• 한국식물생명공학회:학술대회논문집
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• 한국식물생명공학회 2003년도 식물바이오벤처 페스티발
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• pp.162-162
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• 2003

• Biotechnology and Bioprocess Engineering:BBE
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• 제9권6호
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• pp.423-427
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• 2004

A simple purification procedure of bioactive human granulocyte macrophage colony stimulating factor (hGM-CSF) secreted in rice cell suspension culture has previously been described. In this study the protein was purified to apparent homogeneity with an overall yield of $80.1\%$ by ammonium sulfate precipitation and a single chromatographic step involving FPLCanion exchange chromatography. The purified hGM-CSF revealed at least five glycosylated forms ranging from $21.5{\~}29$ kDa, and its biological activity was independent of the glycosylation pattern. This is the first purification report of recombinant hGM-CSF to apparent homogeneity from rice cell suspension cultures.

• Biotechnology and Bioprocess Engineering:BBE
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• 제9권1호
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• pp.12-16
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• 2004

Partitioning of human granulocyte-macrophage colony stimulating factor (hGM-CSF) was achieved in the aqueous two-phase systems (ATPSs) using a crude extract of transgenic tobacco cell suspension culture. This study examined the effects of polyethylene glycol (PEG) molecular weight and concentration and the effects of sodium phosphate concentration in different PEG/sodium phosphate systems on the partition coefficient, K. The best ATPS system was 5% PEG 8,000/1.6 M sodium phosphate after 2 h of incubation at room temperature. In this system, hGM-CSF was partitioned in the PEG-rich phase with a yield of 57.99% and K$\_$hGM-CSF/ of 8.12. In another system, 3% PEG 10,000/1.6 M sodium phosphate, hGM-CSF was also partitioned primarily in the top phase with a yield of 45.66% and K$\_$hGM-CSF/ of 7.64 after 2 h of incubation at room temperature.

• Asian Pacific Journal of Cancer Prevention
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• 제16권1호
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• pp.299-301
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• 2015

Objectives: To assess safety and efficacy of JiSaiXin (Recombinant Human Granulocyte Colony Stimulating Factor Injection manufactured in China, G-CSF) 150ug per day for three days and whether this regimen could reduce the incidence of febrile neutropenia caused by chemotherapy. Method: From July 2014 to December 2014 patients treated by chemotherapy in our hospital were randomly divided into two groups: Group A with prophylactic use of G-CSF (JiSaiXin) 24 hours after chemotherapy for consecutive 3 days; and Group B with G-CSF (JiSaiXin) after neutropenia. Routine blood tests were performed 7 days and 14 days after chemotherapy. Results: A total of 100 patients fulfilled study criteria, and the incidence of severe neutropenia (grade III/IV) and the incidence of febrile neutropenia in Group A were lower than those in Group B. Nine patients were found severe neutropenia (grade III/IV) in Group B, but one in Group A, three febrile neutropenia in Group B, but 0 in Group A. Conclusions: This study suggested that prophylactic use of G-CSF (JiSaiXin) 150ug per day 24 hours after chemotherapy for consecutive 3 days is safe and could be effective for preventing febrile neutropenia in patients with chemotherapy.

• Journal of Microbiology
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• 제40권1호
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• pp.77-81
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• 2002

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a pleiotropic hematspoietic growth factor involved in the development of myeloid cells from bone marrow, and an activator of mature myeloid cells functioning in a variety of antimicrobial and inflammatory responses. Recently, recombinant GM-CSF is increasingly under clinical study for treatment of various diseases including cancer, infectious diseases and hematopoietic diseases as well as for an immune response modulator, In this study, we constructed a recombinant human GM-CSF (rhGM-CSF) expression plasmid with a pelB leader sequence and His. Tag under T7 promoter control. The expression construct was shown to produce a recombinant protein of 20 kDa in the 8M urea preparation, indicating the rhGM-CSF may be expressed as an insoluble inclusion form. The 20 kDa recombinant protein in 8M urea was transformed into the water-so1ub1e form by dialysis against PBS buffer (phosphate buffered saline). The soluble rhGM-CSF protein was shown to stimulate colony formation and cell proliferation in vitro, indicating that the rhGM-CSF could be refolded into its native form to show colony stimulating activity.

