Lung Cancer

Key Points

Molecular Markers in Lung Cancer

The presence of molecular markers in non-small cell lung cancer (NSCLC) is associated with responsiveness to certain targeted therapies. The current markers hold the most importance for nonsquamous histology, particularly for adenocarcinoma subtypes.

• Epidermal Growth Factor Receptor (EGFR)

  Epidermal Growth Factor Receptor (EGFR)
  Pathophysiology	Receptor tyrosine kinase involved in cell proliferation Inhibition of tyrosine kinase activity diminishes tumor growth Two methods for blocking kinase activity Prevent ligand binding to the extracellular domain with a monoclonal antibody OR Inhibit activity with small molecules (tyrosine kinase inhibitors) that block the magnesium-ATP binding pocket of the intracellular TK domain
  Mutation	Certain EGFR mutations are associated with increased sensitivity to tyrosine kinase inhibitors (TKIs) (eg, gefitinib and erlotinib) Lack of EGFR mutations (wild type) is associated with decreased sensitivity to TKIs Mutation status of EGFR does not appear to predict response to treatment with EGFR monoclonal antibody (cetuximab) Mutation rate based on patient characteristics 10-15% of NSCLC have EGFR mutations, predominantly adenocarcinoma Highest rate in Asians, adenocarcinoma histology subtype, lifetime nonsmokers or light smokers (<10 packs per year), female sex T790M mutation associated with decreased sensitivity to TKIs – typical mutation that develops when treatment resistance occurs
  Recommended Testing Strategy ARUP available test: EGFR Mutation Detection by Pyrosequencing 2002440	Mutational analysis is preferred rather than gene amplification If EGFR testing is negative, testing for ALK may be helpful; if EGFR testing is done initially and is positive, testing for KRAS may be helpful

• KRAS

  KRAS
  Pathophysiology	RAS genes regulate signal conduction pathways that control cell growth Three distinct genes – HRAS, NRAS, and KRAS
  Mutation	Almost all RAS mutations in NSCLC are in KRAS gene Mutations in KRAS gene are associated with decreased sensitivity to EGFR TKIs and overall poor prognosis Mutation rate based on patient characteristics 25-30% of adenocarcinomas have mutations of KRAS Highest rate in mucinous subtype, current smokers
  Recommended Testing Strategy ARUP available test: KRAS Mutation Detection 0040248	Mutational analysis for KRAS establishes the reflex pattern of testing for EGFR mutations and ALK rearrangements If KRAS is negative, test for EGFR mutations; if EGFR is negative, test for ALK rearrangements Double mutations in EGFR and KRAS are associated with decreased sensitivity to TKIs despite the presence of EGFR mutation

• Anaplastic Lymphoma Kinase (ALK)

  Anaplastic Lymphoma Kinase (ALK)
  Pathophysiology	Fusion of ALK gene with echinoderm microtubule-associated protein-like-4 (EML-4) or other fusion partners mediates ligand-independent oligomerization of ALK resulting in ALK kinase activation Functions as potent oncogenic driver
  Mutation	Certain ALK rearrangements are associated with increased sensitivity to ALK-inhibitors (eg, crizotinib) but also have decreased sensitivity to EGFR TKIs Mutation status based on patient characteristics Relatively rare mutation in NSCLC (3-7% in selected patients) Highest rate in young patients, lifetime nonsmokers or light smokers (<10 packs per year), adenocarcinoma with signet rings and mucin histology subtype, male sex
  Recommended Testing Strategy ARUP available test: ALK (D5F3) with Interpretation by Immunohistochemistry 2007324	Immunohistochemistry detects protein expression and is a surrogate marker for ALK gene fusion; more cost effective and efficient compared to FISH and RT-PCR RT-PCR is used to detect the most common ALK rearrangements, but may not detect all ALK fusion partners FISH interpretation is challenging due to small inversions involving chromosome 2p (2p21 and 2p23) Presence of ALK rearrangements are generally mutually exclusive of EGFR and KRAS mutations

Diagnosis

Indications for Testing

• New pulmonary mass

Laboratory Testing

• Serum testing is not helpful in diagnosing lung cancer; however, baseline testing (eg, CBC with differential, liver function) may be performed as a general screen for metastases
• Other testing
  • Neuron specific enolase (NSE), serum – may have diagnostic value for small cell lung cancer (SCLC)  

