14 Cytologic Sub-Classification of Lung Cancer: A New Challenge for Practicing Pathologists

Parvin Ganjei-Azar MD Merce Jorda MD, PhD

2011 Annual Meeting – Las Vegas, NV

AMERICAN SOCIETY FOR CLINICAL PATHOLOGY 33 W. Monroe, Ste. 1600

Chicago, IL 60603

14 Cytologic Sub-Classification of Lung Cancer: A New Challenge for Practicing Pathologists This session will present aspects of cytologic sub-classification of lung cancer, focusing on the introduction of target therapy and its associated significant therapeutic implications for patients with advanced stage lung cancer. Included in the presentation will be subclassification of non-small cell carcinomas into adenocarcinomas and squamous cell carcinomas, cytologic samples including fine needle aspiration (FNA), bronchoscopic brushing (BB) and washings (BW), and bronchoalveolar lavages (BAL) to establish the diagnosis of lung cancer and subclassification of these tumors.Molecular testing for lung cancer will also be discussed. Formulate a differential diagnosis based on routinely prepared cytologic slides, based on clinical, imaging and cytologic findings.

• Learn how to identify and mark diagnostic cells properly for further application of immunocytochemistry, select a limited panel of antibodies based on available clinical and cytomorphologic imformation and to interpret immunostain results, and become fam

• Differentiate lung primary carcinomas from those of metastatic origin, accurately diagnose small cell carcinomas of lung and sub classify non small cell carcinomas into those with squamous differentiation and distinguish between malignant mesothelioma and

FACULTY: Parvin Ganjei-Azar MD Merce Jorda MD, PhD Practicing Pathologists Cytopathology Cytopathology (Non-Gynecologic) 2.0 CME/CMLE Credits Accreditation Statement: The American Society for Clinical Pathology (ASCP) is accredited by the Accreditation Council for Continuing Medical Education to provide continuing medical education (CME) for physicians. This activity has been planned and implemented in accordance with the Essential Areas and Policies of the Accreditation Council for Continuing Medical Education (ACCME). Credit Designation: The ASCP designates this enduring material for a maximum of 2 AMA PRA Category 1 Credits™. Physicians should only claim credit commensurate with the extent of their participation in the activity. ASCP continuing education activities are accepted by California, Florida, and many other states for relicensure of clinical laboratory personnel. ASCP designates these activities for the indicated number of Continuing Medical Laboratory Education (CMLE) credit hours. ASCP CMLE credit hours are acceptable to meet the continuing education requirements for the ASCP Board of Registry Certification Maintenance Program. All ASCP CMLE programs are conducted at intermediate to advanced levels of learning. Continuing medical education (CME) activities offered by ASCP are acceptable for the American Board of Pathology’s Maintenance of Certification Program.

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Parvin Ganjei-Azar MD.Merce Jorda MD, PhD

Cytologic Sub-Classification of Lung CancerNew Challenge for Practicing Pathologists

Merce Jorda MD, PhD

I do not have, and have not had, any relevant financial relationship with

any commercial interests within the past 12

Disclosure

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any commercial interests within the past 12 months, as pertaining to this presentation.

Parvin Ganjei-Azar MD.Merce Jorda MD, PhD

Lung cancer is the most frequent cause ofmajor cancer incidence and mortalityworldwide.

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More people die of lung cancer than of colon, breast, and prostate cancers combined.

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• Lung cancer is the second most common cancer in both men (after prostate cancer) and women (after breast cancer)

• In 2011….

221 130 l di d

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• 221,130 newly diagnosed cases

• 156,940 deaths from lung (27% of all cancer deaths)

Incidence

3

Combined data from the National Program of Cancer Registries as submitted to CDC and from the Surveillance, Epidemiology and End Results program as submitted to the National Cancer Institute in November 2009. *Rates are per 100,000 persons and are age-adjusted to the 2000 U.S. standard population

• Black men are about 40% more likely to develop lung cancer than white men

• The rate is about the same in black women and in white women

• The rate of lung cancer has been dropping

3

• The rate of lung cancer has been dropping among men for many years

• The rate of lung cancer is fairly stable among women.

