.H.JONES, 1952 and demonstrated its structure as a plant growth hormone, the first auxin obtained in crystalline form from higher plant tissues, abbreviated IAN. The most neutral plant growth hormone of cruciferous plants is 3-indolylacetonitrile, and the American society for oncology recommends that cruciferous vegetables (including cauliflower, cabbage, Chinese cabbage, radish, etc.) are eaten more frequently to reduce the incidence of tumors. Among them, 3-substituted indoles and isothiocyanates are considered as two main anticancer components existing in the form of glycosides in cruciferous vegetables. The 3-substituted indole compounds mainly comprise: indole-3-methanol (Indole-3-carbinol, I3C); 3,3' -Diindolylmethane (DIM) and 3-Indolacetonitrile (IDA).
Researches show that the indole compounds can effectively inhibit bacterial pathogenic bacteria such as escherichia coli and the like, and indole and derivatives thereof are regarded as potential resistant compounds of antibiotic drug-resistant pathogens, but at present, no relevant report about 3-indole acetonitrile for resisting SARS-CoV-2 virus is found.
Disclosure of Invention
The invention aims to make up the defects of the prior art and provides a small molecular compound 3-indolacetonitrile in the preparation of a medicine for treating or preventing novel coronavirus SARS-CoV-2infectionThe application of the compound in the medicine can provide an effective small molecular compound for clinically treating SARS-CoV-2. The molecular formula of the 3-indolylacetonitrile is C10H8N2Molecular weight of 156.18, and structural formula as follows:
in order to achieve the purpose, the invention adopts the following technical measures:
the application of 3-indole acetonitrile in preparing medicine for treating or preventing SARS-CoV-2infection includes:
the application of 3-indole acetonitrile in preparing medicine for inhibiting SARS-CoV-2;
the application of 3-indole acetonitrile in preparing medicine for treating or preventing SARS-CoV-2 infection;
the application of 3-indole acetonitrile in preparing medicine for relieving body weight reduction after SARS-CoV-2 infection;
the application of 3-indole acetonitrile in preparing medicine for improving the survival rate of the body after SARS-CoV-2 infection;
the application of 3-indole acetonitrile in preparing medicine for reducing virus content in lung of organism after SARS-CoV-2 infection.
The application of 3-indole acetonitrile in preparing medicine for treating or preventing mouse SARS-CoV-2 infection;
the application of 3-indole acetonitrile in preparing medicine for improving the survival rate of mice SARS-CoV-2 infection;
the application of 3-indole acetonitrile in preparing medicine for relieving body weight reduction after mouse SARS-CoV-2 infection;
or 3-indole acetonitrile in preparing medicine for reducing virus content in lung of body after SARS-CoV-2infection of mouse.
In the above applications, preferably, SARS-CoV-2in mouse SARS-CoV-2infection is a novel coronavirus SARS-CoV-2/WBP-1, with the collection number being CCTCC NO: v202031.
Compared with the prior art, the invention has the following advantages:
1. the 3-indole acetonitrile is a small molecular compound, has small toxic effect on Caco-2 cells, can obviously inhibit infection of SARS-CoV-2 on the Caco-2 cells at the in vitro cell level, and inhibits the proliferation of the SARS-CoV-2 cells in the cells, thereby showing that the indole acetonitrile has the characteristics of safety and high efficiency.
2. The 3-indole acetonitrile has effect in a mouse SARS-CoV-2infection model, can obviously reduce the virus content in mouse lung tissues, reduce the weight loss degree of the mouse and improve the survival rate of the mouse, so that the 3-indole acetonitrile has great clinical treatment potential.
Drawings
FIG. 1 is a schematic representation of the cytotoxicity of 3-indolylacetonitrile on Caco-2 cells.