Imaging Studies

• Chest x-ray, CT scan, MRI provide basis for initial testing

Histology

• Invasive testing to obtain tissue necessary for diagnosis
  • Bronchoscopic biopsy
  • Mediastinal node sampling
  • Fine needle aspiration using CT guidance
  • Open lung biopsy
  • Fine needle aspiration using endobronchial ultrasound (EBUS)
• Immunohistochemistry
  • Should be used adjunctly and interpreted in clinical context
  • ALK immunohistochemistry – more cost effective and efficient than ALK by RT-PCR, FISH
    • Uses ALK clone D5F3
      • D5F3 clone – more sensitive than ALK1 clone
  • Primary adenocarcinoma and bronchioalveolar carcinoma
    • Positive for CK 7, napsin A, and thyroid transcription factor-1 (TTF-1); negative for CK 20
    • Primary mucinous adenocarcinoma – positive for CK 7 and CK 20, negative for TTF-1
  • Squamous cell carcinoma (SCC)
    • Usually negative for CK 7, CK 20, TTF-1
    • Does not typically require immunohistochemical staining for diagnosis
  • SCLC and neuroendocrine carcinomas
    • Typically negative for CK 7, CK 20
    • Positive for neuroendocrine markers such as chromogranin A, CD56 (NCAM), synaptophysin, NSE
    • Differentiating poorly differentiated SCC from SCLC
      • p63 and CK 5,6 positive and TTF-1 negative supports SCC
  • Differentiating non-small cell lung cancer (NSCLC) adenocarcinoma from mesothelioma
    • Positive in NSCLC/negative in mesothelioma – CEA-P, b72.3, Ber-EP4, MOC-31, TTF-1
  • Differentiating NSCLC from SCLC
    • MicroRNA (mRNA) expression – may be used to distinguish NSCLC from SCLC
  • Differentiating metastatic adenocarcinomas
    • Pulmonary – CK 7+/CK 20-
    • Colorectal – CK 7-/CK 20+
  • Differentiating primary lung tumors from metastatic gastrointestinal tumors – CDX2 specific for gastrointestinal tumors

Solitary Pulmonary Nodule

• Usually found incidentally on chest x-ray (0.09-0.2% of all x-rays)
  • 35% are malignant
• Histologic diagnosis required in the following situations
  • Patient is ≥35 years
  • Nodule is >10 mm diameter
  • Growth of lesion
  • Lack of calcification
  • Adenopathy
  • Positive PET scan
  • No previous imaging to review in order to determine if nodule has changed in size

Prognosis

• Markers
  • EGFR, KRAS, EML4-ALK – refer to Key Points section
    • Establish eligibility for therapies such as tyrosine kinase inhibitors and ALK inhibitors
• Promising markers without sufficient evidence to suggest helpfulness in prognosis
  • Cytokeratin-19 fragment (CYFRA 21-1)
    • Most sensitive tumor marker for NSCLC (particularly squamous)
      • May also be elevated in urological, gastrointestinal, and gynecological tumors
    • Potential role as independent prognostic factor in both early and late stages of NSCLC; may serve same role in SCLC
    • Potential role for predicting response to therapy
  • NSE
    • High specificity for SCLC
    • May be useful in assessing prognosis in NSCLC and SCLC
    • Currently in clinical use, but prognostic value has not been validated in high-level study
  • p53 – tumor suppressor gene
    • Presence prognostic of short survival and potential benefit from adjuvant chemotherapy in patients with NSCLC
  • Pro GRP – SCLC
  • CA 125 (available as an immunohistochemical stain) – NSCLC
  • STK11 – adenocarcinoma
  • KRAS, NRAS, HRAS – adenocarcinoma
  • HER-2 – NSCLC
  • Tumor M2-PK (TU M2-PK) – all types
  • BCL-2 (available as an immunohistochemical stain) – SCLC
  • ERCC1 expression
    • High level – prognostic of better survival in NSCLC independent of therapy
    • High level – predictive of poor response to platinum-based chemotherapy; low level associated with sensitivity
  • RRM1 expression
    • High level – prognostic of better survival in NSCLC independent of therapy
    • High level – predictive of poor response to gemcitabine-based chemotherapy

Differential Diagnosis

• Solitary pulmonary nodule
  • Granuloma
    • M. tuberculosis
    • Yeasts – Coccidiomycosis, Histoplasma, Cryptococcus
    • Molds – Aspergillus
  • Hamartoma
  • Round pneumonia
  • Bronchogenic cyst
  • Pulmonary infarction
  • Focal hemorrhage
  • Arteriovenous malformation
  • Other malignancy
    • Carcinoid tumor
    • Lung metastases
• Mass
  • Pneumonia
  • Metastatic tumor
  • Fibrotic lung disease
  • Mesothelioma
  • Other lung tumor
    • Lymphoma
    • Carcinoid

Screening

Monitoring

Clinical Background

Lung cancer is the leading cause of cancer-related mortality in the U.S. and worldwide.

