Field cancerization refers to genetic and molecular alterations that occur in histologically normal tissue surrounding tumors. These alterations predispose the tissue to developing additional new cancers. The document discusses two cases presenting with multiple primary tumors in the oral cavity and larynx as examples of field cancerization. It then reviews the original description of field cancerization from 1953 and various theories for how it occurs. The concept of an "etiologic field effect" is introduced, which broadens the understanding of cancer susceptibility at the molecular, cellular and environmental levels. Several examples of field cancerization are described for different cancer types. Clinical tools for detecting field cancerization like iodine staining and toluidine blue staining are also mentioned.

1. FIELD CANCERIZATION & ITS CLINICAL
IMPLICATIONS
DR MUHAMMED MUNEER M
MS GENERAL SURGERY
SGMC & RF
TRIVANDRUM KERALA

2. Introduction
Case 1:
 Carcinoma (L) buccal mucosa : Irradiated in 2001
 Ca (L) Upper alveolus-Hard palate : IPM in 2006
 Ca (R) Buccal Mucosa :WE + SSG in 2015
Case 2:
 Ca Larynx : Radical RT in 2004
 CaTongue :WE + ESOHND in 2015

3. Introduction
Field Cancerization!!
 1953
 Slaughter et al

4. Field Cancerization - 1953 Slaughter et al
783 patients with Oral SCC
 88 (11.2 % ) Instances of independent multiple
tumors
 far beyond the statistical possibility of chance occurrence.
 Microscopic evidence of multicentric origin
 Abnormal and hyperplastic, atypical, epithelium
- surround all oral cancers for varying distances.

5. Introduction
Field Cancerization!!
 1953
 Slaughter et al

7. Theories....
Two theories
1. Multiple genetic abnormalities in the
whole tissue region. Independently of
each other (Slaughter et al)
2. Multiple lesions due to widespread
migration of transformed cells
2A : Micrometastases – eg: saliva
2B : Intraepithelial migration
1. Califano, J., et al : Genetic progression model for head and neck cancer: implications for field cancerization.Cancer Res., 56: 2488–2492,
1996.
2. Bedi, et al Multiple head and neck tumors: evidence for a common clonal origin. Cancer Res., 56: 2484–2487, 1996.

8. Defining....
FC: A field of cellular / molecular alteration, which predisposes to
the development of neoplasms within that territory.
‘Etiologic field effect’ (EFE)
Various etiologic factors and their interactions
- ‘Field Of Susceptibility’
Histologically: Normal, Hyperplastic or Dysplastic
Cancer development, per se, is not a requirement

9. The genesis of the ‘Etiologic
Field Effect’
Molecular Pathological Epidemiology (MPE)
Relationships between etiologic factors and somatic molecular
alterations
 BMI and MSI expression in CRC
 Cigarette smoking and MSI, CpG island & BRAF mutation in CRC
 Cigarette smoking and KRAS mutation in lung tumors
 Epstein–Barr virus and CpG island hypermethylation in gastric cancer
 H. pylori infection and CpG island methylation in gastric epithelial cells
 Viral hepatitis and CpG island hypermethylation in HCC

10. The ‘Etiologic Field Effect’
Broadens the horizons of cancer susceptibility at
 Molecular
 Cellular
 Environmental levels
Etiologic field effect: reappraisal of the field effect concept in cancer predisposition and progression Paul Lochhead1,AndrewT Chan2,3,
Reiko Nishihara4,5, Charles S Fuchs3,4, Andrew H Beck6, EdwardGiovannucci3,5,7 and Shuji Ogino4,7,8

11. FC vs EFE

12. Re-defining “Boundaries”...
Skin
Oral cavity
Esophagus
Breast
Lung
Gall Bladder
Prostate
Vulva & Cervix

13. Lung cancer
Franklin et al
 Sampled tissues from the entire tracheobronchial
tree
 Same individual with 50-pack-years of smoking
 No lung cancer
p53 mutation
 Present in 7/10 sites in both lungs
 Indicating a field change
1. FranklinWA, GazdarAF, Haney J,Wistuba, La Rosa FG, KennedyT, Ritchey DM, MillerYE: Widely dispersed p53 mutation in respiratory
epithelium. A novel mechanism for field carcinogenesis. J Clin Invest 1997, 100:2133-2137.

