1. Estrogen Metabolism Role in Oncology
Tom Archie, MD
SLWRMC Tumor Board
June 19, 2008

2. Early breast cancer – poorly differentiated
50yo Caucasian Female
Early breast cancer – poorly differentiated
MRI negative for contralateral tumor
Receptor Status:
Triple Negative (ER, PR, HER2/neu)
Lumpectomy followed by Taxotere and Cytoxan

3. Estrogen Metabolism Testing
Identify high risk patients for new breast cancer
Others: prostate cancer, leukemia, olfactory tumors, and Parkinson’s Disease (probably more to come)
Identify high risk of recurrence in breast cancer patients (and other cancers)
Modify risk via modification of estrogen metabolism balance
Future: “pre-mammogram” biomarker

5. Catechol Estrogen Quinones to DNA Adducts
Endogenous Estrogens can become carcinogenic via formation of catechol estrogen quinones, which react with DNA to form specific depurinating estrogen-DNA adducts.
The mutations resulting from these adducts can lead to cell transformation and the initiation of breast cancer.
Irregardless of ER status
Mechanism: Sheer volume of DNA “apurinic” sites – DNA repair enzymes make mistakes, leading to single nucleotide polymorphisms (SNP)

6. Catechol Estrogen Quinones to DNA Adducts
4-OH-estrone induces DNA Adduct formation in normal breast epithelium (MCF-10F cells)
Saeed M et al. Int J Cancer Apr 15;120(8):

7. Human Study of Urinary Estrogen Metabolites w/ and w/o Breast CA
Human study comparing healthy controls, breast cancer patients, and “high risk” patients (as determined by oncologists in study)
The levels of the ratios of depurinating DNA adducts to their respective estrogen metabolites/conjugates were significantly higher in high-risk women (p < 0.001) and women with breast cancer (p < 0.001) than in control subjects.
This mechanism represents the best understood and documented initiation step in the formation of any cancer.
Gaikwad NW et al. The molecular etiology of breast cancer: evidence from biomarkers of risk. Int J Cancer May 1;122(9):

9. COMT and CE (Catechol Estrogen)
Quantitatively, the most active CE conjugative pathway is methylation. CE methylation is catalyzed by COMT
Catechol-O-methyltransferase (COMT) a classical phase II enzyme, catalyzes the transfer of methyl groups from S-adenosyl methionine, the enzyme cofactor, to hydroxyl groups of a number of catechol substrates, including the CEs.
Under normal circumstances, CEs are, for the most part, promptly O-methylated by COMT to form 2- and 4-O-methylethers, which are then excreted.
While virtually all catechols are substrates for COMT, the highest affinities for the enzyme are exhibited by the CEs
Journal of the National Cancer Institute Monographs No. 27, 2000

10. Low Functioning COMT and Breast Cancer Risk
Genetic epidemiology studies have proposed a possible correlation between the low activity allele (COMTLL) and increased breast cancer risk
Lavigne JA, et al. An association between the allele coding for a low activity variant of catechol-O-methyltransferase and the risk for breast cancer. Cancer Res 1997;57:5493–5497.
Huang CS, et al. Breast cancer risk associated with genotype polymorphism of the estrogen metabolizing genes CYP17, CYP1A1, and COMT: A multigenic study on cancer susceptibility. Cancer Res 1999;59:4870–4875.
Yim D-S, et al. Relationship between the val158met polymorphism of catechol O-methyl transferase and breast cancer. Pharmacogenetics 2001;11:1–8.

11. COMT and Breast Cancer
COMT protects cells from the genotoxicity and cytotoxicity of catechol estrogens, by preventing their conversion to quinones
Adds methyl group (-CH3) at the -OH site that would otherwise be oxidized by peroxidase enzymes
Low activity of COMT leads to higher levels of depurinating estrogen-DNA adducts that can induce mutations and initiate cancer.
MCF-10F (human breast epithelial cells that are ER neg)
Estrogen-DNA adducts’ carcinogenicity independent of ER status
Zahid M et al. Free Radic Biol Med Dec 1;43(11):
Lu F et al.J Steroid Biochem Mol Biol ; 105(1-5): 150–158.

