Penicillium brevi-compactum을 이용한 면역억제제 Mycophenolic Acid 발효에서 탄소원 및 질소원의 영향

노용택,Rho, Yong-Taek 한국미생물학회 2011 미생물학회지 Vol.47 No.3

본 연구에서는 탄소원 및 질소원의 이용 패턴과 mycophenolic acid의 생성 패턴을 확인하기 위하여 먼저 5 L 발효조에서 mycophenolic acid 발효의 경시적 변화를 조사하였다. 그 다음에는 여러 가지 탄소원들이 세포 생장과 mycophenolic acid 생산에 미치는 효과를 조사하였다. 과당이 mycophenolic acid 발효에 가장 좋은 탄소원이었지만, 값이 고가인 단점이 있어서, 과당을 지니고 있는 설탕이 주성분인 당밀을 탄소원으로 사용한 결과 mycophenolic acid의 산업적 생산에 가장 좋은 것으로 확인되었다. 당밀을 첨가한 실험구는 포도당을 탄소원으로 사용한 대조구에 비하여 발효 생산성이 2배 이상 증가하였다. 양호한 세포 생장과 높은 mycophenolic acid 생산을 얻기 위하여 다양한 무기질소원과 유기질소원에 대한 실험을 실시하였다. 무기질소원들 가운데 요소는 암모늄 형태의 질소원을 천천히 공급함으로써 생장과 mycophenolic acid 생산을 저해하는 배양액의 급격한 pH 하락을 일으키지 않았다. 요소를 첨가한 실험구는 질산암모늄을 첨가한 대조구보다 발효 생산성이 3.6배 증가하였다. 카제인, 펩톤, casamino acid 같은 우유 단백질 유래 유기질소원들은 대조구에 비하여 mycophenolic acid 발효 생산성을 최고 3.4배까지 증진시켰다. 카제인의 가수분해물인 펩톤과 casamino acid는 mycophenolic acid 발효 생산성뿐만 아니라 세포 생장도 촉진하였다. Mycophenolic acid blocking the synthesis of xanthosine monophosphate is a nonnucleoside inhibitor of inosine monophosphate dehydrogenase. Therefore mycopholoic acid is a drug currently used as immunosuppressive agent in transplantation of heart, kidney and liver. Mycophenolic acid has been industrially produced through fermentation process by fungus Penicillium brevi-compactum. In this study, the profile of mycophenolic acid fermentation was observed in 5L-jar fermentor to investigate the utilization of carbon and nitrogen sources and the production of mycophenolic acid. It was investigated that what kind of carbon sources was better to cell growth and mycophenolic acid production. Fructose was the best carbon source for mycophenolic acid fermentation, but it is the most expensive one. Thereafter molasses containing sucrose as the supply source of fructose was confirmed to be the best carbon source for the industrial production. Use of molasses increased the fermentation yield of mycophenolic acid more than two times higher than glucose. It was confirmed that urea was the best inorganic nitrogen source, which did not give rise to sudden drop of culture pH. Addition of urea increased the fermentation yield of mycophenolic acid about 3.6 times higher than addition of ammonium nitrate as control. Casein, peptone and casamino acid originated from milk protein increased the fermentation yield of mycophenolic acid about 3.4 times higher than control. Peptone and casamino acid, which are casein hydrolysates, increased cell growth considerably as well.

Effect of Ginsenoside Rc on the Pharmacokinetics of Mycophenolic Acid, a UGT1A9 Substrate, and its Glucuronide Metabolite in Rats

( So-young Park ),( Ji-hyeon Jeon ),( Su-nyeong Jang ),( Im-sook Song ),( Kwang-hyeon Liu ) 한국질량분석학회 2021 Mass spectrometry letters Vol.12 No.2