14. Lung Cancer
LOH, especially at chromosome 12p12
 Detected in normal bronchial epithelium of long-term
smokers
 Deletion hotspots at two chromosomes (2q35-q36, 12p12p13)
 In NSCLC & Normal bronchial epithelial cells
Suggests
 LOH could indicate susceptibility
 ?Hallmark of progression of phenotypically normal
epithelium1. Pan H, Califano J, Ponte JF, Russo AL, Cheng KH,Thiagalingam A, Nemani P, Sidransky D,Thiagalingam S: Loss of heterozygosity patterns
provide fingerprints for genetic heterogeneity in multistep cancer progression of tobacco smoke-induced non-small cell lung cancer.
Cancer Res 2005, 65:1664-1669.

15. Esophageal cancer
Barrett's esophagus – A model for field cancerization.
Prevo et al. mapped
 213 endoscopic biopsies from 58 patients
 p53 mutation as a clonal marker
 50% were clonal
 Cancer fields ranging from 1 cm to 9 cm
LOH at 9p21 (p16 locus) &17p13 (p53 locus) - Evidence for FC
1. Prevo LJ, Sanchez CA,Galipeau PC, Reid BJ: p53-mutant clones and field effects in Barrett's esophagus. Cancer Res 1999, 59:4784-4787.
2. Galipeau PC, Prevo LJ, Sanchez CA, Longton GM, Reid BJ: Clonal expansion and loss of heterozygosity at chromosomes 9p and 17p in
premalignant esophageal (Barrett's) tissue. J Natl Cancer Inst 1999, 91:2087-2095.

16. Gastric cancer
53% of gastric cancers - Epigenetic silencing via CpG island
hypermethylation of LIMS1
1. LIMS1 methylation was observed in normal-appearing
gastric tissue - Suggesting an early genetic event
2. C-erb amplification was observed in a subset of tumors
and the normal mucosa close to tumor margin
3. Aneuploidy was frequent in normal mucosa at about 3
cm distance from the tumor margin
1. Kim SK, Jang HR, Kim JH, Noh SM, Song KS, Kim MR, Kim SY,YeomYI, Kim NS,Yoo HS, KimYS: The epigenetic silencing of LIMS2 in
gastric cancer and its inhibitory effect on cell migration. Biochem Biophys Res Commun 2006, 349:1032-1040
2. Kim JS, Choi CW, Kim BS, Shin SW, KimYH, MokYJ, Koo BH: Amplification of c-erbB-2 proto-oncogene in cancer foci, adjacent normal,
metastatic and normal tissues of human primary gastric adenocarcinomas. J Korean Med Sci 1997, 12:311-315.
3. Kim JY, Cho HJ: DNA ploidy patterns in gastric adenocarcinoma. J Korean Med Sci 2000, 15:159-166.

17. Vulval and cervical cancers
 Vulval intraepithelial neoplasia (VIN) is often clonal
with vulval squamous cell carcinoma (VSCC).
 This suggestsVIN may be a precursor lesion ofVSCC
X chromosome inactivation analysis
• 9 cases ofVIN
• 10 cases ofVSCC with contiguousVIN
• 11 cases ofVSCC with noncontiguousVIN
• Indicates the majority ofVIN andVSCC were monoclonal

18. Skin cancer
The organ most exposed to environmental carcinogens
 Skin SCC: Signature gene mutations and alterations
Precursor lesions such as actinic keratosis (AK)
 Associated with p53 mutations and p16 expression
SCC is also associated with p53 mutations & increased p16 expression

19. Skin cancer
“The whole neighborhood is affected".
 Mutations in p53 are used as biomarkers of clonality
Kanjilal S et al
• p53 mutations were present NMSC as well as the normal
appearing peri-lesional skin
1. Kanjilal S, Strom SS, Clayman GL,Weber RS, el-Naggar AK, KapurV,Cummings KK, Hill LA, Spitz MR, Kripke ML, et al.: p53 mutations in
nonmelanoma skin cancer of the head and neck: molecular evidence for field cancerization. Cancer Res 1995, 55:3604-3609.

