Chlorophyllin treatment reduces benzo[a]pyrene (BP)-induced CYP1A1 and CYP1B1 expression and BP-DNA adduct formation in normal human mammary epithelial cells (NHMECs).
Keshava-C; Divi-RL; Whipkey-DL; Leonard-SL; Poirier-MC; Weston-A
Proceedings of the American Association for Cancer Research (AACR) 95th Annual Meeting, March 27-31, 2004, Orlando Florida, Abstract 741. Philadelphia, PA: American Association for Cancer Research, 2004 Mar; 46:1
The capacity of cultured NHMECs to activate BP and form BP-DNA adducts has been previously demonstrated. The advantage of these normal breast cells obtained from reduction mammoplasty is that each cell strain reflects the metabolic profile of an individual donor. Recently, we showed wide inter-individual variation in BP-induced CYP1A1 ( 100-fold) and CYP1B1 ( 40-fold) expression in NHMECs. Although levels of CYP1A1 and CYP1B1 induction were highly correlated with each other (r2 = 0.82, p<0.01) neither appeared to be a good predictor of BP-DNA adduct formation (r2 = 0.40 and 0.50, p<0.10 and 0.05, respectively). However, we hypothesized that since the first step in BP activation is dependent on cytochrome p450 (CYP) metabolism, inhibition of CYP induction might result in reduction of BP-DNA adduct formation. In these studies one NHMEC strain was exposed for 24h to 5ÁM chlorophyllin either before exposure to 4ÁM BP for 24h, together with BP exposure (4ÁM for 24h) or both 24h before and during exposure to 4ÁM BP for 24h. After exposure, CYP1A1 and CYP1B1 gene expression was monitored by Affymetrix DNA oligonucleotide microarrays (U133A) and reverse transcription/quantitative real-time polymerase chain reaction (RT-PCR). In addition, DNA was extracted and BP-DNA adducts were measured using an anti-BP-DNA chemiluminescence immunoassay. The data showed that BP-induced CYP1A1 and CYP1B1 expression was mitigated by all three chlorophyllin treatment regimens, but that maximal inhibition of enzyme induction was obtained with chlorophyllin exposure before and during BP-exposure. Using RT-PCR, the CYP1A1 induction was maximally reduced by >85% (p<0.01) and CYP1B1 induction was reduced by >70% (p<0.01). BP-induced expression of several other genes (STAT1, 1F1TM1 and G1P3) was also reduced by chlorophyllin exposure. BP-DNA adduct formation was concomitantly reduced in direct proportion to the levels of CYP1A1 and CYP1B1 reduction (r2 = 0.90, p<0.01), and the DNA adduct levels were maximally reduced by >85% (p<0.01). These data show that chorophyllin substantially blocks the BP-induction of CYP1A1 and CYP1B1 expression and associated BP-DNA adduct formation in NHMECs.
Exposure-levels; Exposure-assessment; Tissue-culture; Cell-cultures; Monitoring-systems; Cancer; Polycyclic-aromatic-hydrocarbons; Disease-prevention; Disease-control
Conference/Symposia Proceedings; Abstract
Proceedings of the American Association for Cancer Research (AACR) 96th Annual Meeting, April 16-20, 2005, Anaheim/Orange County, CA