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• Significant decline in incidence/mortality for African American and white males

• Increase in

Mortality

3

incidence/mortality for females of both races.

Source: North American Association of Central Cancer Registries (NAACCR), the National Cancer Institute (NCI), and the American Cancer Society. Journal of the National Cancer Institute, May 4, 2011.

Cytology of Lung Cancer Sampling Methods

• Sputum• Transthoracic (FNA)• Transbronchial (FNA BB BL)

3

• Transbronchial (FNA, BB, BL)• Transesophageal (FNA)• Body Cavity Fluids

Cytology of Lung Cancer: Goals

• Preventive • Diagnostic

Prognostic

3

• Prognostic• Predictive

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Spiral (helical) C T scans of the lungsChest X-rays

Sputum cytology

Preventive

3

•They can find cancers earlier •There is little evidence that they prevent death from lung cancer

High-risk patients

The International Early Lung Cancer Action Program (I-ELCAP):–48 institutions in 9 countries

H li l C T ith f ll d bi t l

3

–Helical C T scan with followed biopsy or cytology–Curability of stage I lung cancers is 80-90%–Costs of CT screening for lung cancer compare favorably with breast, cervical, and colon cancer screenings

The U.S. Preventive Services Task Force concludes that evidence is insufficient to recommend for or against screening asymptomatic persons for lung cancer with either low dose computerized tomography,

3

chest x-ray, sputum cytology, or a combination of these tests.

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• Preventive• Diagnostic

Pulmonary Cytology: Goals

3

• Prognostic• Predictive

Diagnostic Goals

• Malignant neoplasms• Benign neoplasms

I f ti /I fl t

3

• Infectious/Inflammatory processes

Malignant Neoplasms of Lung

• Primary• Secondary

3

• Secondary

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Primary L C iLung Carcinoma

WHO 1967, 1981, 1999Based on histomorphology

Primary Lung Neoplasms: Classification

3

WHO 2004Based on histomorphology, genetics and

clinical information

International Multidisciplinary ClassificationIASLC/ATS/ERS, 2011

Identification of prognostic and predictive factors

Primary Lung Neoplasms: Classification

3

• International Association for the study of lung cancer (IASLC)

• American Thoracic Society (ATS)• European Respiratory Society (ERS)

J Thorac Oncol. 2011; 6: 244-285

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International Multidisciplinary Classification, 2011

• Applies to – Resection specimens– Biopsies– Cytology samples

• Bronchioloalveolar carcinoma and mixed

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J Thorac Oncol. 2011; 6: 244-285

• Bronchioloalveolar carcinoma and mixed subtypes adenocarcinoma are NOT used

• Provides guidance for diagnosis in cytology samples (70% of lung cancers are diagnosed by cytology)

Primary Lung CarcinomagCytomorphology

Type Frequency 5-year survival

NSCLCAdenocarcinoma Squamous Cell

80 % 17%15%

3

Small Cell 20 % 5%

Large Cell5%

11%

88

Adenocarcinoma:Mixed TypeAcinarPapillarySolid

Bronchioloalveolar (non-mucinous)

Bronchioloalveolar (mucinous)

Adenocarcinoma with lepidic pattern

Mucinous adenocarcinoma

Adenocarcinoma

Adenocarcinoma(describe patterns present: Acinar, Papillary, Solid,Micro-papillary)

WHO, 2004Small Biopsy/CytologyIASLC/ATS/ERS, 2011

3

Mucinous (colloid)

Fetal

Signet ring

Clear cell

No counterpart

Adenocarcinoma with fetal pattern

Adenocarcinoma with colloid pattern

Adenocarcinoma with signet ring cell features

Adenocarcinoma with clear cell features

NSCLC, favor AdenocarcinomaJ Thorac Oncol. 2011; 6: 244-285

AdenocarcinomaCytomorphology

• Cohesive groups• Acinar, papillary, micro‐papillary formations

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papillary formations• Cytoplasmic vacuoles• Mucinous background

Small Biopsy/Cytology IASLC/ATS/ERS, 2011

Gland FormingMicropapillaryLepidic growthM i diff

Cytology

YesYesYesY

YesYesNoY

BiopsyAdenocarcinoma

3

Mucinous diff.“Colloid”Signet-ringClear cell

YesYesYesYes

YesYesYes?