20. Skin cancer
Jonason et al.
 Clones of 60–3000 cells more frequent in sunexposed
than sun-shielded skin
 Genetically AlteredClones + Other Genetic Alterations =
Malignant Phenotype
1. Jonason AS, Kunala S, Price GJ, Restifo RJ, Spinelli HM, Persing JA, Leffell DJ,Tarone RE, Brash DE: Frequent clones of p53-mutated
keratinocytes in normal human skin. Proc Natl Acad Sci U S A 1996, 93:14025-14029.

21. Bladder Cancer
Molecular and histopathologic data comparison
• Suggested monoclonality of multifocal lesions
Hoglund M studied 32 tumors from 6 bladders
 Multiple tumors from the same bladder genetically identical
 Genetically transformed but histologically normal urothelium
1. Denzinger S, Mohren K, Knuechel R,Wild PJ, Burger M,WielandWF, HartmannA, Stoehr R: Improved clonality analysis of multifocal
bladder tumors by combination of histopathologic organ mapping, loss of heterozygosity, fluorescence in situ hybridization, and p53
analyses. Hum Pathol 2006, 37:143-151.
2. Hoglund M: Bladder cancer, a two phased disease? Semin Cancer Biol 2006.

22. Gallbladder cancer
Mitochondrial D310 alterations
(Analysed in Normal, Pre-neoplastic and NeoplasticGB samples)
 Infrequent in normal samples
 Increased in frequency in dysplastic lesions and normal
adjacent epithelium
Mitochondrial genome alterations : pre-malignant
GB lesions
1. Tang M, Baez S, Pruyas M, DiazA, CalvoA, Riquelme E,Wistuba: Mitochondrial DNA mutation at the D310 (displacement loop)
mononucleotide sequence in the pathogenesis of gallbladder carcinoma. Clin Cancer Res 2004, 10:1041-1046.

23. Breast Cancer
LOH in normal breast tissue adjacent to tumor
 8/30 cases
 Same missing allele as the corresponding carcinoma
1. Deng G, et al: Loss of heterozygosity in normal tissue adjacent to breast carcinomas. Science 1996, 274:2057-2059
2. Euhus DM et al: Loss of heterozygosity in benign breast epithelium in relation to breast cancer risk. J Natl Cancer Inst 2002

24. Breast Cancer
Euhus DM et al
FNA biopsy samples
 30 asymptomatic women with known risk of ca breast
 11 with normal cytology
 19 with proliferative cytology
LOH was observed in
2/11 with normal cytology
14/19 with abnormal cytology
Random FNA biopsy sampling of breast tissue for molecular
screening could potentially be useful in individualized medicine
1. Euhus DM et al: Loss of heterozygosity in benign breast epithelium in relation to breast cancer risk. J Natl Cancer Inst 2002

25. Prostate cancer
Genomic instability
Gene expression
Mitochondrial genome alterations
Methylation in GSTP1 and RARbeta2
 Present In
 Absent in normal epithelia from BPH
 Telomere alterations : in normal appearing tissue close
to tumors - good predictor of prostate cancer recurrence
1. Hanson JA, Gillespie JW, GroverA,Tangrea MA, Chuaqui RF, Emmert-Buck MR,Tangrea JA, Libutti SK, Linehan WM,Woodson KG: Gene
promoter methylation in prostate tumor-associated stromal cells. J Natl Cancer Inst 2006, 98:255-261.
2. FordyceCA, Heaphy CM, Joste NE, Smith AY, HuntWC, Griffith JK: Association between cancer-free survival and telomere DNA content
in prostate tumors. J Urol 2005, 173:610-614
Field Cancerization
Prostate cancer
Adjacent stroma
Adjacent normal gland close to tumor

26. Genetic Markers of FC
 LOH - microsatellite alterations
 Chromosomal instability
 Mutations in the TP53 gene
One of the Most reliable markers (van Houten et al., 2000)
Detected by
 DNA amplification techniques
 Immunohistochemistry
 in situ hybridization
1. Tabor, M. P.,et al Persistence of genetically altered fields in head and neck cancer patients: Biological and clinical implications. Clin. Cancer
Res., 7: 1523–1532, 2001.
2. Hittelman,W. N. Genetic instability in epithelial tissues at risk for cancer.Ann. N.Y. Acad. Sci., 952: 1–12, 2001.
3. Brennan, J. A et al Molecular assessment of histopathological staging in squamous-cell carcinoma of the head and neck. N. Engl. J. Med.,
332: 429–345, 1995.

