Skip to main content

Lund University Publications

LUND UNIVERSITY LIBRARIES

Comparison of ROS1-rearrangement detection methods in a cohort of surgically resected non-small cell lung carcinomas

Thurfjell, Viktoria ; Micke, Patrick ; Yu, Hui ; Krupar, Rosemarie ; Svensson, Maria A. ; Brunnström, Hans LU orcid ; Lamberg, Kristina ; Moens, Lotte N.J. ; Strell, Carina and Gulyas, Miklos , et al. (2022) In Translational Lung Cancer Research 11(12). p.2477-2494
Abstract

Background: Patients with non-small cell lung cancer (NSCLC) harboring a ROS proto-oncogene 1 (ROS1)-rearrangement respond to treatment with ROS1 inhibitors. To distinguish these rare cases, screening with immunohistochemistry (IHC) for ROS1 protein expression has been suggested. However, the reliability of such an assay and the comparability of the antibody clones has been debated. Therefore we evaluated the diagnostic performance of current detection strategies for ROS1-rearrangement in two NSCLC-patient cohorts. Methods: Resected tissue samples, retrospectively collected from consecutive NSCLC-patients surgically treated at Uppsala University Hospital were incorporated into tissue microarrays [all n=676, adenocarcinomas (AC) n=401,... (More)

