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E. Thunnissen
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MA12 - Miscellaneous Biology/Pathology (ID 476)
- Event: WCLC 2016
- Type: Mini Oral Session
- Track: Biology/Pathology
- Presentations: 1
- Moderators:B. Dome, W.D. Travis
- Coordinates: 12/06/2016, 14:20 - 15:50, Schubert 1
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MA12.07 - Discussant for MA12.04, MA12.05, MA12.06 (ID 7106)
15:02 - 15:14 | Author(s): E. Thunnissen
- Abstract
- Presentation
Abstract not provided
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P1.02 - Poster Session with Presenters Present (ID 454)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Biology/Pathology
- Presentations: 1
- Moderators:
- Coordinates: 12/05/2016, 14:30 - 15:45, Hall B (Poster Area)
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P1.02-025 - Evaluation of NGS and RT-PCR Methods for ALK Assessment in European NSCLC Patients: Results from the ETOP Lungscape Project (ID 5001)
14:30 - 14:30 | Author(s): E. Thunnissen
- Abstract
Background:
The reported prevalence of ALK rearrangement in NSCLC ranges from 2%-7%, depending on population and detection method. The primary standard diagnostic method is fluorescence in situ hybridization (FISH). Recently, immunohistochemistry (IHC) has also proven to be a reproducible and sensitive technique. Reverse transcriptase-polymerase chain reaction (RT-PCR) has been advocated and most recently the advent of targeted Next-Generation Sequencing (NGS) for ALK and other fusions has become possible. This is one of the first studies comparing all 4 techniques in resected NSCLC from the large ETOP Lungscape cohort.
Methods:
96 cases from the ETOP Lungscape iBiobank (N=2709) selected based on any degree of IHC staining (clone 5A4 antibody, Novocastra, UK) were examined by FISH (Abbott Molecular, Inc.; Blackhall, JCO 2014), central RT-PCR and NGS. H-score 120 is used as cutoff for IHC+. For both RT-PCR and NGS, RNA was extracted from the same formalin-fixed, paraffin-embedded tissues. For RT-PCR, primers were used covering the most frequent ALK translocations. For NGS, the Oncomine™ Solid Tumour Fusion Transcript Kit was used, allowing simultaneous sequencing of 70 ALK, RET and ROS1 specific fusion transcripts associated with NSCLC, as well as novel ALK translocations using 5’-3’ ALK gene expression ‘Imbalance Assay’.
Results:
NGS provided results for 90 cases, while RT-PCR for 77. Overall, 70 cases have results for all 4 methods, with fully concordant 60 (85.7%) cases (49 ALK-, 11 ALK+). Before employing the ‘Imbalance Assay', in 5 of the remaining 10 cases, NGS differs from the other methods (3 NGS-, 2 NGS+), while in the other 5, NGS agrees with RT-PCR in all, IHC in 2, and FISH in 1. Using the concordant result of at least two of the three methods as true negative/positive, the specificity and sensitivity of the fourth is 96/94/100/96% and 94/94/89/72% for IHC/FISH/RT-PCR/NGS, respectively (incorporating imbalance: NGS sensitivity=83%). Imbalance scores are presented here for 18 NGS- cases: 9 ‘NGS-/FISH+/IHC+’, 9 ‘NGS-/FISH-/IHC-‘. Among the ‘NGS-/FISH+/IHC+’, there is strong evidence of imbalance in 4 cases (score’s range: 0.0144-0.0555), uncertain in 5 (range: 0.0030-0.0087), and no evidence (scores≤0.0004) in the 9 negative cases.
Conclusion:
NGS is a useful screening tool for ALK rearrangement status, superior to RT-PCR when RNA yield is limited. When using NGS, it is critically important to integrate the 5’-3’ imbalance assay and to confirm with one or more additional methods in the ‘imbalance’ cases. Data further highlight the possibility of missing actionable rearrangements when only one screening methodology is available.
