Sinil Pharmaceutical Co.

Cheongju, South Korea

Sinil Pharmaceutical Co.

Cheongju, South Korea
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Chi W.-J.,National Institute of Biological Resources Incheon | Kim H.,Sinil Pharmaceutical Co. | Yoo H.,Sinil Pharmaceutical Co. | Kim Y.P.,Sinil Pharmaceutical Co. | Hong S.-K.,Myongji University
Biotechnology and Bioprocess Engineering | Year: 2016

New structural designs of antibody fragments have considerable biotechnological and therapeutic potential. In this study, we describe the construction and functional expression of a cetuximab-based antibody fragment (scFv-CH3, minibody) that exhibits activity against human colon cancer. Heterologous expression in Escherichia coli (E. coli) was improved by optimizing the host cells, signal peptides, induction conditions, and culture media. The recombinant minibody was expressed successfully in the periplasm of E. coli BL21(DE3) and purified by immobilized metal affinity chromatography using a Ni2+-NTA resin. The purified minibody showed high binding affinity to cell-surface epidermal growth factor receptor (EGFR) and exhibited inhibition of EGFR-mediated signal transduction in the human colon cancer cell line HT29 in a similar way by the cetuximab. The minibody also showed significant level of anti-cancer ability in the HT29 colorectal cancer xenograft model, which was lower than that by the cetuximab. © 2016, The Korean Society for Biotechnology and Bioengineering and Springer-Verlag Berlin Heidelberg.


Kim Y.P.,Sinil Pharmaceutical Co. | Park D.,Ajou University | Kim J.J.,Ajou University | Choi W.-J.,Sinil Pharmaceutical Co. | And 11 more authors.
PLoS ONE | Year: 2014

Cetuximab, a chimeric monoclonal antibody developed for targeting the Epidermal Growth Factor Receptor (EGFR), has been intensively used to treat cancer patients with metastatic colorectal cancer and head and neck cancer. Intact immunoglobulin G (IgG) antibody like cetuximab, however, has some limitations such as high production cost and low penetration rate from vasculature into solid tumor mass due to its large size. In attempt to overcome these limitations, we engineered cetuximab to create single chain variable fragments (scFv-CH3; Minibody) that were expressed in bacterial system. Among three engineered minibodies, we found that MI061 minibody, which is composed of the variable heavy (VH) and light (VL) region joined by an 18-residue peptide linker, displays higher solubility and better extraction properties from bacterial lysate. In addition, we validated that purified MI061 significantly interferes ligand binding to EGFR and blocks EGFR's phosphorylation. By using a protein microarray composed of 16,368 unique human proteins covering around 2,400 plasma membrane associated proteins such as receptors and channels, we also demonstrated that MI061 only recognizes the EGFR but not other proteins as compared with cetuximab. These results indicated that engineered MI061 retains both binding specificity and affinity of cetuximab for EGFR. Although it had relatively short half-life in serum, it was shown to be highly significant anti-tumor effect by inhibiting ERK pathway in A431 xenograft model. Taken together, our present study provides compelling evidence that engineered minibody is more effective and promising agent for in vivo targeting of solid tumors. © 2014 Kim et al.


Choi J.N.,Sinil Pharmaceutical Co. | Choi Y.-H.,Sinil Pharmaceutical Co. | Lee J.-M.,Sinil Pharmaceutical Co. | Noh I.C.,Sinil Pharmaceutical Co. | And 3 more authors.
Natural Product Research | Year: 2012

Trachelospermum jasminoides (Apocynaceae) has pharmacological effects that include anti-inflammatory, anti-bacterial and anti-viral activities, which have been observed from various studies. Of these pharmacological effects, the anti-inflammatory capacity of compounds from T. jasminoides is not yet known exactly. In this study, we investigated the compound that can be used for the suppression of lipopolysacchaide (LPS) stimulated inflammatory responses in macrophages among the five isolated compounds. β-sitosterol-β-D- glucoside (1) was found to reduce nitric oxide (NO) production from LPS-induced RAW 264.7 cells the most. In addition, compound 1 strongly inhibited the interleukin 6 (IL-6) activities of stimulated macrophages. Treatment of RAW 264.7 cells with compound 1 reduced secretion of inflammatory elements including tumour necrosis factor - alpha (TNF-α) and interleukin 1 beta (IL-1β). Thus, compound 1 may be a useful candidate for the development of new drugs to treat endotoxemia and inflammation accompanied by the overproduction of NO. © 2012 Copyright Taylor and Francis Group, LLC.


