Jiong Li

Publications of Jiong Li

• In vitro and in vivo anti-tumor activities of anti-EGFR single-chain variable fragment fused with recombinant gelonin toxin.

  Authors: Xikun Zhou, Ji Qiu, Zhen Wang, Nongyu Huang, Xiaolei Li, Qian Li, Yinbing Zhang, Chengjian Zhao, Can Luo, Nannan Zhang, Xiu Teng, Zhongwen Chen, Xiao Liu, Xianlian Yu, Wenling Wu, Yu-Quan Wei, Jiong Li

  Journal of cancer research and clinical oncology. 03/2012;

  PURPOSE: Epidermal growth factor receptor (EGFR) plays an important role in the growth and metastasis of many solid tumors. Strategies that target EGFR hold promising therapeutic potential for thePURPOSE: Epidermal growth factor receptor (EGFR) plays an important role in the growth and metastasis of many solid tumors. Strategies that target EGFR hold promising therapeutic potential for the treatment for non-small cell lung cancer (NSCLC), as EGFR is normally overexpressed in these tumors. This study was designed to determine whether an anti-EGFR immunotoxin has anti-tumor activity against NSCLC, and if so, to further investigate the possible mechanisms of cytotoxicity. METHODS: A fusion protein of anti-EGFR single-chain variable fragment (anti-EGFR scFv) and the plant toxin gelonin (rGel) was constructed, expressed in bacteria, and purified to homogeneity. Cytotoxicity of anti-EGFR scFv/rGel (E/rG) immunotoxin was assessed on A549, HCC827, and H1975 cells (EGFR-overexpressing NSCLC-derived cell lines) and A549 xenografts in nude mice. RESULTS: Cytotoxicity experiments using E/rG on A549, HCC827, and H1975 cells demonstrated that E/rG can specifically inhibit proliferation of these cells, whereas it did not affect the proliferation of Raji cells that do not express EGFR. Treatment for A549 xenografts in nude mice with E/rG resulted in significant suppression of tumor growth compared to controls. Immunofluorescence in frozen tissue sections confirmed that E/rG could specifically bind to tumor tissues in nude mice bearing A549 tumor xenografts, while rGel alone showed no binding activity. Furthermore, E/rG inhibited the growth of A549 cells by cytotoxic effects that blocked tumor proliferation, and the immunotoxin-induced cell death may be mediated by autophagy. CONCLUSIONS: These results showed that E/rG might have significant potential as a novel clinical therapeutic agent against human NSCLC.

• [Antitumoral efficacy by systemic delivery of cationic liposome-plasmid interleukin-15 complexes in murine models of lung metastasis].

  Authors: Xiu Teng, Xi-Kun Zhou, Yin-Bing Zhang, Ji Qiu, Yong-Qiu Mao, Hong-Xin Deng, Jiong Li

  Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology. 02/2012; 28(2):148-52.

  To investigate the therapeutic effect of the plasmid pcDNA3.1-IL15 complexed with cationic liposome (CL-IL15) in the B16-F10 melanoma lung metastasis model. A plasmid with high secretive efficiencyTo investigate the therapeutic effect of the plasmid pcDNA3.1-IL15 complexed with cationic liposome (CL-IL15) in the B16-F10 melanoma lung metastasis model. A plasmid with high secretive efficiency of IL-15 was constructed and the optimum mix ratio was determined to formulate cationic liposome-plasmid complex with the optimal encapsulation. The CHO-K1 cell line was transfected by CL-IL15. The secretion of transfected IL-15 gene was detected by Western blot and its biological function was measured through the proliferation response of CTLL-2 cytotoxic T cell line of murine by MTT assay. The C57BL/6 mice were inoculated intravenously (i.v.) with B16-F10 melanoma lung metastasis cells then treated (i.v.) by CL-IL15 in a therapeutic setting to derermine the tumorigenesis and research the corresponding mechanisms. The pcDNA3.1-IL15 plasmid was successfully constructed and the mass-ratio of optimal condition of cationic liposome-plasmid with perfect entrapment was 1:5 (plasmid: cationic liposome). Western blot analysis displayed the detection of IL-15 both in the medium and the pcDNA3.1-IL15 transfected cells. MTT assay showed that CTLL-2 cells could proliferate with the medium obtained from CHO-K1 cells transfected by CL-IL15. And the administration of CL-IL15 complexes led to the significant inhibition lung metastasis of malignant melanoma (P<0.05). CL-IL15 could inhibit the metastasis of malignant melanoma and the cationic liposome delivered plasmid pcDNA3.1-IL-15 complexes may be an efficient therapeutic strategy for the treating of lung metastasis. And the effective splenic cell-mediated cytotoxicity and the obvious NK cells recruitment may be involved.

• NOXA-Induced Alterations in the Bax/Smac Axis Enhance Sensitivity of Ovarian Cancer Cells to Cisplatin.

