Beijing Key Laboratory for Protein Therapeutics

Laboratory for, China

Beijing Key Laboratory for Protein Therapeutics

Laboratory for, China
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Cao Y.,Tsinghua University | Cao Y.,Beijing Key Laboratory for Protein Therapeutics | Chen Y.,Tsinghua University | Chen Y.,Beijing Key Laboratory for Protein Therapeutics
Tsinghua Science and Technology | Year: 2010

The epitope ELDKWA, which is located in the membrane-proximal external region (MPER) of HIV-1 gp41, is an important neutralizing epitope. The human monoclonal antibody (mAb) 2F5 against this epitope shows broad neutralizing activity toward many HIV strains. However, several reports have shown that the epitope-specific mAbs induced by peptides containing MPER did not exhibit the same neutralizing activities as human mAb 2F5. In this study, four ELDKWA epitope specific mAbs (9E7, 7E10, 6B5, and 2B4) induced by immunization with the ELDKWA epitope in varied molecular contexts, all showed inhibitory activities with different potencies in HIV-1 Env-mediated membrane fusion assays and pseudovirus neutralization assays. This result indicates that though these antibodies recognize the epitope ELDKWA, their characterizations differ from that of neutralizing antibodies, implying that the neutralizing mAbs can be induced but also need to be screened, and the protective ability of a related vaccine antigen depends on the concentration of the neutralizing mAbs in the induced polyclonal antibodies. © 2010 Tsinghua University Press.

Li Z.-G.,Chinas National Center for Safety Evaluation of Drugs | Jia L.,National Engineering Laboratory for Anti tumor Protein Therapeutics | Jia L.,Beijing Key Laboratory for Protein Therapeutics | Jia L.,Tsinghua University | And 13 more authors.
Acta Pharmacologica Sinica | Year: 2015

Aim:M2ES is PEGylated recombinant human endostatin. In this study we investigated the pharmacokinetics, tissue distribution, and excretion of M2ES in rats.Methods: 125I-radiolabeled M2ES was administered to rats by intravenous bolus injection at 3 mg/kg. The pharmacokinetics, tissue distribution and excretion of M2ES were investigated using the trichloroacetic acid (TCA) precipitation method. Results:The serum M2ES concentration-time curve after a single intravenous dose of 3 mg/kg in rats was fitted with a non-compartment model. The pharmacokinetic parameters were evaluated as follows: Cmax =28.3 μg·equ/mL, t1/2 =71.5 h, AUC(0-∞) =174.6 μg·equ·h/mL, Cl=17.2 mL·h-1 ·kg-1, MRT=57.6 h, and Vss =989.8 mL/kg for the total radioactivity; Cmax =30.3 μg·equ/mL, t 1/2 =60.1 h, AUC(0-∞) =146.2 μg·equ·h/mL, Cl=20.6 mL·h-1 ·kg-1, MRT=47.4 h, and Vss =974.6 mL/kg for the TCA precipitate radioactivity. M2ES was rapidly and widely distributed in various tissues and showed substantial deposition in kidney, adrenal gland, lung, spleen, bladder and liver. The radioactivity recovered in the urine and feces by 432 h post-dose was 71.3% and 8.3%, respectively. Only 0.98% of radioactivity was excreted in the bile by 24 h post-dose. Conclusion: PEG modification substantially prolongs the circulation time of recombinant human endostatin and effectively improves its pharmacokinetic behavior. M2ES is extensively distributed in most tissues of rats, including kidney, adrenal gland, lung, spleen, bladder and liver. Urinary excretion was the major elimination route for M2ES. © 2015 CPS and SIMM.

Guo L.,Lanzhou University | Guo L.,National Engineering Laboratory for Anti Tumor Protein Therapeutics | Guo L.,Beijing Key Laboratory for Protein Therapeutics | Guo L.,Tsinghua University | And 21 more authors.
Molecular Medicine Reports | Year: 2015

Hypoxia-inducible factor-1α (HIF-1α) is key in tumor progression and aggressiveness as it regulates a series of genes involved in angiogenesis and anaerobic metabolism. Previous studies have shown that the transcriptional levels of HIF-1α may be downregulated by endostatin. However, the molecular mechanism by which endostatin represses HIF-1α expression remains unknown. The current study investigated the mechanism by which nuclear-translocated endostatin suppresses HIF-1α activation by disrupting Zn(II) homeostasis. Endostatin was observed to downregulate HIF-1α expression at mRNA and protein levels. Blockage of endostatin nuclear translocation by RNA interference of importin α1/β1 or ectopic expression of NLS-deficient mutant nucleolin in human umbilical vein endothelial cells co-transfected with small interfering (si)-nucleolin siRNA compromises endostatin-reduced HIF-1α expression. Nuclear-translocated apo-endostatin, but not holo-endostatin, significantly disrupts the interaction between CBP/p300 and HIF-1α by disturbing Zn(II) homeostasis, which leads to the transcriptional inactivation of HIF-1α. The results reveal mechanistic insights into the method by which nuclear-translocated endostatin downregulates HIF-1α activation and provides a novel way to investigate the function of endostatin in endothelial cells.

