EGFR; HER2; HER3

In a dose-dependent manner, (-)-Epigallocatechin Gallate (EGCG, 10-60 μM) suppresses the development of WRO and FB-2 cells [1]. (-)-Epigallocatechin Gallate (10–60 μM, 0–24 h) raises the expression of p21 and p53 and decreases the phosphorylation of cyclin D1, AKT, and ERK1/2 [1]. The effects of (10–60 μM, 12 hours)-Epigallocatechin Gallate on cell motility and migration are reported [1]. According to a biochemical experiment, (-)-Epigallocatechin Gallate (0 – 20 μM, or roughly 0 – 20 minutes) suppresses GLUD1/2 and IDH1 activities in a concentration- and time-dependent manner [2]. (-)- Epigallocatechin Gallate (0-35 μg/mL, 24-72 hours) promotes cell apoptosis, reduces G0/G1 phase cell proliferation, and suppresses the proliferation of colorectal cancer cells (LoVo, SW480, HT-29, and HCT-8 cells)[3]. In osteoblasts, LPS-induced COX-2 and mPGES-1 mRNA expression as well as prostaglandin E2 synthesis are inhibited by (-)-Epigallocatechin Gallate (30 μM, 3–24 h) [4].

(-)- Epigallocatechin Gallate inhibits the growth of tumors when given intraperitoneally (5–20 mg/kg) once day for 14 days in an orthotopic transplantation paradigm [3]. (-)- The LPS-induced loss of bone mineral density (BMD) is inhibited and reduced when epigallocatechin gallate is injected into the lower gums of mice in an experimental periodontitis model at a single dose of 0.5 mg/mouse [3].

GLUD1/2 and IDH enzymatic assays[2]
IDH1 was expressed as glutathione S-transferase (GST)-fusion protein in pDEST15 and purified on glutathione beads as described. Purified bovine GLUD1/2 was purchased from Serva. Enzyme reactions were initiated by adding 4 μg IDH1 enzyme to a mixture of 100 μM NADP+, 2 mM MgCl2, 0.5 mM isocitrate, and 100 mM Tris-HCl (pH 7.4). GLUD1/2 activity was measured in reactions containing 0.1 U bovine GLUD1/2 enzyme, 500 μM NAD+, 10 mM glutamate, and 2 mM ADP in phosphate buffer (pH 8.0). Stoichiometric production of NADPH and NADH was measured by real-time monitoring of NADPH or NADH absorbance at 340 nm with 20 s intervals on an Omega Fluostar.

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DNA-double strand break (DSB) detection[2]
Cells (cultured with or without AGI-5198) were plated at a density of 300,000 cells/well in 6-well plates and left to adhere overnight. After 24 h incubation with Epigallocatechin gallate (EGCG) (0, 50, or 100 μM), cells were irradiated with 0, 2, or 4 Gy. After 30 min, cytosolic extracts were prepared in 1× RIPA buffer containing 1 mM phenylmethylsulfonyl fluoride (PMSF). Cell extracts were sonicated to release nuclear proteins. Protein samples (25 μg) were electrophoresed on 10% SDS-PAGE gels and electroblotted onto nitrocellulose. Blots were stained with anti-γH2AX antibody and anti-γ-tubulin (C20), followed by appropriate secondary antibodies labeled with IRDye680 or IRDye800. Signals were visualized and quantified using the Odyssey system

Cell Proliferation Assay[1]
Cell Types: FB-2 and WRO cells (serum-starved for 48h)
Tested Concentrations: 10, 40, 60 μM.
Incubation Duration: 4 days
Experimental Results: Inhibited basal cell proliferation (40% in FB-2 and 35% in WRO) at 10 μM, inhibited cell number (by 68% to 73%) at 40 and 60 μM).

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Western Blot Analysis[1]
Cell Types: FB-2 cells
Tested Concentrations: 10, 40, 60 μM.
Incubation Duration: 24 h
Experimental Results: decreased cyclin D1 level, phosphorylation of AKT and ERK1/2. Induced the expression of p21 and p53, and E-cadherin, N-cadherin, Vimentin and α5-integrin.

