Kim, Yeon-Sil;Roh, Kwang-Won;Chae, Soo-Min;Mun, Seong-Kwon;Yoon, Sei-Chul;Jang, Hong-Seok;Chung, Su-Mi
Radiation Oncology Journal
/
v.25
no.4
/
pp.233-241
/
2007
Purpose: We examined the effect of the dual EGFR/HER2 tyrosine kinase inhibitor, GW572016, on EGFR/HER2 receptor phosphorylation, inhibition of downstream signaling and radiosensitization in either an EGFR or HER2 overexpressing human breast cancer xenograft. Materials and Methods: We established SCID mice xenografts from 4 human breast cancer cell line that overexpressed EGFR or HER 2 (SUM 102, SUM 149, SUM 185, SUM 225). Two series of xenografts were established. One series was established for determining inhibition of the EGFR/HER2 receptor and downstream signaling activities by GW572016. The other series was established for determining the radiosensitization effect of GW572016. Inhibition of the receptor and downstream signaling proteins were measured by the use of immunoprecipitation and Western blotting. For determining the in vivo radiosensitization effect of GW572016, we compared tumor growth delay curves in the following four treatment arms: a) control; b) GW572016 alone; c) radiotherapy (RT) alone; d) GW572016 and RT. Results: GW572016 inhibited EGFR, HER2 receptor phosphorylation in SUM 149 and SUM 185 xenografts. In addition, the p44/42 MAPK (ERK 1/2) downstream signaling pathway was inactivated by GW572016 in the SUM 185 xenograft. In the SUM 225 xenograft, we could not observe inhibition of HER2 receptor phosphorylation by GW572016; both p44/42 MAPK (Erk1/2) and Akt downstream signal protein phosphorylation were inhibited by GW572016. GW572016 inhibited growth of the tumor xenograft of SUM 149 and SUM 185. The combination of GW572016 and RT enhanced growth inhibition greater than that with GW572016 alone or with RT alone in the SUM 149 xenograft. GW572016 appears to act as an in vivo radiosensitizer. Conclusion: GW572016 inhibited EGFR/HER2 receptor phosphorylation and downstream signaling pathway proteins. GW572016 modestly inhibited the growth of tumor in the SUM 185 xenograft and showed radiosensitization in the SUM 149 xenograft. Our results suggest that a better predictor of radiation response would be inhibition of a crucial signaling pathway than inhibition of a receptor.
Kim, Ok-Soo;Choi, Jung-Hye;Soung, Young-Hwa;Lee, Seon-Hee;Lee, Jae-Hwa;Ha, Jong-Myung;Ha, Bae-Jin;Heo, Moon-Soo;Lee, Sang-Hyeon
Archives of Pharmacal Research
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v.27
no.9
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pp.906-911
/
2004
In order to evaluate estrogenic compounds in natural products, an in vitro detection system was established. For this system, the human breast cancer cell line MCF7 was stably trans-fected using an estrogen responsive chloramphenicol acetyltransferase (CAT) reporter plas-mid yielding MCF7/pDsCAT-ERE119-Ad2MLP cells. To test the estrogenic responsiveness of this in vitro assay system, MCF7/pDsCAT-ERE119-Ad2MLP cells were treated with various concentrations of 17f3-estradiol. Treatments of 10$^{-8}$ to 10$^{-12}$ M 17$\beta$-estradiol revealed significant concentration dependent estrogenic activities compared with ethanol. We used in vitro assay system to detect estrogenic effects in Puerariae radix and Ginseng radix Rubra extracts. Treat-ment of 500 and 50 $\mu\textrm{g}$/ml of Puerariae radix extracts increased the transcriptional activity approximately 4- and 1.5-fold, respectively, compared with the ethanol treatment. Treatment of 500, 50, and 5 $\mu\textrm{g}$/ml of Ginseng radix Rubra extracts increased the transcriptional activity approximately 3.2-,2.7, and 1.4-fold, respectively, compared with the ethanol treatment. These observations suggest that Puerariae radix and Ginseng radix Rubra extracts have effective estrogenic actions and that they could be developed as estrogenic supplements.
In order to verify the occurrence of an estrogenic compound in natural products, the estrogenic activity was measured using an in vitro detection system. For this system, human breast cancer cell line MCF7 was transfected using an estrogen responsive CAT reporter plasmid. Estrogenic activities of photosynthetic algae spirulina and sea lettuce were evaluated using this system. Estrogenic activities of a $500\mug/ml\; and\; 50 \mug/ml$ ethanol extracts of spirulina were as much as that of $10^{-8}$M standard solution (17$\beta$-estradiol) and activity of $5\mug/ml$ ethanol extract of spirulina was as much as that of $10^{-10}$ M standard solution. However, no significant estrogenic activity was observed using sea lettuce extract. Estrogenic activities of marine animals, such as star fish and shrimp, were also evaluated using this system, however, no significant estrogenic activity was observed in these extracts. In this result, it is confirmed that spirulina extract possesses estrogenic compound.
