Synthesis of Diverse Hydroxycinnamoyl Phenylethanoid Esters Using <i>Escherichia coli</i>

Song, Min Kyung,Cho, A Ra,Sim, GeunYoung,Ahn, Joong-Hoon American Chemical Society 2019 Journal of agricultural and food chemistry Vol.67 No.7

<P>Caffeic acid phenethyl ester (CAPE) is an ester of a hydroxycinnamic acid (phenylpropanoid) and a phenylethanoid (2-phenylethanol; 2-PE), which has long been used in traditional medicine. Here, we synthesized 54 hydroxycinnamic acid-phenylethanoid esters by feeding 64 combinations of hydroxycinnamic acids and phenylethanols to <I>Escherichia coli</I> harboring the rice genes <I>OsPMT</I> and <I>Os4CL</I>. The same approach was applied for ester synthesis with caffeic acid and eight different phenyl alcohols. Two hydroxycinnamoyl phenethyl esters, <I>p-</I>coumaroyl tyrosol and CAPE, were also synthesized from glucose using engineered <I>E. coli</I> by introducing genes for the synthesis of substrates. Consequently, we synthesized approximately 393.4 mg/L <I>p</I>-coumaroyl tyrosol and 23.8 mg/L CAPE with this approach. Overall, these findings demonstrate that the rice PMT and 4CL proteins can be used for the synthesis of diverse hydroxycinnamoyl phenylethanoid esters owing to their promiscuity and that further exploration of the biological activities of these compounds is warranted.</P> [FIG OMISSION]</BR>

Caffeic acid phenethyl ester의 cytochrome P450 저해 활성 평가

류창선(Chang Seon Ryu),박지은(Ji-Eun Park),김상겸(Sang Kyum Kim) 대한약학회 2018 약학회지 Vol.62 No.2

Caffeic acid phenethyl ester (CAPE) is one of the major active components of propolis showing inhibitory activ-ity against cytochrome P450 (CYP). The purpose of this study was to determine CYP inhibitory potential and metabolic sta-bility of CAPE using human liver microsomes (HLM) for characterization of its metabolic properties. Among CYP isoforms including CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4, CAPE exhibited strong inhibitory potential against CYP1A2 and 2C9, moderate against CYP2E1, 2B6, 2C8, 2C19 and 3A4, and weak against CYP 2D6 and 2A6, based on the IC 50 values. The inhibition of CYP1A2 but not CYP2C9 was increased by preincubation with CAPE and NADPH, suggesting that the CAPE-induced CYP1A2 inhibition may be metabolism-dependent. In metabolic stability study using HLM, CAPE was rapidly hydrolyzed to caffeic acid in a NADPH-independent manner. Caffeic acid exhibited week CYP inhibitory activity, relative to CAPE. These results raise the possibility that CAPE plays a role in propolis-mediated CYP inhibition.

Glycosylation Enhances the Physicochemical Properties of Caffeic Acid Phenethyl Ester

( Keum-ok Moon ),( Soyoon Park ),( Myungsoo Joo ),( Ki-tae Ha ),( Nam-in Baek ),( Cheon-seok Park ),( Jaeho Cha ) 한국미생물생명공학회(구 한국산업미생물학회) 2017 Journal of microbiology and biotechnology Vol.27 No.11

In this study, we synthesized a glycosylated derivative of caffeic acid phenethyl ester (CAPE) using the amylosucrase from Deinococcus geothermalis with sucrose as a substrate and examined its solubility, chemical stability, and anti-inflammatory activity. Nuclear magnetic resonance spectroscopy showed that the resulting glycosylated CAPE (G-CAPE) was the new compound caffeic acid phenethyl ester-4-O-α-D-glucopyranoside. G-CAPE was 770 times more soluble than CAPE and highly stable in Dulbecco’s modified Eagle’s medium and buffered solutions, as estimated by its half-life. The glycosylation of CAPE did not significantly affect its anti-inflammatory activity, which was assessed by examining lipopolysaccharide-induced nitric oxide production and using a nuclear factor erythroid 2-related factor 2 reporter assay. Furthermore, a cellular uptake experiment using high-performance liquid chromatography analysis of the cell-free extracts of RAW 264.7 cells demonstrated that G-CAPE was gradually converted to CAPE within the cells. These results demonstrate that the glycosylation of CAPE increases its bioavailability by helping to protect this vital molecule from chemical or enzymatic oxidation, indicating that G-CAPE is a promising candidate for prodrug therapy.