27. Genetic Markers of FC
“Patch – Field – Carcinoma” Concept
Corresponding Proposed Genetic Alterations
HNSCC
1. Califano, J., et al : Genetic progression model for head and neck cancer: implications for field cancerization.Cancer Res., 1996.

28. Markers of FC
HNSCC: By measuring LOH
 One-third(10 of 28) biopsy taken from the macroscopically
normal mucosa adjacent to the tumors have tumor-associated
genetic alterations (Four biopsies in each quadrant surrounding the tumor)
 7 of 10 - genetically altered cells in margins of specimen
 15 microsatellite markers on 6 different chromosomes
1. Tabor, M. P.,et al Persistence of genetically altered fields in head and neck cancer patients: Biological and clinical implications. Clin.
Cancer Res., 7: 1523–1532, 2001

29. Potential biomarkers for
oral carcinogenesis

31. Clinical Tools for FC
Staining
 Iodine Staining
 Toluidine Blue
Chemiluminescence
 TheVizilite Kit
 Tissue FluorescenceVisualization (Fluorescent
Visualization Devices)

32. Clinical Tools..
Iodine
Stains Normal Epithelium
Abnormal Epithelium – No Glycogen – No staining
Toluidine Bule
Acidophilic metachromatic dye of thiazine group
Selectively stains acidic tissue components (sulfates, carboxylates and phosphate
radicals)
Thus staining DNA and RNA
Established diagnostic adjunct in detecting pre malignant oral
lesions
1. E Allegra, N Lombardo, L Puzzo1, A Garozzo.The usefulness of toluidine staining as a diagnostic tool for precancerous and cancerous
oropharyngeal and oral cavity lesions. Acta Otorhinolaryngologica Italica.2009;29:187-90.
2. Lewei Zhang, MicheleWilliams, Catherine F Poh, et al.Toluidine Blue Staining Identifies High-Risk PrimaryOral Premalignant Lesions with
Poor Outcome. Cancer Research.2005;65:8017-21

33. Clinical Tools..
Toluidine Bule
Acidophilic metachromatic dye of thiazine group
Selectively stains acidic tissue components (sulfates, carboxylates
and phosphate radicals)
Thus staining DNA and RNA
Established diagnostic adjunct in detecting pre
malignant oral lesions
1. E Allegra, N Lombardo, L Puzzo1, A Garozzo.The usefulness of toluidine staining as a diagnostic tool for precancerous and cancerous
oropharyngeal and oral cavity lesions. Acta Otorhinolaryngologica Italica.2009;29:187-90.
2. Lewei Zhang, MicheleWilliams, Catherine F Poh, et al.Toluidine Blue Staining Identifies High-Risk PrimaryOral Premalignant Lesions with
Poor Outcome. Cancer Research.2005;65:8017-21

34. Clinical Tools..
 Rinsing with a dilute acetic acid
 Abnormal squamous epithelium - appear
 Low-energy wavelength light
 Normal epithelium will absorb the light and
appear dark

35. Tissue Fluorescence Visualization
FluorescentVisualization Devices)
 Detects cellular, structural, and/or metabolic activity
changes in oral mucosa
 By observing the fluorescence response to light excitation.
 Natural tissue fluorescence is caused by "fluorophores“.
 When fluorophores are excited by light of an appropriate
wavelength (eg: blue), they emit their own light at a longer
wavelength (eg: green).
A fluorophore is a fluorescent chemical compound that can re-emit light upon light excitation.
 Fluorophores typically contain several combined aromatic groups, or plane or cyclic molecules with several π bonds.