FIG. 2 is a schematic diagram showing the effect of Western Blot detection of 3-indolylacetonitrile on cell level against SARS-CoV-2;
wherein the upper panel shows the effect of different concentrations of 3-indolylacetonitrile on the proliferation of SARS-CoV-2 at the cellular level, the weaker the bands indicate the lower the viral content, GADPH is an intracellular reference control, and the lanes show working concentrations of 3-indolylacetonitrile from left to right as 0. mu.M, 75. mu.M, 150. mu.M, 300. mu.M, respectively;
the lower graph shows the ratio of the intensity of SARS-CoV-2 Nucleoprotein (NP) band to the intensity of intracellular reference (GADPH) band, with smaller ratios indicating lower virus content and the horizontal coordinate values from left to right indicating working concentrations of 3-indolylacetonitrile of 0. mu.M, 75. mu.M, 150. mu.M and 300. mu.M, respectively.
FIG. 3 is a schematic diagram showing the effect of 3-indoleacetonitrile on cell level anti-SARS-CoV-2 plaque assay.
FIG. 4 is a graph showing the effect of 3-indolylacetonitrile on the body weight of mice infected with SARS-CoV-2 virus;
wherein the dotted lines represent the SARS-CoV-2 virus-uninfected PBS group and the 3-indolylacetonitrile group.
FIG. 5 is a graph showing the effect of 3-indolylacetonitrile on the survival rate of mice infected with SARS-CoV-2 virus.
Wherein mice in the PBS group and the 3-indolylacetonitrile group which are not infected with the new coronavirus do not die, the survival rate is 100 percent, and lines are overlapped.
FIG. 6 is a schematic illustration of the effect of 3-indolylacetonitrile on the viral load of lung tissue of mice infected with SARS-CoV-2 virus.
FIG. 7 shows the effect of 3-indolylacetonitrile on the pathological changes of lung tissue of mice infected with SARS-CoV-2 virus.
Detailed Description
For a better understanding of the present disclosure, the following further description is provided in conjunction with the specific embodiments, but the present disclosure is not limited to the following examples. Unless otherwise specified, the test methods and conditions in the examples of the present invention are conventional methods. The technical scheme of the invention is a conventional scheme in the field if no special description exists; the reagents or materials are commercially available, unless otherwise specified. All tests related to the live SARS-CoV-2 virus were performed in the biosafety third-level laboratory (ABSL 3).
Currently, antiviral drug evaluation models are largely classified into an in vitro model (in vitro model) and an in vivo model (in vivo model).
The in vitro model mainly uses various cell lines to evaluate the medicine, and has the advantages of providing a large number of cells with the same genetic character as research objects, being convenient to operate, eliminating the influence of other external factors, detecting the toxicity, effective concentration and the like of the medicine, and providing more bases for later mechanism research. In vivo models various models of animal infection are generally used, and the overall effect of the drug in living animals is measured by various phenotypic indicators after drug treatment. The method has the advantage of being capable of carrying out real and systematic evaluation on the effect of the candidate drug in the living body. The invention adopts human colon cancer cell line Caco-2 to measure the in vitro anti-SARS-CoV-2 effect of 3-indole acetonitrile; the in vivo anti-SARS-CoV-2 effect of 3-indolylacetonitrile was evaluated by using a mouse adaptive strain WBP-1 infection model.
Experimental materials:
(1) cell line, experimental animal and virus required by experiment
Cell line: caco-2 cells were stored in the university of Huazhong agriculture laboratory;
experimental animals: SPF grade 6 week old Balb/c mice purchased from the Experimental animals center of the university of three gorges;
strain: the novel coronavirus SARS-CoV-2/WBP-1 has the preservation number of CCTCC NO: v202031, CN111961653A, which in the present invention is abbreviated as novel coronavirus WBP-1 or mouse adapted strain WBP-1;
(2) medicine required by experiment
3-Indolylacetonitrile was purchased from Sigma.
(3) Reagents required for the experiment:
DMEM medium, Fetal Bovine Serum (FBS) were purchased from GIBCO;
cell activity detection kit: TransDetect Cell Counting Kit (CCK) was purchased from Beijing Quanji gold organisms.