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Cell Migration Assay [1]
Cell Types: FB-2 and WRO cells (serum-starved for 48h)
Tested Concentrations: 10, 40, 60 μM.
Incubation Duration: 12 h
Experimental Results: decreased migration activity in FB-2 and WRO cells.

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RT-PCR[4]
Cell Types: Mouse primary osteoblasts (1 ng/ml LPS-treated)
Tested Concentrations: 30 μM
Incubation Duration: 3, 6, 12, 24 h
Experimental Results: Suppressed the LPS-induced expression of COX-2 and mPGES-1 mRNAs, prostaglandin E2 production.

Animal/Disease Models: Orthotopic transplant BALB/c nude mice model[3]
Doses: 5, 10, and 20 mg/kg, one time/day for 14 days.
Route of Administration: Intragastrical administration.
Experimental Results: Inhibited tumors growth with no liver or lung metastases.

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Animal/Disease Models: Model of experimental periodontitis, LPS (25 μg/mouse)[4]
Doses: 0.5 mg/mouse, a single dose.
Route of Administration: Injected into the mouse lower gingiva
Experimental Results: Inhibited the LPS-induced loss of bone mineral density (BMD ) in mice.

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Subcutaneous orthotopic colorectal cancer transplant model and medical treatment[3]
The HT-29 colorectal cancer cell line with green fluorescence was established.7 BALB/c nude mice, 20 male and 20 female, that ranged from 4- to 6-weeks-old were fed in a special pathogenic free animal facility. The feed was sterilized using cobalt 60. As described above, the subcutaneous orthotopic colorectal cancer transplant model was established successfully.
At 2 weeks postsurgery, 39 out of the 40 nude mice presented with tumors. Based on the volume of the tumors, the 39 mice with tumors were divided into four groups: a control group (n = 9); a group that received 5 mg/kg of Epigallocatechin gallate (EGCG) (n = 10); a group that received 10 mg/kg of Epigallocatechin gallate (EGCG) (n = 10); and a group that received 20 mg/kg of Epigallocatechin gallate (EGCG) (n = 10). In the therapeutic groups, Epigallocatechin gallate (EGCG) was administrated intragastrically, and in the control group, 100 uL of physiological saline was administrated intragastrically, once daily for 14 days.
After the treatment of the mice with Epigallocatechin gallate (EGCG) for 4 weeks, the growth and metastasis of the primary tumors were continuously monitored using a fluorescent imaging system. After 4 weeks, the primary tumors were weighed and immediately put into liquid nitrogen (−196°C) and 2 to 3 hours later, these specimens were stored at −80°C. In addition, the other parts of the primary tumor and metastases were fixed in 4% formaldehyde.[3]

Metabolism / Metabolites
(-)-Epigallocatechin gallate has known human metabolites that include (-)-Epigallocatechin gallate, 3p-hydroxy-glucuronide and (-)-Epigallocatechin gallate, 4p-hydroxy-glucuronide.

[1]. Epigallocatechin gallate inhibits growth and Epithelial-to-Mesenchymal Transition in human thyroid carcinoma cell lines. J Cell Physiol. 2013 Oct;228(10):2054-62.

[2]. Isocitrate dehydrogenase 1-mutated cancers are sensitive to the green tea polyphenol epigallocatechin-3-gallate. Cancer Metab. 2019 May 20;7:4.

[3]. Epigallocatechin gallate inhibits the proliferation of colorectal cancer cells by regulating Notch signaling. Onco Targets Ther. 2013;6:145-53.

[4]. Tsukasa Tominari; Epigallocatechin gallate (EGCG) suppresses lipopolysaccharide-induced inflammatory bone resorption, and protects against alveolar bone loss in mice. FEBS Open Bio. 2015 Jun 12;5:522-7.