36. Tissue Fluorescence
Visualization
Abnormal fluorescence patterns aid the clinician in
detecting
 An increase in metabolic activity in the epithelium;
 A breakdown of the fluorescent collagen cross-links in the
connective tissue layer beneath the basement membrane;
 An increase in tissue blood content, (inflammation or
angiogenesis)
 The presence of pigments (e.g., melanin) which absorb
light
 NormalTissue : Pale Green Auto-fluorescence
 AbnormalTissue : Dark

37. Tissue Fluorescence
Visualization
FluorescenceVisualization
Loss (FVL)
FluorescenceVisualization
Retained (FVR)
ClinicallyApparent Lesion

38. Tissue Fluorescence
Visualization
Conclusion:
No dysplasia beyond
the FVR margin

39. Translating Clinically...
Lumerman et al., 1995
• 6.6 to 36% of pre-malignant lesions transformed into SCC
Silverman et al., 1984
• 17.5% incidence of progression of oral cavity dysplasias
 over an average time of 8yrs
Field cancerization would predict the presence of foci of pre-
malignant change surrounding a malignancy.

40. Clinical Translation...
 62.5% of HNSCC second primary tumor recurrences
are from similar clonal fields left behind after
resection
 Field size of over 7 cm has been mapped
1. Tabor MP, Brakenhoff RH, Ruijter-Schippers HJ,Van DerWal JE, Snow GB, Leemans CR, Braakhuis BJ: Multiple head and neck tumors
frequently originate from a single preneoplastic lesion. AmJ Pathol 2002, 161:1051-1060
2. Tabor MP, Brakenhoff RH, Ruijter-Schippers HJ, Kummer JA, Leemans CR, Braakhuis BJ: Genetically altered fields as origin of locally
recurrent head and neck cancer: a retrospective study. Clin Cancer Res 2004, 10:3607-3613.

41. Clinical Translation...
1. p53 positivity in adjacent epithelium : High Recurrence
2. p53-negative in adjacent epithelium : Better OverallSurvival
3. Ki67 was not related
4. No significant association between Ki67 and p53

42. Molecular Definitions...

43. Practical Implications...
1. Risk assessment
2. Early detection
3. Chemoprevention
4. Complemetary Evaluation of Biopsy
5. Sugical Margins

44. Risk assessment & Early
Detection
Future identification of
Biosensors that signal the genesis of disease
Rather than biomarkers of the disease
RiskAssessment
Early Detection
Chemoprevention.

45. For example...
 Random FNA biopsy - LOH in normal breast epithelial cells
Predict breast cancer risk
In this study
 Gail risk model predicted a mean lifetime breast cancer risk
 16.7% for women with no LOH compared
 22.9% for women with LOH
LOH correlate with individual risk
Useful for early detection and risk assessment
1. Euhus DM, et al : Loss of heterozygosity in benign breast epithelium in relation to breast cancer risk. J Natl Cancer Inst 2002,

46. Primary Chemoprevention
Genetic changes present in Normal Appearing cells
Identify & Recruit individuals at risk
Primary chemoprevention
Epigenetic gene silencing
(Promoter Hypermethylation and Transcriptional Repression of tumor-associated genes)
 An Early Event in
 Breast, Prostate, Colorectal, Gastric & OvarianCancers
 Potential for RiskAssessment & Chemoprevenion

47. Primary Chemoprevention
 Knowledge of methylation patterns
 Methylation (Epigenetic changes) reversal agents
 Hydralazine
 5-Aza-2'-deoxycytidine
 Zebularine
 Magnesium valporate
Reversal of Epigenetic Events

48. Chemoprevention
Biofluids (representative of cells) from a particular organ
• Useful noninvasive samples for disease surveillance.
 For eg, genetic changes preceding ca breast development might be
detectable in nipple aspirates
Useful endpoint measures
Targets of secondary chemoprevention
(i.e., to prevent the progression of pre-
malignant lesions to invasive cancers)
Markers in precancerous lesions

49. Complementary Evaluation Of
Biopsy Specimen
An important clinical utility of field cancerization
Currently: HistologyThe Gold Standard
 Absence of abnormal cells  No Cancer?!?!
Histologically normal - but molecular signatures of cancer fields
 Suggest that some tissue are progressing towards malignancy
Such High Risk Patients
Secondary prevention stategies
Close surveillance for early detection