Neuroprotective effect of caffeic acid phenethyl ester in 3-nitropropionic acid-induced striatal neurotoxicity

Jia Bak,Hee Jung Kim,Seong Yun Kim,Yun-Sik Choi 대한생리학회-대한약리학회 2016 The Korean Journal of Physiology & Pharmacology Vol.20 No.3

Caffeic acid phenethyl ester (CAPE), derived from honeybee hives, is a bioactive compound with strong antioxidant activity. This study was designed to test the neuroprotective effect of CAPE in 3-nitropropionic acid (3NP)-induced striatal neurotoxicity, a chemical model of Huntington’s disease (HD). Initially, to test CAPE’s antioxidant activity, a 2,2’-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid (ABTS) antioxidant assay was employed, and CAPE showed a strong direct radicalscavenging eff ect. In addition, CAPE provided protection from 3NP-induced neuronal cell death in cultured striatal neurons. Based on these observations, the <i>in vivo</i> therapeutic potential of CAPE in 3NP-induced HD was tested. For this purpose, male C57BL/6 mice were repeatedly given 3NP to induce HD-like pathogenesis, and 30 mg/ kg of CAPE or vehicle (5% dimethyl sulfoxide and 95% peanut oil) was administered daily. CAPE did not cause changes in body weight, but it reduced mortality by 29%. In addition, compared to the vehicle-treated group, robustly reduced striatal damage was observed in the CAPE-treated animals, and the 3NP-induced behavioral defi cits on the rotarod test were signifi cantly rescued after the CAPE treatment. Furthermore, immunohistochemical data showed that immunoreactivity to glial fibrillary acidic protein (GFAP) and CD45, markers for astrocyte and microglia activation, respectively, were strikingly reduced. Combined, these data unequivocally indicate that CAPE has a strong antioxidant eff ect and can be used as a potential therapeutic agent against HD.

Caffeic acid phenethyl ester activation of Nrf2 pathway is enhanced under oxidative state: Structural analysis and potential as a pathologically targeted therapeutic agent in treatment of colonic inflammation

Kim, Hyunjeong,Kim, Wooseong,Yum, Soohwan,Hong, Sungchae,Oh, Jeong-Eun,Lee, Ji-Woo,Kwak, Mi-Kyoung,Park, Eun Ji,Na, Dong Hee,Jung, Yunjin Elsevier 2013 FREE RADICAL BIOLOGY AND MEDICINE Vol.65 No.-

<P><B>Abstract</B></P> <P>Caffeic acid phenethyl ester (CAPE) is a polyphenolic natural product that possesses numerous biological activities including anti-inflammatory effects. CAPE-mediated nuclear factor-erythroid 2 p45 (NF-E2)-related factor 2 (Nrf2) activation is likely responsible for some of its biological effects. CAPE was chemically modified to yield CAPE analogues that were subjected to experiments examining cellular Nrf2 activity. CAPE and the CAPE analogue with a catechol moiety, but not the other analogues, activated the Nrf2 pathway. In addition, only biotin-labeled CAPE analogues with the catechol moiety precipitated Kelch-like ECH associated protein 1 (Keap1) when incubated with cell lysates and streptavidin agarose beads. Sodium hypochlorite (NaOCl) oxidation of the catechol moiety in CAPE produced an oxidized, electrophilic form of CAPE (Oxi-CAPE) and greatly enhanced the ability of CAPE to activate Nrf2 and to bind to Keap1. Rectal administration of CAPE ameliorated 2,4,6-trinitrobenzene sulfonic acid-induced rat colitis and activated the Nrf2 pathway in the inflamed colon, and incubation of CAPE in the lumen of the inflamed distal colon generated Oxi-CAPE. However, these biological effects and chemical change of CAPE were not observed in the normal colon. Our data suggest that CAPE requires the catechol moiety for the oxidation-enhanced activation of the Nrf2 pathway and has potential as a pathologically targeted Nrf2-activating agent that is exclusively activated in pathological states with oxidative stress such as colonic inflammation.</P> <P><B>Highlights</B></P> <P> <UL> <LI> The catechol moiety in caffeic acid phenethyl ester (CAPE) is required for CAPE activation of Nrf2. </LI> <LI> Oxidized CAPE (most likely an electrophilic <I>o</I>-quinone form) is responsible for the biological activity of CAPE. </LI> <LI> CAPE ameliorates experimental rat colitis and generates the oxidized form of CAPE in the inflamed colonic tissues. </LI> <LI> CAPE activates the Nrf2 pathway in the inflamed colonic tissues but not in the normal colonic tissues. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>[DISPLAY OMISSION]</P>

Neuroprotective effect of caffeic acid phenethyl ester in 3-nitropropionic acid-induced striatal neurotoxicity

Bak, Jia,Kim, Hee Jung,Kim, Seong Yun,Choi, Yun-Sik The Korean Society of Pharmacology 2016 The Korean Journal of Physiology & Pharmacology Vol.20 No.3