  Authors: Chao Lin, Xin-Yu Zhao, Lei Li, Huan-Yi Liu, Kang Cao, Yang Wan, Xin-Yu Liu, Chun-Lai Nie, Lei Liu, Ai-Ping Tong, Hong-Xin Deng, Jiong Li, Zhu Yuan, Yu-Quan Wei

  PloS one. 01/2012; 7(5):e36722.

  Ovarian cancer is the most common cause of death from gynecologic malignancy. Deregulation of p53 and/or p73-associated apoptotic pathways contribute to the platinum-based resistance in ovarianOvarian cancer is the most common cause of death from gynecologic malignancy. Deregulation of p53 and/or p73-associated apoptotic pathways contribute to the platinum-based resistance in ovarian cancer. NOXA, a pro-apoptotic BH3-only protein, is identified as a transcription target of p53 and/or p73. In this study, we found that genetic variants of Bcl-2 proteins exist among cisplatin-sensitive and -resistant ovarian cancer cells, and the responses of NOXA and Bax to cisplatin are regulated mainly by p53. We further evaluated the effect of NOXA on cisplatin. NOXA induced apoptosis and sensitized A2780s and SKOV3 cells to cisplatin in vitro and in vivo. The effects were mediated by elevated Bax expression, enhanced caspase activation, release of Cyt C and Smac into the cytosol. Furthermore, gene silencing of Bax or Smac significantly attenuated NOXA and/or cisplatin-induced apoptosis in chemosensitive A2780s cells, whereas overexpression of Bax or addition of Smac-N7 peptide significantly increased NOXA and/or cisplatin-induced apoptosis in chemoresistant SKOV3 cells. To our knowledge, these data suggest a new mechanism by which NOXA chemosensitized ovarian cancer cells to cisplatin by inducing alterations in the Bax/Smac axis. Taken together, our findings show that NOXA is potentially useful as a chemosensitizer in ovarian cancer therapy.

• Protective effects of IL-4 on Bacillus Calmette-Guerin and lipopolysaccharide induced immunological liver injury in mice.

  Authors: Nan N Zhang, Nong Y Huang, Xi K Zhou, Xiao L Luo, Chang Y Liu, Yan Zhang, Ji Qiu, Yin B Zhang, Xiu Teng, Can Luo, Xian C Chen, Bing Kan, Yong Q Mao, Ai P Tong, Yu Q Wei, Jiong Li

  Inflammation research : official journal of the European Histamine Research Society ... [et al.]. 09/2011; 61(1):17-26.

  Mice injected with Bacillus Calmette-Guérin (BCG) were challenged with lipopolysaccharide (LPS) to induce inflammatory liver injury. This study was performed to explore the protective effects ofMice injected with Bacillus Calmette-Guérin (BCG) were challenged with lipopolysaccharide (LPS) to induce inflammatory liver injury. This study was performed to explore the protective effects of interleukin (IL)-4 against liver injury induced by BCG and LPS in mice. Mice injected with BCG (125 mg/kg) were challenged with LPS (10 μg/kg) to induce the model of inflammatory liver injury. Half an hour after injection of LPS, mice were subcutaneously administered rmIL-4 at 5 and 0.5 μg/kg, respectively. Liver injury was evaluated by serum transaminase assay and H & E staining. Liver cytokine concentrations were determined by enzyme-linked immunosorbent assay, and intrahepatic cytokine and iNOS mRNA levels by reverse transcriptase polymerase chain reaction. Intrahepatic apoptosis was evaluated by terminal deoxynucleotidyl transferase mediated nick end labeling. NF-κB p65 and ERK signal pathway was detected by Western-blotting. NF-κB signal pathway was also detected by electrophoretic mobility shift assay. IL-4 reduced the serum ALT, AST and LDH, alleviated the inflammatory cells infiltration, down regulated the expression of TNF-α, IL-1β, IFN-γ, IL-6 and iNOS mRNA in liver, and alleviated hepatic glutathione depletion (GSH). In addition, IL-4 displayed inhibition of extracellular signal-regulated kinase phosphorylation and NF-κB activation. IL-4 may protect mice against BCG/LPS-induced immune liver injury, besides ERK and NF-κB signal pathways were involved in the effects.

• [The construction of anti-EGFR single chain variable fragment fused with gelonin toxin prokaryotic expressing plasmid and the study of its anti-tumor effects].

  Authors: Ji Qiu, Qian Li, Xiao-Lei Li, Xi-Kun Zhou, Nong-Yu Huang, Jiong Li

  Sichuan da xue xue bao. Yi xue ban = Journal of Sichuan University. Medical science edition. 09/2011; 42(5):621-4.

