Dr Stephen CrowtherTallaght Hospital

Lung CytopathologyIrish Society of Surgical Pathology

First Draft ISSP Saturday morningSATURDAY: Lung Neoplasms9.00 - 9.40: My Holidays Stephen Crowther (sponsored by British

Airways)9.40 - 10.20: Lung Tumours : Louise Burke (TBC)10.20 - 11.00: Mesenchymal Lung tumours: Leona Doyle

ContentsSpecimens, preparation and adequacyProposed classification systemCommon malignancies Pitfalls

What is the role of cytologyLung cancer presents at advanced stageIncreased number of nodules sampled (screening)

DiagnosisStagingMolecular testing

SpecimensSputumBronchoscopic derived specimens

Bronchial brushingsBronchial washings Bronchoalveolar lavage (BAL)Endobronchial ultrasound guided aspiration (lung, lymph nodes)

Percutaneous trans-thoracic CT-guided FNAPleural Fluid

PreparationsDirect smears

Air dried (MGG)Alcohol fixed (Pap, H&E)

Touch imprints – core biopsyConcentration methods

Cytospin (Hanks)Liquid-based preparations (Cytolyte)

Cell block

Sputum

Non invasiveEarly morning, induced, post BronchoscopicPreparation

Pick and smearAdequacy

2 smears with easily identifiable pulmonary macrophages

Bronchial Brush & WashingsCentral tumour exfoliating into the bronchiSquamous cell carcinoma and small cell most sensitiveWashings

5mls of salt solution instilled and then re-aspirated

BrushRolled on a glass slideBefore biopsy

AdequacyPulmonary macrophages, squamous cells, bronchial epithelial cellsAdequate if positive!

Bronchoalveolar LavageUseful in benign disease

Cell differentialCD4:CD8 ratio

Assessment of terminal air spaces50-100 mls of warm salineFlood distal airspace and re-aspiratedAdequacy

Alveolar macrophages (>10 per 10 HPF)<5% BE or Squamous cells

Lung FNAMost effective cytological technique to diagnose pulmonary carcinomaClose radiological correlation requiredOnsite rapid assessment - triage materialDirect smears (AD, IMS)Needle rinse

Hanks/PEG - ICC RPMI - flowCell block – IHC, molecular

Adequate if malignant!

EBUS Lymph node FNADiagnose and Stage (N stage)Preparation

Rapid onsite assessment and smearsCytolyte, Thinprep and cell block

Adequacy Minimum number of lymphocytes (>40 at x40)Pigment laiden macrophagesGerminal centre fragments

Cellblock for molecular

Pleural FluidDiagnose & Stage (M stage)No adequacy criteriaCytospins (2 PAP, 2 MGG)

Peg for immunocytochemistryCell block for molecular

Standardized Terminology and Nomenclature for Respiratory Cytology

I Non diagnosticII Negative (for malignancy)III AtypicalIV Neoplasm, benign neoplasm, and low-grade malignancyV Suspicious for malignancyVI Malignant

The Papanicolaou Society of Cytopathology GuidelinesDiagnostic Cytopathology

Risk of Malignancy

I. Non diagnosticAdequacy

Defined for sputum and LN EBUS-FNANot for brush, washing and Lung FNA (adequate if malignant!)Doesn't mean lesion has been sampled

Poor SamplingMultidisciplinary approachOn-site evaluation

Technical issuesSmearing, fixation, staining

II. Negative for malignancy

AdequateAbsence of cellular atypiaDiagnose a lesion defined radiologically

Infection (abscess, TB, fungi, parasite, viral)InfarctionSarcoidAspirationAmyloidLipoid

Normal tissue – radiology vague (no definite lesion)

PASF

III. Atypical

Limited use of this category – not a wastebasket!Two types

Abnormalities greater than inflam/repair but less than suspicious for malignancyHighly suspicious for a benign neoplasm

Eg bland spindled cells suggestive of SFT

IV. Neoplasm, benign neoplasm and low-grade malignancy

Allows discretion in treatment of elderly/medically unfitAdequately cellular and diagnostic

Solitary fibrous tumourPulmonary hamartomaSquamous papillomaGranular cell tumourPEComa (clear cell sugar tumour)MeningiomaInflammatory myofibroblastic tumourLangerhan’s cell histiocytosis

