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Amy Elizabeth Hanlon Newell



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    MA26 - New Therapies and Emerging Data in ALK, EGFR and ROS1 (ID 930)

    • Event: WCLC 2018
    • Type: Mini Oral Abstract Session
    • Track: Targeted Therapy
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/26/2018, 13:30 - 15:00, Room 201 BD
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      MA26.07 - ROS1 (SP384) Immunohistochemistry Inter-Reader Precision Between 12 Pathologists (ID 12387)

      14:10 - 14:15  |  Author(s): Amy Elizabeth Hanlon Newell

      • Abstract
      • Presentation
      • Slides

      Background

      ROS1 positive non-small cell lung cancer (NSCLC) patients can be treated with specific tyrosine kinase inhibitors including crizotinib. ROS1 positivity is often clinically detected by fluorescence in situ hybridization (FISH), however ROS1 IHC can be used to screen samples prior to FISH confirmation of ROS1 status. The ROS1 (SP384) antibody detects ROS1 with high sensitivity, specificity, and consistency. Consistent interpretation of a ROS1 IHC assay between pathologists is important patient evaluation. Here we present inter-reader precision of 12 pathologists across 60 FFPE cases stained with ROS1 (SP384).

      a9ded1e5ce5d75814730bb4caaf49419 Method

      A retrospective cohort of 60 FFPE NSCLC cases stained with H&E, Rabbit Monoclonal Negative Control Ig, and ROS1 (SP384) were selected to represent positive, negative, and borderline ROS1 IHC status. Twelve practicing lung pathologists independently scored the cases as positive or negative around a cutoff of cytoplasm staining in > 30% tumor cells at a ≥2+ intensity level using Pathotrainer software (Pathomation bvba). Scoring was blinded to other readers and ROS1 status of the cases. Overall percent agreement (OPA), negative percent agreement (NPA), and positive percent agreement (PPA) were calculated in comparison to the group mode. Average overall percent agreement (AOPA), average positive agreement (APA), and average negative agreement (ANA) were calculated pairwise for each reader pair. Following independent assessment, participating pathologists conducted a discordant case review establishing consensus reads for all 60 cases and compared 44 cases to available FISH results.

      4c3880bb027f159e801041b1021e88e8 Result

      OPA of each of the 12 readers to the mode was 96.4% (95% CI 93.9-98.6) with PPA of 96.3% (95% CI 92.7-99.4) and NPA of 96.5% (95% CI 92.8-99.5). Pairwise AOPA between each of the 12 readers was 94.5% (95%CI 91.2-97.7) with APA 94.0% (95% CI 89.5-97.6) and ANA 95.0% (95%CI 91.2-97.9).

      Consensus IHC scores were concordant with FISH 90.0% (40/44 cases).

      8eea62084ca7e541d918e823422bd82e Conclusion

      Inter-reader precision around a cutoff of >30% tumor cells with cytoplasmic staining at a ≥2+ intensity level was high in interpreting ROS1 (SP384) in NSCLC samples. Case review highlighted confirmation with FISH in questionable cases and staining patterns to be considered when interpreting ROS1 (SP384) IHC.

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    P2.09 - Pathology (Not CME Accredited Session) (ID 958)

    • Event: WCLC 2018
    • Type: Poster Viewing in the Exhibit Hall
    • Track:
    • Presentations: 2
    • Moderators:
    • Coordinates: 9/25/2018, 16:45 - 18:00, Exhibit Hall
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      P2.09-09 - Precision and Repeatability of VENTANA DLL3 (SP347) Assay Immunohistochemistry Assay in Fine Needle Aspirations of Small Cell Lung Cancer (ID 12809)

      16:45 - 18:00  |  Author(s): Amy Elizabeth Hanlon Newell

      • Abstract
      • Slides

      Background

      Fine needle aspirations (FNA) are a common type of sample that is obtained for diagnostic purposes in small cell lung cancer (SCLC) patients due to its relatively less invasive nature. Several delta-like protein 3 (DLL3) therapies that target DLL3 in SCLC are under development. We recently developed an immunohistochemistry assay utilizing a rabbit monoclonal antibody that can be used to identify patients that may potentially respond to DLL3 targeted therapies. Due to the high prevalence of FNA samples in SCLC, the robustness of the VENTANA DLL3 (SP347) Assay (DLL3 (SP347)) in FNAs is examined.

