MicroRNA and mRNA profiling of cerebral cortex in a transgenic mouse model of Alzheimer’s disease by RNA sequencing

In a previous study,we found that long non-coding genes in Alzheimer’s disease(AD)are a result of endogenous gene disorders caused by the recruitment of microRNA(miRNA)and mRNA,and that miR-200a-3p and other representative miRNAs can mediate cognitive impairment and thus serve as new biomarkers for AD.In this study,we investigated the abnormal expression of miRNA and mRNA and the pathogenesis of AD at the epigenetic level.To this aim,we performed RNA sequencing and an integrative analysis of the cerebral cortex of the widely used amyloid precursor protein and presenilin-1 double transgenic mouse model of AD.Overall,129 mRNAs and 68 miRNAs were aberrantly expressed.Among these,eight down-regulated miRNAs and seven up-regulated miRNAs appeared as promising noninvasive biomarkers and therapeutic targets.The main enriched signaling pathways involved mitogen-activated kinase protein,phosphatidylinositol 3-kinase-protein kinase B,mechanistic target of rapamycin kinase,forkhead box O,and autophagy.An miRNA-mRNA network between dysregulated miRNAs and corresponding target genes connected with AD progression was also constructed.These miRNAs and mRNAs are potential biomarkers and therapeutic targets for new treatment strategies,early diagnosis,and prevention of AD.The present results provide a novel perspective on the role of miRNAs and mRNAs in AD.This study was approved by the Experimental Animal Care and Use Committee of Institute of Medicinal Biotechnology of Beijing,China(approval No.IMB-201909-D6)on September 6,2019.

Excellent effects and possible mechanisms of action of a new antibody–drug conjugate against EGFR-positive triple-negative breast cancer

Background:Triple-negative breast cancer(TNBC)is the most aggressive subtype and occurs in approximately 15%–20%of diagnosed breast cancers.TNBC is characterized by its highly metastatic and recurrent features,as well as a lack of specific targets and targeted therapeutics.Epidermal growth factor receptor(EGFR)is highly expressed in a variety of tumors,especially in TNBC.LR004-VC-MMAE is a new EGFR-targeting antibody–drug conjugate produced by our laboratory.This study aimed to evaluate its antitumor activities against EGFR-positive TNBC and further studied its possible mechanism of antitumor action.Methods:LR004-VC-MMAE was prepared by coupling a cytotoxic payload(MMAE)to an anti-EGFR antibody(LR004)via a linker,and the drug-to-antibody ratio(DAR)was analyzed by HIC-HPLC.The gene expression of EGFR in a series of breast cancer cell lines was assessed using a publicly available microarray dataset(GSE41313)and Western blotting.MDA-MB-468 and MDA-MB-231 cells were treated with LR004-VC-MMAE(0,0.0066,0.066,0.66,6.6 nmol/L),and the inhibitory effects of LR004-VC-MMAE on cell proliferation were examined by CCK-8 and colony formation.The migration and invasion capacity of MDA-MB-468 and MDA-MB-231 cells were tested at different LR004-VCMMAE concentrations(2.5 and 5 nmol/L)with wound healing and Transwell invasion assays.Flow cytometric analysis and tumorsphere-forming assays were used to detect the killing effects of LR004-VC-MMAE on cancer stem cells(MDA-MB-468 and MDA-MB-231 cells).The mouse xenograft models were also used to evaluate the antitumor efficacy of LR004-VC-MMAE in vivo.Briefly,BALB/c nude mice were subcutaneously inoculated with MDA-MB-468 or MDAMB-231 cells.Then they were randomly divided into 4 groups(n=6 per group)and treated with PBS,naked LR004(10 mg/kg),LR004-VC-MMAE(10 mg/kg),or doxorubicin,respectively.Tumor sizes and the body weights of mice were measured every 4 d.The effects of LR004-VC-MMAE on apoptosis and cell cycle distribution were analyzed by flow cytometry.Western blotting was used to detect the effects of LR004-VC-MMAE on EGFR,ERK,MEK phosphorylation and tumor stemness marker gene expression.Results:LR004-VC-MMAE with a DAR of 4.02 were obtained.The expression of EGFR was found to be significantly higher in TNBC cells compared with non-TNBC cells(P<0.01).LR004-VC-MMAE inhibited the proliferation of EGFRpositive TNBC cells,and the ICvalues of MDA-MB-468 and MDA-MB-231 cells treated with LR004-VC-MMAE for 72 h were(0.13±0.02)nmol/L and(0.66±0.06)nmol/L,respectively,which were significantly lower than that of cells treated with MMAE[(3.20±0.60)nmol/L,P<0.01,and(6.60±0.50)nmol/L,P<0.001].LR004-VC-MMAE effectively inhibited migration and invasion of MDA-MB-468 and MDA-MB-231 cells.Moreover,LR004-VC-MMAE also killed tumor stem cells in EGFR-positive TNBC cells and impaired their tumorsphere-forming ability.In TNBC xenograft models,LR004-VC-MMAE at 10 mg/kg significantly suppressed tumor growth and achieved complete tumor regression on day 36.Surprisingly,tumor recurrence was not observed until the end of the experiment on day 52.In a mechanistic study,we found that LR004-VC-MMAE significantly induced cell apoptosis and cell cycle arrest at G/M phase in MDAMB-468[(34±5)%vs.(12±2)%,P<0.001]and MDA-MB-231[(27±4)%vs.(18±3)%,P<0.01]cells.LR004-VC-MMAE also inhibited the activation of EGFR signaling and the expression of cancer stemness marker genes such as Oct4,Sox2,KLF4 and EpCAM.Conclusions:LR004-VC-MMAE showed effective antitumor activity by inhibiting the activation of EGFR signaling and the expression of cancer stemness marker genes.It might be a promising therapeutic candidate and provides a potential therapeutic avenue for the treatment of EGFR-positive TNBC.

