1.Immunogenicity of a novel vaccine with VLP as vector for Wilm's tumor
Zheng WANG ; Yueqiang DUAN ; Deyan LUO ; Xiliang WANG ; Zhoulu LIU
Chinese Journal of General Surgery 2008;23(6):457-459
Objective To develop a vaccine for Wilm's tumor in order to lay a foundation for effective treatment and substitute for the radiotherapy and chemotherapy of this tumor. Methods DNA fragments encoding the WT1 antigens (including, HLA-A * 2402 and HLA-A * 2402x), and DNA fragments encoding the couple antigens HLA-A * 2402-HBc and HLA-A * 2402x-HBc were cloned and inserted into the eukaryotic expression vector pcDNA3.1(+) in sense orientation and baculovirus expression pFactbac respectively, then transfected into COS-7 cells and Sf 9 cell respectively. The expression of the target proteins in the cells were identified by CPE, SDS-PAGE and electron microscopy. Virus-like particles were observed under electron microscopy. Results The fusion gene of HLA-A * 2402 and HLA-A * 2402x and HBc express virus-like particle protein. Conclusions This preliminary result shows a hopeful future in developing an effective immunotherapy to Wilm's tumor.
2.Preparation of the Coxsackievirus A16 VP1 subunit vaccine and detection of its immunogenicity
Xiaonan LI ; Deyan LUO ; Zhongpeng ZHAO ; Yueqiang DUAN ; Peifeng LI ; Xiliang WANG
Chinese Journal of Microbiology and Immunology 2010;30(3):250-255
Objective To prepare VP1 protein vaccine of Coxsackievirus A16(CA16) and evalu-ate immunngenicity the subunit vaccines of Coxsackievirus (VP1), and to establish foundation for studying CA16 vaccine. Methods CA16 VP1 was amplified by RT-PCR and cloned into pFastBac HT A plasmid, recombinated with Bacmid DNA by transposition reaction and then transfected Sf9 cell, mixed with adjuvant AI(OH)_3. After immunization BALB/c mice, evaluating immune effectiveness after booster injections 2 weeks. Results The expressed protein was analyzed by SDS-PAGE and Western blot, mice immunized with CA16 (VP1) both induced specific IgG antibody and neutralization antibody. The best immunization antigen was 20 μg, IgG antibody was 1: 1600, neutralization antibody was 1:250, typical Th1/Th2 immune response was determined by lymphocyte proliferation assay and cytokine analysis. Conclusion The CA16 VP1 gene was cloned successfully and expressed in Sf9 insect cells, CA16 VP1 protein vaccine induced both humoral and cellular immune response, to lay solid foundation for further study on CA16 vaccine.
3.Generation of cold-adapted, attenuated reassortant human influenza A virus with eight-plasmid system
Penghui YANG ; Xinfu SHI ; Yan YAN ; Deyan LUO ; Yu ZHANG ; Li XING ; Jinxue LONG ; Xiufan LIU ; Xiliang WANG
Chinese Journal of Microbiology and Immunology 2008;28(2):139-143
Objective To set up a technical platform of reverse genetics based on the 8 plasmid.virus rescue system of cold-adapted influenza virus strain. Methods The cold-adapted, temperature sensitive, live attenuated influenza virus strain A/AnnArbor/6/60(H2N2)was chosen as the master donor virus(MDV)for rescue research,and its six internal gene fragments PB2,PB1,PA,NP,M and NS were artificially synthesized. Meanwhile, five amino acid mutations have been introduced as tags. Six fragments were ligated with modified pAD3000 for the construction of rescue plasmid. Six transcription/expression plasmids(pMDV-A-PB2,pMDV-A-PB1,pMDV-A-PA,pMDV-A-NP,pMDV-A-M,and pMDV-A-NS)were obtained, and their sequences were accurate. Results The reassorted virus named as rMDV-A contains HA and NA gene segments derived from PR8 strain along with six gene segments,PB2,PB1,PA,NP,M and NS,from MDV. The COS-1 cells were co-transfected with eight recombinant plasmids. The results showed that a cold-adapted, attenuated reassortant influenza A virus with hemagglutination activity was rescued successfullv bv"6+2" combination of MDV and PR8, and the allanotoic fluid of the injected eggs gave a posigenes of A/AA/6/60 used as backbone has provided experimental materials for further research on the gene function and novel vaccine candidate of cold-adapted, attenuated human influenza virus.