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WHO 2004Bronchoalveolar Carcinoma, non-mucinous

IASLC/ATS/ERS, 2011Adenocarcinoma with lepidic pattern

WHO 2004Bronchoalveolar Carcinoma, non-mucinous

IASLC/ATS/ERS, 2011Adenocarcinoma

Squamous cell carcinomaPapillaryClear cellSmall cell

Squamous cell carcinomaPapillaryClear cellSmall cell

Squamous Cell Carcinoma

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

WHO, 2004

3

Small cellBasaloid

No counterpart NSCLC, favor Squamous Cell Carcinoma

Small cellBasaloid

J Thorac Oncol. 2011; 6: 244-285

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Squamous Cell CarcinomaCytomorphology

• Cohesive groups• Isolated pleomorphic cells

• Hyperchromatic nuclei

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• Hyperchromatic nuclei• Eosinophilic cytoplasm

• Necrotic background

WHO 2004Squamous cell carcinoma

IASLC/ATS/ERS, 2011Squamous cell carcinoma

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WHO 2004No counterpart

IASLC/ATS/ERS, 2011NSCLC, favor Squamous Cell Carcinoma

Small cell carcinoma Small cell carcinoma

Small cell carcinoma

Small Biopsy/CytologyIASLC/ATS/ERS, 2011WHO, 2004

3

Small cell carcinoma Small cell carcinoma

J Thorac Oncol. 2011; 6: 244-285

Small cellCarcinoma 20%

Incidence

3

Non-small cellcarcinoma 80%

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• Isolated small cells / small groups• Crushing artifact

Small Cell CarcinomaSmall Cell CarcinomaCytomorphologyCytomorphology

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• Nuclear molding• Cellular fragmentation• Tumor necrosis

WHO 2004Small Cell Carcinoma

IASLC/ATS/ERS, 2011Small Cell Carcinoma

3

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

Large cell carcinoma Non-small cell carcinoma, NOS

Large cell neuroendocrinecarcinoma (LCNEC)

Non-small cell carcinoma, withneuroendocrine morphology (positiveNE markers) possibly LCNEC

Large Cell Carcinoma

WHO, 2004

3

NE markers) possibly LCNEC

Large cell carcinoma withneuroendocrinemorphology (LCNEM)

Non-small cell carcinoma, withneuroendocrine morphology(negative NE markers); LCNEC issuspected

J Thorac Oncol. 2011; 6: 244-285

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• No squamous or glandular differentiation• Isolated cells • Pleomorphic nuclei

Large Cell CarcinomaCytomorphology

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Pleomorphic nuclei• Macronucleoli• Binucleation

WHO 2004Large Cell Carcinoma

IASLC/ATS/ERS, 2011Non-small cell carcinoma, NOS

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

AdenosquamousCarcinoma

Non-small cell carcinomawith squamous cell andadenocarcinoma patterns

Adenosquamous Carcinoma

WHO, 2004

3

Non-small cell carcinoma,NOS with IHC favoring bothsquamous andadenocarcinoma patterns

No counterpart

J Thorac Oncol. 2011; 6: 244-285

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Small cell carcinomaIASLC/ATS/ERS, 2011

Sarcomatoid Carcinoma Poorly differentiated NSCLC

Sarcomatoid Carcinoma

WHO, 2004

3

with spindle and or giant cellmorphology (with adeno or squamouscomponent)

J Thorac Oncol. 2011; 6: 244-285

Primary Lung Carcinomag

Immunocytochemistry

• Sub-classification of primary lung cancer

••• Distinction between primary vsDistinction between primary vsDistinction between primary vs