Caffeic acid phenethyl ester (CAPE), derived from honeybee hives, is a bioactive compound with strong antioxidant activity. This study was designed to test the neuroprotective effect of CAPE in 3-nitropropionic acid (3NP)-induced striatal neurotoxicity, a chemical model of Huntington's disease (HD). Initially, to test CAPE's antioxidant activity, a 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid (ABTS) antioxidant assay was employed, and CAPE showed a strong direct radical-scavenging effect. In addition, CAPE provided protection from 3NP-induced neuronal cell death in cultured striatal neurons. Based on these observations, the in vivo therapeutic potential of CAPE in 3NP-induced HD was tested. For this purpose, male C57BL/6 mice were repeatedly given 3NP to induce HD-like pathogenesis, and 30 mg/kg of CAPE or vehicle (5% dimethyl sulfoxide and 95% peanut oil) was administered daily. CAPE did not cause changes in body weight, but it reduced mortality by 29%. In addition, compared to the vehicle-treated group, robustly reduced striatal damage was observed in the CAPE-treated animals, and the 3NP-induced behavioral deficits on the rotarod test were significantly rescued after the CAPE treatment. Furthermore, immunohistochemical data showed that immunoreactivity to glial fibrillary acidic protein (GFAP) and CD45, markers for astrocyte and microglia activation, respectively, were strikingly reduced. Combined, these data unequivocally indicate that CAPE has a strong antioxidant effect and can be used as a potential therapeutic agent against HD.

ATF3를 통한 caffeic acid phenethyl ester에 의한 NAG-1 유전자의 발현 증가

박민희(Min-Hee Park),정정욱(Chungwook Chung),이승호(Seong Ho Lee),백승준(Seung Joon Baek),김종식(Jong Sik Kim) 한국생명과학회 2018 생명과학회지 Vol.28 No.1

NAG-1 단백질은 TGF-β superfamily 유전자로서 암세포의 apoptosis를 유도하고 항암 활성과 관련이 있는 것으로 알려져 있다. 본 연구에서는 프로폴리스 유래의 파이토케미칼 CAPE (caffeic acid phenethyl ester)에 의한 항암유전자 NAG-1의 발현과 발현조절에 대해 연구하였다. 인간 대장암 세포주 HCT116에서 CAPE의 처리에 의해 농도의존적, 시간의존적으로 NAG-1의 발현이 증가됨을 확인하였다. 게다가, 다른 대장암 세포주인 LOVO 세포주에서도 농도의존적으로 NAG-1의 발현이 증가됨을 확인하였다. p53-null HCT116세포주를 이용한 실험에서 CAPE에 의한 NAG-1의 발현은 전사조절인자인 p53에 의존하지 않음을 증명하였다. 또한, 3가지 종류의 NAG-1 프로모터 construct를 이용한 실험에서, cis-element 후보가 -474와 -1,086사이에 있음을 증명하였다. CAPE에 의해 전사조절인자인 ATF3와 CREB의 발현이 변화되는 지를 확인한 결과, CREB은 전혀 발현이 증가되지 않는 반면 ATF3는 CAPE 처리에 의해 농도의존적으로 발현이 증가함을 확인하였다. 그리고, pCG-ATF3와 pCREB의 cotransfection 실험에서 CREB은 NAG-1의 발현에 영향을 못 미치는 반면, ATF3의 과대발현에 의해 NAG-1의 발현 이 증가됨을 확인하였다. 결론적으로, CAPE에 의한 NAG-1의 발현은 주로 전사조절인자인 ATF3를 경유하여 일어남을 시사한다. Non-steroidal anti-inflammatory drug-activated gene-1 (NAG-1) is a transforming growth factor beta (TGF-β) superfamily gene associated with pro-apoptotic and anti-tumorigenic activities. In the present study, we investigated if caffeic acid phenethyl ester (CAPE) derived from propolis could induce the expression of anti-tumorigenic gene NAG-1. Our results indicate that CAPE significantly induced NAG-1 expression in a time- and concentration-dependent manner in HCT116 cells. We also found that CAPE induced NAG-1 expression in a concentration-dependent manner in another human colorectal cancer cell line, LOVO. In addition, CAPE triggered apoptosis, which was detected with Western blot analysis using poly-(ADP-ribose) polymerase antibody. NAG-1 induction by CAPE was not dependent on transcription factor p53, which was confirmed with Western blot analysis using p53 null HCT116 cells. The luciferase assay results indicated that the new cis-elements candidates were located between -474 and -1,086 of the NAG-1 gene promoter. CAPE dramatically induced activating transcription factor 3 (ATF3) expression, but not cAMP response element-binding protein (CREB), which shares the same binding sites with ATF3. The co-transfection experiment with pCG-ATF3 and pCREB showed that only ATF3 was associated with NAG-1 up-regulation by CAPE, whereas CREB had no effect. In conclusion, the results suggest that CAPE could induce the expression of anti-tumorigenic gene NAG-1 mainly through ATF3.

