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A. Yoshizawa
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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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P1.02-009 - High Concordance of ALK Rearrangement Testing between ALK RNA-In Situ Hybridization and IHC/FISH in Patients with Lung Adenocarcinoma (ID 4402)
14:30 - 14:30 | Author(s): A. Yoshizawa
- Abstract
Background:
Patients with anaplastic lymphoma kinase (ALK) gene rearrangements manifest good responses to ALK inhibitors and thus, accurate and rapid identification of ALK gene rearrangements is essential for the clinical application of ALK-targeted therapies. The aim of this study was to investigate the diagnostic accuracy of the recently developed ALK RNA-in situ hybridization (RNA-ISH) assay, using formalin fixed paraffin embedded samples of lung adenocarcinoma tissue.
Methods:
We first tested whether ALK RNA-ISH could be performed in 11 resected lung adenocarcinomas in which ALK gene rearrangements were confirmed by immunohistochemistry (IHC, D5F3), and/or fluorescence in situ hybridization (FISH), and also clarified the intra-tumor heterogeneity of ALK RNA-ISH by counting 100 tumor cells in 10 different loci (total, 1000 tumor cells) in each tumor. Secondly, we analyzed the diagnostic accuracy of ALK RNA-ISH in tissue microarrays (TMAs) containing 294 lung adenocarcinoma cores (by counting 100 tumor cells in each core) with no information about ALK gene rearrangements and compared these results with those of conventional IHC and FISH tests.
Results:
ALK mRNA expression was observed in all 11 resected lung adenocarcinomas by the ALK RNA-ISH assay and the median of positive tumor cells was 67.7%, whereas ALK mRNA expression was not observed in normal lung cells in the background. Next, 5 ALK positive cases were found by IHC and/or FISH in the 294 cases of lung adenocarcinoma. The median of positive cells by ALK RNA-ISH in these 5 cases was 75.6% (range: 40-94%), whereas the median of positive cells by ALK RNA-ISH in the remaining 289 cases was 0.3% (range: 0-15%). When the cutoff value was set as 15% based on the first test, the ALK RNA-ISH–positive and ALK RNA-ISH–negative cases were readily distinguishable with 100% sensitivity and specificity compared with the results of IHC and/or FISH.
Conclusion:
Our findings suggest that the ALK RNA-ISH assay is useful for detecting ALK positive lung adenocarcinomas with high sensitivity and specificity compared with the conventional IHC and FISH test. Thus, this study provides important and timely insight into the clinical testing of ALK in lung cancer because the RNA-ISH assay detects the target mRNA easily and rapidly.
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P2.04 - Poster Session with Presenters Present (ID 466)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Mesothelioma/Thymic Malignancies/Esophageal Cancer/Other Thoracic Malignancies
- Presentations: 1
- Moderators:
- Coordinates: 12/06/2016, 14:30 - 15:45, Hall B (Poster Area)
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P2.04-018 - Comprehensive Copy Number Alteration and Gene Expression Analysis of Surgically Resected Thymic Carcinoma (ID 4725)
14:30 - 14:30 | Author(s): A. Yoshizawa
- Abstract
Background:
Thymic carcinoma is rare and comparatively poor prognostic. Due to its rarity, our knowledge of treatments and prognostic biomarkers available for these tumors is limited. Previous reports revealed low genetic mutation frequency of the tumors, and the inverse correlation between the frequency of genetic mutation and copy number alteration (CNA). Based on these reports, we hypothesized tumorigenesis of thymic carcinomas is mostly caused by copy number and transcriptional alterations. To substantiate the hypothesis, we extracted and analyzed CNA and gene expression data from surgical samples to elucidate driver genes, druggable targets and prognostic factors.
Methods:
Between January 2009 and March 2016, patients underwent surgery for thymic epithelial tumor in our institution were reviewed. RNA and DNA of the tumors were extracted from FFPE operative samples, gene expression data were obtained by GeneChip Human Transcriptome Array 2.0 (affymetrix), and CNA were detected by OncoScan (affymetrix). These results were analyzed using Transcriptome analysis console (affymetrix) and Nexus expression for OncoScan (BioDiscovery). Upregulated genes in copy number gained region and down regulated genes in copy number lost region were selected as a candidate of tumorigenesis of thymic carcinoma.