Journal of the Korean Society of Food Science and Nutrition
/
v.44
no.4
/
pp.516-523
/
2015
This study investigated the antimutagenic and anticancer effects of Platycodon grandiflorum extract (PGE) and its fractions against carcinogenic N-nitrosodimethylamine (NDMA) and genotoxicity. The Ames Salmonella mutagenicity test employing histidine mutants of Salmonella Typhimurium TA98 and TA100 was used to examine the mutagenicity of PGE and its fractions. Bacterial reversion assay with S. Typhimurium TA98 and TA100 did not show a significantly increased number of revertant colonies. The same test was used to examine the ability of PGE and its fractions to prevent acquisition of N-methyl-N'-nitro-N-nitrosoguanidine- and 4-introquino-line-1-oxide-induced mutations. PGE and its fractions inhibited mutagenesis in a dose-dependent manner. Among the fractions, ethyl acetate fraction from PGE (PGEA) exhibited a higher antimutagenic effect than other fractions. PGE and its fractions suppressed the growth of cancer cell lines, including human cervical adenocarcinoma, human hepatocellular carcinoma, human breast adenocarcinoma, human lung carcinoma, and transformed primary human embryonic kidney cells. In addition, we evaluated the antitumor activity of PGEA and its fractions in sacorma-180 solid tumor-bearing mice. In vivo anticancer activity results showed that PGE and its fractions could more effectively suppress tumor growth than the control. PGEA showed higher in vitro and in vivo anticancer effects than PGE and other fractions, and PGEA inhibited NDMA formation. Thus, we showed that PGEA has antimutagenic and anticancer activities, making it a candidate anticancer material under these experimental conditions.
The naturally occurring chemical indole-3-carbinol (13C), found in vegetables of the Brassica genus, is a promising anticancer agent that was shown previ- ously to induce a Gl cell cycle arrest of human breast cancer cell lines, independent of estrogen receptor signaling. The anticancer activity of 13C and the possible mechanisms of its action were explored in a human hepatocellular carcinoma cell line, HepG2. Treatment of HepG2 cells with 13C suppressed the growth of the cells. The growth sup- pression caused by 13C ($IC_{50}$/: 444$\mu$M) was found to be partially due to its ability to stop the cell cycle in HepG2 cells. Western blot analysis for the Gl phase artiest demonstrated that the expression-levels of cyclin-dependent kinase (Cdk4, Cdk6) and cyclic D were reduced strongly after treatment of Hep72 cells with 13C (4007M) for 24- 72 hrs. Furthermore, I3C selectively abolished the expression of Cdk6 in a dose- and time-dependent manner, and accordingly, inhibited the phosphorylation of retinoblastoma. Interestingly, after the HepG2 cells reached their max- imal growth arrest, the level of the p21, a well-known Cdk inhibitor, increased significantly. Therefore, it could be considered that the Gl arrest of HepG2 cells treated with 13C was due to the indirect inhibition of Cdk4/6 activities by p21 Western blot analysis for G2/M phase arrest of demonstrated the levels of Cdc2 and cyclin Bl werer reduced dramatically after the treatment of HepG2 cells with 13C ($40\mu$M) for 24-72 hrs. flow cytometry of propidium iodide-stained HepG2 cells revealed that 13C induces a Gl (53%,72hr incubation) and G2 (25%,24hr incubation) cell cycle arrest. Thus, our observations have uncovered a previously undefined antiproliferative pathway for r3C that implicates Cdk4/6 and Cdc2 as a target for cell cycle control in human HepG2 cells. However, the 13C-medi- ated cell cycle arrest and repression of Cdk4/6 production did not affect the apoptotic induction of HepG2 cell.
