Correlation of ROS1 immunohistochemistry with ROS1 fusion status determined by fluorescence in situ hybridization

Context.-The ability to determine ROS1 status has become mandatory for patients with lung adenocarcinoma, as many global authorities have approved crizotinib for patients with ROS1-positive lung adenocarcinoma. Objective.-To present analytical correlation of the VENTANA ROS1 (SP384) Rabbit Monoclonal Primary Antibody (ROS1 [SP384] antibody) with ROS1 fluorescence in situ hybridization (FISH). Design.-The immunohistochemistry (IHC) and FISH analytical comparison was assessed by using 122 non-small cell lung cancer samples that had both FISH (46 positive and 76 negative cases) and IHC staining results available. In addition, reverse transcription-polymerase chain reaction (RT-PCR) as well as DNA and RNA next-generation sequencing (NGS) were used to further examine the ROS1 status in cases that were discrepant between FISH and IHC, based on staining in the cytoplasm of 2+ or above in more than 30% of total tumor cells considered as IHC positive. Here, we define the consensus status as the most frequent result across the 5 different methods (IHC, FISH, RT-PCR, RNA NGS, and DNA NGS) we used to determine ROS1 status in these cases. Results.-Of the IHC scoring methods examined, staining in the cytoplasm of 2+ or above in more than 30% of total tumor cells considered as IHC positive had the highest correlation with a FISH-positive status, reaching a positive percentage agreement of 97.8% and negative percentage agreement of 89.5%. A positive percentage agreement (100%) and negative percentage agreement (92.0%) was reached by comparing ROS1 (SP384) using a cutoff for staining in the cytoplasm of 2+ or above in more than 30% of total tumor cells to the consensus status. Conclusions.-Herein, we present a standardized staining protocol for ROS1 (SP384) and data that support the high correlation between ROS1 status and ROS1 (SP384) antibody. © 2020 College of American Pathologists. All rights reserved. Boehringer Ingelheim; Bristol-Myers Squibb; Pfizer; Novartis; Roche; AbbVie; Merck KGaA Dr Huang, Mr Smith, Dr Menzl, Dr Le, Mr Liu, Dr Ordinario, Dr Manohar, Mr Lee, Mr Rajamani, Mr Truong, Dr Jing Li, Ms Choi, Dr Jingchuan Li, Dr Pati, Dr Hanlon Newell, and Mr Pate are Roche employees and receive a salary from Roche. Dr Bubendorf received research support from Roche and received honoraria related to advisory board meetings (Pfizer, Roche). Dr Buettner is a cofounder and chief scientific officer of Targos Mol. Pathol. Inc, Kassel, Germany. Dr Kerr has received honoraria for advisory work and lectures from Ventana Medical Systems, Roche, and Pfizer; these payments were outside the scope of this work. Dr Lopez-Rios received funding and honoraria from Roche, Pfizer, and Thermo Fischer. Dr Marchetti has received honoraria for advisory work and lectures from Ventana Medical Systems, Roche, and Pfizer; these payments were outside the scope of this work. Dr Marondel is an employee and receives a salary from Pfizer. Dr Nicholson serves as a consultant and receives a fee from Merck, Boehringer Ingelheim, Pfizer (also research grant), Novartis, Astra Zeneca (also payment for lecture), Bristol-Myers Squibb, Roche, and ABBVIE. Dr Pauwels received research grants from Pfizer, Roche, and Astra Zeneca and was paid for attending advisory boards of BMS, MSD Astra Zeneca, Takedo, Roche, and Boehringer. Dr Thunnissen participates in Ventana advisory board meetings. The other authors (Dr Öz, Dr Penault-Llorca, and Dr Rossi) have no relevant financial interest in the products or companies described in this article.