Unit Presentation Dec 08 Short


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Unit Presentation Dec 08 Short

  1. 1. Epigenetics, Diet and Disease<br />Giles Elliott<br />
  2. 2. Diet<br />Epigenetics<br />DNA Methylation<br />Disease<br />Colon Cancer<br />Overview<br />
  3. 3. What is DNA Methylation?<br />C<br />C<br />C<br />C<br />C<br />C<br />C<br />C<br />A<br />A<br />A<br />A<br />A<br />A<br />G<br />G<br />G<br />G<br />G<br />G<br />T<br />T<br />T<br />T<br />T<br />T<br />T<br />T<br />C<br />C<br />C<br />C<br />C<br />C<br />C<br />C<br />A<br />A<br />A<br />A<br />A<br />A<br />G<br />G<br />G<br />G<br />G<br />G<br />T<br />T<br />T<br />T<br />T<br />T<br />T<br />T<br /><ul><li>Covalent DNA modification
  4. 4. Retained through mitosis</li></ul>C<br />C<br />C<br />C<br />C<br />C<br />A<br />A<br />A<br />A<br />A<br />A<br />A<br />A<br />G<br />G<br />G<br />G<br />G<br />G<br />G<br />G<br />T<br />T<br />T<br />T<br />T<br />T<br />C<br />C<br />C<br />C<br />C<br />C<br />A<br />A<br />A<br />A<br />A<br />A<br />A<br />A<br />G<br />G<br />G<br />G<br />G<br />G<br />G<br />G<br />T<br />T<br />T<br />T<br />T<br />T<br />Dnmt3<br />Dnmt1<br />
  5. 5. What does DNA methylation do?<br />Global Methylation<br />Regional Methylation<br /><br /><br />Promoter<br />Gene<br /><ul><li>Low denisity
  6. 6. Genomic stability
  7. 7. Repression of transposable elements
  8. 8. High Density – CpG Islands (CGIs)
  9. 9. Located in promoters and introns
  10. 10. Gene Silencing</li></li></ul><li>DNA methylation in relation to disease - cancer<br /><ul><li>Tumour samples exhibited reduced methylation compared to corresponding normal tissue- Feinberg and Vogelstein, 1983
  11. 11. Due to reduction in global methylation - hypomethylation
  12. 12. Increased genomic instability – increased mutation rate
  13. 13. Reactivation of transposable elements – chromosomal rearrangements
  14. 14. Loss of imprinting</li></li></ul><li>DNA methylation in relation to disease - cancer<br />Hypermethylation of CpG Islands<br /> Silencing of tumour suppressor genes<br /><ul><li>Affects genes involved in: cell cycle, DNA repair, cell-cell interactions, angiogenisis, proliferation, apoptosis...
  15. 15. Present in both hereditary and sporadic types of cancer
  16. 16. Occurs at all stages of cancer development
  17. 17. Interacts with genetic lesions – 2 hit
  18. 18. Underlying causes inducing hypermethylation unknown</li></li></ul><li>Age related hypermethylation<br />
  19. 19. DNA methylation in relation to disease – Colorectal cancer<br />Adenoma<br />Lifetime risk 50%<br />Carcinoma<br />Lifetime risk 5%<br />Healthy Mucosa<br />Somatic mutation…..<br />Dietary and <br />microbiological factors<br />act on the mucosa for<br />decades before the<br />emergence of tumours.<br />APC<br />K-ras<br />p53<br />Epigenetic deregulation…<br />MLH1<br />p16<br />ESR-1<br />Tumour Biology<br />Diet-Related “Field Effects” ?<br />
  20. 20. Evidence for the Field Effect<br />
  21. 21. Evidence for the Field Effect<br />Adenoma<br />Carcinoma<br />Healthy Mucosa<br />Diet-Related “Field Effects” ?<br />
  22. 22. Evidence for the Field Effect<br />Adenoma<br />Carcinoma<br />Healthy Mucosa<br />Diet-Related “Field Effects” ?<br />
  23. 23. Evidence for the Field Effect<br /><ul><li>Multinomial models, based on CGI methylation profiles from normal mucosa correctly identified:
  24. 24. 78.9% of cancer patients
  25. 25. 87.9% of non- cancer patients
  26. 26. informative variables: APC, HPP1, p16, SFRP4, WIF1 and ESR1
  27. 27. CGI methylation of SFRP4, SFRP5 and WIF1 was used to correctly identify:
  28. 28. 61.5% of adanoma patients
  29. 29. 78.9% of neoplasia-free subjects
  30. 30. This indicates that apparently normal mucosa of patients with neoplasia has undergone significant epigenetic modification associated with the onset of carcinogenesis.
