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Cytochrome P450 1A2 Metabolizes 17β-Estradiol to Suppress Hepatocellular Carcinoma.

Ren J, Chen GG, Liu Y, Su X, Hu B, Leung BC, Wang Y, Ho RL, Yang S, Lu G, Lee CG, Lai PB - PLoS ONE (2016)

Bottom Line: The inhibitory effect was significantly enhanced in cells with CYP1A2 overexpression, which effectively conversed E2 to the cytotoxic 2-ME.The anti-HCC effect of E2 was not associated with estrogen receptors ERα and ERβ as well as tumor suppressor P53 but enhanced by the approved anti-HCC drug sorafenib.The reduction of CYP1A2 significantly disrupts this metabolic pathway, contributing the progression and growth of HCC and the gender disparity of this malignancy.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, Faculty of Medicine, The Chinese University of Hong Kong; New Territories, Hong Kong, China.

ABSTRACT
Hepatocellular carcinoma (HCC) occurs more frequently in men than in women. It is commonly agreed that estrogen plays important roles in suppressing HCC development, however, the underlying mechanism remains largely unknown. Since estrogen is mainly metabolized in liver and its metabolites affect cell proliferation, we sought to investigate if the liver-specific cytochrome P450 1A2 (CYP1A2) mediated the inhibitory effect of estrogen on HCC. In this study, the expression of estrogen-metabolizing enzyme CYP1A2 was determined in HCC tissues and cell lines. Cell proliferation and apoptosis were assessed in cells with or without CYP1A2 overexpression. The levels of 17β-estradiol (E2) and its metabolite 2-methoxyestradiol (2-ME) were determined. A xenograft tumor model in mice was established to confirm the findings. It was found that CYP1A2 expression was greatly repressed in HCC. E2 suppressed HCC cell proliferation and xenograft tumor development by inducing apoptosis. The inhibitory effect was significantly enhanced in cells with CYP1A2 overexpression, which effectively conversed E2 to the cytotoxic 2-ME. E2 in combination with sorafenib showed an additive effect on HCC. The anti-HCC effect of E2 was not associated with estrogen receptors ERα and ERβ as well as tumor suppressor P53 but enhanced by the approved anti-HCC drug sorafenib. In addition, HDAC inhibitors greatly induced CYP1A2 promoter activities in cancer cells, especially liver cancer cells, but not in non-tumorigenic cells. Collectively, CYP1A2 metabolizes E2 to generate the potent anti-tumor agent 2-ME in HCC. The reduction of CYP1A2 significantly disrupts this metabolic pathway, contributing the progression and growth of HCC and the gender disparity of this malignancy.

No MeSH data available.


Related in: MedlinePlus

Estrogen suppresses proliferation of liver cancer cells.In A–E HCC cells were seeded at 5x103 cells/per well of 96-well plates and grown for 24 hours. The cells were treated as indicated and analyzed for proliferation rate by MTT. Each column represents mean±s.d. of data obtained from four replicate wells. Consistent results were obtained in three independent experiments. (A) E2 inhibited proliferation of HCC cells. The cells were treated with E2 of the indicated concentrations for 48 hours before MTT assay. The proliferation rate of individual cell line is expressed as the value of treated cells versus that of the untreated cells (= 100%). (B) 2-ME inhibited Hep3B cell growth in a dose-dependent manner. Hep3B was treated with 2-ME of the indicated concentrations for 48 hours before MTT assay. The proliferation rate is expressed as the value of treated Hep3B cells versus that of the untreated Hep3B cells (= 100%). (C) Inhibitory effect of E2 is not influenced by P53. Hep3B cells were transfected with wt p53 DNA or empty vector control. Inset shows the expression of p53 in the transfected cells. The proliferation rate is expressed as the value of treated cells versus that of the untreated cells of the same transfection (= 100%). (D) The Hep3B cells were treated with fulvestrant of indicated concentrations supplemented with or without 30μM E2 for 48 hours before MTT assay. E2 inhibited the HCC cell proliferation in a way basically independent of the increasing doses of fulvestrant. (E) E2 additively enhanced the effect of sorafenib in suppressing the growth of Hep3B cells. (F) E2 and 2-ME induced apoptosis in Hep3B cells. The cells were treated with E2 or ME2 for 48 hours before being harvested for PI staining. Apoptotic cells were identified as the sub-G1 peak of the DNA content profiles.
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pone.0153863.g001: Estrogen suppresses proliferation of liver cancer cells.In A–E HCC cells were seeded at 5x103 cells/per well of 96-well plates and grown for 24 hours. The cells were treated as indicated and analyzed for proliferation rate by MTT. Each column represents mean±s.d. of data obtained from four replicate wells. Consistent results were obtained in three independent experiments. (A) E2 inhibited proliferation of HCC cells. The cells were treated with E2 of the indicated concentrations for 48 hours before MTT assay. The proliferation rate of individual cell line is expressed as the value of treated cells versus that of the untreated cells (= 100%). (B) 2-ME inhibited Hep3B cell growth in a dose-dependent manner. Hep3B was treated with 2-ME of the indicated concentrations for 48 hours before MTT assay. The proliferation rate is expressed as the value of treated Hep3B cells versus that of the untreated Hep3B cells (= 100%). (C) Inhibitory effect of E2 is not influenced by P53. Hep3B cells were transfected with wt p53 DNA or empty vector control. Inset shows the expression of p53 in the transfected cells. The proliferation rate is expressed as the value of treated cells versus that of the untreated cells of the same transfection (= 100%). (D) The Hep3B cells were treated with fulvestrant of indicated concentrations supplemented with or without 30μM E2 for 48 hours before MTT assay. E2 inhibited the HCC cell proliferation in a way basically independent of the increasing doses of fulvestrant. (E) E2 additively enhanced the effect of sorafenib in suppressing the growth of Hep3B cells. (F) E2 and 2-ME induced apoptosis in Hep3B cells. The cells were treated with E2 or ME2 for 48 hours before being harvested for PI staining. Apoptotic cells were identified as the sub-G1 peak of the DNA content profiles.