Zhuo W.,National Engineering Laboratory for Anti tumor Protein Therapeutics | Zhuo W.,Beijing Key Laboratory for Protein Therapeutics | Zhuo W.,Tsinghua University | Chen Y.,National Engineering Laboratory for Anti tumor Protein Therapeutics | And 8 more authors.
Frontiers of Medicine in China | Year: 2011

Endostatin, a 20 kDa C-terminal fragment of collagen XVIII, was first identified as a potent angiogenic inhibitor. The anti-angiogenic function of endostatin has been well documented during the past decade. Recently, several studies demonstrated that endostatin also inhibits tumor lymphangiogenesis and lymphatic metastasis. However, the exact mechanism that endostatin executes its anti-angiogenic and anti-lymphangiogenic functions remains elusive. In the current mini-review, we briefly summarize recent novel findings, including the functions of endostatin targeting not only angiogenesis but also lymphangiogenesis, and the underlying mechanism by which endostatin internalization regulates its biological functions. © 2011 Higher Education Press and Springer-Verlag Berlin Heidelberg.

Wang J.,Tsinghua University | Wang J.,Beijing Key Laboratory for Protein Therapeutics | Wang J.,Protein Science Laboratory of MOE | Xu L.,Tsinghua University | And 8 more authors.
Vaccine | Year: 2011

Induction of mucosal antibodies to prevent HIV infection is an important strategy for the HIV-1 prophylaxis. Here we report an epitope-vaccine based antigen that was able to elicit mucosal antibodies capable of blocking HIV-1 transcytosis. Because the ELDKWA epitope of neutralizing antibody 2F5 plays a crucial role in transcytosis, a series of immunogens that contain tandem copies of ELDKWA were prepared. Mice were immunized with these immunogens intranasally, and received intraperitoneal. +. intranasal boosters. The immunogens that contained more ELDKWA epitopes elicited higher level of mucosal ELDKWA-epitope specific IgAs and systemic IgGs. Although the antisera from the immunized mice exhibited mild neutralizing potency to HIV-1 isolates HXB2 and JRFL, the affinity purified mucosal ELDKWA-epitope specific antibodies could block the transcytosis of cell-free CNE3 (a primary isolate of subtype CRF01_AE) in human tight epithelial models. © 2011 Elsevier Ltd.

Chen Y.,National Engineering Laboratory for Anti tumor Protein Therapeutics | Chen Y.,Beijing Key Laboratory for Protein Therapeutics | Chen Y.,Tsinghua University | Wang S.,National Engineering Laboratory for Anti tumor Protein Therapeutics | And 14 more authors.
Blood | Year: 2011

Specific internalization of endostatin into endothelial cells has been proved to be important for its biologic functions. However, the mechanism of endostatin internalization still remains elusive. In this study, we report for the first time that both caveolae/lipid rafts and clathrin-coated pits are involved in endostatin internalization. Inhibition of either the caveolae pathway or the clathrin pathway with the use of chemical inhibitors, small interfering RNAs, or dominant-negative mutants alters endostatin internalization in vitro. Intriguingly, cholesterol sequestration by nystatin, a polyene antifungal drug, significantly enhances endostatin uptake by endothelial cells through switching endostatin internalization predominantly to the clathrin-mediated pathway. Nystatinenhanced internalization of endostatin also increases its inhibitory effects on endothelial cell tube formation and migration. More importantly, combined treatment with nystatin and endostatin selectively enhances endostatin uptake and biodistribution in tumor blood vessels and tumor tissues but not in normal tissues of tumor-bearing mice, ultimately resulting in elevated antiangiogenic and antitumor efficacies of endostatin in vivo. Taken together, our data show a novel mechanism of endostatin internalization and support the potential application of enhancing the uptake and therapeutic efficacy of endostatin via regulating distinct endocytic pathways with cholesterolsequestering agents. © 2011 by The American Society of Hematology.

Qi F.,National Engineering Laboratory for Anti tumor Protein Therapeutics | Qi F.,Beijing Key Laboratory for Protein Therapeutics | Qi F.,Tsinghua University | He T.,National Engineering Laboratory for Anti tumor Protein Therapeutics | And 29 more authors.
Clinical Cancer Research | Year: 2015

Purpose: Before metastasis, primary tumor can create a pre-metastatic niche in distant organ to facilitate the dissemination of tumor cells. In the premetastatic phase, the permeability of pulmonary vasculatures is increased to accelerate the extravasation of circulating tumor cells. However, it is not clear whether local miRNAs contribute to the vascular hyperpermeability of the premetastatic niche. Experimental Design: The expression of total miRNAs was determined using microarray in series of premetastatic lungs from tumor-bearing mice. Significantly differentially expressed miRNAs were identified and validated with qRT-PCR. Vascular permeability assays, vascular mimic systems, and orthotopic tumor models were used to investigate roles of selected miRNAs and target genes in premetastatic hyperpermeability. Results: We identified a miRNA signature in premetastatic lungs. Among these miRNAs, miR-30a, b, c, d, and e were significantly attenuated. Subsequent investigations elucidated that lung fibroblast-derived miR-30s stabilized pulmonary vessels. Overexpression of miR-30s in lungs postponed metastasis and extended overall survival of B16 tumor-bearing mice. Following studies uncovered that Skp2 was directly targeted by miR-30s. Overexpression of Skp2 could disrupt pulmonary vessels, promote lung metastasis, and decrease overall survival of B16 tumor-bearing mice. Conclusions: These findings illuminate a novel mechanism for the modulation of premetastatic niches by miR-30s, which suggest that miR-30s represent not only promising targets for anti-metastasis therapy but also indicators for metastasis. © 2014 American Association for Cancer Research.

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