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P3.01 - Poster Session with Presenters Present (ID 469)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Biology/Pathology
- Presentations: 2
- Moderators:
- Coordinates: 12/07/2016, 14:30 - 15:45, Hall B (Poster Area)
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P3.01-009 - A Prospective Study of 'Spread through a Knife Surface' (STAKS) in Non-Small Cell Lung Cancer Resection Specimens (ID 4694)
14:30 - 14:30 | Author(s): E. Thunnissen
- Abstract
Background:
An extraneous tissue contaminant on a slide is called a floater. Spread Through Air Spaces (STAS) is in the WHO classification considered as a form of invasion in lung adenocarcinoma. The artifactual spread of tissue fragments during lung specimen sectioning was recently described and termed Spreading Through A Knife Surface (STAKS).1 The purpose of this study was to prospectively examine lung resection specimens for the presence and frequency of STAKS.
Methods:
A prospective, multi-institutional study of NSCLC lobectomy and pneumonectomy resection specimen was performed from January 1 –July 1, 2016. Prosection, sampling and scoring of displaced fragments was undertaken in a systematic manner. The first cut was made with a clean long knife, the second cut was made in a parallel plane to the first cut, without cleaning the knife. Four tissue blocks were sampled: Block 1: first cut, upper part; Block 2: first cut, lower part; Block 3: second cut, upper part; Block 4: second cut, lower part. From these formalin fixed and paraffin embedded tissue blocks a superficial complete H&E stained slide was examined for the presence of displaced tissue fragments at 10x or 20x. A displaced fragment was scored as STAKS if the tissue fragment was at least 0.5 mm from the tumor or if it was on the pleural surface in the plane of the second cut. Benign and malignant STAKS were separately noted.
Results:
A total of 41 resection specimen were included in this study. The mean number of malignant STAKS for blocks 1-4 was 0.36, 1.44, 1.86 and 1.95, respectively and for benign STAKS the mean number was 0.11, 0.11, 0.13 and 0.25, respectively. Almost all STAKS were intra-alveolar. Comparison of malignant STAKS in block 1 (before the tumor was reached) with blocks 2-4 (containing tumor) was significant with p-values (p=0.003 Friedman’s test and post-hoc comparisons p=0.031, p=0.002 and p=0.005, respectively). For benign STAKS no difference was identified (p=0.23). The chance of malignant STAKS seemed to be higher when tumor was cut fresh than when cut after formalin fixation.
Conclusion:
The morphologic definition of STAKS is not different from STAS. This prospective study confirms the presence of benign and malignant STAKS. The presence of malignant STAKS is an artifact and increases with each and every knife cut during tissue sectioning. 1) Thunnissen et al. ArchPatholLabMed2016,140(212-220)
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P3.01-021 - Reproducibility of Comprehensive Histologic Assessment and Refining Histologic Criteria in P Staging of Multiple Tumour Nodules (ID 5365)
14:30 - 14:30 | Author(s): E. Thunnissen
- Abstract
Background:
Multiple tumor nodules (MTNs) are being encountered, with increasing frequency with the 8[th] TNM staging system recommending classification as separate primary lung cancers (SPLC) or intrapulmonary metastases (IM). Pathological staging requires assessment of morphological features, with criteria of Martini and Melamed supplanted by comprehensive histologic assessment of tumour type, predominant pattern, other histologic patterns and cytologic features. With publication of the 2015 WHO classification of lung tumours, we assessed the reproducibility of comprehensive histologic assessment and also sought to identify the most useful histological features.
Methods:
We conducted an online survey in which pathologists reviewed a sequential cohort of resected multifocal tumours to determine whether they were SPLC, IM, or a combination. Specific histological features for each nodule were entered into the database by the observing pathologist (tumour type, predominant adenocarcinoma pattern, and histological features including presence of lepidic growth, intra-alveolar cell clusters, cell size, mitotic rate, nuclear pleomorphism, nucleolar size and pleomorphism, nuclear inclusions, necrosis pattern, vascular invasion, mucin content, keratinization, clear cell change, cytoplasmic granules¸ lymphocytosis, macrophage response, acute inflammation and emperipolesis). Results were statistically analyzed for concordance with submitting diagnosis (gold standard) and among pathologists. Consistency of each feature was correlated with final determination of SPLC vs. IM status (p staging) by chi square analysis and Fisher exact test.
