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A. Montero Fernandez
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ORAL 40 - Biology 1 (ID 154)
- Event: WCLC 2015
- Type: Oral Session
- Track: Thymoma, Mesothelioma and Other Thoracic Malignancies
- Presentations: 1
- Moderators:C. Brambilla, R. Bueno
- Coordinates: 9/09/2015, 16:45 - 18:15, 702+704+706
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ORAL40.02 - Molecular Landscape of Malignant Mesothelioma from Whole Exome Sequencing (ID 2439)
16:45 - 18:15 | Author(s): A. Montero Fernandez
- Abstract
- Presentation
Background:
Whole exome sequencing has revealed key genetic events in several cancer types that have been successfully translated into clinical benefits. These advances are still lacking in malignant mesothelioma (MM), a highly aggressive malignancy with limited effective therapy. Frequent BAP1 mutations occur in a subset of this disease but the full molecular landscape of MM is still poorly characterized.
Methods:
We have therefore conducted whole exome sequencing of tumours from the pleura for 36 cases of MM. DNA from matched blood was available for 7 of the cases and was also sequenced. The variants were identified with GATK tools and annotated with ANNOVAR. Variants were filtered with the following criteria: quality score ≤ 50, present in dbSNP138, 1000 genomes variants and NHLBI ESP 6500 variants. Mutations with deleterious functional consequences predicted by Polyphen-2, SIFT and Mutation Taster tools were confirmed by Sanger sequencing.
Results:
A total of 9,064 variants (3,256 somatic) were identified. We confirmed mutations in genes previously described to be mutated in MM in 5 cases: BAP1 (R227C, Q684X, H141P), NF2 (76_76del, R221X) and TP53 (I195N). In BAP1 wt tumours (6 of the 7 cases with matched blood), we confirmed somatic mutations in 5 genes encoding components of either MAPK or WNT signaling pathways. In addition, we validated somatic mutations in 12 genes across 4 of the 6 cases, many of which are novel in MM and are involved in chromatin modification. We also observed these genes to be mutated in BAP1 wt tumours in the 29 additional unmatched MM cases.
Conclusion:
Thus our data suggests that in addition to BAP1, mutations in genes associated with MAPK, WNT signaling and the chromatin remodeling complex may represent a consistent pattern of molecular alterations in MM.
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P3.04 - Poster Session/ Biology, Pathology, and Molecular Testing (ID 235)
- Event: WCLC 2015
- Type: Poster
- Track: Biology, Pathology, and Molecular Testing
- Presentations: 1
- Moderators:
- Coordinates: 9/09/2015, 09:30 - 17:00, Exhibit Hall (Hall B+C)
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P3.04-011 - A Validation Study for the Use of ROS-1 Immunohistochemistry in Screening for ROS-1 Translocations in Lung Cancer (ID 2826)
09:30 - 17:00 | Author(s): A. Montero Fernandez
- Abstract
Background:
ROS-1 translocations are a rare genetic abnormality in lung cancers that, when identified, are a target for personalised therapy. The current test of choice is FISH, although with a rate of no more than 1-2%, screening using FISH is an expensive proposition. A further possibility is using immunohistochemistry (IHC) as a screening tool and commercial antibodies are now available that identify the ROS-1 protein in tumour cells. We present our data in undertaking a validation study for potential diagnostic usage.
Methods:
Given the relative rarity of the translocation and the fact the most driver mutations occur in isolation, a test cohort of cases was selected from patients recruited to phase 1 of the Cancer Research UK-Stratified Medicine Project (CRUK-SMP), who were identified as negative for EGFR, KRAS and/or BRAF mutations, as well as ALK translocations. Negative cases were then screened with an antibody for ROS-1 (D4D6, Cell Signalling, 1 in 300 dilution) and scored as negative, weakly positive or moderately positive, along with the percentage of positive cells. Cases were then sent for FISH analysis for the ROS-1 translocation, with a cut-off of > or = to15%, and the sensitivity and specificity of positive staining for ROS-1 was generated.
Results:
From 170 patients recruited from our institution into CRUK-SMP phase 1, a total of 103 patients were wild type for the above mutations (90 for all 4 genetic abnormalities. 9 further cases had failed tests for one and 4 for two mutations (6 carcinoids, 38 squamous cell carcinomas, 5 small cell carcinoma, 2 adenosquamous carcinoma, 1 pleomorphic carcinoma, 3 large cell carcinoma, 2 large cell neuroendocrine cell carcinoma, 7 non-small cell carcinoma (on biopsy) and 39 adenocarcinomas). 39 cases were tested (adenocarcinoma = 37, adenosquamous carcinoma = 2) with FISH, and one case was positive (78% positive cells). FISH testing was negative in 35 cases with scores of 1-8%, and three cases failed. The one positive case was positive on IHC (>90% of cells, moderate staining). In the 35 cases negative for FISH, four cases showed variable positivity on IHC (20, 40,50, 90%, moderate staining) and five cases showed weak focal staining (<5, <5, 10, 20, 30%, weak staining). The remainder were negative on IHC. All non-adenocarcinomas were negative on IHC. Several cases show positive staining of entrapped background pneumocytes and alveolar macrophages, making scoring problematic in some adenocarcinomas.
Conclusion:
Moderate staining for ROS-1 using IHC, independent of percentage positive cells, showed high sensitivity (100%) for tumours that contained a high level of translocated cells. However, specificity was at best 50%, even if a cut-off of 50% positive cells was applied. Pathologists also need to be aware of background staining so cases are not interpreted as false positives.