12. Low Functioning COMT Common
25% of US Caucasians are homozygous for the val108/158met polymorphism in the COMT gene
Lachman HM, et al. Pharmacogenetics 1996;6:243–250.
Scanlon PD, et al. Science 1979;203:63–65.
27% Chinese Americans and 34% Japanese Americans
Wu A et al. Cancer Res 2003;63: 7526–7529
Val108/158Met SNP associated with 3-4x reduction in functional enzymatic rate of COMT.
Zhu BT. Curr Drug Metab 2002;3: 321–349

13. E2:E16 Ratio - Breast Cancer Risk
Prospective Study 10,786 women aged with 5 ½ yr followup
Measured urinary estrogen metabolites
144 breast cancer pts w/ 4 matched controls for each cancer
Highest quintile E2:E16 ratio
Premenopausal: OR 0.58 (42% risk red)
Postmenopausal: OR 1.29 (29% risk inc)
Muti et al. Epidemiology Nov;11(6):635-40

14. Broccoli increases E2:E16 ratio
Increase E2:E16 ratio 29.5% with broccoli 500gr/day
Cruciferous vegetables cause the upregulation of Cyp1A2 (19%) and Cyp1A1 and inhibit Cyp2E1
Indole 3 Carbinol (glucocinolate)
Sulforaphane (isothiocyanate)
Diindolylmethane (glucocinolate)
Calcium D Glucarate

16. Interpretation Low 2-Hydroxyestrone/16α-Hydroxyestrone Ratio
Premenopausal female
Increased risk of ongoing carcinogensis leading to treatment failure

17. Interpretation Poor methylation capacity – she is a “slow methylator”
4-Methoxyestrone is undetectable
4-Hydroxyestrone is not being methylated adequately.
4-Hydroxyestrone level is high.
This is associated with increased levels of 4-catechol estrogen DNA adducts, which are strongly associated with the initiation of breast and prostate cancer.
COMT is likely genetically slow
Principal agent for eliminating catechol estrogens

18. Interpretation
Interestingly, the methylation of 2OHE is adequate, whereas methylation of 4OHE is not
I find no literature citing methylation preferences for 2OHE vs. 4OHE
Fact remains that additional methylation support is needed

19. Treatment Goals Enhance methylation Decrease Cyp1B1 activity
Increase E2:E16 ratio
Cruciferous vegetables

20. Improve Methylation Increase substrate for COMT (SAMe)
Add methyl donors
Folate, methylcobalamin (B12)
Trimethylglycine (Betaine)
Vit B6 (to discourage the accumulation of homocysteine and encourage the formation of glutathione via synthesis of cysteine)
There is no physiological mechanism to suggest an adverse interaction between methylation and the metabolism of either taxotere or cytoxan

21. Improve Methylation
COMT

22. Cyp1B1 inhibition (ie: reduction of DNA adducts)
Reduce xenobiotic pollutant exposure
N-acetyl Cysteine
Sulforaphane (glucosinolate from broccoli) induces quinone reductase, which takes CEQs back to catechol estrogens, reducing the potential for the creating of DNA adducts.
Hwang. J Med Food Summer;8(2):
Glutathione conjugates are not playing much of a role in protecting against DNA adducts.

