GOOD EVENING

DIAGNOSTIC AIDS FOR THE DETECTION OF ORAL CANCER

Presented by
Ridhima Dhamija Pankaj Bhansal Samyutha Balasubramaniam Sheethal Srinivas M.S.RAMAIAH DENTAL COLLEGE AND HOSPITAL, BANGALORE (DEPT. OF ORAL MEDICINE AND RADIOLOGY)

INTRODUCTION
A neoplasm is an abnormal mass of tissue, the growth of which exceeds and is uncoordinated with that of the normal tissues and persists in the same excessive manner after cessation of the stimulus which evoked the change*.

* Willis RA: The Spread of Tumors in the Human Body. London, Butterworth & Co, 1952

EPIDEMIOLOGY

Study of cancer patterns in population can contribute substantially to knowledge about the origin of cancer.

WORLDWIDE

Oral carcinoma is one of the most prevalent cancer. Oral cancer is one of the 10 most common cause of death. Sixth most common cause of cancer related deaths. Oral Cancer overall Mortality: 50-55%

Cancer of the oral cavity and pharynx is the first and third commonest cancer in Indian men and women, respectively. One of the major(91%) reasons for the high incidence of oral cancer is the continued use of tobacco.

Etiology and risk factors

 Familial and Genetic  Tobacco and tobacco related products  Occupation  Immunosuppression  Alcohol

 Viruses  Diet and nutrition  Atmospheric pollution  Dental factors

Poor prognosis is likely due to .. REASONS????

 Oral cancer is frequently associated with the development of multiple primary tumors.

 Advanced extent of the disease at the time of diagnosis, with over 60% of patients presenting in stages III and IV.

TYPES OF DIAGNOSTIC AIDS

CLINICAL EXAMINATION TOLUIDINE BLUE STAINING COLPOSCOPY LIGHT BASED DETECTION SYSTEM (USING CHEMILUMINESCENCE) VIZILITE MICROLUX DL  LIGHT BASED DETECTION SYSTEM (AUTOFLUORESCENCE) VELSCOPE NUCLEAR MEDICINE BONE SCAN

BIOPSIES CONVENTIONAL INCISIONAL EXCISIONAL PUNCH CYTOLOGY EXFOLIATIVE FINE NEEDLE ASPIRATION CYTOLOGY BRUSH

ORAL SCREENING
 Is defined as the application of a test or tests to people who are apparently free from the disease in question in order to sort out those who probably have the disease from those who probably do not

CRITERIA FOR SCREENING AND FOR SCREENING TESTS

The national screening committee(UK) has enlisted certain criteria for the implementation of a screening programmeThe disease must be an important health problem An accepted treatment must be available for patients with recognised disease Facilities for diagnosis and treatment must be available

There must be a recognizable early or latent stage A suitable test must be available. The natural history of the disease of the condition should be adequately understood The screening test should be cost effective. The screening test should be a continuing process and not a once and for all project.

CHARACTERISTICS OF A GOOD SCREENING TEST: Be simple,safe and acceptable to the public. Detect disease early in its natural history Detect lesions which are treatable or where an intervention will prevent progression. Preferentially detect those lesions which are likely to progress.

BASIC MEASURES OF ACCURACY OF DIAGNOSTIC TESTS

POSITIVE PREDICTIVE VALUE(PPV)

SPECIFICITY

SENSITIVITY

NEGATIVE PREDICTIVE VALUE(NPV)

 SENSITIVITY - The probability that someone who has the target disease will generate a positive result.  SPECIFICITY - The probability that someone who does not have the target disease will generate a negative test finding.

 POSITIVE PREDICTIVE VALUE - The probability that a person with positive test results actually has a the target disease  NEGATIVE PREDICTIVE VALUE - The probability that a person with negative test results doesn t have the disease.

ONE IMPORTANT CRITERION IN ASSESSING THE VALIDITY OF A TEST IS COMPARING IT WITH GOLD STANDARD
The GOLD STANDARD diagnostic test for oral mucosal lesions that are suggestive of premalignancy or malignancy remains TISSUE BIOPSY and HISTOPATHOLOGICAL EXAMINATION

CLINICAL EXAMINATION

It has long been the standard method for oral cancer screening.