Epidemiology

• Incidence – 62.5/100,000
  • 160,000 deaths from lung cancer estimated in 2012 (NCCN, 2012)
• Age – peak incidence is 65-74 years; median is 71 years
• Sex – M>F, minimal
  • Female prevalence has increased; male prevalence has stabilized
• Ethnicity – highest incidence in African American males

Risk Factors

• Tobacco use – 85-90% of all lung cancers attributed to tobacco use
  • 13-fold increase in risk for primary user
  • Secondhand exposure – 20-30% increased risk for those who live/lived with smokers
• Radon/uranium exposure
• Asbestos exposure – cumulative risk; estimated to cause 3-4% of cases
  • Risk increased if patient also smokes
• Occupational exposure to carcinogens (eg, bis(chloromethyl)ether, polycyclic aromatic hydrocarbons, chromium, nickel, organic arsenic)
• Previous chest irradiation
• Genetic – positive family history combined with tobacco use increases the risk

Pathophysiology

• Any tumor arising from respiratory epithelium or pneumocytes
• Two main types
  • Non-small cell lung cancer (NSCLC) – ~85% of all lung cancers
    • Adenocarcinoma (non-squamous cell) – most common
      • Occurs in glandular tissue of lung lining
    • Squamous (epidermoid) cell carcinoma (SCC)
    • Large cell (large cell anaplastic)
  • Small cell lung cancer (SCLC) – ~15% of all lung cancers
    • Epithelial tumor of small cells
      • 95% arise in lung; may also arise from extrapulmonary sites (eg, nasopharynx, gastrointestinal tract, genitourinary tract)
    • Nearly all cases result from smoking
    • Two subtypes
      • Small cell carcinoma (oat cell)
      • Combined small cell carcinoma
• Other tumors (rare)
• Undifferentiated bronchial-gland tumors, sarcomas, neuroendocrine tumors

Clinical Presentation

• 20% of cases incidentally identified while asymptomatic by chest x-ray for other reasons
• Symptoms based on area of tumor growth
  • Central – cough, wheeze, hemoptysis, stridor, dyspnea, postobstructive pneumonia
  • Peripheral – pleural/chest wall pain, cough, dyspnea
  • Invasion and obstruction of adjacent structures
    • Tracheal obstruction – dyspnea, wheezing
    • Esophageal compression – dysphagia
    • Recurrent laryngeal nerve invasion – hoarseness
    • Phrenic nerve invasion – diaphragmatic paralysis
    • Sympathetic nerve invasion – Horner syndrome
      • Ptosis
      • Miosis
      • Enophthalmos
      • Unilateral loss of sweating
    • Invasion of lung apex – Pancoast tumor, superior vena caval syndrome
  • Distant metastases
    • Superior vena caval syndrome
    • Pericardial tamponade
    • Pleural effusions
    • Pathologic bone fractures
    • Adrenal insufficiency (rare) 
  • Paraneoplastic syndromes – common; may be first presenting symptoms of lung cancer
    • Endocrine syndromes
      • Ectopic parathyroid hormone
        • Usually squamous cell
        • Hypercalcemia, hypophosphatemia
      • Antidiuretic hormone (ADH)
        • Usually SCLC
        • Syndrome of inappropriate secretion of ADH
        • Hyponatremia
      • Adrenocorticotropic hormone (ACTH)
        • Usually SCLC
        • Usually not Cushingoid in appearance
        • Hypokalemia
    • Skeletal/connective tissue syndromes
      • Clubbing
        • 30% incidence
        • Usually NSCLC
      • Hypertrophic pulmonary osteoarthropathy
        • Usually adenocarcinoma
    • Neurologic/myopathic syndromes
      • Eaton-Lambert syndrome
        • Usually SCLC
        • Myasthenia gravis symptoms
      • Retinal blindness
        • Usually SCLC
      • Peripheral neuropathy
      • Subacute cerebellar degeneration
      • Cortical degeneration
      • Polymyositis
    • Hematologic syndromes
      • Migratory thrombophlebitis – Trousseau sign
      • Nonbacterial endocarditis – marantic endocarditis
      • Disseminated intravascular coagulation
    • Dermatologic syndromes
      • Uncommon
      • Dermatomyositis
      • Acanthosis nigricans
    • Systemic syndromes
      • Unknown etiology
      • Cachexia, anorexia, fever, weight loss, suppressed immunity

Indications for Laboratory Testing

• Tests generally appear in the order most useful for common clinical situations
• Click on number for test-specific information in the ARUP Laboratory Test Directory

Click the plus sign to expand the table of additional tests.

Test Name and Number	Comments
EGFR Gene Amplification by FISH 0049234 Method: Fluorescence in situ Hybridization	Preferred test is EGFR mutation analysis
ALK Gene Rearrangements in NSCLC for Crizotinib Eligibility by FISH 2006102 Method: Fluorescence in situ Hybridization	Preferred test is ALK (D5F3) by IHC; interpretation is challenging in FISH for rearrangements due to small inversions involving chromosome 2p
Lung Cancer Assessment and Early Detection (EarlyCDT) 2007513 Method: Semi-Quantitative Enzyme-Linked Immunosorbent Assay	May be useful as screening for patients deemed high risk for lung cancer
Solid Tumor Mutation Panel by Next Generation Sequencing 2007991 Method: Massively Parallel Sequencing	Prognosis/treatment of individuals with solid tumor cancers at initial diagnosis or with refractory disease

Diagnostic Algorithm

  Lung Cancer Molecular Testing Algorithm