Results:
We had 10 thymic squamous cell carcinoma samples. As a comparison, we use three samples of type A thymoma. CNA data from thymic squamous cell carcinoma showed similar characteristics, chromosome 1q gain, 6p and q loss, and 16q loss, corresponding with previous reports. Gene expression analysis of thymic carcinoma in comparison with type A thymoma revealed down regulation of the genes, BRD2, HSP90AB1, FOXO3, and MARCKS in chromosome 6, and MTSS1L in chromosome 16q. Figure 1
Conclusion:
We reported the results of genome wide gene expression and CNA analysis. We extracted some candidate genes, but farther validations are needed.
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P3.01 - Poster Session with Presenters Present (ID 469)
- Event: WCLC 2016
- Type: Poster Presenters Present
- Track: Biology/Pathology
- Presentations: 3
- Moderators:
- Coordinates: 12/07/2016, 14:30 - 15:45, Hall B (Poster Area)
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P3.01-018 - Reproducibility in Classification of Small Lung Adenocarcinomas: An International Interobserver Study (ID 5222)
14:30 - 14:30 | Author(s): A. Yoshizawa
- Abstract
Background:
The 2015 WHO classification for lung adenocarcinoma (ACA) provides criteria for diagnosis of adenocarcinoma in-situ (AIS), minimally invasive adenocarcinoma (MIA), and invasive adenocarcinoma (INV). Differentiating these entities can be difficult, and as understanding of prognostic significance increases, inconsistent classification is problematic.
Methods:
Sixty cases of lung ACAs (<2cm) were reviewed by an international panel of 6 lung pathologists. One slide reflecting overall morphology of each case was digitally scanned to an internet browser-based viewer. In round one, the panel independently reviewed each case to assess predominant pattern, invasive component size, and final diagnosis (AIS, MIA or INV). After a consensus conference among participants, a second round of independent review of the cases was performed. Additionally, a discussion on interpretation of elastic stain for evaluation of invasion will precede a third round of review with assessment of a concomitant elastic stain for each case. Statistical analysis was performed for each round.
Results:
In round one, the overall kappa value for AIS versus MIA and INV was 0.34 (fair agreement), and that for AIS and MIA versus INV was 0.44 (moderate agreement). The raters had 100% agreement on final diagnosis in 10 cases (AIS, n=2; MIA, n=2; INV, n=6). In 28 cases with poor agreement on final diagnosis and invasive measurement, inconsistent measurement of multifocal invasion led to wide variance in 5 cases, and subjectivity in pattern recognition led to variance in 23 cases. Misinterpretation of the WHO criteria for MIA resulted in 18 instances of misclassification across all raters. A case with a predominant mucinous lepidic pattern had a range of diagnoses (AIS, n=1; MIA, n=1; INV, n=4). In round two, the overall kappa value for AIS versus MIA and INV is 0.40 (fair agreement), and that for AIS and MIA versus INV is 0.36 (moderate agreement). The raters had 100% agreement on final diagnosis in 12 cases (AIS, n=3; MIA n=4; INV, n=5). Misinterpretation of the WHO criteria for MIA was seen in 6 instances. The intraobserver kappa coefficient ranged widely from 0.259 to 0.859.
Conclusion:
Interobserver agreement on diagnosis of small lung ACAs between raters was fair to moderate, with minimal improvement after a consensus conference. Inconsistent measurement of multifocal invasion, subjectivity in pattern recognition, misinterpretation of the WHO criteria, and subjective interpretation of mucinous ACA have contributed to interobserver discordance. A third round of evaluation is currently ongoing to assess for improvement and the utility of elastic stains.
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P3.01-026 - Clinical and Pathological Reappraisal of Primary Lung Carcer with Lymphoepithelioma-Like Carcinoma Morphology (ID 5978)
14:30 - 14:30 | Author(s): A. Yoshizawa
- Abstract
Background:
Lymphoepithelioma-like carcinoma (LELC) is a rare form of lung cancer, usually encountered in Chinese patients. Similar to nasopharyngeal carcinoma which is strongly associated with Epstein-Barr virus (EBV) infection, LELC is defined as a poorly differentiated carcinoma reveals EBER-positive neoplastic cells and marked lymphocyte infiltrate by 2015 WHO classification; however, EBER-negative carcinomas showing LELC-like morphology are present and such cases might be classified as adenocarcinoma or squamous cell carcinoma based on the results of immunohistochemistry staining, such as p40 or TTF-1.