Malignant Neoplasms in LungImmunocytochemistry

Applications

3

••• Distinction between primary vs. Distinction between primary vs. Distinction between primary vs. secondary carcinomassecondary carcinomassecondary carcinomas

••• Distinction between lung primary and Distinction between lung primary and Distinction between lung primary and malignant mesotheliomamalignant mesotheliomamalignant mesothelioma

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TTF-1 Positive

CK 7 Positive

AdenocarcinomaAdenocarcinoma Squamous CellSquamous Cell Small CellSmall Cell

p63 Positive

TTF-1 Negative

TTF-1 Positive

CK Positive (dot-like)

Immunocytochemistry in Primary Lung Immunocytochemistry in Primary Lung CarcinomasCarcinomas

3

p63 Negative

CDX-2(*) Negative (*)

CK 20 Negative

(*) positive in intestinal type

Synapto Positive

Chromo Positive/Negative

p63 Negative

Ganjei, Jorda, Krishan. Effusion Cytology: A Practical Guide to Cancer Diagnosis. Demos 2011

NSCLCPrognostic and Predicti ePrognostic and Predictive

Molecular Markers

• A prognostic biomarker is a biomolecule that is indicative of patient survival independent of the treatment received

• A predictive biomarker is a biomolecule

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• A predictive biomarker is a biomolecule that is indicative of therapeutic efficacy

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• EGFR mutation:– 15% of Caucasians– 30% of Asians

• EGFR high copy number

NSCLCPrognostic and Predictive Biomarkers

3

EGFR high copy number– 40% Caucasians (FISH,IHC)

• KRAS mutation: 25% • EML4-ALK translocation: 6%• BRAF mutation: 3%

•Patients with EGFR mutation respond to TKI (Erlotinib, Getifinib).

•Patients with KRAS or BRAF mutation do not

NSCLC: Target Therapy

3

respond to TKI, ALKI

•Patients with EML4-ALK fusion, respond to ALK inhibitor (Crizotinib)

Secondary L C iLung Carcinoma

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Secondary Lung Carcinomas

• Metastatic carcinomas are the most common malignant neoplasms found in lung

• Adenocarcinomas are the most common secondary neoplasmsSit f i i B t l t l d t i

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• Sites of origin : Breast, colorectal , endometrium, others

• Distinction between lung primary and secondary carcinomas can not be made , based on cytomorphology alone

Secondary Lung Carcinomas

• Final diagnosis should be based on:–Clinical

3

–Imaging–Cytomorphology–Immunocytochemistry

••• SubSubSub---classification of primary lung classification of primary lung classification of primary lung cancer cancer cancer

• Distinction between primary vs

Malignant Neoplasms in LungImmunocytochemistry

Applications

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• Distinction between primary vs. secondary carcinomas

••• Distinction between lung primary and Distinction between lung primary and Distinction between lung primary and malignant mesotheliomamalignant mesotheliomamalignant mesothelioma

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• TTF-1• Napsin• TGB• PSA• S100, HMB45• ER PR

Primary vs. SecondaryCommonly Used Markers

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• ER, PR• CK 7 and CK20• P63• CDX-2• HCA, RCC• Endocrine markers• Lymphoid markers

• Adenocarcinomas• Squamous cell carcinomas

Primary vs. Secondary

3

• Small cell carcinomas• Large cell carcinomas

Adenocarcinoma with lepidic pattern

Mucinous adenocarcinoma

Adenocarcinoma(describe patterns present: Acinar, Papillary, Solid,Micro-papillary)

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

Secondary Adenocarcinomas

3

Adenocarcinoma with fetal pattern

Adenocarcinoma with colloid pattern

Adenocarcinoma with signet ring cell features

Adenocarcinoma with clear cell features

NSCLC, favor Adenocarcinoma

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Adenocarcinoma

MaleMale Lung Upper GI Colorectal

TTF-1 + - -

CDX-2 - / + + / - +

CK 7 + + -

3

•Colorectal •Upper GI•Others

CK 20 - - +

Cancer Cytopathology 2002

TTF-1

Lung FNA; 76 M History of Colon Ca.