50. Tumor margins
Complementary Molecular profiling of Margins Reduces Reccurences
1. Kim J, et al Unfavourable prognosis associated with K-ras gene mutation in pancreatic cancer surgical margins. Gut 2006, 55:1598
Molecular Profiling Of Surgical Margins
Help Reduce Scar Recurrences
Multiple Independent Patches Of Cancer Fields - In the same organ exposed to the same
insult – Even Clean Molecular Margins Need not prevent SPT within the same organ

51. Molecular Tumor margins –
Clinical Translation...Brennan et al.
 Used p53 gene mutation in the primary tumor and the surgical
margins
 52% of patients with histopathologically negative margins had
malignant cells (p53 mutated) at the margin
 38% of these cases with positive margins after molecular analysis
Local Recurrence
 No patient with negative molecular margins failed treatment at the
local site
1. Brennan, J. A., Mao, L., Hruban, R. H., Boyle, J. O., Eby,Y. J., Koch,W. M., Goodman, S. N., and Sidransky, D. Molecular assessment of
histopathological staging in squamous-cell carcinoma of the head and neck. N. Engl. J. Med., 332: 429–435, 1995.

52. Mol. Tumor margins -
Clinical Significance...
Data to suggest
If p53 mutations are present in single / multiple
biopsies of distant mucosa
Second PrimaryTumors  More frequent
1. Nees, M., et al Expression of mutated p53 occurs in tumor-distant epithelia of head and neck cancer patients: a possible molecular basis for
the development of multiple tumors. Cancer Res., 53: 4189–4196, 1993.
2. Waridel, F. Et al Field cancerisation and polypolyclonalp53 mutation in the upper aerodigestive tract. Oncogene, 14:163–169, 1997.

53. Looking at the Margins..

54. Treatment of FC??
1. Treatment of the entire field
2. Should efficiently remove all clinical lesions
3. Reduce the potential risk for the appearance of new
lesions

55. H&N FC
13 cis-retinoic acid
Genetic alterations in mucosal fields remain unchanged
This implies  Definitive therapy
 Targeted ablation of altered clonal populations
 Repair of genetic damage in affected cells
1. ArmstrongWB, Meyskens FL Jr.Chemoprevention of head and neck cancer. Otolaryngol Head Neck Surg 2000;12:728-735.
2. Lotan R et al. Suppression of retinoic acid receptor-beta in premalignant oral lesions and its up-regulation by isotretinoin. N Engl J Med
1995;332:1405-1410.
3. HongWK et al. 13-cis-retinoic acid in the treatment of oral leukoplakia. N Engl J Med 1986;315:1501-1505.
•Good clinical response in pre-malignant lesions
•Regression in leukoplakia
•Prevention of second primary tumors

56. Cutaneous FC
3 groups:
1. Topical drugs
1. 5- fluorouracil
2. Imiquimod
3. Diclofenac gel
2. Photodynamic therapy
Level B recommendations for treatment of FC (European
guidelines for topical photodynamic therapy)
3. Resurfacing surgical procedures
 Ablative lasers
 Dermabrasion
 Chemical peelings
Level A Recommendation
(British Dermatology Asssociation)

57. Photodynamic Therapy
 Application of a 5-ALA agent
 Converted to the photosensitizing molecule PPIX (protoporphyrin IX)
in the preneoplastic cell
 Rendering the lesion susceptible to visible blue or red spectrums.
 Reactive O2 radicals,
 Cure rates are as high a 75% to 92% after one or two sessions
 Cosmetic effect is considered excellent in all studies.
• Level B recommendations for treatment of FC (European guidelines for topical
photodynamic therapy)

59. Conclusions
Need for substantive prospective studies to compare the sensitivity
of light microscopy and new molecular approaches to detect
residual disease and establish the prognostic significance.
Molecular diagnostics can be combined with conventional
prognostic factors to provide a basis for identifying high risk
patients
Future targeted therapeutic agents directed against the molecular
changes, which can revert the genotypic changes, rather than the
phenotypic reversal.

60. Thank You
ThankYou