Ham, Young-An;Choi, Hyun-Jin;Kim, Soo-Hyun;Chung, Mi-Ja;Ham, Seung-Shi
Journal of the Korean Society of Food Science and Nutrition
/
v.38
no.1
/
pp.25-31
/
2009
This study was carried out to investigate the mutagenic, antimutagenic, cytotoxicity and antitumor effects of Adenophora triphylla (AT). AT was extracted with 70% ethanol and then further fractionated to hexane, chloroform, ethyl acetate, butanol and water. Antimutagenic, cytotoxicity and antitumor effects of AT extracts were measured by using Ames test, SRB method, and the tumor growth inhibition test. AT extracts did not show any mutagenicity in the Ames test; however, 70% ethanol extracts and its fractions had strong antimutagenic effects against mutation induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and 4-nitroquinoline-1-oxide (4NQO). The ethyl acetate fraction of AT (200 ${\mu}g$/plate) showed approximately 66.5% inhibitory effect on the mutagenesis induced by 4NQO against TA98 strain, whereas 83.3% and 75.1% inhibitions were observed on the mutagenesis induced by MNNG and 4NQO against TA100 strain. In anticancer effects, the cytotoxicity of AT extract and its fractions against cancer cell lines including human cervical adenocarcinoma (HeLa), human hepatocellular carcinoma (Hep3B), human breast adenocarcinoma (MCF-7), human gastric carcinoma (AGS), human lung carcinoma (A549) and transformed primary human embryo kidney (293) were investigated. The treatment of 1 mg/mL AT ethyl acetate faction had the highest cytotoxicity of 79.9%, 74.9%, 66.0%, 71.0% and 74.3% against HeLa, Hep3B, MCF-7, AGS and A549 cells, respectively. In contrast, the extract and its fractions showed only $3{\sim}36%$ cytotoxicity for a normal human kidney cell line (293). In vivo anti-cancer effect of Adenophora triphylla extract was tested using Balb/c mice transplanted sarcoma-180 cells. Adenophora triphylla ethyl acetate fraction showed the highest inhibition rate of 37.2% at the 50 mg/kg concentration.
Lee, Su Yeon;Ju, Min Kyung;Jeon, Hyun Min;Kim, Cho Hee;Park, Hye Gyeong;Kang, Ho Sung
Journal of Life Science
/
v.27
no.11
/
pp.1245-1255
/
2017
Most cancer cells produce ATP predominantly through glycolysis instead of through mitochondrial oxidative phosphorylation, even in the presence of oxygen. The phenomenon is termed the Warburg effect, or the glycolytic switch, and it is thought to increase the availability of biosynthetic precursors for cell proliferation. EMTs have critical roles in the initiation of the invasion and metastasis of cancer cells. The glycolytic switch and EMT are important for tumor development and progression; however, their correlation with tumor progression is largely unknown. The Snail transcription factor is a major factor involved in EMT. The Snail expression is regulated by distal-less homeobox 2 (Dlx-2), a homeodomain transcription factor that is involved in embryonic and tumor development. The Dlx-2/Snail cascade is involved in Wnt-induced EMTs and the glycolytic switch. This study showed that in response to Wnt signaling, the Dlx-2/Snail cascade induces the expression of PFKFB2, which is a glycolytic enzyme that synthesizes and degrades fructose 2, 6-bisphosphate (F2,6BP). It also showed that PFKFB2 shRNA prevents Wnt-induced EMTs in the breast-tumor cell line MCF-7. The prevention indicated that glycolysis is linked to Wnt-induced EMT. Additionally, this study showed PFKFB2 shRNA suppresses in vivo tumor metastasis and growth. Finally, it showed the PFKFB2 expression is higher in breast, colon and ovarian cancer tissues than in matched normal tissues regardless of the cancers' stages. The results demonstrated that PFKFB2 is an important regulator of EMTs and metastases induced by the Wnt, Dlx-2 and Snail factors.
Kim, Soo-Hyun;Choi, Hyun-Jin;Chung, Mi-Ja;Cui, Cheng-Bi;Ham, Seung-Shi
Journal of the Korean Society of Food Science and Nutrition
/
v.38
no.10
/
pp.1295-1301
/
2009
This study was carried out to investigate the mutagenic, antimutagenic, cytotoxicity and antitumor effect of Codonopsis lanceolata (CL). CL was extracted with 70% ethanol and then further fractionated to hexane, chloroform, ethyl acetate, butanol and water. Antimutagenic, cytotoxicity and antitumor effects of CL extracts were measured by using Ames test, SRB method, and the tumor growth inhibition test. CL extracts did not show any mutagenicity in the Ames test; however, 70% ethanol extracts and its fractions had strong antimutagenic effects against mutation induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and 4-nitroquinoline-1-oxide (4NQO). The ethyl acetate fraction of CL (200 ${\mu}g$/plate) showed approximately 72.1% inhibitory effect on the mutagenesis induced by 4NQO against TA98 strain, whereas 69.6% and 67.0% inhibitions were observed on the mutagenesis induced by MNNG and 4NQO against TA100 strain. In anticancer effects, the cytotoxicity of CL extract and its fractions against cancer cell lines including human cervical adenocarcinoma (HeLa), human hepatocellular carcinoma (HepG2), human breast adenocarcinoma (MCF-7), human lung carcinoma (A549) and transformed primary human embryo kidney (293) were investigated. The treatment of 1 mg/mL CL ethyl acetate fraction had the highest cytotoxicity of 74.5%, 70.7% and 80.3% against HeLa, MCF-7 and A549 cells, respectively. In contrast, the extract and its fractions showed only 2$\sim$31% cytotoxicity for a normal human kidney cell line (293). In vivo anticancer effect of CL extract was tested using Balb/c mice transplanted sarcoma-180 cells. CL ethyl acetate fraction showed the highest inhibition rate of 56.4% at the 50 mg/kg concentration.