Example 1:
toxicity test of 3-indolylacetonitrile on cells
Caco-2 cells were used in the present invention, and 96-well plates were seeded until the cell density reached 80%, and the cells were incubated with 3-indolylacetone (final concentration: 0. mu.M, 3.5. mu.M, 7. mu.M, 17.5. mu.M, 35. mu.M, 70. mu.M, 175. mu.M, 350. mu.M, 700. mu.M) for 48 hours. Cell activity was measured using a TransDetect cell counting cassette, and absorbance at OD450nm was measured according to the kit instructions. The specific implementation process is as follows:
1. cell culture
After 2 passages, the frozen and recovered Caco-2 cells are subjected to expanded culture by using a DMEM medium containing 10% fetal calf serum and double antibodies (penicillin 100U/ml and streptomycin 100 ug/ml).
2. Toxicity test of 3-indolylacetonitrile on cells
Digesting and passaging Caco-2 cells which grow well, and adjusting the cell density to be 2 multiplied by 10 by using cell growth liquid (DMEM culture medium + 10% fetal calf serum + double antibody)6Inoculating 96-well plates with each well being 100 mul; adding 100 μ l of 3-indolylacetonitrile prepared from culture medium (DMEM medium + 10% serum + double antibody) into each well, and mixing. The drug is set to 6 concentration gradients, each gradient is set to 3 multiple wells, and the final concentration is 0 μ M, 3.5 μ M, 7 μ M, 17.5 μ M, 35 μ M, 70 μ M, 175 μ M, 350 μ M and 700 μ M. Setting cell control at the same time, placing at 37 deg.C and 5% CO2Culturing in an incubator. Culture No. 4After 8h, cell activity was measured using a TransDetect cell counting cassette, 10. mu.l of a reagent containing CCK was added to the cells in the 96-well plate after the culture was treated, the cells were incubated at 37 ℃ for 2h in the absence of light, and OD450nm reading was detected by a microplate reader to calculate the cell survival rate.
Cell viability (%) ═ drug treated group OD450 nm/untreated control group OD450nm × 100%
And (3) test results:
the results are shown in FIG. 1, and the cell viability is measured to reflect the toxic effect of 3-indolylacetone on Caco-2 cells, and it can be seen from the figure that the cell viability is maintained above 88% compared with the control group after 3-indolylacetone is treated for 48h at the concentration of 350. mu.M, which shows that the cytotoxicity of 3-indolylacetone is lower at the concentration.
Example 2:
inhibition of novel coronaviruses at cellular level by 3-indolylacetonitrile:
western Blot and plaque forming unit (plaque forming unit) are adopted to detect the influence of 3-indole acetonitrile on the proliferation of SARS-COV-2 virus (WBP-1) at the cellular level, and the specific steps are as follows:
1. 3-Indolylacetonitrile treatment of cells and infection with New coronavirus WBP-1
1) Digesting and passaging Caco-2 cells with good growth state, and adjusting cell density to 1 × 10 with cell culture medium5Per ml, 1ml per well was seeded in 12-well plates up to a monolayer.
2) Adding 3-indolylacetonitrile with different contents, wherein the final acting concentrations of the 3-indolylacetonitrile are 0 mu M, 75 mu M, 150 mu M and 300 mu M respectively, and the working concentration of 0 mu M is the no-drug control.
3) After the cells are treated for 24 hours by using 3-indolylacetonitrile with different concentrations, the cells are infected with new coronavirus, the virus (MOI is 1) is incubated for 1 hour, virus liquid is discarded, and the cells are continuously cultured by using the DMEM cell maintenance liquid containing the 3-indolylacetonitrile with different concentrations.
4) And collecting supernatant and cell lysate 24 hours after virus infection, detecting SARS-COV-2 virus NP protein in the lysate by Western Blot, and detecting the virus titer in the supernatant by a plaque experiment.
2. Western Blot for detecting proliferation of SARS-COV-2 on cells under action of 3-indolylacetonitrile
The Western Blotting experiment is adopted to detect the influence of 3-indole acetonitrile on the proliferation of the new coronavirus in a cell lysate infected with the new coronavirus, and the specific steps are as follows:
1) adding 5x protein loading into the collected virus infected cell lysate, boiling in boiling water for 10min, and centrifuging for standby.