Previous in vitro studies have demonstrated that ginsenoside Rc inhibits UGT1A9, but there are no available data to indicate that ginsenoside Rc inhibits UGT1A9 in vivo. The effect of single and repeated intravenous injection of ginsenoside Rc was evaluated on the pharmacokinetics of mycophenolic acid. After injection of ginsenoside Rc (5 mg/kg for one day or 3 mg/kg for five days), 2-mg mycophenolic acid was intravenously injected, and the pharmacokinetics of mycophenolic acid and mycophenolic acid-β-glucuronide were determined. Concentrations of mycophenolic acid and its metabolite from rat plasma were analyzed using a liquid chromatography-triple quadrupole mass spectrometry. Single or repeated pretreatment with ginsenoside Rc had no significant effects on the pharmacokinetics of mycophenolic acid (P > 0.05): The mean difference in maximum plasma concentration (C_max) and area under the concentration-time curve (AUC_inf) were within 0.83- and 0.62-fold, respectively, compared with those in the absence of the ginsenoside Rc. These results indicate that ginsenoside Rc has a negligible effect on the disposition of mycophenolic acid in vivo despite in vitro findings indicating that ginsenoside Rc is a selective UGT1A9 inhibitor. As a result, ginsenoside Rc has little possibility of interacting with drugs that are metabolized by UGT1A9, including mycophenolic acid.

Monitoring of Mycophenolic Acid Trough Concentration in Kidney Transplant under Cyclosporine Is Beneficial in Reducing Acute Rejection within 1 Year

유진수,이교원,박재범,김성주 대한이식학회 2018 Korean Journal of Transplantation Vol.32 No.4

Background: This study was designed to analyze the clinical usefulness of mycophenolic acid trough concentration monitoring in kidney transplantation patients who were maintained with cyclosporine. Methods: The data of patients who underwent mycophenolic acid trough concentration monitoring after their first kidney transplant between November 2006 and August 2013 and were prescribed with cyclosporine, mycophenolate, and methylprednisolone were reviewed retrospectively. Cox analysis was used to analyze the risk factors for acute rejection within 1 year post-transplantation. Results: Among 90 patients, 41 (45.6%) achieved both the target levels of cyclosporine and mycophenolic acid, while three patients (3.3%) failed to achieve the target level of either cyclosporine or mycophenolic acid. Nine patients (10.0%) only achieved the mycophenolic acid target level and 37 patients (41.1%) only achieved the cyclosporine target level. While patients who achieved only the mycophenolic acid target concentration had no statistically increased risk compared to patients who achieved both target levels (hazard ratio [HR], 1.569; 95% confidence interval [CI], 0.316 to 7.778; P=0.581), patients who only achieved the cyclosporine target concentration showed an increased risk of rejection compared to the both achievement group (HR, 4.112; 95% CI, 1.583 to 10.683; P=0.004). Patients who had no achievement in the target levels showed significantly increased rejection risk compared to the patients who achieved both target levels (HR, 17.811; 95% CI, 3.072 to 103.28; P=0.001). Conclusions: Mycophenolic acid trough concentration monitoring combined with cyclosporine trough concentration monitoring is useful for avoiding acute cellular rejection if the first 1 year post-transplantation.

Mycophenolic Acid가 사람 Jurkat 세포의 자멸사에서 활성 산소 생성에 미치는 영향

이동규(Dong Kyu Lee),이호균(Ho Kyun Lee),최수진나(Soo Jin Na Choi),정상영(Sang Young Chung) 대한외과학회 2008 Annals of Surgical Treatment and Research(ASRT) Vol.75 No.3

Purpose: Mycophenolic acid (MPA) is the active agent of mycophenolate mofetil (MMF), which is an immunosuppressive drug. MPA is a selective inhibitor of inosine monophosphate dehydrogenase. The aim of this study was for demonstrate that mycophenolic acid induces apoptosis in human Jurkat cells via the generation of reactive oxygen species (ROS). Methods: The cells were cultured in the presence or absence of MPA. Flow cytometric analysis was performed after propidium iodide staining. Western blotting for caspase 3, Bcl-2 and Bax proteins was also performed. Results: MPA decreased the viability of Jurkat cells in a dose- and time-dependent manner. The MPA induced apoptotic cell death displayed nuclear fragmentation and sub G0/G1 phase arrest in the Jurkat cells. The expression of caspase-3 proteases in the MPA treated-Jurkat cells increased in a time-dependent manner. Treatment with MPA resulted in increased ROS generation in the Jurkat cells. There was a decreased expression of Bcl-2 and an increased expression of Bax protein in the MPA treated Jurkat cells. Conclusion: This result suggests that MPA-induced cytotoxicity is associated with a direct increase of both ROS generation and the expression of Bax protein.

