Personalized Medicine: Lung Biopsy and Tumor
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1 Personalized Medicine: Lung Biopsy and Tumor Mutation Testing Elizabeth H. Moore, MD Personalized Medicine: Lung Biopsy and Tumor Mutation Testing Genomic testing has resulted in a paradigm shift in the diagnosis and treatment of lung cancer Elizabeth H. Moore, M.D. Demand for greater amounts of tumor tissue The needle aspiration lung biopsy is dead --Tumor board oncologist November 2013 Radiologists and oncologists both need to understand: The role of mutation testing in the treatment and prognosis of lung cancer How and when biopsy specimens can and should be procured for testing The tissue requirements of specific genomic testing vendors 49 year old female smoker with seizure Coaxial needle biopsy was performed (2 FNAs and 3 cores) Imaging workup reveals a lung mass, brain metastases t 288
2 Cytological evaluation: Adenocarcinoma Smears (Papanicolaou stain) Cell Block (H&E stain) Immunohistochemistry panel: Subtype of lung cancer Tissue of origin i for metastases t Thyroid transcription factor 1 (TTF-1) positive c/w lung adenocarcinoma Core specimens confirmed adenocarcinoma Cores are not significantly better than FNA for diagnosis of malignant lung lesions Small cell AdenoCa Squamous Clinical Stage IV NSCLC was diagnosed d and our patient t was treated with platinum based chemotherapy but progressed Large cell How do tumor cells differ from normal cells? Platinum based drugs work by cross-linking DNA preventing repair/replication in 2 nd half of cell cycle Dose is limited by high toxicity to normal tissues Modest survival benefits Modified from T. SANDAL, The Oncologist 2002;7:73-81 Hanahan D and Weinberg RA, Cell 2000; 100:
3 Mutations that cause tumor behaviors primarily act in the first half of the cell cycle Oncogenes Growth factors Tumor suppressor genes (inactivated) These mutations offer unique targets for therapy not found in normal cells, with low toxicity to normal tissues Modified from T. SANDAL, The Oncologist 2002;7:73-81 Tumor mutation ti basics: Wild-type : Normal, non-mutated gene Germline mutations (all cells in body) Somatic mutations (tumor only) Most tumors have multiple mutations, with one dominant driver mutation Carcinogen induced cancers have more mutations Tumors can be heterogeneous with different mutational clones Various types of mutations can be seen in NSCLC Oncogene: Oncogene: Oncogene: Tumor Suppressor Loss: EGFR MT KRAS MT ALK p53 (Epidermal Growth Factor Receptor) rearrangement Point mutation Single base pair mutation Single base pair mutation ALK fusion Deletions in chromosome 17p Adenocarcinomas Adenocarcinomas Histologic Subtypes AKT BRAF VEGFR ALK HER2 EPHA/B PDGFR FGFR INSR Druggable mutations AKT BRAF VEGFR ALK HER2 EPHA/B PDGFR FGFR INSR Other 11% EGFR KRAS MAPK PI3K Drugs in development EGFR KRAS MAPK PI3K Squamous 34% Adenoca 55% Squamous Cell Carcinomas Squamous Cell Carcinomas FGFR1 Amp FGFR1 Amp EGFRvIII EGFRvIII Unknown PI3KCA Unknown PI3KCA EGFR TK EGFR TK DDR2 DDR2 Li, Gandara et al: JCO 2013 Li, Gandara et al: JCO
4 EGFR mutations (Epidermal Growth Factor Receptor) Common in many cancers Most common actionable mutation in NSCLC Most common in females, non-smokers, Asians, but must test These cell surface receptors have an intracellular lar component and are activated by binding with ligands such as Epidermal Growth Factor (EGF) This stimulates their intrinsic intracellular tyrosine kinase activity resulting in autophosphorylation and downstream activation of growth signaling cascades Tyrosine kinase activating mutations cause EGFR to always be switched on and the tumor cell requires this constant t growth signaling to survive Potential therapeutic agents: Monoclonal antibodies (cetuximab, panitumumab) Tyrosine kinase inhibitors (TKI) (erlotinib, gefitinib, afatinib) Downstream kinase inhibitors The EGFR is a validated target for NSCLC therapy TK-activating mutations of EGFR occur in 10-15% 15% of NSCLC tumors In EGFR-mutated tumors, EGFR TKIs are highly effective with >70% response rate and prolonged survival In EGFR wild-type tumors, EGFR TKIs are far less effective Li et al. Targ Oncol (2009) 4:
5 Phase III IPASS Trial: Efficacy of TKIs vs. platinum based chemotherapy based on EGFR Mutation Status EGFR mutation positive EGFR mutation negative Response after 2 months of TKI Gefitinib (Tarceva) Mok et al. N Engl J Med 2009; 361: TKIs produce few serious side effects but rarely can cause severe interstitial lung disease TKI resistance develops in all patients Often from new or pre-existing T790M mutation Genomic retesting after treatment failure Irreversible TKIs are being developed Biopsy EGFR MT+ After gefitinib Relapse, rebiopsy ALK rearrangements: (Anaplastic Lymphoma Kinase) Another druggable druggable target t but only 4% of NSCLC Most common rearrangement seen in NSCLC is a fusion: EML4-ALK Adenocarcinomas, non-smokers or light smokers Will not respond to EGFR specific TKIs 60% respond to crizotinib, an ALK specific kinase inhibitor ALK Fusion Genes in Lung Cancer Crizotinib 1. Barreca A, et al. J Mol Endocrinol 2011;47:R11R23; 2. Bang Y-J. Ther Adv Med Oncol 2011;3:279291; 3. Wong DW, et al. Cancer 2009;115: ; 4. Rodig SJ, et al. Clin Cancer Res 2009;15: ; 5. Boland JM, et al. J Thorac Oncol 2013;doi: /JTO.0b013e318287c
6 PROFILE 1001 (Crizotinib first-in-human phase I study) Waterfall plots of best percent change in target lesions from baseline Variable Study Subjects (N=143) n(%) Complete response 3 (2.1) 56 year old female never smoker break-apart apart fluorescent in situ hybridization (FISH) EML4-ALK fusion+ Baseline 2 cycles Crizotinib Partial response 84 (58.7) Objective response (CR + PR) [95% CI] 87 (60.8) [ ] Median duration of response, weeks [95% CI] Median time to response, weeks [range] 49.1 [ ] 7.9 [ ] Camidge et al., The Lancet Oncology 13 (10): , 2012 Drug resistance develops 8 cycles Crizotinib 10 cycles Crizotinib KRAS mutation (Kirsten rat sarcoma viral oncogene) Present in 1/3 of human malignancies A loss of function mutation Leads to constant activation of downstream signaling pathways independent of EGFR KRAS mutations are associated with poor prognosis regardless of treatment Smokers, Caucasians Ongoing drug development of specific kinase inhibitors active downstream to KRAS Our patient s genomic tumor testing revealed: EGFR mutation negative Selumetinib: (targets MEK or MAPK kinases) ALK fusion negative KRAS mutation positive (smoker) No FDA approved drug yet Graphic published in: Gregory J. Riely; Jenifer Marks; William Pao; Proc Am Thorac Soc 2009, 6, The American Thoracic Society 293
7 Which patients may benefit from biopsy and genomic testing? No surgical specimen forthcoming Second primary vs. metastasis staging issue Re-testing after treatment failure to look for: New driver mutation Drug resistance mutation New dominant tumor clone All advanced adenocarcinomas should be tested, prioritizing EGFR and ALK (College of American Pathologists) DNA can be extracted from a variety of biopsy specimens Paraffin embedded cell block or cores Fresh frozen tissue Slides from FNA/cell block Macrodissection of larger samples or laser microdissection from slides to isolate pure tumor tissue Tissue requirements for genomic testing High percentage of cells nucleated At least 20% of cells are tumor tissue (otherwise false negatives) Larger amounts of tissue more likely to yield accurate results Amount of tissue required dependent on vendor s method of testing Instructions on vendor website Representative vendor instructions: 3-5 cores requested Methods for genomic testing vary widely among vendors Serial testing by a variety of methods for a limited panel of likely and/or actionable mutations (Lower cost) Next generation sequencing: Simultaneously extract all known oncogenes from mechanically fragmented tumor DNA using probes, PCR, and gene sequencing with computer analysis (Higher cost) Next generation sequencing (NGS) for 182 cancer related genomic alterations ti Tissue requirement: 1 cubic mm 294
8 Why test for hundreds of mutations when so few are actionable? In all tumor types, many carry unknown mutations Further goals of research and drug development Unexpected druggable mutations may be found Graphic from Garraway L A JCO 2013;31: Lung biopsy for mutation testing Start with FNA to confirm presence of viable tumor before taking core biopsies i Avoid areas of necrosis Biopsy different parts of a solid mass Larger amounts of tissue increase the chances of positive results Confine the throw of the needle to the mass; avoid including normal lung Safety concerns for core biopsy: Hemorrhage and air embolism Core biopsy of smaller lesions may hemorrhage h into adjacent lung parenchyma Larger lesions tamponade bleeding Avoid including adjacent normal lung in core specimen Examine thin section images of a contrast CT before planning biopsy approach and avoid large vessels in and adjacent to the tumor 295
9 A preponderance p of recent air embolism cases in the literature and the legal sphere involved core biopsies Summary Test all advanced/progressing adenocarcinomas for EGFR and ALK mutations Only small minority of patients with NSCLC will have a druggable target Biologics will have better response rates and lower toxicity than standard chemotherapy, but resistance will develop Vendor specific requirements for tissue testing vary; determine prior to biopsy 296
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