Background: Patients with non-small cell lung cancer (NSCLC) harboring a ROS proto-oncogene 1 (ROS1)-rearrangement respond to treatment with ROS1 inhibitors. To distinguish these rare cases, screening with immunohistochemistry (IHC) for ROS1 protein expression has been suggested. However, the reliability of such an assay and the comparability of the antibody clones has been debated. Therefore we evaluated the diagnostic performance of current detection strategies for ROS1-rearrangement in two NSCLC-patient cohorts. Methods: Resected tissue samples, retrospectively collected from consecutive NSCLC-patients surgically treated at Uppsala University Hospital were incorporated into tissue microarrays [all n=676, adenocarcinomas (AC) n=401, squamous cell carcinomas (SCC) n=213, other NSCLC n=62]. ROS1-rearrangements were detected using fluorescence in situ hybridization (FISH) (Abbott Molecular; ZytoVision). In parallel, ROS1 protein expression was detected using IHC with three antibody clones (D4D6, SP384, EPMGHR2) and accuracy, sensitivity, and specificity were determined. Gene expression microarray data (Affymetrix) and RNA-sequencing data were available for a subset of patients. NanoString analyses were performed for samples with positive or ambiguous results (n=21). Results: Using FISH, 2/630 (0.3% all NSCLC; 0.5% non-squamous NSCLC) cases were positive for ROS1 fusion. Additionally, nine cases demonstrated ambiguous FISH results. Using IHC, ROS1 protein expression was detected in 24/665 (3.6% all NSCLC; 5.1% non-squamous NSCLC) cases with clone D4D6, in 18/639 (2.8% all NSCLC; 3.9% non-squamous NSCLC) cases with clone SP384, and in 1/593 (0.2% all NSCLC; 0.3% non-squamous NSCLC) case with clone EPMGHR2. Elevated RNA-levels were seen in 19/369 (5.1%) cases (Affymetrix and RNA-sequencing combined). The overlap of positive results between the assays was poor. Only one of the FISH-positive cases was positive with all antibodies and demonstrated high RNA-expression. This rearrangement was confirmed in the NanoString-assay and also in the RNA-sequencing data. Other cases with high protein/RNA-expression or ambiguous FISH were negative in the NanoString-assay. Conclusions: The occurrence of ROS1 fusions is low in our cohorts. The IHC assays detected the fusions, but the accuracy varied depending on the clone. The presumably false-positive and uncertain FISH results questions this method for detection of ROS1-rearrangements. Thus, when IHC is used for screening, transcript-based assays are preferable for validation in clinical diagnostics.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; ; ; ; ; ; and , et al. (More)
; ; ; ; ; ; ; ; ; ; ; ; and (Less)
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Crizotinib, fusion gene detection, molecular pathology, ROS proto-oncogene 1 (ROS1), targeted therapy
in
Translational Lung Cancer Research
volume
11
issue
12
pages
18 pages
publisher
AME Publishing Company
external identifiers
  • scopus:85147779135
  • pmid:36636421
ISSN
2218-6751
DOI
10.21037/tlcr-22-504
language
English
LU publication?
yes
id
40a23515-ca04-4915-b981-96281c92fb3a
date added to LUP
2023-02-20 12:22:43
date last changed
2024-02-01 23:20:01
@article{40a23515-ca04-4915-b981-96281c92fb3a,
  abstract     = {{<p>Background: Patients with non-small cell lung cancer (NSCLC) harboring a ROS proto-oncogene 1 (ROS1)-rearrangement respond to treatment with ROS1 inhibitors. To distinguish these rare cases, screening with immunohistochemistry (IHC) for ROS1 protein expression has been suggested. However, the reliability of such an assay and the comparability of the antibody clones has been debated. Therefore we evaluated the diagnostic performance of current detection strategies for ROS1-rearrangement in two NSCLC-patient cohorts. Methods: Resected tissue samples, retrospectively collected from consecutive NSCLC-patients surgically treated at Uppsala University Hospital were incorporated into tissue microarrays [all n=676, adenocarcinomas (AC) n=401, squamous cell carcinomas (SCC) n=213, other NSCLC n=62]. ROS1-rearrangements were detected using fluorescence in situ hybridization (FISH) (Abbott Molecular; ZytoVision). In parallel, ROS1 protein expression was detected using IHC with three antibody clones (D4D6, SP384, EPMGHR2) and accuracy, sensitivity, and specificity were determined. Gene expression microarray data (Affymetrix) and RNA-sequencing data were available for a subset of patients. NanoString analyses were performed for samples with positive or ambiguous results (n=21). Results: Using FISH, 2/630 (0.3% all NSCLC; 0.5% non-squamous NSCLC) cases were positive for ROS1 fusion. Additionally, nine cases demonstrated ambiguous FISH results. Using IHC, ROS1 protein expression was detected in 24/665 (3.6% all NSCLC; 5.1% non-squamous NSCLC) cases with clone D4D6, in 18/639 (2.8% all NSCLC; 3.9% non-squamous NSCLC) cases with clone SP384, and in 1/593 (0.2% all NSCLC; 0.3% non-squamous NSCLC) case with clone EPMGHR2. Elevated RNA-levels were seen in 19/369 (5.1%) cases (Affymetrix and RNA-sequencing combined). The overlap of positive results between the assays was poor. Only one of the FISH-positive cases was positive with all antibodies and demonstrated high RNA-expression. This rearrangement was confirmed in the NanoString-assay and also in the RNA-sequencing data. Other cases with high protein/RNA-expression or ambiguous FISH were negative in the NanoString-assay. Conclusions: The occurrence of ROS1 fusions is low in our cohorts. The IHC assays detected the fusions, but the accuracy varied depending on the clone. The presumably false-positive and uncertain FISH results questions this method for detection of ROS1-rearrangements. Thus, when IHC is used for screening, transcript-based assays are preferable for validation in clinical diagnostics.</p>}},
  author       = {{Thurfjell, Viktoria and Micke, Patrick and Yu, Hui and Krupar, Rosemarie and Svensson, Maria A. and Brunnström, Hans and Lamberg, Kristina and Moens, Lotte N.J. and Strell, Carina and Gulyas, Miklos and Helenius, Gisela and Yoshida, Akihiko and Goldmann, Torsten and Mattsson, Johanna Sofia Margareta}},
  issn         = {{2218-6751}},
  keywords     = {{Crizotinib; fusion gene detection; molecular pathology; ROS proto-oncogene 1 (ROS1); targeted therapy}},
  language     = {{eng}},
  number       = {{12}},
  pages        = {{2477--2494}},
  publisher    = {{AME Publishing Company}},
  series       = {{Translational Lung Cancer Research}},
  title        = {{Comparison of ROS1-rearrangement detection methods in a cohort of surgically resected non-small cell lung carcinomas}},
  url          = {{http://dx.doi.org/10.21037/tlcr-22-504}},
  doi          = {{10.21037/tlcr-22-504}},
  volume       = {{11}},
  year         = {{2022}},
}