 INSPECTION  PALPATION

VERMLLION BORDER OF LIP

SIDE OF THE TONGUE

Recently many adjunctive techniques have emerged with claims of enhanced mucosal examinations and facilitating the detection & distinctions between oral benign,oral premalignant and malignant lesions (OPML).

TOLUIDINE BLUE STAINING AND LIGHT BASED DETECTION SYSTEMS AS DIAGNOSTIC AIDS IN ORAL CANCER DETECTION

TOLUIDINE BLUE STAINING

Toluidine blue(tolonium chloride)is a vital METACHROMATIC DYE that stains nucleic acids and abnormal tissues. It has been used for decades as an aid to the identification of mucosal abnormalities of the oral cavity as well as the cervix It demarcates the extent of lesion prior to excision.

METHOD
Rinse oral cavity with water Rinse with 1% toluidine blue or direct appln. Rinse again with water Rinse with 1% acetic acid or direct appln Observe for darkly stained areas

PREMALIGNANT LESION STAINED WITH TOLUIDINE BLUE

MINOR SALIVARY GLAND LESION STAINED WITH TOLUIDINE BLUE

ADVANTAGES
Helps determine the extent of biopsy site Easy to perform Non-invasive Inexpensive Helps to monitor treated cancer patients for recurrence Sensitivity of 72 100% Specificity of 45 93%

DISADVANTAGES
Not recommended for patients with physical or mental disability Acetic acid may irritate the mucosa Equivocal dye retention Variable mode of application 30% false positive results noted

COLPOSCOPY ORAL DIRECT MICROSCOPY

Saliva removed using a cotton swab

1 3% acetic acid applied

Comparison of vascular pattern

Examine using colposcope (focal length 200nm)

Decision taken regarding biopsy

Typical findings on direct oral microscopy.:-

 Punctation

vessels(dilated,twis
ted,hairpin like cappilaries)

Mosaic vessels
(seen in sharply demarcated areas)

Atypical vessels
(doesn t resemble any pattern)

Advantages:
1.Used to follow mucosal lesions 2.High correlation with biopsy findings 3.Only technique which detects vascular changes.

Disadvantages:
1.More expensive equipment 2.Complexity of technique 3.Requires more clinical research

LIGHT BASED REFLECTANCE SYSTEMS CHEMILUMINESENCE

PRINCIPLES
Dysplastic epithelial cell shows altered characteristics after application Of 1% acetic acid in comparison to normal oral mucosa. The acid applied dissolves the surface glycoprotein layer and alters the nucleocytoplasmic ratio of cells. Dysplastic area gets Readily identified when viewed under bluish white light and appear as acetowhite in comparison to normal diffuse blue color of oral mucosa.

APPARATUS
THE VIZILITE KIT CONTAINS:1%ACETIC ACID SOLUTION A CAPSULE(WHICH EMITS LIGHT) A RETRACTOR AND MANUFACTURERS INSTRUCTION

ADVANTAGES OF USING VIZILITE

Increase the ability of clinicians to detect oral lesions,mainly white lesions and those with white and red areas . Borders observed were usually more extensive than those detected in the visual examination.s

DRAWBACKS
Low specificity and the high rate of false positives. High cost and its inability to indicate the appropriate site for a biopsy. Detection by an expert clinician remains essential.

Author, year and reference

Type of article Pilot study Cross sectional study Cross sectional study Cross sectional study Cross sectional study Cross sectional study

Sample 150

Sens itivit y

Specif icity

Main conclusions
Epithelium behaviour similar to that of the uterine cervix under chemiluminescent illumination

Huber et al., 2004 (7) Ram and Siar , 2005 (6) Epstein et al., 2006 (10) Epstein et al., 2007 (9) Farah and McCullough, 2007 (8) Oh and Laskin, 2007 (5)

