Comparison of Molecular Testing Modalities for Detection of ROS1 Rearrangements in a Cohort of Positive Patient Samples

Davies, Kurtis D.; Le, Anh‐Tuan; Sheren, Jamie; Nijmeh, Hala; Gowan, Katherine; Jones, Kenneth L.; Varella‐Garcia, Marileila; Aisner, Dara L.; Doebele, Robert C.

doi:10.1016/j.jtho.2018.05.041

Cited by 141 publications

(150 citation statements)

References 30 publications

Supporting

Mentioning

132

Contrasting

Unclassified

Order By: Relevance

“…This assay uses a proprietary Anchored Multiplex PCR (AMP™)‐based enrichment to detect all fusions associated with the genes in a single sequencing assay, without prior knowledge of fusion partners or breakpoints. Nucleic acids were extracted and processed from formalin‐fixed paraffin‐embedded material as reported previously . RNA libraries were sequenced on an Illumina MiSeq instrument using v3 chemistry (Illumina, San Diego, CA).…”

Section: Methodsmentioning

confidence: 99%

Targeted fusion analysis can aid in the classification and treatment of pediatric glioma, ependymoma, and glioneuronal tumors

Lake

Donson

Prince

et al. 2019

Pediatric Blood & Cancer

Self Cite

View full text Add to dashboard Cite

Background The use of next‐generation sequencing for fusion identification is being increasingly applied and aids our understanding of tumor biology. Some fusions are responsive to approved targeted agents, while others have future potential for therapeutic targeting. Although some pediatric central nervous system tumors may be cured with surgery alone, many require adjuvant therapy associated with acute and long‐term toxicities. Identification of targetable fusions can shift the treatment paradigm toward earlier integration of molecularly targeted agents. Methods Patients diagnosed with glial, glioneuronal, and ependymal tumors between 2002 and 2019 were retrospectively reviewed for fusion testing. Testing was done primarily using the ArcherDx FusionPlex Solid Tumor panel, which assesses fusions in 53 genes. In contrast to many previously published series chronicling fusions in pediatric patients, we compared histological features and the tumor classification subtype with the specific fusion identified. Results We report 24 cases of glial, glioneuronal, or ependymal tumors from pediatric patients with identified fusions. With the exception of BRAF:KIAA1549 and pilocytic/pilomyxoid astrocytoma morphology, and possibly QKI‐MYB and angiocentric glioma, there was not a strong correlation between histological features/tumor subtype and the specific fusion. We report the unusual fusions of PPP1CB‐ALK, CIC‐LEUTX, FGFR2‐KIAA159, and MN1‐CXXC5 and detail their morphological features. Conclusions Fusion testing proved to be informative in a high percentage of cases. A large majority of fusion events in pediatric glial, glioneuronal, and ependymal tumors can be identified by relatively small gene panels.

show abstract

Section: Methodsmentioning

confidence: 99%

Targeted fusion analysis can aid in the classification and treatment of pediatric glioma, ependymoma, and glioneuronal tumors

Lake

Donson

Prince

et al. 2019

Pediatric Blood & Cancer

Self Cite

View full text Add to dashboard Cite

show abstract

“…Detailed information regarding RNA quality assessment has been published previously. 12 Briefly, for a sample to pass quality metrics, the average number of unique read start sites originating from primers to four housekeeping genes must exceed a defined threshold.…”

Section: Rna-based Ngsmentioning

confidence: 99%

DNA-Based versus RNA-Based Detection of MET Exon 14 Skipping Events in Lung Cancer

Davies

Lomboy

Lawrence

et al. 2019

Journal of Thoracic Oncology

Self Cite

123

125

View full text Add to dashboard Cite

Introduction: Genomic variants that lead to MET proto-oncogenem receptor tyrosine kinase (MET) exon 14 skipping represent a potential targetable molecular abnormality in NSCLC. Consequently, reliable molecular diagnostic approaches that detect these variants are vital for patient care.Methods: We screened tumor samples from patients with NSCLC for MET exon 14 skipping by using two distinct approaches: a DNA-based next-generation sequencing assay that uses an amplicon-mediated target enrichment and an RNA-based next-generation sequencing assay that uses anchored multiplex polymerase chain reaction for target enrichment.Results: The DNA-based approach detected MET exon 14 skipping variants in 11 of 856 NSCLC samples (1.3%). The RNA-based approach detected MET exon 14 skipping in 17 of 404 samples (4.2%), which was a statistically significant increase compared with the DNA-based assay. Among 286 samples tested by both assays, RNA-based testing detected 10 positives, six of which were not detected by the DNA-based assay. Examination of primer binding sites in the DNA-based assay in comparison with published MET exon 14 skipping variants revealed genomic deletion involving primer binding sequences as the likely cause of false negatives. Two samples positive via the DNA-based approach were uninformative via the RNA-based approach due to poor-quality RNA.Conclusions: By circumventing an inherent limitation of DNA-based amplicon-mediated testing, RNA-based analysis detected a higher proportion of MET exon 14 skipping cases. However, RNA-based analysis was highly reliant on RNA quality, which can be suboptimal in some clinical samples.