Mycophenolic Acid가 올레산으로 유도한 흰쥐 혈관 평활근 세포의 증식에 미치는 영향

안형준(Hyung Joon Ahn),박제현(Jehyun Park),송재숙(Jae Sook Song),주만기(Man Ki Ju),김명수(Myoung Soo Kim),하헌주(Hunjoo Ha),송기호(Ki Ho Song),김유선(Yu Seun Kim) 대한외과학회 2007 Annals of Surgical Treatment and Research(ASRT) Vol.72 No.3

Purpose: Vascular smooth muscle cell (VSMC) proliferation plays an important role in the development and progression of chronic allograft vasculopathy. Mycophenolic acid (MPA) inhibits various mesenchymal cell proliferation, and reactive oxygen species (ROS) are involved in the anti-pro-liferative effect of MPA. In this study, we investigated the effects of MPA on oleic acid (OA)-induced VSMC proliferation and also the role of ROS in these processes. Methods: Primary cultured rat VSMCs from Sprague-Dawley were stimulated with OA 100μM. MPA 0.1∼10μM and N-acetylcystein (NAC) 5 mM were administered 1 hour before adding the OA. Cell proliferation was measured by Methylthiazoletetrazolium (MTT) assay, proliferating cell nuclear antigen (PCNA) expression by Western blot analysis, and dichlorofluorescein (DCF)-sensitive cellular ROS by flow cytometry. Results: OA at 100μM significantly increased MTT level by 1.6-fold as well as PCNA expression at 48 hours in rat VSMCs. OA also induced DCF-sensitive cellular ROS by 1.6-fold at 5 minutes and the increment of cellular ROS remained for up to 1 hour. MPA at above 1μM inhibited OA- induced VSMC proliferation and cellular ROS in a dosedependent manner. NAC 5 mM also inhibited OA-induced rat VSMC activation. Conclusion: These results suggest that MPA inhibits OA-induced VSMC proliferation partially through the inhibition of cellular ROS.

Mycophenolic Acid가 PDGF에 의해 증식 유도된 흰쥐 혈관평활근 세포에서 NAD(P)H Oxidase, 세포내 활성산소족 및 Mitogen-activated Protein Kinases에 미치는 영향

박제현 ( Je Hyun Park ),하헌주 ( Hun Joo Ha ),김명수 ( Myoung Soo Kim ),허규하 ( Kyu Ha Huh ),김유선 ( Yu Seun Kim ) 대한신장학회 2004 Kidney Research and Clinical Practice Vol.23 No.4

Mycophenolic Acid에 의해 유도된 Jurkat 세포주 세포자멸사에서 Heme Oxygenase-1의 발현이 미치는 영향

이호균(Ho Kyun Lee),최수진나(Soo Jin Na Choi) 대한외과학회 2010 Annals of Surgical Treatment and Research(ASRT) Vol.78 No.6

Purpose: This study demonstrates that pharmacologic induction of heme oygenase-1 (HO-1) along with catalytic activation significantly modulated apoptosis of Jurkat cells induced by mycophenolic acid (MPA). Methods: Cells were cultured with the presence or absence of MPA. Flow cytometric analysis was performed after propidium iodide staining. Western blotting of HO-1, Bcl, and Bax was also performed. Cells were stained 4’-6-Diamidino-2-phenylindole (DAPI) and measured by flow cytometry in the absence or presence of CoPPIX. Results: Treatment of MPA decreased cell viability in a dose- and time-dependent manner. MPA-induced cell death was confirmed as apoptosis characterized by sub G0/G1 phase arrest. Expression of HO-1 assumes a pattern of decline after rising at the initial phase. CoPPIX, HO-1 inducer, significantly inhibited the cisplatin-induced apoptosis. Treatment of MPA resulted in reactive oxygen species (ROS) generation in Jurkat cells. CoPPIX attenuated ROS production in MPA-treated cells. Conclusion: This result suggests that the protective mechanism of HO-1 on MPA-induced cytotoxicity is associated with direct inhibition of ROS generation and mitochondrial permeability transition.

Hydroxamic Acid Derivatives of Mycophenolic Acid Inhibit Histone Deacetylase at the Cellular Level

BATOVSKA, Daniela I.,KIM, Dong Hoon,MITSUHASHI, Shinya,CHO, Yoon Sun,KWON, Ho Jeong,UBUKATA, Makoto Japan Society for Bioscience, Biotechnology, and A 2008 Bioscience, biotechnology, and biochemistry Vol.72 No.10