40

100 14.2% Diagnostic aid and follow-up of patients with precancerous lesions and cancer Facilitates the detection of lesions of the oral mucosa, mainly white ones It may improve the visual identification of malignant and premalignant oral lesions It does not help in the identification of malignant and premalignant lesions of the oral mucosa Acetic acid mouthwash may be useful but not chemiluminescent light

134

84

55

100 0%

100

BIBLIOGRAPHY

Advantages and difficulties of toluidine blue in the identification of early oral cancerSmaroula Divani1, Maria Exarhou2, Leonidas-Nectarios Theodorou2, Dimitrios Georgantzis1, Haralambos Skoulakis3 Toluidine Blue Staining Identifies High-Risk Primary Oral Premalignant Lesions with Poor Outcome - Lewei Zhang1,4, Michele Williams2, Catherine F. Poh1,2, Denise Laronde1, Joel B. Epstein2, Scott Durham2,4, Hisae Nakamura3, Ken Berean4, Alan Hovan2, Nhu D. Le2, Greg Hislop2, Robert Priddy1, John Hay2, Wan L. Lam2 and Miriam P. Rosin2,3 Utility of Toluidine Blue Staining and Brush Biopsy in Precancerous and Cancerous Oral Lesions - Anurag Gupta, MBBS, Mamta Singh, M.D., Rahela Ibrahim, M.Sc., and Ravi Mehrotra, M.D., M.I.A.C. Direct oral microscopy and its value in diagnosing mucosal lesions Goran W. Gynther, Bjorn Rozell, Anders Heimdahl, Visby and Huddinge Analysis of new diagnostic methods in suspicious lesions of the oral mucosa Anna Trullenque-Eriksson, Marta Munoz-Corcuera, Julian Campo-Trapero, Jorge Cano-Sanchez, Antonio Bascones-Martinez

AUTOFLUORESCENCE AS A DIAGNOSTIC AID IN ORAL CANCER DETECTION

By Samyutha Balasubramaniam

WHAT IS AUTOFLUORESCENCE
It is the intrinsic property possessed by certain substances by which they emit fluorescent light when excited by light of a certain wavelength, without any adjunctive staining techniques. Initially thought to be interference conventional fluorescent microscopy

CONVENTIONAL FLUORESCENT MICROSCOPY

Uses fluorescent dyes to label specific structures fluoresce under light of certain wavelength Autofluorescence - fluorescence of substances other than the fluorophore of interest Named biological autofluorescence considered to be a potentially useful diagnostic tool.

BIOLOGICAL AUTOFLUORESCENCE
Molecules in cells endogenous fluorophores Light of a certain wavelength used

Excitation of endogenous fluorophores Emission of fluorescent light

ENDOGENOUS FLUOROPHORES
INTRACELLULAR Mitochondria Lysosomes NADPH Flavin co-enzymes Aromatic amino acids Lipopigments EXTRACELLULAR Collagen Elastin

PRINCIPLE OF AUTOFLUORESCENCE
Light of a certain wavelength trained upon the tissue light gets absorbed released as light of greater wavelength When certain tissues are illuminated with light energy from short wavelength (380 - 430 nm) light, the absorbed energy is emitted as light at a longer wavelength (475-800 nm) - this is observed as fluorescent light of a different color.

 The fluorescent light can be observed using special optical filters designed to block the background light and allow the fluorescent light to be viewed.

APPLICATION IN DIAGNOSTICS
Since connective tissues and surface epithelia have background autofluorescence (AF), pathologic lesions that grow on the surface of an epithelial layer may stand out compared with normal tissue when viewed in this manner by having a different light pattern than the normal tissue.

 Cellular alterations (dysplastic tissues or in malignancies) - causes a change in the concentrations of endogenous fluorophores. This affects the scattering and absorption of light in the tissue This causes changes in the colour of the fluoresced light - can be observed visually. Dysplastic tissues cause a reduction in the wavelength of emitted light as compared to normal tissues.