  To construct a prokaryotic expressing plasmid for recombinant immunotoxin which fused anti-EGFR scFv together with gelonin toxin, express and verify its function. The gene fragments coding anti-EGFRTo construct a prokaryotic expressing plasmid for recombinant immunotoxin which fused anti-EGFR scFv together with gelonin toxin, express and verify its function. The gene fragments coding anti-EGFR single chain fragment were amplified with PCR and cloned into pET32a vector which contains gelonin toxin. The new plasmid was transformed into BL21 (DE3) cells. The induced inclusion bodies were denatured, refolded and purified through SP Sepharose Fast Flow Column. The purified immunotoxin rEG was identified by western blot analysis, and the bioactivity was identified using cell immnuohistochemistry and MTT assay. The expressing vector pET32a-rEG has been constructed correctly, confirmed by restriction endonuclease digestion and sequencing. The recombinant immunotoxin rEG was purified after denaturing the inclusion bodies, refolding and cationic exchange chromatograph. The purified protein rEG had the right immunology specificity, rEG could efficiently target to EGFR positive cells identified by cell immnuohistochemistry. And the result of MTT assay showed rEG could specifically kill EGFR positive cells. The recombinant immunotoxin rEG with high purity and biologic activity was prepared in this study, which would become the basic for the further study of the biologic function of rEG.

• PUMA Chemosensitizes Intrinsically Resistant Ovarian Cancer Cells to Cisplatin by Lowering the Threshold Set by Bcl-x(L) and Mcl-1.

  Authors: Zhu Yuan, Kang Cao, Chao Lin, Lei Li, Huan-Yi Liu, Xin-Yu Zhao, Lei Liu, Hong-Xin Deng, Jiong Li, Chun-Lai Nie, Yu-Quan Wei

  Molecular medicine (Cambridge, Mass.). 08/2011;

  Ovarian cancer is the number one cause of death from gynecologic malignancy. Defective p53 pathway is a hallmark of ovarian carcinoma. p53 mutation correlates significantly with resistance toOvarian cancer is the number one cause of death from gynecologic malignancy. Defective p53 pathway is a hallmark of ovarian carcinoma. p53 mutation correlates significantly with resistance to platinum-based chemotherapy, early relapse, and shortened overall survival in ovarian cancer patients. PUMA, a BH3-only Bcl-2 family protein, was recently identified as a transcriptional target of p53 and a potent apoptosis inducer in various cancer cells. In this study, we showed that the induction of PUMA by cisplatin was abolished in p53-deficient SKOV3 cells. Elevated expression of PUMA induced apoptosis and sensitized A2780s and SKOV3 ovarian cancer cells to cisplatin, and the combination of PUMA and low-dose cisplatin significantly suppressed xenograft tumor growth in vivo through enhanced induction of apoptosis compared with treatment with PUMA or cisplatin alone. The effects of PUMA were mediated by enhanced caspase activation and release of cytochrome c and Smac into the cytosol. Furthermore, PUMA chemosensitized intrinsically resistant SKOV3 cells to cisplatin through downregulation of Bcl-x(L) and Mcl-1. PUMA-mediated Bcl-x(L) downregulation mainly happened at the transcription level while PUMA-induced Mcl-1 downregulation was associated with caspase-dependent cleavage and proteasome-mediated degradation. To our knowledge, these data suggest a new mechanism by which overexpression of PUMA enhances sensitivity of SKOV3 cells to cisplatin by lowering the threshold set simultaneously by Bcl-x(L) and Mcl-1. Taken together, our findings indicate that PUMA is an important modulator of therapeutic responses of ovarian cancer cells, and is potentially useful as a chemosensitizer in ovarian cancer therapy.

• Proteomic analysis of the interleukin-4 (IL-4) response in hepatitis B virus-positive human hepatocelluar carcinoma cell line HepG2.2.15.

  Authors: Yuqin Yao, Jiong Li, Zejun Lu, Aiping Tong, Wei Wang, Xiaolan Su, Yan Zhou, Bo Mu, Shijie Zhou, Xiaoan Li, Lijuan Chen, Lantu Gou, Hongbing Song, Jinliang Yang, Yuquan Wei

  Electrophoresis. 08/2011; 32(15):2004-12.

  Hepatitis B virus (HBV) infection is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. In recent decades, significant progress toward understanding the molecular virologyHepatitis B virus (HBV) infection is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. In recent decades, significant progress toward understanding the molecular virology and pathogenesis of HBV infection has been made. In addition, multiple treatment modalities have been developed for persons with HBV infection. In the present study, we demonstrated that IL-4 inhibits the expression of hepatitis B surface antigen and hepatitis B e antigen in a HBV stably transfected hepatocellular carcinoma cell line (HepG2.2.15). To reveal the anti-HBV mechanism of IL-4 by proteomics, 2-DE and MS technology were utilized to profile global changes in protein expression in HepG2.2.15 cells after IL-4 treatment. A total of 56 differentially expressed proteins were identified in IL-4-treated HepG2.2.15 cells. To find out the interaction of these changed proteins by bioinformatics, signaling network analysis with the STRING tool showed that the identified proteins are primarily involved in transcription and proteolysis. Taken together, these results offer valuable clues for understanding the molecular mechanisms of the IL-4-mediated anti-HBV response.

• Antitumoral efficacy by systemic delivery of heparin conjugated polyethylenimine-plasmid interleukin-15 complexes in murine models of lung metastasis.