Results:
Seventeen pathologists evaluated 126 tumors from 48 patients. Kappa score on overall assessment of primary v. metastatic status was 0.60. There was good agreement as measured by Cohen’s Kappa (0.64, p<0.0001) between WHO histological patterns in individual cases with SPLC or IM status but proportions for histology and SPT or IM status were not identical (McNemar's test, p<0.0001) and additional histological features were assessed. There was marked variation in p values among the specific histological features. The strongest correlations (<0.05) between p staging status and histological features were with nuclear pleomorphism, cell size, acinus formation, nucleolar size, mitotic rate, nuclear inclusions, intra-alveolar clusters and necrosis pattern. Correlation between lymphocytosis, mucin content, lepidic growth, vascular invasion, macrophage response, clear cell change, acute inflammation keratinization and emperipolesis did not reach a p value of 0.05.
Conclusion:
Comprehensive histologic assessment shows good reproducibility between practicing lung pathologists. In addition to main tumour type and predominant patterns, nuclear pleomorphism, cell size, acinus formation, nucleolar size, and mitotic rate appear to be useful in distinguishing between SPLC and IM.
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SC02 - Multifocal Lung Cancer (ID 326)
- Event: WCLC 2016
- Type: Science Session
- Track: Radiology/Staging/Screening
- Presentations: 1
- Moderators:G. Veronesi, S. Cicenas
- Coordinates: 12/05/2016, 11:00 - 12:30, Strauss 1
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SC02.01 - Multiple Primary Lung Cancers Versus Lung Metastases: Pathological Differential Diagnosis (ID 6604)
11:00 - 11:20 | Author(s): E. Thunnissen
- Abstract
- Presentation
Abstract:
Introduction The pathological differential diagnosis of in a patient with synchronous multiple tumors has a similar thought process for classic morphology as for DNA analysis. Metachronous multiple lung cancers are not discussed here, as the provided title implies a clinical situation at moment in time. In the end of the text the usefulness of clinical context is mentioned.[1–5] Morphology In the setting of obvious metastatic dissemination, histologic appearance is generally conserved. This implies that most metastases look alike and in most instances also have a similar morphologic appearance as the primary tumor. However, dedifferentiation occasionally occurs in metastases. Thus it is reasonable to conclude that lesions of different histological types, e.g. squamous cell and adenocarcinoma, are two separate primary cancers. Recently, it was suggested that differences in a comprehensive histologic assessment may provide an argument for separate primary cancers. However, resection specimen analysis is required for this assessment, making this approach only useful in about 15-20% of the cases. Moreover, an evidence based data set supporting comprehensive histologic assessment is currently lacking. Reproducibility has to be taken into account.[6] In contrast to different morphological types, in case of two tumors with similar morphology a conclusion for same lineage (primary tumor- metastases relation) or different lineage (two primary tumors) is not easy to reach. The reason for this is that within an individual the genetic predisposition and etiologic factors are the same and may lead to separate tumors with the same morphology, which may still represent two lineages (thus two primary tumors) or alternatively, the same morphology in metastatic setting is one lineage. In case of multiple adenocarcinoma in-situ lesions, they are assumed to be separate primaries. In summary , in comparing two tumors differences in morphologic types is conclusive for second primary tumors, but demonstration of the same morphology is not sufficient for lineage analysis. DNA changes Demonstration of specific driver mutations by widely available PCR sequencing techniques may have use in establishing lineage.[7]In case of different driver mutations between two tumors, this obviously provides a strong argument for two primary (lung) cancers. However the frequency of discordant driver mutations is not so high. Noteworthy is that the demonstration of different passenger mutations does not have any use for lineage determination. In case of equal driver mutations a conclusion about lineage is not easy to reach.[8,9]Not only because the genetic predisposition and etiologic factors are the same, but also because in certain genes hotspot mutations occur. Thus simple demonstration of the same mutation does not provide definite evidence for lineage analysis. On this matter more research is needed with posterior probability analysis involving a relevant number of similar mutations. A detailed genetic assessment such as in comparative genomic hybridisation (CGH) may have greater discriminative power but has been used in only a few small studies.