DYSPLASTIC TISSUES SHOWING REDUCED AUTOFLUORESCENCE

NORMAL TISSUES SHOWING INCREASED AUTOFLUORESCENCE

INDICATIONS
Difficult to determine which abnormal tissues in the oral cavity are a cause for concern Oral cancer has an abysmal 5 survival rate of 50% Disease morbidity is directly related to the cancer stage at the time of diagnosis Early detection impacts disease mortality and facilitates minimally invasive procedures

 Oral Squamous Cell Carcinoma is believed to progress from :

Oral premalignant lesions

Hyperplasia

Increasing dysplasia (mild, moderate, severe)

Invasive Squamous cell Carcinoma Stage I - IV

Carcinoma in situ

 A significant portion of these cancers are diagnosed at Stage III or IV Huge potential to include oral cancer screening tests as part of regular dental examinations More acceptable than screening tests for other forms of cancer Need was felt for a quick, non-invasive diagnostic tool with good sensitivity and specificity

VELscope
VELscope Handheld imaging device jointly developed by LED Medical Diagnostics and British Columbia Cancer Agency (BCCA) The device is meant to help dentists in the screening of early tissue changes (Oral premalignant lesions) which might not be apparent under normal light

VELscope Unit

COMPONENTS OF VELscope UNIT

1. Fibreoptic probe 2. Nitrogen pumped dye lasers 3. Optical multichannel analyser

1 mm diameter section of tissue selected

Excitation light of 410 nm delivered

Sent to a polychromator and then a series of diodes

Emitted fluorescent light collected by fibreoptic probe

Diodes digitize fluoresence produce emission spectrum

CLINICAL USAGE OF VELscope

NORMAL TONGUE UNDER INCANDESCENT LIGHT

NORMAL TONGUE UNDER VELscope

TISSUE APPEARING CLINICALLY NORMAL

SAME TISSUE UNDER VELscope SHOWING DYSPLASTIC CHANGES

Advantages
Easy to use Non-invasive Inexpensive Can be used for mass screening Detects dysplastic tissues which are clinically inapparent Can be incorporated into a routine dental examination

BONE SCINTIGRAPHY AS A DIAGNOSTIC TOOL IN ORAL CANCER DETECTION

INTRODUCTION
Scintigraphy is also known as bone scan. It is an imaging test that shows areas of increased or decreased bone metabolism. It is a diagnostic test that is primarily used to diagnose or help diagnose a number of abnormalities relating to bones Eg Cancers of the bone, metastases involving the bone, inflammation, infection and fracture of bone.

INDICATIONS
Diagnosis of cancer involving the bone Staging of cancer involving the bone Check for metastasis To select biopsy site Pre and post palliative therapy Chemo and radiotherapy

PRINCIPLE
It involves the injection of a radioactive material (radiotracer) into a vein. The substance travels through the bloodstream and gives off radiation This radiation is detected by a gamma camera Areas of high metabolic activity, eg. malignant sites take up more of the radioactive tracer and appear as hot spots

BONE SCINTIGRAPHY MACHINE

RADIOPHARMACEUTICALS
The radioactive tracer used is Technetium (99mTc) It is a gamma ray emitting nucleotide It is labelled with phosphate or phosphonate, which has affinity to bone Adult dose is 15 25 mCi Children depends on age and weight

PATIENT PREPARATION
Patient should be asked to remove all jewellery and metal objects Patient should void bladder just before the scan Patient asked to remove all prostheses No restrictions as to food intake or driving Patient is told in detail about the procedure

PROCEDURE
The tracer substance is injected I.V. Patient is scanned after 2 to 3 hours A gamma camera fitted with a high resolution collimator is used The whole body is scanned The information is converted into images by the use of suitable processors

ADVANTAGES
More sensitive than radiographs Increased sensitivity Less radiation exposure as compared to radiographs Allows scan of entire body to check for metastasis Non-invasive procedure

DISADVANTAGES
Time consuming procedure Contraindicated in pregnancy and lactation Less specificity Risk of hypersensitivity Patient may feel slight discomfort due to lying still for a prolonged period of time