2) SDS-PAGE electrophoresis: preparing 10% polyamide gel as separation gel, performing sample application electrophoresis, keeping constant pressure of concentrated gel at 80V for 30min, and keeping constant pressure of separation gel at 120V for 1-1.5 h.
3) Film transfer: and pre-cooling the membrane transferring liquid in advance during membrane transferring, putting the PAGE gel subjected to electrophoresis into a membrane transferring groove, and transferring the membrane for 1h at constant pressure of 100V.
4) And (3) sealing: and taking out the NC membrane after the membrane is transferred, and sealing the NC membrane for 1h by using sealing liquid.
5) After the primary antibody and secondary antibody incubation is finished, color development is carried out: color development was performed using Thermo's ECL color development kit.
3. Determination of virus titer in new coronavirus proliferation liquid by plaque experiment
1) The Caco-2 cells with good growth state are subcultured to 12-well cell culture plates at 3X 105/ml～5×105Ml/well, the next day a dense monolayer was grown.
2) Continuously diluting cell culture supernatant collected after the new coronavirus infected cells by 10 times by using serum-free DMEM medium in a 24-well plate, and specifically comprising the following steps:
(1) 900 μ l DMEM was added to each well.
(2) Adding 100 μ l of the supernatant obtained in step 1, 4) to the first row of wells, i.e., the total volume of the liquid in each well is 1000 μ l, and mixing by gentle shaking.
(3) Taking 100 mu l of the mixture from the first row to the second row by using a row gun, blowing, beating, shaking and uniformly mixing; the tip was changed and 100. mu.l was taken from the second row to the third row, and the operation was repeated until the sixth row.
3) The culture medium in the Caco-2 cell culture plate is aspirated, and 500 mul of virus diluent is added; the mixture was incubated at 37 ℃ for 1 hour in an incubator, and the mixture was aspirated, 1ml of DMEM + 2.5% FBS + 0.9% methyl cellulose was added to each well, and the mixture was placed in the incubator for 3 days.
4) Add 1ml 8% formaldehydee-PBS per well and fix for more than 1 h.
5) The fixative was poured off and washed once with single distilled water.
6) Adding 0.5% crystal violet for dyeing for 10min, and washing with single distilled water for three times to obtain several plaques.
And (3) test results:
the results are shown in FIG. 2, 3-indolylacetonitrile significantly inhibited the replication of the new coronavirus. After virus infection, the grey value of the virus NP protein in the 3-indolylacetone treatment cell sap with the concentration of 300 μ M is significantly lower than that of the control group without the added medicine, and it can be known from FIG. 2 that the inhibition effect of the 3-indolylacetone on the novel coronavirus is dose-dependent, and the inhibition effect on the virus is better with the increase of the medicine concentration. In addition, the plaque counting result also shows that the 3-indole acetonitrile has a significant inhibition effect on the proliferation of the new coronavirus, and the inhibition effect on the proliferation of the new coronavirus is correspondingly improved along with the increase of the dosage of the 3-indole acetonitrile (table 1 and figure 3).
TABLE 1 Effect of different concentrations of 3-indolylacetonitrile on the proliferation of SARS-CoV-2 Virus
3-Indolylacetonitrile (μ M)
SARS-CoV-2 content (PFU/ml)
0
2.07×105
75
1.40×105
150
7.83×104
300
1.00×104
Example 3:
evaluation of antiviral Effect of 3-Indolylacetonitrile in mouse lethal infection model
Mouse experiment procedure:
1) dividing female BALB/c mice of 6 weeks old into 2 big groups randomly, wherein the first big group is a group without infecting new coronavirus, and the first big group is a control group (PBS) (5 mice) and a 3-indolylacetonitrile group (5 mice) respectively; the second major group is the SARS-CoV-2 virus infection group, which is PBS + SARS-CoV-2infection group (10), 3-indolylacetonitrile + SARS-CoV-2infection group (10).