      a9ded1e5ce5d75814730bb4caaf49419 Method

      Formalin-fixed, paraffin-embedded (FFPE) FNA samples of SCLC were used in a series of studies addressing precision of performance of DLL3 (SP347). FNA SCLC samples were stained with the VENTANA DLL3 (SP347) Assay and assessed by a pathologist for percent tumor cell staining (%TC), at any stain intensity at ≤4X magnification. The samples were placed into quartile bins (0-24, 25-49, 50-74 and 75-100 %TC) and evaluated for intra-day repeatability and inter-day precision. For each case, the modal staining result based on %TC bin was determined and the result from each test sample was compared to its respective case-level modal staining result and deemed concordant or discordant. Results were aggregated across cases and the overall percent agreement (OPA) was calculated for each study.

      4c3880bb027f159e801041b1021e88e8 Result

      Both studies (Table 1) showed >90% OPA for DLL3 concordance to the %TC bins.

      Table 1: Precision and Repeatability of FNA SCLC Samples
      Study # of Cases

      # DLL3

      Observations

      OPA
      Intra-day 21 63 100%
      Inter-day 21 124 98.4

      8eea62084ca7e541d918e823422bd82e Conclusion

      The presented data shows that VENTANA DLL3 (SP347) Assay can precisely evaluate DLL3 expression in FNA SCLC samples.

      6f8b794f3246b0c1e1780bb4d4d5dc53

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      P2.09-13 - Correlation of ROS1 (SP384) Immunohistochemistry with ROS1 Rearrangement Determined by Fluorescence in Situ Hybridization (ID 12808)

      16:45 - 18:00  |  Author(s): Amy Elizabeth Hanlon Newell

      • Abstract
      • Slides

      Background

      With the approval of crizotinib as treatment for ROS1 positive patients, identification of ROS1 status has become standard of care for non-small cell lung cancer (NSCLC). Immunohistochemistry (IHC) can be used to detect aberrant ROS1 expression; however, current commercially available IHC assays do not have optimized recommended protocols, or data correlating ROS1 protein expression to fluorescence in situ hybridization (FISH). Hence, the ROS1 (SP384) Rabbit Monoclonal Primary Antibody (ROS1 SP384) was developed, and here we present correlation data to ROS1 FISH status across multiple scoring algorithms using our optimized IHC staining protocol.

      a9ded1e5ce5d75814730bb4caaf49419 Method

      120 formalin-fixed, paraffin-embedded (FFPE) NSLC cases with known FISH status were procured. 4μm sections were cut and stained with H&E, Rabbit Monoclonal Negative Control Ig, and ROS1 SP384. The slides were evaluated for percent positivity of tumor cells in each intensity level (evaluated separately for staining within the nuclear, cytoplasmic, and membranous compartments). The intensity level of staining was evaluated on an integer scale of 0-3: 0 = no, 1 = weak, 2 = moderate and 3 = strong staining intensity. The data was analyzed several scoring algorithms to assess its correlation with FISH status.

      4c3880bb027f159e801041b1021e88e8 Result

      ROS1 SP384 showed high correlation with FISH status, with multiple scoring algorithms (Table 1) examined. Of note, a positive percent agreement of 97.8% and negative percent agreement of 89.2% was achieved when comparing ROS1 SP384 IHC staining ≥2+ at a cutoff of > 30% staining in the cytoplasm of tumor cells with FISH status.

      Table 1. VENTANA ROS1 (SP384) Rabbit Monoclonal Primary Antibody IHC vs FISH analytical comparison data

      wclc abstract ros1 correlation table.jpg

      8eea62084ca7e541d918e823422bd82e Conclusion

      Herein, we present evidence thatROS1 (SP384) Rabbit Monoclonal Primary Antibody may provide an effective means of stratifying cases by aberrant ROS1 protein expression prior to confirmation with orthogonal methods.

      6f8b794f3246b0c1e1780bb4d4d5dc53

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