CK20Adenocarcinoma of Lung

CK7

Ganjei & NadjiSpringer 2007

2020

Lung FNA : Patient with history of colorectal carcinoma

3

CK20

CDX2

3

Adenocarcinoma

FemaleFemale Lung Breast Colorectal FGT

TTF-1 + - / + - -

ER - /+ +/- - + /-

CK 7 + + - +CK 20 + - + + /-

3

•Breast•Colorectal•Female Genital tract•Others

CK 20 + + + /CA 125 - - - +P16 - - - +

2121

PF of a woman with history of breast cancer

“Cellular evidence of adenocarcinoma. ICC for further subclassification to

follow”

ER(+) “This immunophenotype is consistent with metastasis from the patient’s known primary breast carcinoma”

ER1D5

Secondary Squamous Cell

C i

Squamous cell carcinomaPapillaryClear cellSmall cell

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

3

Carcinomas

NSCLC, favor Squamous Cell Carcinoma

Small cellBasaloid

Squamous cell Carcinoma

Male Male Lung Head & Neck

Esophagus Urothelial

TTF-1 - - - -

P63 + + + +

3

•Lung•Head & Neck•Esophagus•Urothelial•Others

P16 - - / + - - / +

2222

Squamous cell Carcinoma

Female Female Lung Head & Neck

Esophagus Uterine Cervix

Urothelial

TTF-1 - - - - -

P63 + + + + +

P16 / /

3

•Lung•Head & Neck•Esophagus•Uterine Cervix•Urothelial •Others

P16 - - /+ - + - / +

3

2323

Squamous Cell Carcinomap63p63

Secondary Small Cell Carcinomas

Small Biopsy/CytologyIASLC/ATS/ERS, 2011

Small cell carcinoma

3

Cell Carcinomas

Small Cell Carcinoma

MaleLung Skin (Merckel)

TTF-1 + -

CK 20 - +

Female

3

•Skin•Others

CK 20 - +

•Skin•Others

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Differential DiagnosisSmall Cell Carcinoma vs.

Other Small Cell Neoplasms

CK CD45 DES CD99 SYN TTF-1

Small Cell Ca + - - - + +

3

Small Cell Ca + - - - + +

Lymphoma - + - -/+ - -

Rhabdomyosarcoma - - + -/+ - -

PPNET - - - + +/- -

••• SubSubSub---classification of primary lung classification of primary lung classification of primary lung cancer cancer cancer

••• Distinction between primary vsDistinction between primary vsDistinction between primary vs

Malignant Neoplasms in LungImmunocytochemistry

Applications

3

••• Distinction between primary vs. Distinction between primary vs. Distinction between primary vs. secondary carcinomassecondary carcinomassecondary carcinomas

• Distinction between lung primary and malignant mesothelioma

Lung Primary vs.Malignant Mesothelioma

CEA TTF-1 Calretinin p63

L Ad i

3

Lung Adenocarcinoma + + - -

Malignant Mesothelioma - - + -

Squamous Cell Carcinoma +/Squamous Cell Carcinoma +/-- -- -- ++

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FNA: Pleural- Based Mass, 62 Male

3

• Calretinin is the most useful marker for mesothelial cells

• Expression is both nuclear & cytoplasmic

• Calretinin does diff i

3

not differentiate benign from malignant mesothelial cells

Ganjei & NadjiSpringer 2007

• Sarcomatoid mesotheliomas are negative for calretinin

• Lung spindle cell carcinomas & sarcomatoid mesotheliomas express

Remember….

3

CK but are negative for TTF-1• Lung spindle cell carcinomas may

express p63, whereas mesotheliomas are p63 negative

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• Formulate a differential diagnosis based on clinical, imaging and cytologic findings

• Differentiate lung primary carcinomas from those of metastatic origin, and sub-classify primary carcinomas

Intended Learning Outcomes

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primary carcinomas • Application of immunocytochemistry in

cytologic material • Applicability of molecular assays