<P>Mycophenolic acid (MPA, <B>1</B>), an inhibitor of IMP-dehydrogenase (IMPDH) and a latent PPARγ agonist, is used as an effective immunosuppressant for clinical transplantation and recently entered clinical trials in advanced multiple myeloma patients. On the other hand, suberoylanilide hydroxamic acid (SAHA), a non-specific histone deacetylase (HDAC) inhibitor, has been approved for treating cutaneous T-cell lymphoma. MPA seemed to bear a cap, a linker, and a weak metal-binding site as a latent inhibitor of HDAC. Therefore, the hydroxamic acid derivatives of mycophenolic acid having an effective metal-binding site, mycophenolic hydroxamic acid (MPHA, <B>2</B>), 7-<I>O</I>-acetyl mycophenolic acid (7-<I>O</I>-Ac MPHA, <B>3</B>), and 7-<I>O</I>-lauroyl mycophenolic hydroxamic acid (7-<I>O</I>-L MPHA, <B>4</B>) were designed and synthesized. All these compounds inhibited histone deacetylase with IC<SUB>50</SUB> values of 1, 0.9 and 0.5 μ<SMALL>M</SMALL>, and cell proliferation at concentrations of 2, 1.5 and 1 μ<SMALL>M</SMALL>, respectively.</P>

Simultaneous Determination of Mycophenolic Acid and Its Metabolites by HPLC and Pharmacokinetic Studies in Rat Plasma and Bile

Jun-wei Gao,Zhi-hai Peng,Xiao-yu Li,Bo Sun,Yan-kun Guo,Gao-lin Liu 대한약학회 2011 Archives of Pharmacal Research Vol.34 No.1

In this study, we determined the pharmacokinetics of mycophenolic acid (MPA) and its metabolites mycophenolic acid glucuronide (MPAG) and acyl glucuronide (AcMPAG) in rat plasma and bile, using a newly developed HPLC method. Protein precipitation and liquid-liquid extraction were employed in sample preparation of plasma and bile, respectively. The HPLC methods included a gradient elution consisting of acetonitrile and phosphate buffer at a flow rate of 1.2 mL/min, with UV detection at 254 nm. The HPLC method was found to be sensitive and linear (r^2 ≥ 0.9991, 1.0-128.0 and 0.25-32.0 mg/L for MPA; 1.0-128.0 and 0.5-64.0 mg/L for MPAG; 0.25-32.0 and 1.0-128.0 mg/L for AcMPAG in rat plasma and bile, respectively), precise (both the intra- and inter-day variability were ≤ 6.8%), and accurate (both the intra- and inter-day accuracy were between 92.2% and 105.4%). The average extraction efficiencies for MPA, MPAG and AcMPAG were 85.3%, 100.1%, and 94.7% in plasma, and 88.0%, 67.3%, and 68.3% in bile, respectively. The method was successfully employed for pharmacokinetic studies in plasma and bile after oral administration of mycophenolate mofetil (prodrug of MPA) in rats.

P197 : Mycophenolic acid restores IFNg-induced collapse of immune privilege in human dermal papilla cells but fails to regulate immune privilege in C3H/HeJ mice

( Ki Min Sohn ),( Kwan Ho Jeong ),( Hong Jin Joo ),( Bo In Lee ),( Jung Eun Kim ),( Hoon Kang ) 대한피부과학회 2014 대한피부과학회 학술발표대회집 Vol.66 No.2

Background: Alopecia areata (AA) is an autoimmune disease that self-attacks anagen hair follicles. Gene analysis revealed that genes involved in immune privilege (IP) were increased both in interferon gamma (IFNg)-induced C3H/HeJ mice or human dermal papilla cells (hDPC). Mycophenolic acid (MPA) has been widely used as an immunosuppressive agent, through the inhibition of T and B lymphocytes. Objectives: In this study, we examined the effectiveness of MPA on IFNg-induced hDPCs and to investigate the hair growth stimulating effects of MPA in the IFNg-induced C3H/HeJ mice. Methods: Cultured DP cells were treated with various concentrations of MPA (0-500 nM) and analyzed by MTT assay. Hair growth and IP-related genes were examined by RT-PCR and Western blot analysis. Additionally, hair re-growth was monitored in IFNg-induced C3H/HeJ mice after treatment in 1μM or 1mM of MPA and hematoxylin and eosin staining was performed. Results: We found that MPA significantly showed protective effects on hDPCs against IFNg. However, MPA failed to induce hair re-growth in IFN-induced mice. Conclusion: IP restroating effect of MPA on hDPCs is valuable findings for the development of therapeutic options for AA in human.