(-)-epigallocatechin 3-gallate is a gallate ester obtained by the formal condensation of gallic acid with the (3R)-hydroxy group of (-)-epigallocatechin. It has a role as an antineoplastic agent, an antioxidant, a Hsp90 inhibitor, a neuroprotective agent, a plant metabolite, a geroprotector and an apoptosis inducer. It is a gallate ester, a polyphenol and a member of flavans. It is functionally related to a (-)-epigallocatechin.
Epigallocatechin gallate has been investigated for the treatment of Hypertension and Diabetic Nephropathy.
(-)-Epigallocatechin gallate has been reported in Camellia sinensis, Eschweilera coriacea, and other organisms with data available.
Epigallocatechin Gallate is a phenolic antioxidant found in a number of plants such as green and black tea. It inhibits cellular oxidation and prevents free radical damage to cells. It is under study as a potential cancer chemopreventive agent.

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Well-differentiated papillary and follicular thyroid carcinoma are the most frequent types of thyroid cancer and the prognosis is generally favorable however, a number of patients develops recurrences. Epigallocatechin-3-gallate (EGCG), a major catechin in green tea, was shown to possess remarkable therapeutic potential against various types of human cancers, although data on thyroid cancer cells are still lacking. The aim of this study was to investigate the effect of EGCG on the proliferation and motility of human thyroid papillary (FB-2) and follicular (WRO) carcinoma cell lines. Our results demonstrate that EGCG (10, 40, 60 μM) treatment inhibited the growth of FB-2 and WRO cells in a dose-dependent manner. These changes were associated with reduced cyclin D1, increased p21 and p53 expression. Furthermore, EGCG suppressed phosphorylation of AKT and ERK1/2. In addition EGCG treatment results in reduction of cell motility and migration. Changes in motility and migration in FB-2 were associated with modulation in the expression of several proteins involved in cell adhesion and reorganization of actin cytoskeleton. After 24 h EGCG caused an increase of the E-cadherin expression and a concomitant decrease of SNAIL, ZEB and the basic helix-loop-helix transcription factor TWIST. Besides expression of Vimentin, N-cadherin and α5-integrin was down-regulated. These data well correlate with a reduction of MMP9 activity as evidenced by gelatin zymography. Our findings support the inhibitory role of EGCG on thyroid cancer cell proliferation and motility with concomitant loss of epithelial-to-mesenchymal cell transition markers.[1]

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Background: Mutations in isocitrate dehydrogenase 1 (IDH1) occur in various types of cancer and induce metabolic alterations resulting from the neomorphic activity that causes production of D-2-hydroxyglutarate (D-2-HG) at the expense of α-ketoglutarate (α-KG) and NADPH. To overcome metabolic stress induced by these alterations, IDH-mutated (IDH mut ) cancers utilize rescue mechanisms comprising pathways in which glutaminase and glutamate dehydrogenase (GLUD) are involved. We hypothesized that inhibition of glutamate processing with the pleiotropic GLUD-inhibitor epigallocatechin-3-gallate (EGCG) would not only hamper D-2-HG production, but also decrease NAD(P)H and α-KG synthesis in IDH mut cancers, resulting in increased metabolic stress and increased sensitivity to radiotherapy.

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Methods: We performed 13C-tracing studies to show that HCT116 colorectal cancer cells with an IDH1 R132H knock-in allele depend more on glutaminolysis than on glycolysis for the production of D-2-HG. We treated HCT116 cells, HCT116-IDH1 R132H cells, and HT1080 cells (carrying an IDH1 R132C mutation) with EGCG and evaluated D-2-HG production, cell proliferation rates, and sensitivity to radiotherapy.
Results: Significant amounts of 13C from glutamate accumulate in D-2-HG in HCT116-IDH1 wt/R132H but not in HCT116-IDH1 wt/wt . Preventing glutamate processing in HCT116-IDH1 wt/R132H cells with EGCG resulted in reduction of D-2-HG production. In addition, EGCG treatment decreased proliferation rates of IDH1 mut cells and at high doses sensitized cancer cells to ionizing radiation. Effects of EGCG in IDH-mutated cell lines were diminished by treatment with the IDH1mut inhibitor AGI-5198.
Conclusions: This work shows that glutamate can be directly processed into D-2-HG and that reduction of glutamatolysis may be an effective and promising new treatment option for IDH mut cancers.[2]