  Authors: Xikun Zhou, Xiaolei Li, Maling Gou, Ji Qiu, Jing Li, Chuanjiang Yu, Yinbin Zhang, Nannan Zhang, Xiu Teng, Zhongwen Chen, Can Luo, Zhen Wang, Xiao Liu, Guobo Shen, Li Yang, Zhiyong Qian, Yuquan Wei, Jiong Li

  Cancer science. 07/2011; 102(7):1403-9.

  Gene therapy shows promising application in cancer therapy, but the lack of an ideal gene delivery system is still a tough challenge for cancer gene therapy. Previously, we prepared a novel cationicGene therapy shows promising application in cancer therapy, but the lack of an ideal gene delivery system is still a tough challenge for cancer gene therapy. Previously, we prepared a novel cationic nanogel, heparin-polyethylenimine (HPEI), which had potential application in gene delivery. In the present study, we constructed a plasmid with high expression efficiency of interleukin-15 (IL15) and investigated the effects HPEI-plasmid IL15 (HPEI-pIL15) complexes on the distribution level of the lung. We then evaluated the anticancer effect of HPEI-pIL15 complexes on lung metastases of B16-F10 melanoma and CT26 colon carcinoma. These results demonstrated that intravenous injection of the HPEI-pIL15 complex exhibited the highest plasmid distribution level in the lung compared with that of PEI2K-pIL15 and PEI25K-pIL15, and mice treated with HPEI-pIL15 had a lower tumor metastasis index compared with other treatment groups. Moreover, the number of natural killer cells, which were intermingled among the tumor cells, and the level of tumor necrosis factor-α and interferon-γ in the serum also increased in the pIL15-treated mice. Furthermore, the cytotoxic activity of spleen cells also increased significantly in the HPEI-pIL15 group. In addition, induction of apoptosis and inhibition of cell proliferation in lung tumor foci in the HPEI-pIL15 group was observed. Taken together, treating lung metastasis cancer with the HPEI nanogels delivered by plasmid IL15 might be a new and interesting cancer gene therapy protocol.

• Intravenous liposomal delivery of the short hairpin RNAs against Plk1 controls the growth of established human hepatocellular carcinoma.

  Authors: Hongxin Deng, Qingyuan Jiang, Yang Yang, Shuang Zhang, Yongping Ma, Gang Xie, Xiang Chen, Zhiyong Qian, Yanjun Wen, Jiong Li, Jinliang Yang, Lijuan Chen, Xia Zhao, Yuquan Wei

  Cancer biology & therapy. 02/2011; 11(4):401-9.

  Polo-like kinase 1 (Plk1) is a key cell cycle regulator that is frequently overexpressed in human hepatocellular carcinomas. Blockade of the Plk1 pathway has been reported to be capable of inducingPolo-like kinase 1 (Plk1) is a key cell cycle regulator that is frequently overexpressed in human hepatocellular carcinomas. Blockade of the Plk1 pathway has been reported to be capable of inducing anti-tumor effect. Here, plasmids containing U6 promoter-driven shRNAs against human Plk1 were constructed and transfected in human hepatocellular carcinoma cell line HepG2. ShRNA targeting Plk1 almost completely reduced Plk1 expression in HepG2 hepatocellular carcinoma cells, as confirmed by RT-PCR and Western blot. As a consequence, HepG2 cells exhibited reduced proliferation and enhanced apoptosis in vitro. Most importantly, Treatment with Plk shRNA-DOTAP:Chol complex significantly suppressed the growth of HepG2 xenografts, accompanied with phenotypic changes in tumor cells, including proliferation inhibition and apoptosis induction. Our study suggested that shRNA-mediated silencing of Plk1 might be a novel therapeutic approach against human hepatocellular carcinoma by inhibiting tumor cells proliferation and inducing apoptosis.

• Retinoic acid and dimethyl sulfoxide promote efficient delivery of transgenes to mouse skin by topically transdermal penetration.

  Authors: Xiangjun Chen, Yinbing Zhang, Changyong Liu, Yan Zhang, Xikun Zhou, Taoyou Zhou, Yongqiu Mao, Bing Kan, Yu-Quan Wei, Jiong Li

  Drug delivery. 08/2010; 17(6):385-90.

  Simple and efficient gene transfer to the skin would facilitate many local and systemic gene therapy applications. This study reports a novel approach that allows expression of plasmid DNA inSimple and efficient gene transfer to the skin would facilitate many local and systemic gene therapy applications. This study reports a novel approach that allows expression of plasmid DNA in epidermis and hair follicle cells with dimethyl sulfoxide (DMSO) after pre-treatment with depilation and retinoic acid (RA) for the purposes of gene therapy. This study investigated the transdermal efficacy of gene to mouse skin when utilizing DMSO after RA pre-treatment. Retinoic acid pre-treatment can increase the efficiency of transfection. This finding indicates that one can more effectively and much less expensively make use of genes therapy to treat diseases of the hair and skin.

• A novel transdermal plasmid-dimethylsulfoxide delivery technique for treatment of psoriasis.