[10]In array CGH the number of data points is orders of magnitude greater than in mutation analysis. Array CGH encompasses the predisposition and etiologic factor related copy number variations (CNV) as well as lineage specific CNV. In this sense comparison of exact breakpoints in gene rearrangements is useful. Although the data are limited as the assessment was in the past complex, nowadays arrayCGH in the form of shallow sequencing can reliable be performed on small biopsies as well. To which extend NGS with a large mutation panel may be useful remains to be seen. In summary , different driver mutations is conclusive for second primary tumors, but demonstration of the same driver mutation is not sufficient for lineage analysis. Clinical context Adding clinical context provides interesting arguments. 1) Imaging may show multiple ground glass opacities (mGGO) and lack of enlarged lymph nodes. Although the morphology may be similar (lepidic pattern with AIS, MIA, Invasive adenocarcinoma) the mGGO lesions are considered to be separate primaries. In case of part solid component the morphological equivalent is usually infarction (=benign) or invasive cancer. 2) Imaging may show multiple consolidations (pneumonic type) with mostly the morphological correlate of invasive mucinous adenocarcinoma. 3) Abundance of nodular lesions, provides an argument for metastases (although rare exceptions exist, e.g. DIPNECH, Carney’s triad), while lack of nodal or systemic metastases provides an argument of two primary lung cancers. 4) Clinical follow-up is in hindsight only partly useful: lack of nodal or systemic metastases provides an argument of two primary lung cancers, while presence of local and/or distant metastases may be due to one of the two or both lung cancers. Conclusion If morphological and/or DNA analysis provides differences between two tumors it is relatively easy to establish that these pulmonary foci of cancer are separate primary tumors. Many commonly used characteristics are associated with a substantial rate of misclassification. Careful review by a multidisciplinary tumor board, considering all available information, is recommended. References 1. Detterbeck, F. C. et al. The IASLC Lung Cancer Staging Project: Summary of Proposals for Revisions of the Classification of Lung Cancers with Multiple Pulmonary Sites of Involvement in the Forthcoming Eighth Edition of the TNM Classification. J. Thorac. Oncol. 11, 639–50 (2016). 2. Detterbeck, F. C. et al. The IASLC Lung Cancer Staging Project: Background Data and Proposals for the Classification of Lung Cancer with Separate Tumor Nodules in the Forthcoming Eighth Edition of the TNM Classification for Lung Cancer. J. Thorac. Oncol. 11, 681–92 (2016). 3. Detterbeck, F. C. et al. The IASLC Lung Cancer Staging Project: Background Data and Proposals for the Application of TNM Staging Rules to Lung Cancer Presenting as Multiple Nodules with Ground Glass or Lepidic Features or a Pneumonic-Type of Involvement in the Forthcoming Eighth. J. Thorac. Oncol. 11, 666–680 (2016). 4. Kozower, B. D., Larner, J. M., Detterbeck, F. C. & Jones, D. R. Special treatment issues in non-small cell lung cancer: Diagnosis and management of lung cancer, 3rd ed: American College of Chest Physicians evidence-based clinical practice guidelines. Chest 143, e369S–99S (2013). 5. Detterbeck, F. C. et al. The IASLC Lung Cancer Staging Project: Background Data and Proposed Criteria to Distinguish Separate Primary Lung Cancers from Metastatic Foci in Patients with Two Lung Tumors in the Forthcoming Eighth Edition of the TNM Classification for Lung Cancer. J. Thorac. Oncol. 11, 651–65 (2016). 6. Thunnissen, E. et al. Reproducibility of histopathological subtypes and invasion in pulmonary adenocarcinoma. An international interobserver study. Mod. Pathol. 25, 1574–83 (2012). 7. Vignot, S. et al. Next-generation sequencing reveals high concordance of recurrent somatic alterations between primary tumor and metastases from patients with non-small-cell lung cancer. J. Clin. Oncol. 31, 2167–72 (2013). 8. Arai, J. et al. Clinical and molecular analysis of synchronous double lung cancers. Lung Cancer 77, 281–7 (2012). 9. de Bruin, E. C. et al. Spatial and temporal diversity in genomic instability processes defines lung cancer evolution. Science (80-. ). 346, 251–256 (2014). 10. Macintyre, G., Ylstra, B. & Brenton, J. D. Sequencing Structural Variants in Cancer for Precision Therapeutics. Trends Genet. 32, 530–42 (2016).
Only Members that have purchased this event or have registered via an access code will be able to view this content. To view this presentation, please login, select "Add to Cart" and proceed to checkout. If you would like to become a member of IASLC, please click here.