Choi Y.-H.,Sinil Pharmaceutical Co. | Yoo H.-J.,Sinil Pharmaceutical Co. | Noh I.C.,Sinil Pharmaceutical Co. | Lee J.-M.,Sinil Pharmaceutical Co. | And 3 more authors.
Archives of Pharmacal Research | Year: 2012

The inhibition of Interleukin-1beta (IL-1β) is of substantial interest for the treatment of rheumatoid arthritis. Using an in vitro assay with RAW 264.7 cells, oxo-acetic acid 2-ethoxy-4-(3- hydroxy-2-oxopropyl) phenyl ester (1) was isolated from the roots of Paeonia suffruticosa Andrews as an inhibitor of IL-1β with an IC50 value of 56 μM. Compound 1 is a novel phenylesteric compound from P. suffruticosa Andrews. Compound 1 was shown to inhibit the production of pro-inflammatory cytokines in RAW 264.7 cells. Thus, a possible new action of novel compound is provided explaining the anti-rheumatoid arthritic properties of P. suffruticosa Andrews.


PubMed | Ajou University, Sinil Pharmaceutical Co. and Konkuk University
Type: Journal Article | Journal: Applied microbiology and biotechnology | Year: 2016

Fragment engineering of monoclonal antibodies (mAbs) has emerged as an excellent paradigm to develop highly efficient therapeutic and/or diagnostic agents. Engineered mAb fragments can be economically produced in bacterial systems using recombinant DNA technologies. In this work, we established recombinant production in Escherichia coli for monovalent antigen-binding fragment (Fab) adopted from a clinically used anticancer mAB drug cetuximab targeting epidermal growth factor receptor (EGFR). Recombinant DNA constructs were designed to express both polypeptide chains comprising Fab in a single vector and to secrete them to bacterial periplasmic space for efficient folding. Particularly, a C-terminal engineering to confer an interchain disulfide bond appeared to be able to enhance its heterodimeric integrity and EGFR-binding activity. Conformational relevance of the purified final product was validated by mass spectrometry and crystal structure at 1.9 resolution. Finally, our recombinant cetuximab-Fab was found to have strong binding affinity to EGFR overexpressed in human squamous carcinoma model (A431) cells. Its binding ability was comparable to that of cetuximab. Its EGFR-binding affinity was estimated at approximately 0.7nM of Kd in vitro, which was quite stronger than the binding affinity of natural ligand EGF. Hence, the results validate that our construction could serve as an efficient platform to produce a recombinant cetuximab-Fab with a retained antigen-binding functionality.


Chi W.-J.,Myongji University | Chi W.-J.,Sinil Pharmaceutical Co. | Park J.-S.,Myongji University | Kang D.-K.,Dankook University | Hong S.-K.,Myongji University
Applied Biochemistry and Biotechnology | Year: 2014

A Gram-negative, aerobic, motile, rod-shaped, agarolytic bacterium, designated as H7, was isolated from a coastal seawater sample. This strain grows at pH 6.0-8.0, temperature of 15-40°C, and at an NaCl concentration of 1-7 % (w/v). Ubiquinone-8 was the predominant respiratory quinone, and the DNA G+C content was 45.82 mol%. Analysis of the 16S rRNA sequence suggests that strain H7 belongs to the genus Pseudoalteromonas. DNA-DNA hybridization analysis showed DNA relatedness of as low as 55.42 and 40.27 % with its nearest phylogenetic neighbors Pseudoalteromonas atlantica IAM12927T and Pseudoalteromonas espejiana NCIMB2127T, respectively, which led us to name H7 Pseudoalteromonas hodoensis sp. nov. The type strain is H7T (=DSM25967T = KCTC23887T). An agarase (AgaA7) was purified to homogeneity from the cell-free culture broth of H7 through many steps of chromatography. Purified AgaA7 had an apparent molecular weight of 35 kDa, with a distinct NH2-terminal sequence of Ala-Asp-Ala-Thr-X-Pro (X, any amino acid) from the reported proteins, implying that it is a novel enzyme. The optimum pH and temperature for agarase activity were 7.0 and 45°C, respectively. Thin-layer chromatography analysis, mass spectrometry, and enzyme assay using p-nitrophenyl-α/β-D-galactopyranoside revealed that AgaA7 is both an exo- and endo-type β-agarase that degrades agarose into neoagarotetraose, neoagarohexaose, and neoagarooctaose (minor). © 2014 Springer Science+Business Media.


