Publications by authors named "Lun Ou"

9 Publications

  • Page 1 of 1

Development and Validation of a Simoa Assay for Determination of Recombinant Batroxobin in Human Serum.

Curr Med Sci 2021 Jun 25;41(3):618-625. Epub 2021 Jun 25.

State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center of Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing, 102206, China.

Recombinant batroxobin (S3101) is a thrombin-like serine protease that binds to fibrinogen or is taken up by the reticuloendothelial system. A literature survey showed no adequate method that could determine sufficient concentrations to evaluate pharmacokinetic parameters for phase I clinical studies. Therefore, a sensitive method is urgently needed to support the clinical pharmacokinetic evaluation of S3101. In this study, a sensitive bioanalytical method was developed and validated, using a Quanterix single molecular array (Simoa) assay. Moreover, to thoroughly assess the platform, enzyme-linked immunosorbent assay and electrochemiluminescence assay were also developed, and their performance was compared with that of this novel technology platform. The assay was validated in compliance with the current guidelines. Measurements with the Simoa assay were precise and accurate, presenting a valid assay range from 6.55 to 4000 pg/mL. The intra- and inter-run accuracy and precision were within -19.3% to 15.3% and 5.5% to 17.0%, respectively. S3101 was stable in human serum for 280 days at -20°C and -70°C, for 2 h prior to pre-treatment and 24 h post pre-treatment at room temperature (22°C-28°C), respectively, and after five and two freeze-thaw cycles at -70°C and -20°C, respectively. The Simoa assay also demonstrated sufficient dilution linearity, assay sensitivity, and parallelism for quantifying S3101 in human serum. The Simoa assay is a sensitive and adequate method for evaluating the pharmacokinetic parameters of S3101 in human serum.
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http://dx.doi.org/10.1007/s11596-021-2382-6DOI Listing
June 2021

A randomized phase I clinical trial comparing the pharmacokinetic, safety, and immunogenicity of potential biosimilar recombinant human HER2 monoclonal antibody for injection and trastuzumab in healthy Chinese adults.

Expert Opin Investig Drugs 2020 Jul 27;29(7):755-762. Epub 2020 Jun 27.

Department of Pharmacy, Peking University People's Hospital , Beijing, China.

Objectives: Recombinant human HER2 monoclonal antibody for injection (AK-HER2) is a potential biosimilar of trastuzumab (Herceptin®). This phase Ⅰ study aimed to demonstrate the pharmacokinetic (PK) equivalence between AK-HER2 and trastuzumab in healthy volunteers. Besides, safety and immunogenicity were investigated.

Research Design And Methods: This was a randomized, double-blind phase Ⅰ trial in 96 healthy adults who received a single intravenous infusion of AK-HER2 or trastuzumab at 6 mg/kg. The primary PK endpoints were area under the serum concentration curve (AUC) from time 0 to the last time point (AUC) and peak concentration in serum (C). The PK bioequivalence was confirmed using the standard equivalence margins of 80%-125%.

Results: The PK profiles of AK-HER2 and trastuzumab displayed high similarity. The geometric mean ratios (90% confidence intervals) of primary PK endpoints were within 80%-125%. The C  and AUC  of female subjects in the AK-HER2 group were greater than those of male subjects (P <0.05). No infusion-related reactions (IRRs) or anti-drug antibody-positivity was observed after dosing.

Conclusions: AK-HER2 was demonstrated to have highly similar PK to trastuzumab in healthy Chinese adults. Both drugs showed comparable safety and immunogenicity using dexamethasone as premedication to prevent IRRs..
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http://dx.doi.org/10.1080/13543784.2020.1770226DOI Listing
July 2020

Gankyrin is essential for hypoxia enhanced metastatic potential in breast cancer cells.

Mol Med Rep 2014 Mar 12;9(3):1032-6. Epub 2013 Dec 12.

Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing, P.R. China.