  Authors: Yan Zhang, Jiong Li, Chang-Yong Liu, Xi-Kun Zhou, Ji Qiu, Yin-Bing Zhang, Nong-Yu Huang, You Li, Xiang-Jun Chen, Xiao-Lei Li, Yong-Sheng Wang, Han-Suo Yang, Xian-Cheng Chen, Bing Kan, Yong-Qiu Mao, Hong-Xin Deng, Li Yang, Yan-Jun Wen, Xia Zhao, Yu-Quan Wei

  Dermatology (Basel, Switzerland). 08/2010; 221(1):84-92.

  Psoriasis is a chronic and relapsing inflammatory skin disease associated with various immunologic abnormalities. Repeated subcutaneous injection of interleukin-4 (IL-4) has been established as anPsoriasis is a chronic and relapsing inflammatory skin disease associated with various immunologic abnormalities. Repeated subcutaneous injection of interleukin-4 (IL-4) has been established as an effective treatment to counteract psoriasis. We investigated whether gene therapy using IL-4 expression plasmid (pIL-4) via transdermal delivery was an alternative treatment for psoriasis. In our experiment, dimethylsulfoxide (DMSO) was used as a penetration enhancer. At first, the penetration efficiency of the complex of reporter plasmid accompanied by DMSO was investigated both in vitro and in vivo. Then, the antipsoriasis efficiency of the treatment with pIL-4-DMSO was tested in mice. The expression of the reporter gene was detected in epidermis and dermis both in vitro and in vivo. More importantly, the psoriasis symptoms were relieved, and significant reductions in some psoriasis-associated factors were observed after pIL-4-DMSO treatment. We conclude that the topical application of pIL-4-DMSO can treat psoriasis to a significant extent.

• Gene therapy for psoriasis in the K14-VEGF transgenic mouse model by topical transdermal delivery of interleukin-4 using ultradeformable cationic liposome.

  Authors: Jiong Li, Xia Li, Yan Zhang, Xi K Zhou, Han S Yang, Xian C Chen, Yong S Wang, Yu Q Wei, Li J Chen, Huo Z Hu, Chang Y Liu

  The journal of gene medicine. 06/2010; 12(6):481-90.

  Topical transdermal gene delivery to the skin shows great potential for painless, non-invasive administration of vaccines and therapeutic agents. Interleukin (IL)-4 strategies have shown a goodTopical transdermal gene delivery to the skin shows great potential for painless, non-invasive administration of vaccines and therapeutic agents. Interleukin (IL)-4 strategies have shown a good antipsoriatic effect in clinic trials. To date, no information has been acquired on the effectiveness of gene therapy for psoriasis in the K14-VEGF transgenic mouse model by topical transdermal penetration of murine IL-4 (mIL-4) using ultradeformable cationic liposome (UCL). In the present study, we synthesized an UCL and determined a suitable formula for transdermally delivering plasmid DNA to mouse skin. We then tested the antipsoriatic efficacy in the K14-VEGF transgenic mouse model by transdermal delivery of mIL-4 using UCL. We found that plasmid DNA was transdermally delivered to vicinal sites of epidermis and hair follicles using this optimized formula. Plasmid DNA expression was detected in ear skin. Twenty-four hours after topical application, plasmid DNA was not detected in blood serum and liver, which may decrease the risk of insertion of promoter from plasmid to genomic DNA. Mice treated with UCL/mIL-4 displayed a mild psoriasis phenotype. Histological analysis of pathological score using the Baker scoring system revealed an antipsoriatic effect. Immunohistochemical analysis revealed that hyperplastic and inflamed vessels were suppressed. These observations provide evidence of antipsoriatic efficacy by topical transdermal delivery of mIL-4. Therefore, topical transdermal gene transfer is attractive and offers future potential for application in human patients with other dermatogic diseases.

• A vector-based short hairpin RNA targeting Aurora A inhibits breast cancer growth.

  Authors: Yang Wan, Anliang Huang, Yang Yang, Gang Xie, Xiang Chen, Jia Hu, Xiancheng Chen, Li Yang, Jiong Li, Lijuan Chen, Yu Jiang, Xia Zhao, Yuquan Wei, Hongxin Deng

  International journal of oncology. 05/2010; 36(5):1121-8.

  Aurora A plays an essential role in centrosome maturation, separation and in the formation of the mitotic bipolar spindle. Overexpression or amplication of Aurora A gene has been detected in manyAurora A plays an essential role in centrosome maturation, separation and in the formation of the mitotic bipolar spindle. Overexpression or amplication of Aurora A gene has been detected in many cancer cell lines and various tumor tissues, including breast cancer, suggesting that Aurora A might be drug target for breast cancer treatment. In the current study, short hairpin RNA targeting Aurora A was cloned into pGenesil-2 plasmid vector and then transfected into MDA-MB-435S and ZR-75-30 human breast cancer cells using cationic liposome. Reduced expression of Aurora A was detected by RT-PCR and Western blot. The effect of pGenesil-2-shAURKA plasmid on tumor growth in MDA-MB-435S xenogenic implantation model was studied. pGenesil-2-shAURKA plasmid inhibited tumor growth significantly by systemantic administration. To further study the underlying mechanisms, cell apoptosis and proliferation were investigated by flow cytometric analysis, propidium iodide staining, TUNEL and Ki-67 immunostaining respectively. Increased apoptosis and reduced cell proliferation were detected in vitro and in vivo studies. In summary, our results suggested that specific knockdown of Aurora A expression by vector based shRNA may be a potential therapy for human breast cancer.

