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Amanjit Bal

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    ES09 - How I Do It - Real World Issues in the Diagnosis and Treatment of Metastatic NSCLC (ID 12)

    • Event: WCLC 2019
    • Type: Educational Session
    • Track: Treatment in the Real World - Support, Survivorship, Systems Research
    • Presentations: 1
    • Now Available
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      ES09.01 - How I Optimize Tissue Specimen Processing for Histopathological and Molecular Profiling (Now Available) (ID 3199)

      13:30 - 15:00  |  Presenting Author(s): Amanjit Bal

      • Abstract
      • Presentation
      • Slides


      Lung cancer is the most common cancer in males worldwide, and the most common cause of cancer related mortality in both sexes. There have been significant advances in the treatment of non-small cell lung cancer (NSCLC) in the last several years, which require careful biopsy sample acquisition and processing. Approximately 70% lung cancer patients present in advanced stage, thus most diagnosis of lung cancer are offered on cytology specimens and small biopsies.

      Specimens for diagnosing lung cancer

      For diagnosis of lung cancer, the specimens received in pathology laboratory includes; Effusions and FNAC samples from which cell blocks can be prepared, endobronchial biopsies, transbronchial lung biopsies, CT/PET guided core biopsies and biopsies from metastatic sites like lymph nodes, bone etc. Since tissue is an issue in diagnosis and management of lung cancer, thus strategies to manage limited tissues have been developed:

      The pre-analytical steps involved in handling, fixing and processing these precious tissue samples are critically controlled to obtain accurate and meaningful biomarker tests.

      For molecular testing two block setting is used: One block for diagnostic immunostains and second block is reserved for molecular workup.

      Pathological analysis of NSCLC on small biopsies:

      Histological Sub-typing of NSCLC:The subtyping of NSCLC on small biopsies has poor inter-observer agreement. WHO 2015 classification recommends use of ancillary techniques like mucin histochemsitry and immunohistochemistry for cases that cannot be classified based on light microscopy and minimize the use of term NSCLC-NOS.1To refine separation of squamous from adenocarcinoma; use of one adenocarcinoma marker and one squamous marker is suggested to preserve tissue for further molecular testing.For adenocarcinoma, TTF-1, Keratin 7 and napsin and for squamous, P40, p63 and keratin5/6 are used. Cocktails of nuclear and cytoplasmic antibodies like, TTF1/CK5/6 and napsin/p63 are also available. For neuroendocrine tumours, the specific markers include; neuron-specific enolase, chromogranin, synaptophysin, and CD56. With the use of immunohistochemistry the NSCLC-NOS category has been reduced from 10% to 5% in our setup.2

      Morphological Patterns of Adenocarcinoma:The histologic patterns of adenocarcinoma described in WHO 2015 classification include; lepidic predominant, acinar predominant, papillary predominant, solid predominant and micropapillary predominant pattern. These histological patterns though described for resection specimens, should be reported on small biopsies as they provide prognostic information; Lepidic pattern is associated with favorable prognosis, acinar and papillary with intermediate prognosis whereas the solid and micropapillary patterns have poor prognosis.3

      Genetics for personalized medicine in lung cancer

      Epidermal Growth Factor Receptor gene (EGFR) mutation:Most common mutations in young Asian females and/or never smokers and involves exons 18-21 of EGFR gene, the encoding a portion of the tyrosine kinase domain. Ninety percent of EGFR mutations are exon 19 deletions or missense point mutation in L858P in exon 21. Additional mutations in EGFR (T790M in exon 20) as well as mutations in other genes such as MET have been responsible for resistance to EGFR inhibitors.

      Anaplastic Lymphoma Kinase (ALK) rearrangement:EML4-ALK translocation of the transcription activation domain of ALK and the dimerization domain of EML4, leads to interstitial inversion in the short arm of chromosome 2. EML4-ALK translocation is seen in 5-8% of adenocarcinomas and is detected by break apart FISH or by IHC (D5F3 clone on Ventana system).

      Overall incidence of EGFR mutations in our set up is 22.3% and for ALK re-arrangement is 9.5%.3,4

      ROS1 re-arrangement: Similar to ALK mutations, ROS1 re-arranged tumours respond to tyrosine kinase inhibitor therapyandaccounts for 1-2% of pulmonary adenocarcinomas. ROS1 expression is screened by immunohistochemistry (D4D6 clone from Cell signaling) and break-apart FISH confirms the positive cases.

      Rare genetic changes:Other rare mutations reported in lung cancer includes; HER2 mutations (1%); BRAF mutations (2%); RET and NTRK rearrangements reported in 1.9% and 0.9%, respectively


      In addition to targetable mutations, immune checkpoint inhibitors have revolutionized the treatment of lung cancer. The programmed death 1 (PD-1) and programmed death ligand 1 (PD-L1) have monoclonal antibodies directed towards them, which have shown promise with regards to overall survival in advanced NSCLC. PD-L1 expression status as assessed by immunohistochemistry is important in predicting response to PD-1/PD-L1 inhibitors. However, there are challenges in PD-L1 testing that includes; four different IHC clones on different staining platforms, due to limited tissue all assays cannot be performed, heterogeneity of staining and the need for standardization of interpretation and scoring criteria.

      Recent advances

      Since tissue is an issue in molecular testing of lung cancer, targeted next generation sequencing panels are available for testing all relevant molecular changes in one go. Oncomine Dx is one such panel for lung cancer that has got FDA approval. In absence of available tissue especially in setting of relapse, cell free DNA is an alternative for detecting molecular alterations.


      The exact histological subtyping of NSCLC, thus reducing the ‘NOS’ rate, analysis of genetic alterations for targeted therapy and evaluation of targets for immune checkpoint inhibitors has significantly impacted the treatment and prognosis of lung cancer patients. Thus procurement of adequate tissue and its judicious use is of utmost importance. As the number of clinically significant targetable mutations and chromosomal rearrangements continues to grow, the next generation sequencing becomes the need of the day.


      Travis WD, Brambilla E, Nicholson AG et al. The 2015 World Health Organization classification of lung tumors: Impact of genetic, clinical and radiologic advances since the 2004 classification.J Thorac Oncol 2015;10(9):1243 60.

      Kaur H, Sehgal IS, Bal A et al. Evolving epidemiology of lung cancer in India: Reducing non-small cell lung cancer-not otherwise specified and quantifying tobacco smoke exposure are the key. Indian J Cancer 2017;54:285-90.

      Maturu VN, Singh N, Bal A, et al. Relationship of epidermal growth factor receptor activating mutations with histologic subtyping according to International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society 2011 adenocarcinoma classification and their impact on overall survival.Lung India 2016;33:257-66.

      Bal A, Singh N, Agarwal P et al. ALK gene rearranged lung adenocarcinomas: molecular genetics and morphology in cohort of patients from North India.APMIS. 2016;124(10):832-8.

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