Hypoxia, a critical regulator of tumor growth and metastasis, induces the transcriptional activation of several pathways involved in proliferation, migration and invasion. Gankyrin was found to be overexpressed, and also promoted the metastasis in breast cancer cells, which is also involved in the regulation of hypoxia inducible factor‑1 and hypoxia‑inducible factor‑1α. The present study showed that gankyrin mRNA and protein expression were increased under hypoxic conditions in the BT474 breast cancer cell line, accompanied with increased ability of cell migration and invasion. Lentivirus‑mediated siRNA targeting gankyrin was transfected into BT474 cells. Wound‑healing and transwell experiments showed that gankyrin deletion abrogated the increased migration and invasion of BT474 cells due to hypoxia. In addition, E‑cadherin was found to be involved in the gankyrin induced invasion of breast cancer cells due to hypoxia. The present study indicated that gankyrin deletion abrogated the increased metastatic potential of breast cancer cells under hypoxic conditions partly through regulating E‑cadherin, suggesting that an improved understanding of gankyrin may offer a potential therapeutic target for the treatment of human breast cancer metastasis.
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http://dx.doi.org/10.3892/mmr.2013.1860DOI Listing
March 2014

Immunostimulatory and anti-neoplasm effects of a novel palindrome CpG oligodeoxynucleotide in mice.

Acta Pharmacol Sin 2012 Aug 25;33(8):1047-54. Epub 2012 Jun 25.

Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, China.

Aim: DNAs containing unmethylated CpG motifs can stimulate innate and adaptive immunity. The aim of this study was to investigate the immunostimulatory and anti-neoplasm effects of a novel CpG oligodeoxynucleotide, ODN10, in tumor-bearing mice.

Methods: B16 melanoma-bearing C57BL/6 mice were administered ip or sc with ODN10 or conventional CpG ODN1826 on the indicated days post inoculation. The animal survival rate and the inhibitory effect on tumor growth were observed in vivo. B and T lymphocyte proliferation, natural killing cell cytotoxicity and the phagocytic ability of peritoneal macrophages from the animals were determined using [(3)H]-thymidine incorporation assay, 4-h (51)Cr release assay and neutral red chromometry method, respectively. The serum levels of IL-12, IL-4 and IgE were quantified using ELISA assays. Histological examination of tumor tissues was performed after HE staining, and the expression of PCNA, CD63, and CD80 in tumor tissues was analyzed with immunohistochemistry.

Results: ODN10 (1, 5 and 25 mg/kg) significantly inhibited the growth and metastasis of the tumor, and significantly prolonged the survival of tumor-bearing mice, as compared with ODN1826. The immune status was suppressed in tumor-bearing mice. Both ODN10 and ODN1826 significantly reversed the suppressed immunoactivities in tumor-bearing mice, which included promoting B and T lymphocyte proliferation, enhancing NK cell and peritoneal macrophage activities, inducing IL-12 secretion and inhibiting IL-4 and IgE secretion. Further, CpG ODNs decreased PCNA and CD63 expression while induced expression of CD80. ODN10 presented more potent activity, and displayed the most prominent immunostimulatory potential.

Conclusion: ODN10 produces prominent immunomodulatory effects on cellular immunity in tumor-bearing mice, which might help reverse the established Th2-type responses to the Th1-type responses, thus may be used as a potent anti-tumor immunotherapy agent or adjuvant.
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http://dx.doi.org/10.1038/aps.2012.54DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4011330PMC
August 2012

Enzyme-linked bridging assay method for the quantification of oligonucleotide-based drugs in biological matrices.

Nucleic Acid Ther 2011 Dec 7;21(6):403-13. Epub 2011 Nov 7.

Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing, People's Republic of China.