• Suppression of human MDA-MB-435S tumor by U6 promoter-driven short hairpin RNAs targeting focal adhesion kinase.

  Authors: An-liang Huang, Yang Wan, Dian-ying Liao, Huo-zhen Hu, Lin Wei, Xian-huo Wang, Yan-jun Wen, Jiong Li, Li-juan Chen, Bing Kan, Ping Chen, Yong-sheng Wang, Xiang Chen, Xia Zhao, Hong-xin Deng, Yu-quan Wei

  Journal of cancer research and clinical oncology. 02/2010; 136(8):1229-42.

  Focal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase implicated in cancer cell survival, proliferation, and in various steps in the metastatic cascade. In the present study, we tookFocal adhesion kinase (FAK) is a non-receptor protein tyrosine kinase implicated in cancer cell survival, proliferation, and in various steps in the metastatic cascade. In the present study, we took advantage of a cationic liposome as gene carrier and targeted FAK function through both in vitro and in vivo approaches. We utilized a plasmid-encoded hairpin RNA targeting the human FAK mRNA (pGensil2-shRNA/FAK), as a means to inhibit FAK expression for evaluating its anti-tumor effect in vitro and in vivo. Human MDA-MB-435S breast cancer cells were transfected with pGensil2-shRNA/FAK and examined for apoptosis by propidium iodide staining, DNA ladder, and flow cytometric analysis. For in vivo study, subcutaneous breast carcinomatosis models in nude mice were established to evaluate the therapeutic potential of pGensil2-shRNA/FAK. Assessments of proliferation (Ki-67), apoptosis (TUNEL) and angiogenesis (CD31) were done using immunohistochemical analysis. Transcripts expressed from plasmid both in vitro and in vivo were identified by northern blot analysis. pGensil2-shRNA/FAK effectively down-regulated the expression of FAK as demonstrated in vitro by real time RT-PCR and western blot analysis, whereas by real time RT-PCR and IHC staining of MDA-MB-435S tumors growing subcutaneously. Breast cancer cells lacking FAK expression undergo apoptosis in vitro. Systemic delivery of cationic liposome-complexed plasmids targeting FAK, resulted in the diminishment of subcutaneous tumor growth beyond the effects observed with liposomes carrying a non-specific shRNA. This diminishment in growth was associated with elevated levels of apoptosis (TUNEL staining), decreased cell proliferation (Ki-67 staining) and diminished endothelial cell density (CD31 staining). These results indicate that the systemic delivery of plasmid DNA targeting FAK function using cationic liposome as a gene carrier, represents a promising avenue for breast cancer therapy.

• Antitumor effect of mSurvivinThr34 --> Ala in murine colon carcinoma when administered intravenously.

  Authors: Hong-Xia Li, Xin-Yu Zhao, Lian Wang, Yong-Sheng Wang, Bin Kan, Jian-Rong Xu, Jiong Li, Yan-Jun Wen, Xing-Chen Peng, Xiang Chen, Fei Yan, Bin Ye, Xiao-Bo Du, Ju-Mei Zhao, Tao Yi, Xian-Cheng Chen, Xiao-Xia Du, Yu-Quan Wei, Xia Zhao

  Medical oncology (Northwood, London, England). 12/2009;

  Colorectal cancer is one of the most common cancers. Survivin is strongly immunogenic in a fraction of colorectal cancer patients. The present study was designed to determine whether full-lengthColorectal cancer is one of the most common cancers. Survivin is strongly immunogenic in a fraction of colorectal cancer patients. The present study was designed to determine whether full-length mouse Survivin dominant-negative mutant SurvivinT34A has the antitumor activity in a murine colon carcinoma model. The complex of cationic liposome (DOTAP/Chol) to plasmid pORF9-mSurvivin T34A was administered intravenously in a mouse subcutaneous (S. C.) CT 26 tumor model. Apoptotic cells and anti-angiogenesis were evaluated by fluorescent in situ TUNEL assay and by immunohistochemistry with an antibody reactive to CD31, respectively. A 4 h (51)Cr release assay was performed to determine Survivin-specific cytotoxicity. The adoptive transfer of CD8(+) or CD4(+) T-lymphocytes assay was to further explore the roles of immune cell subsets. We demonstrated the complex of cationic liposome (DOTAP/Chol) to plasmid pORF9-mSurvivin T34A when administered intravenously induced an efficient antitumor activity in a S. C. CT26 tumor model in mice. The main mechanism is involved in three aspects: triggering the apoptosis of tumor cells, inhibiting angiogenesis, and inducing Survivin-specific immune response. Our observations may have potential implications for the further exploration of the treatment of human colorectal cancer by intravenous delivery of dominant-negative mutant Survivin T34A.

• Prokaryotic expression, purification and characterization of a novel pro-apoptosis protein hPNAS-4.

