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Zeligs MA, Fulfs JC, Peterson R, Wilson SM, McIntyre L, Sepkovic DW, Bradlow HL. Absorbable Diindolylmethane (BioResponse-DIM® [BR-DIM]) is a patented, orally active formulation of 3,3'-diindolylmethane (DIM), available as a dietary supplement. DIM is found in cruciferous vegetables, but also under investigation for its chemopreventive and pro-apoptotic activities. Based on DIM's known growth-inhibition of endometrial(1) and cervical cancer cell lines(2), oral BR-DIM was tested in vivo in a model of estrogen-driven uterine growth(3) and during monitored use in humans. Groups of immature female rats were implanted with placebo or androstenedione pellets and fed either control, or BR-DIM (250 mg/kg/day [62.5 mg/kg/day DIM]) containing diets. Androstenedione served as a source of aromatase-derived estrogen. When sacrificed after 72 hrs, the BR-DIM group revealed significantly lower uterine/body weight indexes (p=0.04) compared to control rats. Thus, BR-DIM reduced estrogen-related uterine growth in rats. In monitored human use, BR-DIM, at the label-recommended dose of 300 mg/day (providing 75 mg/day of DIM), was taken as a dietary supplement by a 41 year old with treated, Stage I breast cancer on Tamoxifen (20 mg/day). Dysfunctional uterine bleeding and Tamoxifen-related endometrial hyperplasia were present before BR-DIM use, as confirmed by transvaginal ultrasound and endometrial biopsy. Following 1 month of BR-DIM use, resolution of bleeding was noted. Normalization of endometrial contour and thickness were documented during 3 months of BR-DIM use, and lasting 1 month following cessation of its use (from 16.0 mm before, to 6.6 mm during, and 5.0 mm after DIM). 2 months after discontinuing BR-DIM, endometrial thickness had returned to the pre-treatment level (17.2 mm). In testing urinary estrone metabolites in women with treated breast cancer, estrogen 2-hydroxylation was shown to not be influenced by Tamoxifen alone. During combined Tamoxifen use with BR-DIM, a shift to greater estrogen 2-hydroxylation and less 16-hydroxylation was demonstrated. Therefore, Tamoxifen did not inhibit promotion of estrogen 2-hydroxylation by BR-DIM. Women with treated breast cancer responded to BR-DIM with the same increased 2-hydroxylation of estrogen as previously demonstrated in healthy women(4). Supported by these observations, BR-DIM deserves further evaluation as a potential uterine protective agent during combined use with Tamoxifen(5) and during estrogen exposure from HRT(6). References 1. Chen DZ, Qi M, Auborn KJ, Carter TH. Indole-3-carbinol
and diindolylmethane induce apoptosis of human cervical cancer cells and
in murine HPV16-transgenic preneoplastic cervical epithelium. J Nutr.
2001 Dec;131(12):3294-302.
Leong H, Firestone GL, Bjeldanes LF. 3,3'-Diindolylmethane (DIM), a major in vivo product of indole-3-carbinol (I3C), is a promising anticancer agent derived from vegetables of the Brassica genus including broccoli, Brussels sprouts and cabbage. We report here that DIM has a potent cytostatic effect in cultured human Ishikawa endometrial cancer cells. A combination of northern blot and quantitative PCR analyses revealed that DIM induced the level of TGF-alpha transcripts by approximately 4-fold within 24 h of indole treatment. DIM also induced a 4-fold increase in the activity of the estrogen response marker, alkaline phosphatase (AP). Co-treatment of cells with the estrogen receptor (ER) antagonist ICI, or with the inhibitor of PKA-mediated activation of the ER, H89, ablated the DIM induction of both TGF-alpha expression and AP activity. Furthermore, DIM increased the maximum stimulatory effect of estrogen on TGF-alpha expression. Co-treatment with the protein synthesis inhibitor, cycloheximide, abolished the inductive effects of DIM, indicating differences in the mechanistic requirements of DIM and estrogen. DIM treatment also stimulated levels of secreted TGF-alpha protein by >10-fold. The ectopic addition of TGF-alpha inhibited the growth of Ishikawa cells, whereas incubation with a TGF-alpha antibody partially reversed the growth inhibitory effects of DIM. Taken together, these results extend our previous findings of the ligand independent estrogen receptor agonist activity of DIM, and uncover an essential role for the stimulation in TGF-alpha expression and the TGF-alpha activated signal transduction pathway in the potent cytostatic effects of DIM in endometrial cancer cells. Click for FREE full text article
Chen DZ, Qi M, Auborn KJ, Carter TH. Dietary indole-3-carbinol (I3C) has clinical benefits for both cervical cancer and laryngeal papillomatosis, and causes apoptosis of breast cancer cells in vitro. We asked whether I3C and its major acid-catalyzed condensation product diindolylmethane (DIM), which is produced in the stomach after consumption of cruciferous vegetables, could induce apoptosis of cervical cancer cell lines. We also asked whether this effect could be observed in vivo. In vitro, both I3C and DIM caused accumulation of DNA strand breaks in three cervical cancer cell lines. Induction of apoptosis was confirmed by nuclear morphology, nucleosome leakage, altered cytoplasmic membrane permeability and caspase 3 activation. Neither I3C nor DIM caused apoptotic changes in normal human keratinocytes. In C33A cervical cancer cells, DIM was more potent than I3C [dose at which the number of viable cells was 50% of that in untreated cultures (LD(50)) = 50-60 micromol/L for DIM and 200 micromol/L for I3C in a mitochondrial function assay] and faster acting. Furthermore, I3C reduced Bcl-2 protein in a time- and dose-dependent manner. In HPV16-transgenic mice, which develop cervical cancer after chronic estradiol exposure, apoptotic cells were detected in cervical epithelium by TdT-mediated dUTP nick-end labeling staining and by immunohistochemical staining of active caspase 3 only in mice exposed to 17beta-estradiol (E2) and fed I3C. Rare apoptotic cells were also observed by hematoxylin and eosin staining in the spinous layer of the cervical epithelium in both control and transgenic mice. Estradiol reduced the percentage of these late-stage apoptotic cells in the cervical epithelium of transgenic, E2-treated mice, but this reduction was prevented by I3C. These data confirm the pro-apoptotic action of I3C on transformed
cells in vitro, extend the observations to cervical cancer cells and to
DIM and show for the first time that dietary I3C results in increased
apoptosis in target tissues in vivo.
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