This study was conducted to evaluate the estrogen activity of silkworm (Bombyx mori) pupa extracts and their fractions. Powdered samples of freeze-dried silkworm pupa were extracted at room temperature (RT), $40^{\circ}C$, $60^{\circ}C$, $80^{\circ}C$, and $100^{\circ}C$ in water (D.W), chloroform, ethyl acetate, and methanol for 6h and then filtered (0.45 um). The extracts were then freeze-dried. The estrogenic activity of these extracts was then investigated by competition binding assays using estrogen receptor ${\alpha}\;(ER{\alpha})$ and $ER{\beta}$, and by evaluating their effects on the proliferation of the human breast cancer cell line, MCF-7. Among the extracts evaluated, water extracts prepared at RT showed the highest binding affinity to $ER{\alpha}$ ($IC_{50}$, 1.76 ug/ml) and $ER{\beta}$ ($IC_{50}$, 0.07 ug/ml). In addition, MCF-7 cells that were treated with 62.5 ug/ml of the RT extract showed the greatest increase in proliferation (2-fold; 1291.79%) when compared to control cells (659.82%). Next, the water extract that was prepared at RT (sample 1) was dissolved in D.W. and further fractionated using a Dowex 50W - 8X ($H^+$) column. The flow-through and wash were then pooled together and freeze-dried (sample 2). The bound materials were then eluted with 20 mM NaCl, after which they were applied to a Dowex 1X2 - 200 ($Cl^-$) column and washed with D.W. to remove the sodium ions. The eluants were then freeze-dried (sample 3). Of these fractions, sample 2 showed the highest binding affinity to ER{\alpha} ($IC_{50}$, 1.44 ug/ml) and $ER{\beta}$ ($IC_{50}$, 1.18 ug/ml). In addition, MCF-7 cells that were treated with sample 2 (15.6 ug/ml) showed the largest increase in growth (1159.39%) when compared to control cells (525.26%). Taken together, these results suggest that the fraction of the RT water extract of silkworm pupa referred to as sample 2 may be useful as a phytoestrogen.
Kim, Mihyang;Kim, Bokyung;Kim, Jae-Deog;Kang, A-Ram;Lee, Chang-Eun;Seo, Jungmin;Lee, Dong-Geun;Jo, Jung-Kwon;Kim, Yuck Yong;Yu, Ki Hwan;Lee, Sang-Hyeon
Journal of Life Science
/
v.27
no.4
/
pp.456-463
/
2017
The purpose of this study was to investigate the effect of 3 types of medicinal herbs (Glycyrrhizae radix, Astragali radix and Dioscorea rhizoma) extracts on estrogen-like activities, proliferation and differentiation in osteoblast. Human breast cancer cell line MCF7 was transfected using an estrogen responsive luciferase reporter plasmid for measure the estrogen-like activity. Estrogen-like activities of extracts were in the range of 1.11~5.73 fold to that of negative control. The extract of G. radix showed the strongest estrogen-like activities. The estrogen-like activities of 50 and $500{\mu}g/ml$ extracts of G. radix were similar to that of $10^{-8}$ and $10^{-7}$ M standard solution ($17{\beta}-estradiol$), respectively. G. radix extract showed no cytotoxicity against osteoblast MC3T3-E1 cells at $1{\sim}1,000{\mu}g/ml$. The extract of A. radix showed no significant proliferation of osteoblast. However, the extract of G. radix and D. rhizome showed maximum 148% and 133% proliferation effects. The extract of G. radix also increased alkaline phosphatase activity and the maximum was 122% at $100{\mu}g/ml$ compared to that of control. The nodule formation by the method of the Alizarin red S staining increased compared to control. These results suggest that G. radix is able to perform the bone formation and prevent osteoporosis.
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