  Authors: Hongxin Deng, Qingyuan Jiang, Shufang Liang, Fei Yan, Shengyan Hou, Zhiyong Qian, Jiong Li, Yanjun Wen, Jinliang Yang, Yuquan Wei

  Biotechnology and applied biochemistry. 11/2009;

  Recently human PNAS-4 (hPNAS-4) is reported to be a novel pro-apoptotic protein in mammalian cells. However, there is little information about hPNAS-4 characterization and its molecular mechanisms soRecently human PNAS-4 (hPNAS-4) is reported to be a novel pro-apoptotic protein in mammalian cells. However, there is little information about hPNAS-4 characterization and its molecular mechanisms so far. In our present studies, the hPNAS-4 gene was first cloned into the pGEX-6p-1 vector with GST tag to solubly express in E.coli induced with 0.5 mM IPTG at 25,and the recombinant hPNAS-4 was purified to near homogeneity with 96% purity by affinity chromatography and anion-exchange chromatography. The purified hPNAS-4 protein was further identified by liquid chromatography-electrospray ionization-mass spectrometry analysis and used to raise polyclonal antibodies which could specifically recognize the hPNAS-4 protein. In addition, hPNAS-4 is localized in cytoplasm and perinuclear. Furthermore the overexpression of hPNAS-4 in human A549 cells resulted in decrease of cell viability via apoptosis. This study represented an important step to investigate the characterization for the new pro-apoptosis protein hPNAS-4, which aids to discover new drug targets for development of effective therapeutic approaches to cancer in future.

• Intratumoral Expression of Mature Human Neutrophil Peptide-1 Mediates Antitumor Immunity in Mice.

  Authors: Yong-Sheng Wang, Dan Li, Hua-Shan Shi, Yan-Jun Wen, Li Yang, Ning Xu, Xian-Cheng Chen, Xiang Chen, Ping Chen, Jiong Li, Hong-Xin Deng, Chun-Ting Wang, Gang Xie, Shan Huang, Yong-Qiu Mao, Li-Juan Chen, Xia Zhao, Yu-Quan Wei

  Clinical cancer research : an official journal of the American Association for Cancer Research. 10/2009;

  PURPOSE: Human neutrophil peptides (HNP1-3), small molecular antimicrobial peptides, are expressed within tumors and associated with tumor necrosis and inhibition of angiogenesis. RecentPURPOSE: Human neutrophil peptides (HNP1-3), small molecular antimicrobial peptides, are expressed within tumors and associated with tumor necrosis and inhibition of angiogenesis. Recent investigations have suggested that HNP1-3 are likely to be involved in the host immune responses to tumors. EXPERIMENTAL DESIGN: We used recombinant pSec-HNP1, which expresses a secretable form of HNP1, to obtain expression of HNP1 in the tumor milieu in immunocompetent mice to explore the possible roles of HNP1 in tumor immunity. The antitumor effects were investigated in established CT26 colon cancer and 4T1 breast cancer models. RESULTS: HNP1-mediated chemotactic and activating effects on immature dendritic cells were detected both in vitro and in vivo. Intratumoral expression of HNP1 resulted in not only significant tumor growth inhibition but also increased CTL infiltration within tumors. Adoptive transfer of splenocytes and a (51)Cr release assay revealed specific cellular immune responses. Furthermore, increased antibodies were also found in sera from pSec-HNP1-treated mice supporting specific humoral immune responses. Increased apoptosis and decreased angiogenesis were also shown in treated tumors. CONCLUSIONS: These findings indicate that HNP1 can exert multiple antitumor effects through different mechanisms; more importantly, HNP1 mediates host immune responses to tumors in situ through the recruitment and subsequent activation of immature dendritic cells and thus shows promising potential in cancer therapy. (Clin Cancer Res 2009;15(22):OF1-11).

• Recombinant Murine Interleukin 4 Protein Therapy for Psoriasis in a Transgenic VEGF Mouse Model.

  Authors: Xiaoxiao Ren, Jiong Li, Xikun Zhou, Xiaolei Luo, Nongyu Huang, Yongsheng Wang, Xiancheng Chen, Yuquan Wei

  Dermatology (Basel, Switzerland). 09/2009;