show abstract

“…10 Such regions of homology may actually be excluded from hybrid capture sequencing assay design so as to maintain higher levels of overall sequencing quality. Davies et al 8 describe this phenomenon in relation to ROS1 intron 31, which is one of the breakpoint hotspots for this gene. In other instances, repetitive regions may be captured but map poorly to the reference genome and thus may be excluded by software programs designed to identify rearrangements and other structural variants.…”

mentioning

confidence: 97%

“…In addition, comprehensive sequencing approaches have been extended for use in plasma specimens with use of circulating tumor DNA so that, ostensibly, patients can forego tumor biopsies for dedicated genomic testing. 7 As Davies et al 8 show in this issue of the Journal of Thoracic Oncology, however, no single assay, whether it be focused wholly on one gene alteration or on a broader range of diverse targets, offers the perfect solution for detection of ROS1 rearrangements in clinical samples. Davies et al 8 have compared three different platforms-FISH, DNA-based targeted hybrid capture sequencing, and RNA-based anchored multiplexed PCR-for detection of ROS1 fusion events in 23 previously characterized clinical samples.…”

mentioning

confidence: 99%

“…7 As Davies et al 8 show in this issue of the Journal of Thoracic Oncology, however, no single assay, whether it be focused wholly on one gene alteration or on a broader range of diverse targets, offers the perfect solution for detection of ROS1 rearrangements in clinical samples. Davies et al 8 have compared three different platforms-FISH, DNA-based targeted hybrid capture sequencing, and RNA-based anchored multiplexed PCR-for detection of ROS1 fusion events in 23 previously characterized clinical samples. Two of 21 FISH-tested samples (or close to 10%) were falsely negative-an alarming but not unexpected outcome that can result from cryptic or complex rearrangements.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Recognizing the Challenges of Oncogene Fusion Detection: A Critical Step toward Optimal Selection of Lung Cancer Patients for Targeted Therapies

Sholl

2018

Journal of Thoracic Oncology

View full text Add to dashboard Cite

Activating ROS1 gene rearrangements represent an important oncogenic event in a minor subset of human tumors. Fusion events between a partner gene and the 3 0 end of ROS1, including its kinase domain, are sufficient to drive tumorigenesis in 1% to 2% of NSCLCs and, importantly, predict sensitivity to tyrosine kinase inhibitors such as crizotinib. 1,2 In patients with advanced NSCLC with ROS1 rearrangement determined by fluorescence in situ hybridization (FISH) or reverse-transcriptase polymerase chain reaction (PCR), crizotinib therapy can lead to rapid and dramatic tumor shrinkage and progressionfree survival often surpassing that seen after targeted therapy for other oncogenic drivers, including EGFR mutations and ALK receptor tyrosine kinase gene (ALK) rearrangements. 2,3 Although ROS1 rearrangement is more common in NSCLC, arising in light smokers or never-smokers, it is not exclusive to these populations and when present, it will predict benefit from targeted inhibitors irrespective of the patient demographics. 4,5 Therefore, current guidelines recommend that all nonsquamous NSCLCs, irrespective of clinical features, undergo testing for ROS1 rearrangement. 5 The U.S. Food and Drug Administration approved crizotinib for use in patients with ROS1-rearranged NSCLC in 2016 but has not identified a particular assay to serve as a companion diagnostic. In practice, FISH is often the favored approach for detection of ROS1 rearrangements because of its demonstrated utility for selecting responders to crizotinib in clinical trials, the availability of commercial probes, and the fact that many laboratories have established infrastructure and expertise for FISH-based fusion detection of other rearrangements in NSCLC (such as in ALK). FISH has its limitations, however, including its not inconsequential failure rates in real-life practice. 6 It requires that tumor nuclei be well preserved and sufficiently numerous (typically about 50 are required) to attain a confident positive or negative result based on the laboratory's validated cutpoints. Most laboratories accept only formalin-fixed paraffinembedded (FFPE) tissue blocks cut into sections of 5 mm or less, thus excluding cytologic preparations of whole tumor nuclei from analysis. As a screening assay, FISH can be costly in terms of both reagents and time, particularly when the expected rates of a positive finding are low. For all these reasons, alternative methods for detection of ROS1 rearrangement are likely to be preferred by both laboratories and clinicians.The evolution of massively parallel sequencing techniques has enabled widespread implementation of targeted panels in clinical practice to simultaneously identify a diverse set of genomic alterations in cancer samples. Clinical sequencing laboratories may pursue a comprehensive approach to detection of mutations, copy numbers, and structural variants (including rearrangements) by using hybrid capture-based technology to interrogate sample DNA; alternatively, many laboratories, particularly those with a lower samp...

show abstract

Comparison of Molecular Testing Modalities for Detection of ROS1 Rearrangements in a Cohort of Positive Patient Samples

Abstract: ROS1 rearrangement/fusion detection in the clinical setting is complex and all methodologies have inherent limitations of which users must be aware to correctly interpret results.

Cited by 141 publications

References 30 publications

Targeted fusion analysis can aid in the classification and treatment of pediatric glioma, ependymoma, and glioneuronal tumors

Targeted fusion analysis can aid in the classification and treatment of pediatric glioma, ependymoma, and glioneuronal tumors

DNA-Based versus RNA-Based Detection of MET Exon 14 Skipping Events in Lung Cancer

Recognizing the Challenges of Oncogene Fusion Detection: A Critical Step toward Optimal Selection of Lung Cancer Patients for Targeted Therapies

Contact Info

Product

Resources

About