  31. 31. Methylation of the genes selected by the models may play a role in the earliest stages of the development of colorectal neoplasia.</li></ul>What influences the field effect?<br />Diet?<br />
  32. 32. Age related hypermethylation<br />Inter-individual variation possibly due to diet?<br />
  33. 33. Evidence for the involvement of diet<br /><ul><li>Older twins exhibited more differences in epigenetic status than young twins
  34. 34. Due to environmental not genetic factors</li></li></ul><li>Effect of individual dietary components<br />Deficiency in methyl donors (folate, choline, vit B12) can induce global hypomethylation<br /><ul><li>Majority of studies performed in cancer cell lines or animal models
  35. 35. What is going on in humans?</li></ul>Treatment of cells with polyphenols can reduce hypermethylatiion by inhibition of dnmt1<br />
  36. 36. Diet and colon cancer in man<br /><ul><li>CRC predominantly a western disease
  37. 37. Impact of western diet highlighted by increased CRC incidence in Japan
  38. 38. Risk of CRC increased by obesity
  39. 39. Inflammatory bowel disease increases risk of CRC</li></li></ul><li>Increased age related hypermethylation in UC<br />Hypothesis:<br />Inflammatory signals due to dietary (or microbial) factors lead to aberrant DNA methylation resulting in a predisposition to CRC<br />
  40. 40. Aim: To use faecal DNA methylation as a surrogate marker of colonic methylation<br /><ul><li>Quantify faecal methylation levels
  41. 41. Determine if faecal and tissue methylation correlate?
  42. 42. Are there significant differences in faecal methylation between patient groups?
  43. 43. Predicting disease status
  44. 44. Could surrogate markers be used in dietary intervention studies?</li></li></ul><li>Previous work<br />
  45. 45. Summary of early studies<br /><ul><li>Hypermethylated CGIs detected more frequently in cancer than non cancer patients faecal sample
  46. 46. Indicated that a panel of genes may be necessary
  47. 47. Majority of studies performed using non- or semi-quantitative assays
  48. 48. little attempt to correlate to tissue </li></li></ul><li>Study design<br /><ul><li>Matching tissue and faecal samples from:
  49. 49. 19 Cancer patients
  50. 50. 18 Adenoma patients
  51. 51. 20 Patients with no-neoplasia
  52. 52. Faecal samples from 169 health individuals
  53. 53. ~10 males and 10 females/decade until 80 years old
  54. 54. Measured methylation of 8 CGIs</li></li></ul><li>Analysis of faecal methylation levels in patients<br />* p ≤ 0.05, ** p ≤ 0.01, ***p ≤ 0.001<br />
  55. 55. Predicting disease status<br />Multinomial modelling<br />Age + ESR1<br /> Overall = 77%<br /> Sensitivity = 63% , Specificity = 84% <br />ESR1 and p14<br /> Overall = 71%<br /> Sensitivity = 42%, Specificity = 86% <br />
  56. 56. Correlation between tissue and faecal samples<br />ESR1 was the only CGI to correlate<br />
  57. 57. Summary<br /><ul><li>Some differences in faecal methylation seen between patient groups
  58. 58. Unable to significantly predict disease status from faecal methylation profiles
  59. 59. Faecal methylation profiles do not correlate with corresponding tissue samples</li></li></ul><li>Study limitations<br /><ul><li>Number of patients
  60. 60. Age difference between patient groups
  61. 61. Use of patients with no-neoplasia as controls
  62. 62. Comparison to age and sex matched volunteers to patient groups</li></li></ul><li>Age and Sex matched Patients vs. Volunteers<br />
  63. 63. <ul><li>Quantify faecal methylation levels
  64. 64. Determine if faecal and tissue methylation correlate?
  65. 65. Are there significant differences in faecal methylation between patient groups?
  66. 66. Predicting disease status
  67. 67. Could surrogate markers be used in dietary intervention studies?</li></ul><br /><br /><br /><br />?<br />
  68. 68. Acknowledgments<br />IFR<br />Nigel Belshaw<br />Ian Johnson<br />Liz Lund<br />KasiaPrzybylska<br />Jack Dainty<br />Newcastle university<br />John Mathers<br />Amanda Coupe<br />Wendy Bal<br />Wansbeck Hospital<br />D M Bradburn<br />TNO Netherlands<br />Robert Kleeman<br />Funding<br />FSA<br />BBSRC CSG<br />NuGO<br />