With ongoing efforts to develop oligonucleotide-based (ODN-based) therapeutics, there is a need for a sensitive, high-throughput method of quantification of ODN-based drugs in biological matrices. To overcome the insufficient sensitivity and time-consuming sample extraction procedures involved in conventional capillary gel electrophoresis (CGE) and high-performance liquid chromatography (HPLC), we developed a nucleic acid hybridization-based enzyme-linked bridging assay (ELBA), which shows significant advantages over CGE methods in evaluating ODN-based drugs in plasma and tissue: (1) It has higher sensitivity; (2) it involves easier sample extraction procedures; (3) it is suitable for many ODN-based drugs, even those with different secondary structures and modifications, including phosphorothioate oligonucleotide (PSODN), mixed backbones with 2'-O-Me (MBO), locked nucleic acid (LNA) modifications, and B- and C-type CpG sequences; and (4) it is highly selective, even during simultaneous quantification, with regard to intact ODNs and their 3'-metabolites. This universal design produces a rapid, sensitive, specific assay with minimal method development time. It is well suited to high-throughput analysis of various ODN-based drugs.
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http://dx.doi.org/10.1089/nat.2011.0319DOI Listing
December 2011

An improved on-line solid phase extraction coupled HPLC-MS/MS system for quantification of sifuvirtide in human plasma.

J Chromatogr B Analyt Technol Biomed Life Sci 2010 Jul 19;878(21):1893-8. Epub 2010 May 19.

Laboratory of Drug Metabolism and Pharmacokinetics, Beijing Institute of Radiation Medicine, Beijing 100850, China.

An improved liquid chromatographic method with on-line solid phase extraction (SPE) and tandem mass spectrometric detection was optimised for quantification of the anti-HIV peptide Sifuvirtide in human plasma. The SPE sorbents, loading buffer composition and other aspects of the on-line SPE column were investigated in detail for efficiently extracting the interesting peptides and simultaneously discarding the large amount of proteins. The gradient elution program was optimised on the analysis column to decrease the matrix effect and obtain excellent selectivity. The multiple charge ion at m/z 946.4 of Sifuvirtide was quantified by a linear ion trap mass spectrometer, operating in the positive mode, and selective reaction monitoring (SRM) acquisition. Method validation results demonstrated that the linear calibration curve covered a range of 6.1-6250 ng/mL, and the correlation coefficients (r(2)) were above 0.992. The lower limit of detection (LLOD) with a signal-to-noise (S/N) ratio higher than 10 was 6.1 ng/mL. The accuracy ranged from -7.6 to 10.6%, and the intra- and inter-batch precisions were less than 8.7% and 5.5%, respectively. Finally, more than nine hundred of samples from a clinical trial was completely analyzed using this on-line SPE coupled HPLC-MS/MS system in one single week, due to the rapid run-time of individual sample (6.5 min).
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http://dx.doi.org/10.1016/j.jchromb.2010.05.015DOI Listing
July 2010

Enhancing efficacy and mucosa-tropic distribution of an oral HIV-PsV DNA vaccine in animal models.

J Drug Target 2009 Dec;17(10):803-12

Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing, People's Republic of China.

A strategy combined the oral delivery route and bovine papillomavirus (BPV) pseudovirus (PsV)-based human immunodeficiency virus (HIV) DNA vaccine, which has been proven to enhance the mucosal immunization compared with the systemic immunization and in general does not induce effective mucosal immune responses. In this study, the immune responses against the BPV expressing HIV gp41 epitopes (ELDKWA, NWFDIT) after oral administration in Cynomolgus monkeys (Macaca fascicularis) were assessed, and the biodistribution of plasmid DNA encapsulated in the papillomavirus-like particles (VLPs) were evaluated in murine models. Results showed that oral immunization with the HIV-PsV DNA vaccine in monkey generated p24 and gp41 epitopes-specific serum IgG. Importantly, these induced antibodies had been shown to neutralize HIV-1 primary strain. In addition, the advantage of VLPs as vehicles delivering genes had been first revealed in biodistribution results. Therefore, orally administered HIV-PsV DNA vaccine was well-tolerated, enhanced the mucosa targeting property of the plasmid DNA, and reduced the nontargeting distribution, which indicate that it would reduce stress associated with systemic vaccination.
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http://dx.doi.org/10.3109/10611860903089768DOI Listing
December 2009

Quantification of puerarin in plasma by on-line solid-phase extraction column switching liquid chromatography-tandem mass spectrometry and its applications to a pharmacokinetic study.