BIBLIOGRAPHY
Understanding the Biological Basis of Autofluorescence Imaging for Oral Cancer Detection: HighResolution Fluorescence Microscopy in Viable Tissue 1. Ina Pavlova1, 2. Michelle Williams2, 3. Adel El-Naggar2, 4. Rebecca Richards-Kortum4 and 5. Ann Gillenwater3 Noninvasive Diagnosis of Oral Neoplasia Based on Fluorescence Spectroscopy and Native Tissue Autofluorescence Ann Gillenwater, MD; Rhonda Jacob, DDS, MS; Ravi Ganeshappa, MD; Bonnie Kemp, MD; Adel K. ElNaggar, MD, PhD; J. Lynn Palmer, PhD; Gary Clayman, MD, DDS; Michele Follen Mitchell, MD; Rebecca Richards-Kortum, PhD Fluorescence Visualization Detection of Field Alterations in Tumor Margins of Oral Cancer Patients Catherine F. Poh1,2,3, et al Diagnostic aids in the screening of oral cancer Stefano Fedele

Wikipedia, the free encyclopedia

ORAL BIOPSIES AND CYTOLOGY AS DIAGNOSTIC TOOLS IN ORAL CANCER DETECTION

By Sheethal.S

INTRODUCTION
Biopsies are important diagnostic tool for the diagnosis of lesions ranging from simple periapical lesions to malignancies. Planning prior to biopsy is important. It will be beneficial to the receiving pathologist in reaching a helpful and meaningful diagnosis.

BIOPSY
Definition:biopsy is a surgical procedure to obtain tissue from a living organism for its microscopical examination, usually to perform a diagnosis. Objectives-to define a lesion to establish a prognosis to facilitate the prescription of specific treatment To help assess the efficacy of treatment.

INDICATIONS
For diagnostic confirmation of precancerous and malignant lesions. Lesions that interfere with oral function. Lesions of unclear etiology,especially associated with pain. Apparent inflammatory lesions that do not improve within two weeks of removal of local irritants.

TYPES OF BIOPSIES
According to the procedures applied,oral biopsy can be classified by a) Features of the lesion - Direct biopsy Indirect biopsy b) Area of surgical removal - Incisional biopsy Excisional biopsy c) By the timing of biopsy - Pre-operative Intra-operative Post-operative

BIOPSY TECHNIQUE
1. 2. 3. 4. 5. 6. It consists of 6 steps Selection of area of biopsy Preparation of the surgical field Local anaesthetia Incision Handling of the specimen Suturing of the resulting wound

1. Selection of area of biopsy-when dealing with small sized lesions,excisional biopsy is preferred wheareas incisional biopsy is prefered in lesions more than 1cm. toluidine blue staining can be used as an adjunct to select the representative areas when in doubt about the malignant character of the lesion The sample must include healthy tissue at the margin of the lesion

2. Prepation of the surgical field-the surgical area is disinfected with quaternary ammonium compounds. A 0.12%-0.20% chlorhexidine solutionis preferred. 3. Local anaesthesia-an amide containing local anaesthetic with vasoconstrictor should be used and administered away from the lesion to avoid artefacts in the sample.

4. The incision-A clean and defined incision is performed to obtain a slice of the tissue when aiming at an incisional biopsy. Soft tissue incisions should be elliptical in shape producing a v-shaped wedge that includes both the lesions and healthy margins. If various lesions are present,multiple biopsies should be chosen.

Biopsy of minor salivary glands

5. Tissue handling-the specimen is introduced into the fixing solution. The fixing solution preserves the cellular architecture of the tissues. The best fixing solution is 10% formalin. 70% ethanol can also be used. The volume of the fixing agent should exceed 10-20 fold the volume of the sample.

6. Suture-the suture should achieve good hemostasis, facilitate healing and should be after 6-8 days.

Container with a fixing solution

TYPES OF BIOPSIES
Incisional biopsy - Consists of removal of a representative sample of the lesion and the normal adjacent tissue with the preservation of the histological architecture. Incision is made through the entire dermis down to the subcutaneous fat. Advantages- 1. Long and deep incisional biopsy allow a large amount of tissue to be harvested with minimal tension on the surgical wound. 2. Hemostasis can be done more easily due to better visualisation.