Mentions: To examine the effects of E2 and 2-ME on the proliferation of HCC cells, cells were treated with different doses of E2 “Fig 1A” or 2-ME “Fig 1B” for 48 h. The treatment time was determined by carrying out several optimization experiments in which we found that both of short treatment time and high cell confluence attenuated the inhibitory effects of E2 and 2-ME, indicating that only dividing cells responded to the reagents. At the end of the treatment, cell proliferation was measured by MTT assay. Both E2 and 2-ME were able to inhibit HCC cell proliferation. However different cell lines responded differently to E2 treatment. For example, 30μM E2 could lead to 40% inhibition of Hep3B cell proliferation whereas only 20% in HepG2 cells. It was noted that the suppressive effect of E2 only appeared at high doses of E2 though as low as 10nM E2 was able to activate the estrogen receptors (S2 Table, S1 Fig).


Cytochrome P450 1A2 Metabolizes 17β-Estradiol to Suppress Hepatocellular Carcinoma.

Ren J, Chen GG, Liu Y, Su X, Hu B, Leung BC, Wang Y, Ho RL, Yang S, Lu G, Lee CG, Lai PB - PLoS ONE (2016)

Estrogen suppresses proliferation of liver cancer cells.In A–E HCC cells were seeded at 5x103 cells/per well of 96-well plates and grown for 24 hours. The cells were treated as indicated and analyzed for proliferation rate by MTT. Each column represents mean±s.d. of data obtained from four replicate wells. Consistent results were obtained in three independent experiments. (A) E2 inhibited proliferation of HCC cells. The cells were treated with E2 of the indicated concentrations for 48 hours before MTT assay. The proliferation rate of individual cell line is expressed as the value of treated cells versus that of the untreated cells (= 100%). (B) 2-ME inhibited Hep3B cell growth in a dose-dependent manner. Hep3B was treated with 2-ME of the indicated concentrations for 48 hours before MTT assay. The proliferation rate is expressed as the value of treated Hep3B cells versus that of the untreated Hep3B cells (= 100%). (C) Inhibitory effect of E2 is not influenced by P53. Hep3B cells were transfected with wt p53 DNA or empty vector control. Inset shows the expression of p53 in the transfected cells. The proliferation rate is expressed as the value of treated cells versus that of the untreated cells of the same transfection (= 100%). (D) The Hep3B cells were treated with fulvestrant of indicated concentrations supplemented with or without 30μM E2 for 48 hours before MTT assay. E2 inhibited the HCC cell proliferation in a way basically independent of the increasing doses of fulvestrant. (E) E2 additively enhanced the effect of sorafenib in suppressing the growth of Hep3B cells. (F) E2 and 2-ME induced apoptosis in Hep3B cells. The cells were treated with E2 or ME2 for 48 hours before being harvested for PI staining. Apoptotic cells were identified as the sub-G1 peak of the DNA content profiles.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4836701&req=5