J Chromatogr B Analyt Technol Biomed Life Sci 2008 Feb 4;863(1):55-63. Epub 2008 Jan 4.

Laboratory of Drug Metabolism and Pharmacokinetic, Beijing Institute of Radiation Medicine, Beijing 100850, China.

A highly precise, automatic and rapid method for quantification of puerarin in canine and human plasma using an on-line solid-phase extraction (SPE) column switching procedure combined with liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS) was developed. The eluent of SPE column consisted of acetonitrile/methanol/0.1% formic acid (25/25/50) at a flow rate of 0.2mLmin(-1). Puerarin was analyzed by a linear ion trap mass spectrometer, LTQ-MS, operating in the negative ion and selective reaction monitoring (SRM) acquisition mode. Method validation results demonstrated that the linear calibration curve covered a wide range of 0.39-400.00ngmL(-1), the correlation coefficients (r(2)) were above 0.999. The lower limit of detection (LLOD) with the signal-to-noise (S/N) ratio higher than 12 was 0.39ngmL(-1). The intra- and inter-batch precisions were less than 7.61% and 6.42%, respectively. The accuracy was well within the accept limit. The on-line SPE column switching HPLC-MS system was applied to pharmacokinetic (PK) study of puerarin after a single orally dose in beagles. And the optimum conditions were successfully utilized to quantify puerarin in human plasma, which indicated the feasibility and the reliability of this method for application in preclinical and clinical PK studies of isoflavone drugs.
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http://dx.doi.org/10.1016/j.jchromb.2007.12.023DOI Listing
February 2008

Structure-efficacy relationships of immunostimulatory activity of CpG-containing oligodeoxynucleotides on mouse spleen cells.

Acta Pharmacol Sin 2007 Oct;28(10):1637-44

Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, China.

Aim: To study the relationship between primary structures of oligodeoxynucleotides (ODN) containing unmethylated deoxycytidyldeoxyguanosine (CpG) dinucleotide motifs and their immunostimulatory activities in mouse spleen cells.

Methods: A series of CpG ODN with different primary structures were synthesized. Their capabilities to stimulate mouse spleen cell proliferation were determined by [3H]thymidine incorporation assay. Cytokine (interleukin [IL]-6, IL-12, and IFN-alpha) secretion spectra induced by CpG ODN were assessed by ELISA. The ability of CpG ODN to activate natural killer cells was evaluated by standard 4 h (51)Cr-release assay. Flow cytometry was utilized to examine the expressions of various lymphocyte surface molecules on diverse immunocytes. An effective CpG ODN for murine, ODN1826, was set as the template of modification and the positive control.

Results: The immunostimulatory activities of CpG ODN with different sequences and compositions varied markedly, both in character and in extent. It was useless for improving the immunostimulatory activity of ODN1826 by simply increasing the functional hexameric CpG motif number, modifying the site of CpG motifs, or changing the distance between multi-CpG motifs. However, an addition of a self-complementary palindrome structure at the 3'-end, but not the 5'-end of CpG ODN, aroused marked improvement in its activity. Several designed ODN had superior comprehensive immunostimulatory properties compared to ODN1826.

Conclusion: The immunostimulatory activity of a CpG ODN was relevant to its primary structure. It was useless for promoting immunostimulatory activity to simply change CpG motif number, space, or distance. The 3'-end palindrome structure of CpG ODN is associated with enhanced immunostimulatory activity.
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http://dx.doi.org/10.1111/j.1745-7254.2007.00628.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7091584PMC
October 2007
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