4.Development and Identification of a Live Attenuated Influenza B Virus Vaccine Candidate
Penghui YANG ; Wenqi AN ; Xinfu SHI ; Yueqiang DUAN ; Deyan LUO ; Pengfei ZHANG ; Chong TANG ; Li XING ; Yujing ZHANG ; Xiufan LIU ; Xiliang WANG
Progress in Biochemistry and Biophysics 2009;36(3):358-363
A cold-adapted (ca), temperature sensitive (ts), live attenuated influenza B virus strain B/Ann Arbor/1/66 was chosen for influenza virus rescue research, in which six internal gene segments, PB1, PB2, PA, NP, M, NS, were fully synthesized and nine amino acid substitutions were artificially alter by human intervention. The resultant B/Ann Arbor/1/66 plasmids were named as pAB121-PB1, pAB122-PB2, pAB123-PA, pAB124-HA, pAB125-NP, pAB126-NA, pAB127-M and pAB128-NS, respectively. A recombinant influenza A virus was previously generated entirely from cloned cDNA. An infectious recombinant influenza B virus was generated here, and designated as rMDV-B, by plasmid-based reverse genetics. The rMDV-B virus contained HA and NA genes from an epidemic influenza B vires strain B/Malaysia/2506/2004 in the background of internal genes derived from influenza B virus strain B/Ann Arbor/1/66. HA titer of rMDV-B in MDCK cells and embryonated chicken eggs ranged from 1 : 64 to 1 : 512. The results may allow an effective live influenza B vaccine to be produced from a single master strain, providing a model for the design of future live human influenza vaccines.
5.Construction of Attenuated Influenza Virus Vaccines of Vaccines of-2006-2007
Penghui YANG ; Yi YE ; Xinfu SHI ; Deyan LUO ; Yu ZHANG ; Yan YAN ; Pengfei ZHANG ; Yueqiang DUAN ; Li XING ; Xiufan LIU ; Xiliang WANG
Progress in Biochemistry and Biophysics 2008;35(3):312-319
Six gene segments,PB1,PB2,PA,NP,M and NS,were fully synthesized which derived from the master donor virus (MDV),cold-adapted(ca),temperature sensitive(ts),live attenuated influenza virus strain A/Ann Arbor/6/60(MDV-A).Meanwhile,five amino acid substitutions (PB1-391E,58lG,661T,PB2-265S,NP-34G) were artificially altered by human intervention.HA and NA fragments derived from the 2006-2007 circulating strain A/New Caledonia/20/99 (H1N1).Eight fragments were ligated with modified pAD3000 for rescue plasmid construction.Eiight transcription/expression plasmids were named as pMDV-A-PB2,pMDV-A-PB1,pMDV-A-PA,pMDV-A-NP,pMDV-A-M,pMDV-A-NS,pMDV-A-HA,pMDV-A-NA,respectively.The COS-l cells were co-transfected with eight plasmids representing 6 internal viral backbone of the strain A/AA/6/60 and two plasmids containing the CDNA of the HA and NA segments of the strain A/New Caledonia/20/99 (H1N1),the results showed that cold-adapted,attenuated reassortant influenza A virus Was rescued successfully.Titers of a reassorted influenza A virus in embryonated chicken eggs mnged from 1:29to l:210.The rescue system of six intemal genes used as backbone opens the way for further research on gene function and neotype vaccine candidate of cold-adapted,live attenuated human influenza virus.
6.Construction of Attenuated Influenza Virus Vaccines of 2006~2007
Penghui YANG ; Yi YE ; Xinfu SHI ; Deyan LUO ; Yu ZHANG ; Yan YAN ; Pengfei ZHANG ; Yueqiang DUAN ; Li XING ; Xiufan LIU ; Xiliang WANG
Progress in Biochemistry and Biophysics 2006;0(03):-
Six gene segments, PB1, PB2,PA, NP, M and NS, were fully synthesized which derived from the master donor virus(MDV), cold-adapted(ca),temperature sensitive(ts), live attenuated influenza virus strain A/Ann Arbor/6/60(MDV-A). Meanwhile, five amino acid substitutions (PB1-391E, 581G, 661T, PB2-265S, NP-34G) were artificially altered by human intervention. HA and NA fragments derived from the 2006~2007 circulating strain A/New Caledonia/20/99 (H1N1). Eight fragments were ligated with modified pAD3000 for rescue plasmid construction. Eight transcription/expression plasmids were named as pMDV-A-PB2, pMDV-A-PB1, pMDV-A-PA, pMDV-A-NP, pMDV-A-M, pMDV-A-NS, pMDV-A-HA, pMDV-A-NA, respectively. The COS-1 cells were co-transfected with eight plasmids representing 6 internal viral backbone of the strain A/AA/6/60 and two plasmids containing the cDNA of the HA and NA segments of the strain A/New Caledonia/20/99 (H1N1), the results showed that cold-adapted, attenuated reassortant influenza A virus was rescued successfully. Titers of a reassorted influenza A virus in embryonated chicken eggs ranged from 1∶29 to 1∶210. The rescue system of six internal genes used as backbone opens the way for further research on gene function and neotype vaccine candidate of cold-adapted, live attenuated human influenza virus.
7.Discovery of highly selective and orally available benzimidazole-based phosphodiesterase 10 inhibitors with improved solubility and pharmacokinetic properties for treatment of pulmonary arterial hypertension.