Does not include neuroendocrine tumours (carcinoids)

V. Suspicious for malignancyNot diagnostic of malignancy – inform your clinicans!Carcinomas (NSCLC, SCLC, NEC), Carcinoids, Lymphoma, MetastasesInter-observer variability & experience

Scant cellularityPeripheral samplingCavitationContamination of adjacent materialHigh level of differentiation in adca

VI. MalignantNon small cell carcinoma – need to further classify

AdenocarcinomaSquamous cell carcinomaLarge cell carcinoma

Small cell carcinomaNeuroendocrine (TC, AC, LCNEC)Adenoid cystic carcinomaMucoepidermoidLymphomaMetastases

Squamous cell carcinomaSingle cell and flat sheets with well defined cell membranesCytoplasm: polygonal, oval, spindled and irregular cell contourswith dense or keratinised cytoplasmNuclei: oval, rectangular, irregular contours, centrally situated, coarse to pyknotic like dark chromatinNucleoli often inconspicuous

Washings

FNA

FNA

PitfallsUpper RTI with reactive changes (sputum, washings)Reactive changes in cavities (aspergillus), bronchiectasis, infection,radiationContamination – EBUS LNNecrosis can mimic keratinisationIn situ v invasive

AdenocarcinomaMostly flat to three-dimensional aggregates and variable number of individual tumour cellsAcinar and glandular structures in aggregatesCytoplasm: abundant, delicate, granular to vacuolated, mucinNuclei: eccentric round to oval structures with minor membrane irregularities and fine chromatinProminent nucleoli

Washings

FNA

FNA

PitfallsReactive bronchial epithelium (washings)Pseudoglandular squamous cellMetastasesLepidic

Adenocarcinoma with lepidic patternAdenocarcinoma-in-situMinimally invasive adenocarcinomaInvasive adenocarcinoma with lepidic pattern

Can you go any further on cytology??Adequate classification can require resection

ImmunocytochemistryTissue preservation importantCocktail of two markers

TTF1, ck5/6P63(p40)Nap

Special stainsMucin for adenocarcinoma

International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society International Multidisciplinary Classification of Lung Adenocarcinoma

Morphologic adenocarcinoma patterns clearly presentAdenocarcinoma

Morphologic adenocarcinoma patterns not present (supported by special stains)

Non-small cell carcinoma, favour adenocarcinomaMorphologic squamous cell patterns clearly present

Squamous cell carcinomaMorphologic squamous cell patterns not present (supported by stains)

Non-small cell carcinoma, favour squamous cell carcinoma

? Large cell carcinomaGroups of large clearly malignant cellsPleomorphic single cell populationHigh N/C ratioNecrotic backgroundImmunos equivocal/negativeHistological (resection) diagnosisCyto dx – Non-small cell carcinoma, NOSLarge cell neuroendocrine carcinoma – NE markers

Non small cell carcinoma NOS

Small cell carcinomaSingle and loose clusters of cellsScant cytoplasmGranular/clumped chromatin Indistinct nucleoliNuclear mouldingNecrosis and Azzopardi affect

Washings

Washings

Washings

FNA

FNA

FNA

PitfallsDegenerate bronchial cells (washings)Lymphocytes (washings)Poorly differentiated squamous cell carcinoma Lymphoma – CD56

CD56

CD3AE1/3

CarcinoidUniform small cells, rounded nuclei, stippled chromatinPalisades, trabeculaeBare nucleiNecrosis rareBrushings and FNARarely exfoliates into washings

Atypical carcinoidNeuroendocrine cytologyIncreased pleomorphism, mitoses, necrosis (v TC)No moulding, smearing, no abundant mitoses & necrosis (v SCC)Cell block – MIB 1 helps v SCC

FNA

Adenoid cysticPrimary or metastaticLarge acellular basement membrane spheres (MGG)Background of small cells with uniform nuclei

NA

FNA

Metastatic urothelial carcinoma

FNA

Pleural fluid

Pleural fluid

CD56 BerEp4

Pleural fluid – cell block CD56

TTF1

Concluding remarksProposed new classificationTerminology

Non small cell NOS!!Consider cellblockThink molecular