The Protective Effect And Mechanism Of Ginseng Against Myelosuppression Induced By Niraparib

Objective:Poly ADP-ribose Polymerase inhibitor(PARPi)inhibitor has shown good effect in the maintenance treatment of some ovarian cancer,but the myelosuppression caused by it may affect the dosage and course of treatment.Traditional Chinese medicine has been proved to have an effect on myelosuppression caused by drug treatment of malignant tumors,but the concern that drug interactions may affect drug metabolism and efficacy limits the combined use of traditional Chinese medicine and PARPi in clinical practice.Different from the metabolic pathways of other PARPis,niraparib is metabolized through the carboxylesterase pathway,and few cases will happen that cannot be combined with drugs.The purpose of this study was to explore the role of traditional Chinese medicine in the myelosuppression of niraparib benzenesulfonate.Through data mining and clinical research data,it is found that ginseng may have the effect of preventing and treating myelosuppression.The myelosuppression mouse model was established by using niraparib benzenesulfonate,and ginseng was given by gavage at the same time to analyze the preventive and therapeutic effect and mechanism of ginseng on the myelosuppression,and explore the influence of ginseng on the anti-tumor effect of niraparib benzenesulfonate,so as to provide an experimental basis for future clinical application.Method:1.Exploration of the effect of ginseng on myelosuppressionCollect and sort out the clinical studies on traditional Chinese medicine to prevent and treat myelosuppression related to tumor treatment published in Chinese databases(CNKI,Wanfang,VIP)and English databases(Pubmed)from January 1,2012 to May 31,2022.By sorting out and analyzing the data,we can count the four qi,five flavor,meridian tropism frequency and use frequency of drugs,and then conduct the cluster analysis of the prescriptions.Then,we summarize the basic therapeutic principles of traditional Chinese medicine for myelosuppression.According to the principles of traditional Chinese medicine,literature research and clinical experience,we intend to explore the potential role of ginseng in preventing myelosuppression.2.Study on the effect of ginseng in preventing and treating myelosuppression induced by niraparib benzenesulfonate①To explore the conditions of myelosuppression induced by niraparib in mice.Female C57BL/6 mice were randomly divided into three groups:blank group(Control),niraparib low dose group(NIR-L)and niraparib high dose group(NIR-H),with 6 mice in each group.Control group was given 0.1ml/10g distilled water by gavage;NIR-L group and NIR-H group were given niraparib solution by gavage at doses of 40mg/Kg(0.1ml/10g)and 80mg/Kg(0.1ml/10g)respectively,once a day for three consecutive days.The samples were collected at D4 and D11 respectively,and the peripheral blood WBC,NEU,HgB and PLT were counted to assess the change trend of peripheral blood,and the bone marrow of sternum was stained with HE to assess the bone marrow hematopoietic area.②Analysis of the effect of ginseng on preventing and treating myelosuppression.Female C57BL/6 mice were divided into 5 groups:control group,tumor group(Tumor),model group(Model),low-dose ginseng group(GS-L)and high-dose ginseng group(GS-H),with 12 mice in each group.All mice except the control group were injected with ID8 ovarian cancer cells subcutaneously for 21 days to establish the transplanted tumor model.In the morning of D1-14,the mice in Control group,Tumor group and Model group were given 0.1ml/10g distilled water respectively;GS-L group and GS-H group were given ginseng solution at the dose of 100mg/Kg(0.1ml/10g)or 200mg/Kg(0.1ml/10g)respectively;On D5-7 in the afternoon:Control group and Tumor group were given 0.1ml/10g of distilled water,while Model group,GS-L group and GS-H group were given 80mg/Kg(0.1ml/10g)of niraparib solution.Six mice were collected from each group of D8 and D15,and peripheral blood cell count,HE staining of sternal bone marrow tissue,and bone marrow nucleated cell count were performed to evaluate the hematopoietic capacity of mice;Hematopoietic progenitor cells were cultured and counted,and the proliferation activity of bone marrow cells was evaluated by flow cytometry.3.The mechanism of ginseng in preventing and treating myelosuppression induced by niraparib benzenesulfonate①Network pharmacological analysis of ginseng in prevention and treatment of myelosuppression.The active ingredients and target genes of ginseng were collected by searching the traditional Chinese medicine systematic pharmacology database and analysis platform and other organic molecular biological activity databases.The network of"ginseng active ingredient-disease intersection target" was constructed by network pharmacology method to analyze the key targets,biological function and signal pathways of ginseng.