Yuncong YANG ; Sirui ZHANG ; Qian ZHOU ; Chen ZHANG ; Yuqi GAO ; Hao WANG ; Zhe LI ; Deyan WU ; Yinuo WU ; Yi-You HUANG ; Lei GUO ; Hai-Bin LUO
Acta Pharmaceutica Sinica B 2020;10(12):2339-2347
Optimization efforts were devoted to discover novel PDE10A inhibitors in order to improve solubility and pharmacokinetics properties for a long-term therapy against pulmonary arterial hypertension (PAH) starting from the previously synthesized inhibitor
8.A novel inhibitor of N 6-methyladenosine demethylase FTO induces mRNA methylation and shows anti-cancer activities.
Guoyou XIE ; Xu-Nian WU ; Yuyi LING ; Yalan RUI ; Deyan WU ; Jiawang ZHOU ; Jiexin LI ; Shuibin LIN ; Qin PENG ; Zigang LI ; Hongsheng WANG ; Hai-Bin LUO
Acta Pharmaceutica Sinica B 2022;12(2):853-866
N 6-methyladenosine (m6A) modification is critical for mRNA splicing, nuclear export, stability and translation. Fat mass and obesity-associated protein (FTO), the first identified m6A demethylase, is critical for cancer progression. Herein, we developed small-molecule inhibitors of FTO by virtual screening, structural optimization, and bioassay. As a result, two FTO inhibitors namely 18077 and 18097 were identified, which can selectively inhibit demethylase activity of FTO. Specifically, 18097 bound to the active site of FTO and then inhibited cell cycle process and migration of cancer cells. In addition, 18097 reprogrammed the epi-transcriptome of breast cancer cells, particularly for genes related to P53 pathway. 18097 increased the abundance of m6A modification of suppressor of cytokine signaling 1 (SOCS1) mRNA, which recruited IGF2BP1 to increase mRNA stability of SOCS1 and subsequently activated the P53 signaling pathway. Further, 18097 suppressed cellular lipogenesis via downregulation of peroxisome proliferator-activated receptor gamma (PPARγ), CCAAT/enhancer-binding protein alpha (C/EBPα), and C/EBPβ. Animal studies confirmed that 18097 can significantly suppress in vivo growth and lung colonization of breast cancer cells. Collectively, we identified that FTO can work as a potential drug target and the small-molecule inhibitor 18097 can serve as a potential agent against breast cancer.
9.Free energy perturbation (FEP)-guided scaffold hopping.
Deyan WU ; Xuehua ZHENG ; Runduo LIU ; Zhe LI ; Zan JIANG ; Qian ZHOU ; Yue HUANG ; Xu-Nian WU ; Chen ZHANG ; Yi-You HUANG ; Hai-Bin LUO
Acta Pharmaceutica Sinica B 2022;12(3):1351-1362
Scaffold hopping refers to computer-aided screening for active compounds with different structures against the same receptor to enrich privileged scaffolds, which is a topic of high interest in organic and medicinal chemistry. However, most approaches cannot efficiently predict the potency level of candidates after scaffold hopping. Herein, we identified potent PDE5 inhibitors with a novel scaffold via a free energy perturbation (FEP)-guided scaffold-hopping strategy, and FEP shows great advantages to precisely predict the theoretical binding potencies ΔG FEP between ligands and their target, which were more consistent with the experimental binding potencies ΔG EXP (the mean absolute deviations
10.Nobiletin and its derivatives overcome multidrug resistance (MDR) in cancer: total synthesis and discovery of potent MDR reversal agents.
Senling FENG ; Huifang ZHOU ; Deyan WU ; Dechong ZHENG ; Biao QU ; Ruiming LIU ; Chen ZHANG ; Zhe LI ; Ying XIE ; Hai-Bin LUO
Acta Pharmaceutica Sinica B 2020;10(2):327-343
Our recent studies demonstrated that the natural product nobiletin (NOB) served as a promising multidrug resistance (MDR) reversal agent and improved the effectiveness of cancer chemotherapy . However, low aqueous solubility and difficulty in total synthesis limited its application as a therapeutic agent. To tackle these challenges, NOB was synthesized in a high yield by a concise route of six steps and fourteen derivatives were synthesized with remarkable solubility and efficacy. All the compounds showed improved sensitivity to paclitaxel (PTX) in P-glycoprotein (P-gp) overexpressing MDR cancer cells. Among them, compound exhibited water solubility 280-fold higher than NOB. A drug-resistance A549/T xenograft model showed that at a dose of 50 mg/kg co-administered with PTX (15 mg/kg), inhibited tumor growth more effective than NOB and remarkably increased PTX concentration in the tumors P-gp inhibition. Moreover, Western blot experiments revealed that inhibited expression of NRF2, phosphorylated ERK and AKT in MDR cancer cells, thus implying of multiple mechanisms to reverse MDR in lung cancer.