INCISIONAL BIOPSY

INCISIONAL BIOPSY DONE ON THE DORSUM OF THE TONGUE

Excisional biopsy - It is usually done when the lesion is smaller than 1cm. The entire lesion or tumor is removed. Ideal for diagnosing small melanomas.

Excisional biopsy of nodular lesion of palate

Punch biopsy-this is done with a round shaped knife ranging in size from 1mm to 8mm. It is suggested to reduce artefacts. Advantages-1.1mm punch is ideal for locations where cosmetic appearance is of importance. 2.Minimal bleeding is seen with 1mm punch Disadvantages-1.tissue obtained by 1mm punch is difficult to see due to the smaller size. 2.It might tear the tissues in vesiculobullous lesions.

Biopsy punch

Specimens obtained by punch biopsy

 Fine needle aspirate - This is done with a rapid stabbing motion of the hand, guiding a needle tipped syringe and the rapid sucking motion applied to the syringe. It is the method used to diagnose tumor deep in the skin or lymphnodes under the skin. The cellular aspirate is mounted on a glass slide and immediate diagnosis can be done with proper staining.

Fine needle aspiration needles

EXFOLIATIVE CYTOLOGY
Method-Lesion stroked gently, firmly with a wet wooden tongue blade or a cotton tip applicator. Collected cells are smeared on a frosted glass slide. Immediately fixed with alchohol ether spray After drying,slide is stained and processed and the histological features are examined.

BRUSH CYTOLOGY
The oral brush cytology was introduced to the dental profession in 1999 to overcome the limitations of traditional cytology. It was designed for the interrogation of clinical lesions that would otherwise not be subjected to biopsy because the level of suspicion for carcinoma, based upon clinical features, was low.

Limitations of traditional cytology

Deepest layers of the lesion not sampled Sensitivity is often dependant on tedious visual search on the microscope slide. Higher rate of epithelial turnover especially in the mouth, numerically dilutes the few abnormal cells seen in the smear. Number of abnormal cells available for sampling reduced by keratinized nature of many lesions.

PRINCIPLE
The brush biopsy utilises a brush to obtain a transepithelial biopsy specimen with cellular representation from each of the three layers of the lesion : Basal Intermediate Superficial

 Unlike cytology instruments, which collect only exfoliated superficial cells, the biopsy brush penetrates to the basement membrane, removing tissue from all three epithelial layers of the oral mucosa. The brush biopsy doesn t require topical or local anaesthetic. It causes minimal bleeding or pain.

METHOD
The brush biopsy instrument has two surfaces: 1. Flat surface 2. Circular border Either surface may be used to obtain the specimen

BRUSH BIOPSY INSTRUMENTS

ORALCDX BRUSH BIOPSY

The improved efficacy of oralCDx brush biopsy is due to the following factors: The entire thickness of the lesion is sampled. The analysis of the specimen is aided by highly specialised neural network-based image processing system. It can detect one to two abnormal cells scattered among tens of thousands of normal cells.

Bibliography
Anderson, J. B., Webb, A.J.: Fine-Needle Aspiration Biopsy and the Diagnosis of Thyroid Cancer. British Journal of Surgery 74:292-6, 1987 Sausville, Edward A. and Longo, Dan L.: Principles of Cancer Treatment: Surgery, Chemotherapy, and Biologic Therapy in Harrison's Principles of Internal Medicine, 16th Ed. Kaspar, Dennis L. et al., editors. p.446 (2005) ^ Friedman, S. and Blumberg, R.S.: Inflammatory Bowel Disease in Harrison's Principles of Internal Medicine, 16th Ed. Kaspar, Dennis L. et al., editors. pp. 1176-1789 (2005) Saibeni, S., Rondonotti, E., Iozzelli, A., Spina, L., Tontini, G.E., Cavallaro, F., Ciscato, C., de Franchis, R., Sardanelli, F., Vecchi, M.: Imaging of the Small Bowel in Crohn's Disease: A Review of Old and New Techniques World Journal of Gastroenterology 13(24): 3279-87, 2007

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