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Aims: To explore the inhibitory effects of epigallocatechin gallate (EGCG) on the proliferation of colorectal cancer cells and on the gene expression of Notch signaling.
Methods: The colorectal cancer cells and orthotopic colorectal cancer transplant model were treated with EGCG, and MTT assay was used to test the inhibitory role of EGCG in the proliferation of colorectal cancer cells.
Results: MTT assay indicated that EGCG inhibited the proliferation of these four cell lines when the time and concentration increased, and EGCG enhanced the apoptotic rate of these four cell lines. The dosage was positively correlated to the apoptotic rate, and EGCG inhibited the proliferation of colorectal cancer cells by influencing cell cycle. In-vivo study suggested that on the seventh day, the volume of tumors reduced after administrating with 5, 10 and 20 mg/kg of EGCG. At the twenty-eighth day, the volume of tumors was significantly different in three EGCG treatment groups as compared to the control group (P < 0.05), and TUNEL assay indicated that the apoptosis of cancer cells in EGCG treated groups was markedly higher than that in the control group (P < 0.05). In these cell lines, the expressions of HES1 and Notch2 in EGCG treated groups were remarkably lower than that in the control group (P < 0.05). The expression of JAG1 decreased in SW480 cells (P =0.019), HT-29 cells and HCT-8 cells, but increased in LoVo cells at mRNA level. The expression of Notch1 was upregulated in these four cell lines, but its expression was significantly upregulated only in LoVo and SW480 cells (P < 0.05).
Conclusion: In-vitro and in-vivo studies showed that EGCG inhibited the proliferation, induced the apoptosis and affected the cell cycle of colorectal cancer cells. After treating with EGCG, the expressions of HES1 and Notch2 was obviously inhibited, this indicated that EGCG inhibited colorectal cancer by inhibiting HES1 and Notch2.[3]

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Epigallocatechin gallate (EGCG), a major polyphenol in green tea, possesses antioxidant properties and regulates various cell functions. Here, we examined the function of EGCG in inflammatory bone resorption. In calvarial organ cultures, lipopolysaccharide (LPS)-induced bone resorption was clearly suppressed by EGCG. In osteoblasts, EGCG suppressed the LPS-induced expression of COX-2 and mPGES-1 mRNAs, as well as prostaglandin E2 production, and also suppressed RANKL expression, which is essential for osteoclast differentiation. LPS-induced bone resorption of mandibular alveolar bones was attenuated by EGCG in vitro, and the loss of mouse alveolar bone mass was inhibited by the catechin in vivo.[4]

C22H18O11

458.37

458.084

C, 57.65; H, 3.96; O, 38.40

989-51-5

(-)-Epigallocatechin;970-74-1;(-)-Gallocatechin gallate;4233-96-9;(-)-Epigallocatechin Gallate (Standard);989-51-5;(+/-)-Epigallocatechin Gallate-13C3

65064

Off-white to pink solid powder

1.9±0.1 g/cm3

909.1±65.0 °C at 760 mmHg

222-224°C

320.0±27.8 °C

0.0±0.3 mmHg at 25°C

1.857

polyphenol in green tea

2.08

8

11

4

33

667

2

O1C2=C([H])C(=C([H])C(=C2C([H])([H])[C@]([H])([C@@]1([H])C1C([H])=C(C(=C(C=1[H])O[H])O[H])O[H])OC(C1C([H])=C(C(=C(C=1[H])O[H])O[H])O[H])=O)O[H])O[H]

WMBWREPUVVBILR-WIYYLYMNSA-N

InChI=1S/C22H18O11/c23-10-5-12(24)11-7-18(33-22(31)9-3-15(27)20(30)16(28)4-9)21(32-17(11)6-10)8-1-13(25)19(29)14(26)2-8/h1-6,18,21,23-30H,7H2/t18-,21-/m1/s1

[(2R,3R)-5,7-dihydroxy-2-(3,4,5-trihydroxyphenyl)chroman-3-yl] 3,4,5-trihydroxybenzoate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.08 mg/mL (4.54 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (4.54 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (4.54 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 9.09 mg/mL (19.83 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication (<60°C).

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 (Please use freshly prepared in vivo formulations for optimal results.)