Park D.Y.,Myongji University | Chi W.-J.,Sinil Pharmaceutical Co. | Park J.-S.,Myongji University | Chang Y.-K.,Korea Advanced Institute of Science and Technology | Hong S.-K.,Myongji University
Applied Biochemistry and Biotechnology | Year: 2014

An agarase gene (agaH71) was identified from Pseudoalteromonas hodoensis, an agar utilizing marine bacterium. The nucleotide sequence revealed that AgaH71 had significant homology to glycosyl hydrolase (GH) 16 agarases. agaH71 encodes a primary translation product (32.7 kDa) of 290 amino acids, including a 21-amino-acid signal peptide. The entire AgaH71 was expressed in a fused protein with glutathione-S-transferase (GST) at its N-terminal (GST-AgaH71) in Escherichia coli. Purified GST-AgaH71 had an apparent molecular weight of 59 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), which was consistent with the calculated molecular weight (58.7 kDa). Agarase activity of the purified protein was confirmed by zymogram assay. GST-AgaH71 could hydrolyze p-nitrophenyl-β-d-galactopyranoside, but not p-nitrophenyl-α-d-galactopyranoside. The optimum pH and temperature for GST-AgaH71 were 6.0 and 45 °C, respectively. GST-AgaH71 retained more than 95 and 90 % of its initial activity at 40 and 45 °C after heat treatment for 60 min, respectively. The Km and Vmax for agarose were 28.33 mg/ml and 88.25 U/mg, respectively. GST-AgaH71 did not require metal ions for its activity, but severe inhibition by divalent metal ions was observed. Thin-layer chromatography (TLC) analysis, mass spectrometry, and nuclear magnetic resonance (NMR) spectrometry of the GST-AgaH71 hydrolysis products revealed that GST-AgaH71 is an endo-type β-agarase that hydrolyzes agarose into predominantly neoagarotetraose and small proportions of neoagarobiose and neoagarohexaose. © 2014, Springer Science+Business Media New York.


The present invention relates to a pharmaceutical composition for preventing and treating an inflammatory disease, including a mixture of extracts from Trachelospermi Caulis and Paeonia Suffruticosa Andrews as an active ingredient and method for preparing the same. Also, a quasi-drug composition, a health functional food composition and a cosmetic composition, all based on the mixture, are provided for preventing or improving inflammation. In addition, the present invention relates to a method for treating an inflammatory disease by administering the pharmaceutical composition to a subject suspected of having the inflammatory disease. Containing the extract mixture, the composition exhibits excellent anti-inflammatory activity and edema-suppressing activity, compared to individual extracts, and thus can be applied to the prevention, treatment or improvement of an inflammatory disease. As natural materials, the extracts can be used as a safe therapeutic relatively free of fungal infection or other side-effects, compared to synthetic medicines. In addition, the known physiological activities of the extracts from Trachelospermi Caulis and Paeonia Suffruticosa Andrews including antibacterial activity, bone reinforcement, antiphlogistic activity, blood nourishment, vigoration, etc. may bring about a synergistic effect on the prevention, treatment and improvement of an inflammatory disease.


Trachelospermum jasminoides (Apocynaceae) has pharmacological effects that include anti-inflammatory, anti-bacterial and anti-viral activities, which have been observed from various studies. Of these pharmacological effects, the anti-inflammatory capacity of compounds from T. jasminoides is not yet known exactly. In this study, we investigated the compound that can be used for the suppression of lipopolysacchaide (LPS) stimulated inflammatory responses in macrophages among the five isolated compounds. -sitosterol--D-glucoside (1) was found to reduce nitric oxide (NO) production from LPS-induced RAW 264.7 cells the most. In addition, compound 1 strongly inhibited the interleukin 6 (IL-6) activities of stimulated macrophages. Treatment of RAW 264.7 cells with compound 1 reduced secretion of inflammatory elements including tumour necrosis factor - alpha (TNF-) and interleukin 1 beta (IL-1). Thus, compound 1 may be a useful candidate for the development of new drugs to treat endotoxemia and inflammation accompanied by the overproduction of NO.


PubMed | Sinil Pharmaceutical Co.
Type: Journal Article | Journal: Archives of pharmacal research | Year: 2012

The inhibition of Interleukin-1beta (IL-1) is of substantial interest for the treatment of rheumatoid arthritis. Using an in vitro assay with RAW 264.7 cells, oxo-acetic acid 2-ethoxy-4-(3-hydroxy-2-oxopropyl) phenyl ester (1) was isolated from the roots of Paeonia suffruticosa Andrews as an inhibitor of IL-1 with an IC(50) value of 56 M. Compound 1 is a novel phenylesteric compound from P. suffruticosa Andrews. Compound 1 was shown to inhibit the production of pro-inflammatory cytokines in RAW 264.7 cells. Thus, a possible new action of novel compound is provided explaining the anti-rheumatoid arthritic properties of P. suffruticosa Andrews.

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