Abstract 470: Evaluation of two receptor tyrosine kinase (ROS1) clones by immunohistochemistry
Authors: Masabanda Julio S. , Scoggin Tatiana , Vargas Joseph , Ramos Jason 
Published: July 2021

Introduction: ROS1 is a receptor tyrosine kinase which regulates growth and cell division. ROS1 chromosomal rearrangements have been identified in a variety of human malignancies including non-small-cell lung carcinoma (NSCLC). However, patients diagnosed specifically with ROS1+ lung adenocarcinoma accounts for 1-2 percent of all NSCLC cases. Increased expression of ROS1 may suggest a ROS1 gene rearrangement. This is similar to ALK+ NSCLC tumors. Some ALK inhibitors can also be effective in tumors having the ROS1 mutation. Therefore, the development of targeted therapies has enhanced the importance of an accurate classification of the cancer subtypes.

Materials and Methods: Immunohistochemistry: Both, the Biocare ROS1 rabbit monoclonal antibody (clone EPMGHR2) and the Roche VENTANA ROS1 (SP384) rabbit antibody were used in a ready-to-use format. ROS1 antibodies were run on the intelliPATH FLX, VALENT and the Ventana BenchMark Ultra platforms with in-house control and commercially available tissues from US Biolabs. Detection strategies utilized an HRP 2-step polymerized approach on the Biocare instruments, whereas the Optiview DAB IHC Detection kit was used on the Ventana platform. Fluorescence in Situ Hybridization (FISH): FISH was performed on a fully automated ONCORE Pro autostainer using the ROS1 (6q22) Break Apart (BA) (Orange/Green) FISH probe diluted in Biocare zipFISH hybridization buffer.

Results: Initially, the NSCLC tissues were screened with a ROS1 BA FISH probe to identify the ROS1 mutation. Results indicated that the mutation was present on tumor cells throughout the tissue. When compared to FISH results on adjacent sections, a direct correlation between the ROS1 antibody staining and FISH results showed a good staining overlap. IHC staining on the intelliPATH FLX exhibited better signal-to-noise ratio for the Biocare ROS1 antibody compared to the Ventana antibody. A more concentrated Biocare antibody was required to obtain equivalent or better staining on the intelliPATH FLX than the Ventana ROS1 on the VENTANA ULTRA platform. However, the Biocare ROS1 in certain regions showed better staining than the Ventana ROS1. Biocare ROS1 at the same concentration produced a comparable staining on both the IntelliPATH FLX and the VALENT platforms.

Conclusions: The availability of high quality ROS1 antibodies from alternative sources can be beneficial for labs performing lung cancer screening. Our rabbit monoclonal ROS1 antibody has been shown to produce strong and consistent signals with exceptional signal-to-noise. These results have been verified by using highly sensitive FISH probes in combination with a well-characterized commercially available ROS1 antibody.

Citation Format: Julio S. Masabanda, Tatiana Scoggin, Joseph Vargas, Jason Ramos. Evaluation of two receptor tyrosine kinase (ROS1) clones by immunohistochemistry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 470.