Methods:
We retrospectively reviewed the medical records of 5 LELC patients who underwent pulmonary resection in Kyoto University Hospital between 2005 and 2016. All five cases were primary lung tumors with histologic features of carcinoma characterized by poorly differentiated morphology admixed with heavy lymphocyte infiltrates which fit the criteria for the diagnosis of LELC as morphologic findings.
Results:
There were 4 men and 1 woman who ranged in age from 65 to 78 years, with a median age of 70. Three patients had lymph node metastasis and underwent surgical resection, followed by adjuvant chemotherapy.One patient died of second primary lung cancer (small cell carcinoma) but four patients were alive without tumor recurrence 4 months to 8 years and 11 months.Four patients (80%) were negative for EBV, suggesting no association between EBV and LELC in our institution study group.In immunohistochemistry staining, 4 cases were positive for p40 and one case was for TTF-1.patient Age Sex Smoking Location pStage Treatment EBER TTF-1 p40 Outcome 1 78 M Yes RUL T1aN0M0 Surg＋adjuvant rad negative negative positive 3y6m dead 2 68 M Yes LLL T1aN2M0 Surg＋adjuvant chemo negative negative positive 8y11m alive 3 71 F None LLL T3N1M0 Surg＋adjuvant chemo positive negative positive 6y2m alive 4 65 M Yes RUL T1bN1M0 Surg＋adjuvant chemo negative positive negative 5y11m alive 5 73 M Yes RUL T1aN0M0 Surg only negative negative positive 0y4m alive
Conclusion:
Our reexamination revealed that most LELCs were negative for EBER and were classified as squamous cell carcinoma by IHC study. This results might imply that EBER is not a requisite factor in the lung carcinoma with LELC-like morphology.
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P3.01-048 - Cigarette Smoking is Associated with Epithelio-Mesenchymal Transition in Human Adenocarcinoma (ID 5531)
14:30 - 14:30 | Author(s): A. Yoshizawa
- Abstract
Background:
Cigarette smoking (CS) is well known to cause lung cancer. In addition to the mechanisms of tumorigenesis of lung cancer with CS, a lot of evidences are currently accumulating that CS induces epithelio-mesenchymal transition (EMT) in tumor cells. The correlation of CS with the malignant properties of lung cancer remained elusive in clinical settings. Here we examined the clinical significance of CS with regard to EMT and malignant progression in human lung adenocarcinoma.
Methods:
Clinical samples were obtained from the 239 cases of resected lung adenocarcinoma which were consecutively operated from January 2001 to December 2007 in our institution. Pathological stage distribution of the cases by TNM classification (WHO, 7th edition) was below: 1A: 118, 1B: 71, 2A: 22, 2B: 4, 3A: 23, 3B: 1. Smoking history was taken from all the patients, then their smoking status were classified into 3 groups according to the Brinkman Index (Non;0, n=109: Light;1~400, n=29: Heavy;401~, n=101). The samples were immunostained against E-cadherin and Vimentin using tissue microarrays of resected specimens to assess the activation level of EMT. Then, we classified into 3 groups: the group ‘N’, E-cadherin(E+) and Vimentin(V-), “null” EMT activation; the group ‘F’, E-/V+, ‘full’ EMT; the group ‘P’, E-/V- or E+/V+, ‘partial’ EMT. The numbers of the group F/ P/ N were 38/ 93/ 108, respectively. Furthermore, DNA samples were extracted from frozen surgical samples and the mutations for the hot-spot exons of EGFR, K-ras, and p53 were detected by SSCP or direct sequencing methods. The differences of survival duration, pathological invasive factors, DNA mutations and EMT activation level were statistically analysed among smoking groups.
Results:
Significant difference was found in 5-year survival rate among 3 smoking groups: Non, 89.1%; Light, 89.5%; Heavy, 61.7%. Smoking status was significantly associated with EMT activation, DNA mutation status, local invasive factors, and lymph-node metastasis. In the tumors harboring either wild-type K-ras or wild-type p53, heavy smoking was associated with EMT activation (p<0.0001, p=0.0002, respectively), whereas no correlation with regard to EGFR staus.
Conclusion:
Smoking amounts had a significant association with EMT activation level and malignant progression of human adenocarcinoma. Heavy smoking was related with EMT activation of the tumors either with wild-type K-ras or p53.