Total synthesis of cajanine and its antiproliferative activity against human hepatoma cells

Cajanine,a constituent of Cajanaus cajan L.,used in traditional Chinese medicine,is a promising drug candidate because of its broad range of bioactivities.However,the total synthesis of cajanine and its derivatives has never been reported.Herein,we report the total synthesis of cajanine in nine steps with an overall yield of 10%together with its analog,longistyline A,in 8%yield.The antiproliferative activity of the two compounds against a human hepatoma cell line is also reported.

The effect of five-knee-point acupuncture combined with herbal package warm compress for knee osteoarthritis

Objective:To observe the therapeutic effect differences among five-knee-point acupuncture combined with Chinese medication package warm compress therapy of Shēntòng Zhúyū Decoction(身痛逐瘀汤 generalized pain stasis-expelling decoction),simple five-knee-point acupuncture and simple Chinese medication package warm compress therapy of Shēntòng Zhúyū Decoction in treating knee osteoarthritis(KOA).Methods:A total of 126 KOA patients were randomized into a five-knee-point acupuncture combined with Chinese medication package warm compress therapy group(combined treatment group),where there were 42 cases,including 28 cases of unilateral KOA and 14 cases of bilateral KOA,totally 56 affected knees involved,a Chinese medication package warm compress therapy group(medication package group,42 cases,including 22 cases of unilateral KOA,20 cases of bilateral KOA,totally 62 affected knees involved)and a five-knee-point acupuncture group(five-knee-point group,42 cases,including 27 cases of unilateral KOA,15 cases of bilateral KOA,totally 57 affected knees involved).The basic health education was provided in all of the groups.Additionally,in the combined treatment group,acupuncture was applied to the five knee points on the affected side for 30 min.The warm compress with herbal package of Shēntòng Zhúyū Decoction was given for 10 to 15 min.In the medication package group,the warm compress with Shēntòng Zhúyū Decoction was exerted on the affected area for 10 to 15 min.In the fiveknee-point group,acupuncture was applied to SP 10,ST 34,EX-LE 2,EX-LE 4 and ST 35 and the needles were retained for 30 min.The treatment in each group was given once a day,consecutively for 2 weeks.Before and after treatment,the visual analogue scale(VAS)and Lysholm knee scale were adopted to evaluate the pain degree and knee joint motor function in KOA patients.The clinical therapeutic effects were evaluated too.Results:A total of 121 cases accomplished the final observation and 5 cases were dropped out in the three groups,in which,2 cases(2 affected knees)were dropped out in the combined treatment group,1 case(2 affected knees)in the medication package group and 2 cases(3 affected knees)in the fiveknee-point group.VAS scores after treatment were all lower than those before treatment in the three groups and the scores of Lysholm knee scale were all higher than those before treatment,indicating the significant differences(all P<0.05).The total effective rate was 98.1%(53/54)in the combined treatment group,which was higher than 86.7%(52/60)in the medication package group and 92.6%(50/54)in the five-knee-point group,indicating the significant differences(all P<0.05).After treatment,VAS score(2.24±1.24)in the combined medication group was lower than(2.48±1.08)in the medication package group and(2.63 ± 1.44)in the five-knee-point group,presenting the significant difference(all P<0.05).The score of Lysholm knee scale was(60.50±13.76)in the combined medication group,higher than(52.23±11.65)in the medication package group and(52.14±11.77)in the five-knee-point group,indicting the significant differences(all P<0.05).Conclusion:Compared with the simple application of Chinese medication package warm compress therapy of Shēntòng Zhúyū Decoction or the five-knee-point acupuncture therapy,five-knee-point acupuncture combined with Shēntòng Zhúyū Decoction relieves the clinical symptoms of KOA patients more effectively and achieves a better clinical therapeutic effect.