23. Cyp 1B1 Inhibition to decrease DNA Adduct Formation
Increased methylation of catechol estrogens leads to feedback inhibition of Cyp1B1
Dawling et al. Cancer Res Jun 15;63 (12):

24. Cyp 1B1 Inhibition to decrease DNA Adduct Formation
Reduced Lipoic Acid
N-acetyl Cysteine
Resveratrol
Melatonin (minimal but positive effect)
Zahid M. et al. Inhibition of depurinating estrogen-DNA adduct formation by natural compounds. Chem Res Toxicol Dec;20(12): Epub 2007 Nov
Chen et al. Resveratrol inhibits TCDD-induced expression of CYP1A1 and CYP1B1 and catechol estrogen-mediated oxidative DNA damage in cultured human mammary epithelial cells Carcinogenesis vol.25 no.10 pp , 2004
doi: /carcin/bgh183

25. Synergism b/t Paclitaxel and Broccoli Glucosinolate
Diindolylmethane in combination with paclitaxel synergistically inhibits growth of Her2 / neu human breast cancer cells through G2M phase cell-cycle arrest and induction of apoptosis / necrosis
McGuire KP, et al. J Surg Res May 15;132(2): Epub 2006 Mar 31. 

26. Broccoli and Antitumor Effects
Sulforaphane inhibits breast cancer growth and induces Quinone Reductase
Hwang. J Med Food Summer;8(2):
I3C induces Br CA cell cycle arrest

27. Potential Risk Uncertain effect on Cyp3A4
(60% of drugs)
Sulforaphane inhibtis
Diindolylmethane has no effect
Could affect concentration of these drugs and theoretically increase adverse drug events or decrease efficacy
Taxotere metabolized by Cyp3A4
Consider avoiding near time of infusion

28. Prostate CA
Small study of urine estrogen metabolites in men with prostate cancer vs. benign urological d/o vs. healthy controls
4-OHE1-DNA Adducts detected at higher levels in samples from subjects with prostate cancer and benign urological conditions compared to healthy males
This is the first demonstration that CEQ-derived DNA adducts are present in urine samples from subjects with prostate cancer.
Markushkin Y et al. Potential biomarker for early risk assessment of prostate cancer. Prostate Oct 1;66(14):

29. Extension to Other Cancers
This mechanism is also involved in
Initiation of leukemia by benzene
Rat olfactory tumors by naphthalene
Neurodegenerative diseases such as Parkinson's disease by dopamine.
Estrogens and Human Diseases. Volume 1089 published November 2006 Ann. N.Y. Acad. Sci. 1089: 286–301 (2006). doi: /annals

30. Conclusion
No human intervention trials on manipulation of estrogen metabolism in patients w/ active breast cancer
Epidemiologic studies support cruciferous vegetables and methyl donors to decrease breast cancer risk
In vitro studies showing anticancer effects of brassica
Risk of non-action vs. action?
Enhance methylation now
Increase E2:E16 ratio now but reduce likelihood of possible interaction w/ metabolism of taxotere by avoiding for 1 week prior and 2 days after administration of taxotere
Inhibit Cyp1B1 now

31. In a word….

32. One Last Thought for Future Discussion
Multifocal Angiostatic Therapy

33. Multifocal Angiostatic Therapy
curcmin, artemsia, mistletoe, ginger scutellaria, resveratrol, grapeseed extract, green tea, gingko, squalamine, Vit D silymarin, glycine,
Multifocal Angiostatic Therapy
VEGFR
EGCG, silymarin, quercetin, resveratrol, soy isoflavones, curcumin, EPA
Cu antagonists
curcmin, scutellaria, cartilege, silymarin, green tea
ginger artemsia mistletoe curcumin scutellaria
bFGFR and TNF-1:
Cu antagonists
Growth
Factors
NFkB
COX-2
VEGF, AKs, bFGF, IL8, MMPs,
TNF-1, heparinases, collagenases
Silymarin, Glycine, Ginger
Anti- NFkB: poria, coriolus, ginger, resveratrol, green tea, artemsia, quercetin, carnosol, panax ginseng, silymarin, salicylates, curcumin, picentannol, basil, Cu antagonists rosemary
green tea quercetin magnolia resveratrol, soy, curcumin holy basil rosemary ganoderma licorice Vit E
Anti-COX-2: quercetin, scutellaria, EPA/DHA, licorice, ginger, resveratrol, grapeseed extract, curcumin, salicylates, garlic, green tea, panax ginseng, silymarin, bilberry, antioxidants, boswellia, aloe