  Background: Psoriasis is a typical autoimmune disease caused by a deregulation of the Th1/Th2 balance, and immunotherapy for psoriasis has been shown to be clinically efficacious. VascularBackground: Psoriasis is a typical autoimmune disease caused by a deregulation of the Th1/Th2 balance, and immunotherapy for psoriasis has been shown to be clinically efficacious. Vascular endothelial growth factor (VEGF) is a potent mediator of angiogenesis. Evidence suggests that the chronic delivery of VEGF to the skin can result in a profound inflammatory condition with many of the cellular and molecular hallmarks of human psoriasis. In this study, we investigated whether the transgenic VEGF mouse is a suitable model for antipsoriatic studies. Aim: To determine the effect of a recombinant murine interleukin 4 (rmIL-4) in the transgenic VEGF mouse model. Methods: Fifteen homozygous K14-VEGF transgenic mice were injected subcutaneously with rmIL-4 protein for 30 consecutive days with a prospective dose escalation of 0.5, 2 or 5 mug/kg. Hematoxylin-eosin staining, immunohistochemistry and real-time polymerase chain reaction analyses were performed with ear samples. Results: The rmIL-4 protein therapy was well tolerated. Tissue sections from treated skin showed improvements upon morphological and histological examinations: diminution of erythematous appearance and regression of epidermal thickness were observed, and T lymphocyte infiltration decreased significantly. The expressions of adhesion molecules, such as vascular cell adhesion molecule 1 and intracellular adhesion molecule 1, were found reduced. The level of IL-4 mRNA also increased while the level of gamma-interferon mRNA decreased, resulting in a 10-fold increase in the ratio of Th1/Th2. Conclusions: Our results reveal that rmIL-4 has clinical efficacy for the treatment of K14-VEGF transgenic mice. Angiogenesis and inflammation were ameliorated by therapy with rmIL-4.

• Improved therapeutic efficacy against murine carcinoma by combining honokiol with gene therapy of PNAS-4, a novel pro-apoptotic gene.

  Authors: Zhu Yuan, Huanyi Liu, Fei Yan, Yongsheng Wang, Lantu Gou, Chunlai Nie, Zhenyu Ding, Songtao Lai, Yuwei Zhao, Xinyu Zhao, Jiong Li, Hongxin Deng, Yongqiu Mao, Lijuan Chen, Yuquan Wei, Xia Zhao

  Cancer science. 07/2009;

  PNAS-4, a novel pro-apoptotic gene activated during the early response to DNA damage, can inhibit proliferation via apoptosis when overexpressed in some tumor cells. Recent studies have indicatedPNAS-4, a novel pro-apoptotic gene activated during the early response to DNA damage, can inhibit proliferation via apoptosis when overexpressed in some tumor cells. Recent studies have indicated that honokiol can induce apoptosis, inhibit angiogenesis, and suppress tumor growth. In the present study, we investigated whether mouse PNAS-4 (mPNAS-4) could augment the apoptosis of tumor cells induced by honokiol in vitro, and whether the antiangiogenic activity of honokiol and induction of apoptosis by mPNAS-4 could work cooperatively to improve the antitumor efficacy in vivo. In vitro, mPNAS-4 inhibited proliferation of murine colorectal carcinoma CT26 and Lewis lung carcinoma LL2 cells through induction of apoptosis, and significantly augmented the apoptosis of CT26 and LL2 cells induced by honokiol. Compared with treatment with mPNAS-4 or honokiol alone, in vivo systemic administration of an expression plasmid encoding mPNAS-4 and low-dose honokiol significantly suppressed tumor growth through the enhanced induction of apoptosis and the augmented inhibition of angiogenesis. Our data suggest that the combined treatment with mPNAS-4 plus honokiol augments antitumor effects in vitro and in vivo, and that the improved antitumor activity in vivo may be associated with enhanced induction of apoptosis and augmented inhibition of angiogenesis. The present study may provide a novel and effective method for the treatment of cancer. (Cancer Sci 2009).

• [The construction of recombinant mouse interleukin 4 prokaryotic expressing plasmid and the expression and purification of target protein]

  Authors: Xiao-Lei Luo, Xiao-Xiao Ren, Nong-Yu Huang, Ji Qiu, Qian Li, Xi-Kun Zhou, Jiong Li

  Sichuan da xue xue bao. Yi xue ban = Journal of Sichuan University. Medical science edition. 07/2009; 40(4):579-83.

  OBJECTIVE: To construct Recombinant Mouse Interleukin 4 prokaryotic expressing plasmid, express it in E. coli strain BL21 (DE3), purify and identify the expressed cytokine. METHODS: The optimizedOBJECTIVE: To construct Recombinant Mouse Interleukin 4 prokaryotic expressing plasmid, express it in E. coli strain BL21 (DE3), purify and identify the expressed cytokine. METHODS: The optimized mIL-4 cDNA fragment was cloned into the prokaryotic expressing vector pET-32a (+) to generate pET32/rmIL-4 and transformed into BL21 (DE3) cells. After induction, the expressed protein wasfound to be in the inclusion of E. coli cells. The induced product was purified through Q Sepharose Fast Flow Column and Gel Filtration Column under renaturing condition. The purified protein was identified by Western blot analysis, and the biologic activity was identified by the generation of mIL-4 dependence cell CTLL-2 and in vivo experiment of mouse psoriasis model. RESULTS: The recombinant plasmid pET32/rmIL-4 has been constructed correctly. The inclusion body was washed with 3 mol/L guanidine hydrochloride and denaturized in 7 mol/L guanidine hydrochloride. Then, the denaturized protein was gradient dialysis in the condition of pH 9. 5. The protein we purified has the right immunology specificity and biologic activity. CONCLUSION: The recombinant mouse interleukin-4 with high purity and biologic activity was prepared in this study,which will become the basis for the further study of the biologic activity of IL-4.