Caffeic Acid Phenethyl Ester-Mediated Cytotoxicity in MC-3 Human Mucoepidermoid Carcinoma Cell Line

유현주,조남표,조성대 대한구강악안면병리학회 2014 대한구강악안면병리학회지 Vol.38 No.6

Caffeic acid phenethyl ester (CAPE), a component of propolis, was reported to possess anti-inflammatory, anti-bacterial, anti-viral,and anti-tumor activities. Our aim was to investigate the effect of CAPE on apoptosis in cultured human mucoepidermoid carcinoma(MEC) cell line, MC-3. Apoptotic effects of CAPE were measured by cell viability assays, Western blotting, 4’-6-diamidino-2-phenylindole(DAPI) staining and Live/Dead assay. The result of cell viability assay showed that CAPE displayed a strong growth-inhibitory effectin a concentration-dependent manner against MC-3 cells. Consumption of CAPE resulted in pronounced increase in the cleavage ofcaspase-3 and PARP, induced nuclear condensation and fragmentation and clearly increased the number of dead cells in MC-3 cells. CAPE also caused the increase in truncated Bid (t-Bid) and the cleavage of caspase-8 and this phenomenon was regulated by deathreceptor 5 (DR5). In addition, Phosphorylation of AKT and ERK were downregulated by CAPE. Taken together, these results suggestthat CAPE is a potent apoptosis-inducing agent in MC-3 cells.

Caffeic acid phenethyl ester-mediated Nrf2 activation and IκB kinase inhibition are involved in NFκB inhibitory effect: Structural analysis for NFκB inhibition

Lee, Youna,Shin, Dong-ha,Kim, Ji-Hye,Hong, Sungchae,Choi, Daekyu,Kim, Yung-Jin,Kwak, Mi-Kyoung,Jung, Yunjin Elsevier 2010 european journal of pharmacology Vol.643 No.1

<P><B>Abstract</B></P><P>Caffeic acid phenethyl ester (CAPE) is an active component of propolis from honeybee. We investigated potential molecular mechanisms underlying CAPE-mediated nuclear factor kappa beta (NFκB) inhibition and analyzed structure of CAPE for its biological effect. CAPE attenuated expression of NFκB dependent luciferase stimulated with TNF-α or LPS and suppressed LPS-mediated induction of iNOS, a target gene product of NFκB. In HCT116 cells, CAPE interfered with TNF-α dependent IκBα degradation and subsequent nuclear accumulation of p65, which occurred by direct inhibition of inhibitory protein kappaB kinase (IKK). CAPE increased the expression of Nrf2-dependent luciferase and <I>heme oxygenase-1</I>, a target gene of Nrf2, and elevated the nuclear level of Nrf2 protein, indicating that CAPE activated the Nrf2 pathway. In HCT116 cells with stable expression of Nrf2 shRNA, CAPE elicited a reduced inhibitory effect on TNF-α-activated NFкB compared to scramble RNA expressing control cells. On the other hand, the NFκB inhibitory effect of CAPE was diminished by removal or modification of the Michael reaction acceptor, catechol or phenethyl moiety in CAPE. These data suggest that CAPE inhibits TNF-α-dependent NFκB activation via direct inhibition of IKK as well as activation of Nrf2 pathway, in which the functional groups in CAPE may be involved.</P>

Caffeic Acid Phenethyl Ester-Mediated Cytotoxicity in MC-3 Human Mucoepidermoid Carcinoma Cell Line

유현주,조남표,조성대 대한구강악안면병리학회 2014 대한구강악안면병리학회지 Vol.38 No.6

Caffeic acid phenethyl ester (CAPE), a component of propolis, was reported to possess anti-inflammatory, anti-bacterial, anti-viral, and anti-tumor activities. Our aim was to investigate the effect of CAPE on apoptosis in cultured human mucoepidermoid carcinoma (MEC) cell line, MC-3. Apoptotic effects of CAPE were measured by cell viability assays, Western blotting, 4’-6-diamidino-2-phenylindole (DAPI) staining and Live/Dead assay. The result of cell viability assay showed that CAPE displayed a strong growth-inhibitory effect in a concentration-dependent manner against MC-3 cells. Consumption of CAPE resulted in pronounced increase in the cleavage of caspase-3 and PARP, induced nuclear condensation and fragmentation and clearly increased the number of dead cells in MC-3 cells. CAPE also caused the increase in truncated Bid (t-Bid) and the cleavage of caspase-8 and this phenomenon was regulated by death receptor 5 (DR5). In addition, Phosphorylation of AKT and ERK were downregulated by CAPE. Taken together, these results suggest that CAPE is a potent apoptosis-inducing agent in MC-3 cells.