• Journal of Microbiology and Biotechnology
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• 제15권6호
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• pp.1267-1272
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• 2005

The effects of coexpression of GroEL/ES and DnaK/DnaJ/GrpE chaperones on the productivity of the soluble form of human granulocyte colony stimulating factor (hG-CSF) in E. coli were examined. Recombinant hG-CSF protein was coexpressed with DnaK/DnaJ/GrpE or GroEL/ES chaperones under the control of the araB or Pzt-1 promoter, respectively. The optimal concentration of L-arabinose for the expression of DnaK/DnaJ/GrpE was found to be 1 mg/ml. When L-arabinose was added at $OD_{600}$=0.2 (early-exponential phase), soluble hG-CSF production was greatly increased. In addition, it was observed that the DnaK/DnaJ/GrpE and GroEL/ES chaperones had no synergistic effects on preventing aggregation of hG-CSF protein. Consequently, by coexpression of the DnaK/DnaJ/GrpE chaperone, the signal intensity of the hG-CSF protein band in the soluble fraction of cell lysate was increased from $3.5\%\;to\;13.9\%$, and Western blot analysis also revealed about a 4-5-fold increase of production of soluble hG-CSF over the non-induction case of DnaK/DnaJ/GrpE.

• Biomolecules & Therapeutics
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• 제6권1호
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• pp.89-94
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• 1998

CJ50001 is a recombinant human granulocyte colony-stimulating facto, (rHuG-CSF) that stimulates the formation of neutrophils from bone marrow stem cells. It was produced in E. colt and purified through refolding and several processes. We produced CS970125(300) using purified C150001 and additives in order to test the stability of CJ50001. When CS970125(300) was stored at 50'S for more than 1 week, high molecular weight proteins were formed and those proteins were detected by non-reducing SDS-PAGE, gel filtration HPLC, and Western blot. Those proteins showed single band at the same position of CJ50001 in reducing SDS-PAGE. These data indicated that those high molecular weight proteins were the multimers of C150001. In biological assays, iu viro and in viro, the multimers did not have biological activity and inhibitory action to that of CJ 50001. The mutimers did not induce toxicity in mice and rats in acute toxicity test. These results suggest that if Cs970125(300) containing CJ50001 is stored at 5$0^{\circ}C$, CJ50001 will be the multimers that do not have biological activity and inhibitory effect to CJ50001 and do not induce acute toxicity.

• Clinical and Experimental Pediatrics
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• 제45권4호
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• pp.439-448
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• 2002

목 적 : 신생아 호중구 감소증이 합병된 신생아 패혈증 환아의 치료에 있어서 부가적으로 rhG-CSF(recombinant human granulocye colony-stimulating factor)를 투여함에 있어서 서로 다른 용량의 rhG-CSF를 투여함으로써 나타나는 신생아 패혈증에 합병된 호중구 감소증의 치료와 환아들의 생존율에 미치는 영향을 평가, 비교하려고 하였다(group I/II형 연구). 방 법: 1995년 10월부터 1996년까지 신생아 호중구 감소증이 합병된 신생아 패혈증 환아는 모두 10명으로 이들에게는 $10{\mu}g/kg$을 피하주사 하였고(rhG-CSF $10{\mu}g/kg$ 투여군), 1996년 10월부터 1997년 9 월까지는 신생아 호중구 감소증이 합병된 신생아 패혈증 환아는 모두 12명으로 이들에게는 rhG-CSF를 $5{\mu}g/kg$ 피하주사 하였다(rhG-CSF $5{\mu}g/kg$ 투여군). 각 군의 호중구 증가 정도와 임상적 결과를 서로 비교하였다. 결 과 : RhG-CSF $10{\mu}g/kg$ 투여군은 조발형 신생아 패혈증 1명과 지발형 신생아 패혈증 9명으로 이루어졌고, 모두에게 호중구 감소증이 합병되었다. rhG-CSF $5{\mu}g/kg$ 투여군은 조발형 신생아 패혈증 1명과 지발형 신생아 패혈증 11명이 대상이 되었고, 이들 모두 호중구 감소증이 합병되었다. 두 군간에 출생체중, 재태주령, 항생제 사용, 신생아 패혈증 시기에 기계적 환기요법 투여, 승압제로 dopamine 투여 또는 다른 지지적 요법의 투여에 있어서 차이가 없었다. rhG-CSF 투여 전의 순 호중구 수(ANC)는 rhG-CSF $10{\mu}g/kg$ 투여군이 $1,065{\pm}89$($mean{\pm}SEM$), $5{\mu}g/kg$ 투여군이 $1,053{\pm}131$로 차이가 없었다. 투여 후의 ANC의 증가는 rhG-CSF $10{\mu}g/kg$ 투여군과 $5{\mu}g/kg$ 투여군에서 각각 투여 후 24시간에 7배, 6배, 투여 후 48시간에 10배, 6배, 투여 후 72시간에 8배, 4배, 투여 후 120시간에 8배, 4배로 투여 전에 비하여 두 군 모두에서 각 시간에 의미 있는 증가를 보였다(repeated measure ANOVA와 Kruskall-Wallis test, within subjects effect). 그러나 두 군 간의 차이는 투여 후 48시간에 ANC 최고치에서만 의미 있는 차이를 보였다(student t-test와 Wilcoxon rank sum test). 단핵구 수도 이 기간 동안 의미 있게 증가하였으나 정상범위를 넘지는 않았다. rhG-CSF $10{\mu}g/kg$ 투여군에서 1명의 환아가 자의 퇴원하였고, 1명의 환아가 사망하여 신생아 패혈증에서 회복하여 문제없이 퇴원한 생존율은 자의 퇴원 환아를 제외한 9명 중 8명으로 88.9％였고, rhG-CSF $5{\mu}g/kg$ 투여 군은 12명 중 10명이 생존하여 생존율은 83.3%였다. 두 군 모두에서 특별한 독성이나 부작용은 관찰되지 않았다. 결 론 : RhG-CSF의 투여는 호중구 감소증이 합병된 극심한 신생아 패혈증 환아에서 호중구의 증가를 일으켰다. 두가지 투여 용량에 따르는 효과는 거의 동일하였으며, 단지 투여 후 48시간에 ANC 최고치에서만 의미 있는 차이가 있었다. 두 군의 생존율은 80%이상이었다. 이와 같은 호중구 감소증이 합병된 신생아 패혈증에서 rhG-CSF의 투여 효과는 골수 억압이나 호중구 소모에 의하여 호중구 감소증이 합병된 신생아 패혈증에서 시기적으로 적절히 투여하면 효과적인 치료를 이룰 수 있다는 것을 시사한다. 향후 rhG-CSF의 효능과 부작용에 대하여 무작위 대조실험이 필요시 된다.