2) 3-indole acetonitrile is prepared into 4mg/mL aqueous solution, and each mouse is injected with PBS or 3-indole acetonitrile solution 100 mu L in tail vein.
3) 2 hours before SARS-CoV-2infection, 3-indolylacetone (3-indolylacetone + SARS-CoV-2infection group) or PBS (PBS + SARS-CoV-2infection group) is injected into tail vein once, and then 5 half of lethal dose (5LD50) The novel coronavirus mouse adapted strain SARS-CoV-2/WBP-1(CCTCC NO: v202031), the infectious dose is 50 μ l/mouse, and the mice are injected with the drug (3-indolylacetonitril + SARS-CoV-2 infected group) or PBS (PBS + SARS-CoV-2 infected group) by tail vein once every other day after being infected with the new coronavirus for 4 times in total. And mice were observed and recorded daily for weight and survival after infection with new coronavirus.
4) Randomly dividing 20 female BALB/c mice with age of 6 weeks into 2 groups, namely a PBS infected group (10 mice) and a 3-indolylacetonitrile infected group (10 mice), wherein administration before infection and infection operation are the same, the test group is injected with 3-indolylacetonitrile at intervals of one day caudal vein after being infected with new coronavirus, the control group is injected with PBS at intervals of one day caudal vein after being infected with new coronavirus, the total number is 2, the injection dose of each mouse is 100 mu L, a lung sample is taken from the dissected mouse on the 5 th day after being infected with SARS-CoV-2 virus, the collected lung tissue sample is cut into two halves, 1mL of sterile PBS (1mL PBS/lung) is added into one half of the lung tissue sample, a homogenizer is used for homogenizing and breaking, and 0.1mL of homogenate is taken for extracting RNA for virus load detection; the other half of the lung tissue samples were fixed with tissue fixative and prepared into histopathological sections for histopathological analysis.
5) And drawing a weight change curve, a survival rate curve and a tissue virus vector graph according to the statistical result.
And (3) test results:
the results of the weight change of the mice are shown in fig. 4 and table 2, and in the mice without virus infection, the weight of the mice in the 3-indolylacetone injection group is not obviously different from that of the mice in the PBS group, which indicates that the 3-indolacetone has low toxicity in animals. In mice infected with SARS-CoV-2 virus, after administration, the average weight loss of the mice in PBS group was higher than that of the mice in 3-indolylacetonitrile treated group at day 3, and on day 7 after infection, all the mice in PBS group died, and the weight loss was more than 30%. However, the 3-indolylacetonitrile-treated mice started on day 7 post-infection, the weight loss slowed, the weight loss stopped on day 9, and the weight began to rise again by day 10 post-infection.
TABLE 23 Effect of indolylacetonitrile on the body weight of mice infected with SARS-CoV-2 Virus
As shown in fig. 5 and table 3, the survival rate of the virus-infected control mice was all dead at day 7 after infection, while the survival rate of the 3-indolylacetonitrile-treated mice was 40% after infection with neocorona.
TABLE 33 Effect of Indolylacetonitrile on the survival of mice infected with SARS-CoV-2 Virus
The results of the lung tissue viral load test are shown in fig. 6: the 3-indole acetonitrile can obviously reduce the virus content in the lung tissue after the organism is infected with the novel coronavirus SARS-CoV-2.
On day 5 after infection of mice with SARS-CoV-2 virus, severe inflammatory cell infiltration and pulmonary congestion were observed in the lungs of control mice, whereas the lungs of mice injected with indole-3-acetonitrile via tail vein were significantly attenuated (FIG. 7), and thus, the pathological lesions of lungs of mice infected with the new coronavirus were alleviated by 3-indolacetone via tail vein injection.
The results show that the 3-indolylacetonitrile has obvious antiviral effect in mice. In view of the above results, 3-indolylacetonitrile can be used for the preparation of a medicament for the treatment of SARS-CoV-2 infection.