②Verification of the mechanism of ginseng in preventing and treating myelosuppression.Immunofluorescence detection was used to assess the expression of γ-H2AX to evaluate the degree of DNA double strand breaks in bone marrow cells.Western blot was used to detect p53,p21,p27,CDK4,cyclinD1,cyclinA2,cyclinB1,cyclinE1,pRb,Rb,γ-H2AX to explore the role of p53 pathway mediated cell cycle pathway in the prevention and treatment of myelosuppression by ginseng.4.To explore the effect of ginseng on the anti-tumor effect of niraparibThe grouping,establishment of tumor model and administration scheme of mice are detailed in ②.The tumor volume and body weight of mice were monitored during the administration period;Tumor tissues of Model group,GS-L group and GS-H group mice were collected on D15 for γ-H2AX and Ki-67 immunohistochemistry to evaluate the effect of ginseng on DNA double strand damage and proliferation activity of tumor.Result1.Exploration of the effect of ginseng on myelosuppressionA total of 160 articles were included in the study,involving 158 traditional Chinese medicines,which are mostly sweet and warm in nature and enter the spleen,liver and kidney meridians.20 core medicines were listed,most of which are Qi tonifying and blood tonifying medicines.According to the cluster analysis,the traditional Chinese medicines can be grouped into three categories:the drugs mainly for supplementing qi and promoting blood circulation;The drugs mainly for tonifying the kidney and spleen;The drugs mainly used to replenish blood and nourish qi.It is concluded that the method of“invigorating spleen,supplementing qi and nourishing blood”plays an important role in the prevention and treatment of myelosuppression.According to the therapeutic principles of traditional Chinese medicine,literature research results and preliminary clinical research results,it is speculated that ginseng may have the effect of prevention and treatment of myelosuppression.2.Study on the effect of ginseng in preventing and treating myelosuppression induced by niraparib benzenesulfonate①To explore the conditions of myelosuppression induced by niraparib in mice.D4 Results:Compared with the control group,the WBC,NEU and HgB in the NIR-H group were significantly lower(P<0.05);Hematopoietic area of bone marrow decreased.D11 Results:Compared with the control group,the HgB value in the NIR-H group was significantly lower(P<0.05),and the WBC,NEU,and bone marrow hematopoietic area were all recovered.It is found no difference in NIR-L and control group in D4 and D11(P>0.05).In conclusion,the dose concentration of NIR-H group can preliminarily establish the myelosuppression model.②Analysis of the effect of ginseng on preventing and treating myelosuppression.D8 Results:Compared with Tumor group,WBC,NEU and HgB of peripheral blood in Model group were significantly lower(P<0.05);The number of nucleated cells in bone marrow decreased significantly(P<0.05);The hematopoietic area of bone marrow decreased.Compared with the model group,the WBC and HgB values were significantly higher(P<0.05)and the cell cycle proliferation index was significantly higher(P<0.05)in GS-L and GS-H groups.In addition,compared with the Model group,the HgB value of GS-H group was significantly higher(P<0.05);The number of nucleated cells in bone marrow was higher(P<0.05);The hematopoietic area of bone marrow was larger,and the number of hematopoietic progenitor cells was in a larger number(P<0.05).D15 Results:Compared with Tumor group,the HgB value of peripheral blood in Model group decreased significantly(P<0.05).Compared with the Model group,the HgB value of peripheral blood in GS-L group and GS-H group was significantly higher(P<0.05).In addition,compared with Model group,GS-H group had more erythroid burst colony forming unit cells(P<0.05);The cell cycle proliferation index was significantly higher(P<0.05).To sum up,ginseng can prevent and treat the myelosuppression induced by niraparib to a certain extent,and the effect is more significant with the increase of concentration.3.The mechanism of ginseng in preventing and treating myelosuppression induced by niraparib benzenesulfonate①Network pharmacological analysis of ginseng in prevention and treatment of myelosuppression.In this study,22 active components of Panax ginseng were obtained;779 potential targets and 1095 genes related to myelosuppression were predicted.Through Wayne analysis,151 cross targets of Panax ginseng for prevention and treatment of myelosuppression were mapped and 20 core target proteins including TP53,SRC,MAPK1 were screened through protein interaction network analysis.Through KEGG analysis,it is indicated that the targets is enriched in the cell cycle pathway.Combined with literature research and network pharmacological analysis,the prevention and treatment of myelosuppression by ginseng may be related to the p53 pathway mediated cell cycle pathway.②Verification of the mechanism of ginseng in preventing and treating myelosuppression.Compared with Tumor group,the expression of γ-H2AX of bone marrow nucleated cells in model group is higher(P<0.05),the expression of p53,p21 and p27 is up-regulated(P<0.05),and the expression of cyclinE1 is down-regulated(P<0.05),suggesting that niraparib may aggravate DNA double strand damage and activate p53 pathway to reduce the activity of bone marrow nucleated cells.Compared with the model group,the expression of y-H2AX,p53 and p27 decreased in GS-L group and GS-H group(P<0.05);In addition,compared with the model group,the expression of p21 was down-regulated(P<0.05),and the expression of pRb and cyclinE1 was up-regulated(P<0.05)in GS-H group.There was no difference in CDK4,cyclinD1,cyclinA2 and cyclinB1 among groups(P>0.05).To sum up,the prevention and treatment of myelosuppression by ginseng may be related to its protective effect of DNA double-strand stability and inhibition of p53 mediated pathways.4.To explore the effect of ginseng on the anti-tumor effect of niraparib.Compared with the Model group,there was no significant difference in tumor volume and body weight(P>0.05)in GS-L group and GS-H group;the expression of Ki-67 expression and y-H2AX had no significant difference(P>0.05)in GS-L group and GS-H group at each time point.To sum up,there is no evidence that ginseng show tumor progression.ConclusionGinseng can prevent and treat myelosuppression caused by niraparib effectively and safely and the mechanism of which may be related to its protective effect on double-strand DNA stability and the inhibition of p53 mediated pathway.