• Animal cells and systems
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• 제16권3호
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• pp.207-214
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• 2012

Burns are one of the most devastating forms of trauma and wound healing is a complex and multicellular process, which is executed and regulated by signaling networks involving numerous growth factors, cytokines, and chemokines. Recombinant human granulocyte macrophage colony-stimulating factor (rhGM-CSF) was specifically produced from rice cell culture through use of a recombinant technique in our laboratory. The effect of rhGM-CSF on promotion of deep second-degree burn wound healing on the back skin of a hamster model was evaluated through a randomized and double-blind trial. As macroscopic results, hamster skins of the experimental groups showed earlier recovery by new epidermis than the control groups. Immunohistochemical reactions of proliferating cell nuclear antigen and transforming growth factor-b1, which are indicators of cell proliferation, were more active in the experimental group, compared with the control group. On electron microscopy, basal cells in the epidermis of the experimental group showed oval nuclei, prominent nucleoli, numerous mitochondria and abundant free ribosomes. In addition, fibroblasts contained well-developed rough endoplasmic reticulum with dilated cisternae. Bundles of collagen fibrils filled the extracellular spaces. Particularly, ultrastructural features indicating active metabolism for regeneration of injured skin at 15 days after burn injury, including abundant euchromatin, plentiful free ribosomes, and numerous mitochondria, were observed. These findings suggest that use of rhGM-CSF could result in accelerated deep second-degree burn wound healing in animal models.

• Journal of Pharmaceutical Investigation
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• 제31권2호
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• pp.89-94
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• 2001

The pharmacokinetics of recombinant human granulocyte colony stimulating factor (rhG-CSF) following intravenous (i.v.), intramuscular (i.m.) and subcutaneous (s.c.) administration of HM1041l-lyo and HM10411-liq (lyophilized and liquid formulations of rhG-CSF, recently under development by Hanmi Pharmaceutical Company) were studied in rats, and compared with that of Filgrastim (conventional formulation of rhG-CSF on market). The plasma concentration of rhG-CSF was quantified using a specific ELISA. The pharmacokinetic parameters of rhG-CSF, after i.v., i.m. and s.c. administration of Filgrastim, HM1041l-lyo and HM1041l-liq to rats at a rhG-CSF dose of $10\;{\mu}g/kg$, were almost identical among the three formulations. No dose-dependency was observed in the pharmacokinetic parameters of rhG-CSF following i.v. administration in the dose range of $5{\sim}100\;{\mu}g/kg$. rhG-CSF, after i.v. administration of the three preparations at a dose of $10\;{\mu}g/kg$ to rats, was detected at low levels in all of the body tissues with highest tissue/plasma ratio of $0.46{\sim}0.51$ for the kidney at 30 min after the administration. The pharmacokinetics of rhG-CSF, after i.v. administration to mice at a dose of $10\;{\mu}g/kg$, were comparable among the three formulations. In conclusion, HM10411-lyo and HM10411-liq exhibited similar pharmacokinetics for rhG-CSF with Filgrastim regandless of animal species. Considering the fact that HM10411 series, contrary to Filgrastim, are proteins lacking a methionine residue, the methionine moiety in rhG-CSF molecule does not appear to influence the pharmacokinetics of the protein significantly.