pone.0153863.g001: Estrogen suppresses proliferation of liver cancer cells.In A–E HCC cells were seeded at 5x103 cells/per well of 96-well plates and grown for 24 hours. The cells were treated as indicated and analyzed for proliferation rate by MTT. Each column represents mean±s.d. of data obtained from four replicate wells. Consistent results were obtained in three independent experiments. (A) E2 inhibited proliferation of HCC cells. The cells were treated with E2 of the indicated concentrations for 48 hours before MTT assay. The proliferation rate of individual cell line is expressed as the value of treated cells versus that of the untreated cells (= 100%). (B) 2-ME inhibited Hep3B cell growth in a dose-dependent manner. Hep3B was treated with 2-ME of the indicated concentrations for 48 hours before MTT assay. The proliferation rate is expressed as the value of treated Hep3B cells versus that of the untreated Hep3B cells (= 100%). (C) Inhibitory effect of E2 is not influenced by P53. Hep3B cells were transfected with wt p53 DNA or empty vector control. Inset shows the expression of p53 in the transfected cells. The proliferation rate is expressed as the value of treated cells versus that of the untreated cells of the same transfection (= 100%). (D) The Hep3B cells were treated with fulvestrant of indicated concentrations supplemented with or without 30μM E2 for 48 hours before MTT assay. E2 inhibited the HCC cell proliferation in a way basically independent of the increasing doses of fulvestrant. (E) E2 additively enhanced the effect of sorafenib in suppressing the growth of Hep3B cells. (F) E2 and 2-ME induced apoptosis in Hep3B cells. The cells were treated with E2 or ME2 for 48 hours before being harvested for PI staining. Apoptotic cells were identified as the sub-G1 peak of the DNA content profiles.
Mentions: To examine the effects of E2 and 2-ME on the proliferation of HCC cells, cells were treated with different doses of E2 “Fig 1A” or 2-ME “Fig 1B” for 48 h. The treatment time was determined by carrying out several optimization experiments in which we found that both of short treatment time and high cell confluence attenuated the inhibitory effects of E2 and 2-ME, indicating that only dividing cells responded to the reagents. At the end of the treatment, cell proliferation was measured by MTT assay. Both E2 and 2-ME were able to inhibit HCC cell proliferation. However different cell lines responded differently to E2 treatment. For example, 30μM E2 could lead to 40% inhibition of Hep3B cell proliferation whereas only 20% in HepG2 cells. It was noted that the suppressive effect of E2 only appeared at high doses of E2 though as low as 10nM E2 was able to activate the estrogen receptors (S2 Table, S1 Fig).

Bottom Line: The inhibitory effect was significantly enhanced in cells with CYP1A2 overexpression, which effectively conversed E2 to the cytotoxic 2-ME.The anti-HCC effect of E2 was not associated with estrogen receptors ERα and ERβ as well as tumor suppressor P53 but enhanced by the approved anti-HCC drug sorafenib.The reduction of CYP1A2 significantly disrupts this metabolic pathway, contributing the progression and growth of HCC and the gender disparity of this malignancy.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, Faculty of Medicine, The Chinese University of Hong Kong; New Territories, Hong Kong, China.

ABSTRACT
Hepatocellular carcinoma (HCC) occurs more frequently in men than in women. It is commonly agreed that estrogen plays important roles in suppressing HCC development, however, the underlying mechanism remains largely unknown. Since estrogen is mainly metabolized in liver and its metabolites affect cell proliferation, we sought to investigate if the liver-specific cytochrome P450 1A2 (CYP1A2) mediated the inhibitory effect of estrogen on HCC. In this study, the expression of estrogen-metabolizing enzyme CYP1A2 was determined in HCC tissues and cell lines. Cell proliferation and apoptosis were assessed in cells with or without CYP1A2 overexpression. The levels of 17β-estradiol (E2) and its metabolite 2-methoxyestradiol (2-ME) were determined. A xenograft tumor model in mice was established to confirm the findings. It was found that CYP1A2 expression was greatly repressed in HCC. E2 suppressed HCC cell proliferation and xenograft tumor development by inducing apoptosis. The inhibitory effect was significantly enhanced in cells with CYP1A2 overexpression, which effectively conversed E2 to the cytotoxic 2-ME. E2 in combination with sorafenib showed an additive effect on HCC. The anti-HCC effect of E2 was not associated with estrogen receptors ERα and ERβ as well as tumor suppressor P53 but enhanced by the approved anti-HCC drug sorafenib. In addition, HDAC inhibitors greatly induced CYP1A2 promoter activities in cancer cells, especially liver cancer cells, but not in non-tumorigenic cells. Collectively, CYP1A2 metabolizes E2 to generate the potent anti-tumor agent 2-ME in HCC. The reduction of CYP1A2 significantly disrupts this metabolic pathway, contributing the progression and growth of HCC and the gender disparity of this malignancy.

No MeSH data available.


Related in: MedlinePlus