RET Fusion Testing in Patients With NSCLC: The RETING Study
Esther Conde1,2,3,4, Susana Hernandez1,3, Jose Luis Rodriguez Carrillo5
1Hospital Universitario 12 de Octubre, Madrid, Spain.
Introduction:
RET inhibitors with impressive overall response rates are now available for patients with NSCLC, yet the identification of RET fusions remains a difficult challenge. Most guidelines encourage the upfront use of next-generation sequencing (NGS), or alternatively, fluorescence in situ hybridization (FISH) or reverse transcriptase-polymerase chain reaction (RT-PCR) when NGS is not possible or available. Taken together, the suboptimal performance of single-analyte assays to detect RET fusions, although consistent with the notion of encouraging universal NGS, is currently widening some of the clinical practice gaps in the implementation of predictive biomarkers in patients with advanced NSCLC.
Methods:
This situation prompted us to evaluate several RET assays in a large multicenter cohort of RET fusion-positive NSCLC (n = 38) to obtain real-world data. In addition to RNA-based NGS (the criterion standard method), all positive specimens underwent break-apart RET FISH with two different assays and were also tested by an RT-PCR assay.
Results:
The most common RET partners were KIF5B (78.9%), followed by CCDC6 (15.8%). The two RET NGS-positive but FISH-negative samples contained a KIF5B(15)-RET(12) fusion. The three RET fusions not identified with RT-PCR were AKAP13(35)-RET(12), KIF5B(24)-RET(9) and KIF5B(24)-RET(11). All three false-negative RT-PCR cases were FISH-positive, exhibited a typical break-apart pattern, and contained a very high number of positive tumor cells with both FISH assays. Signet ring cells, psammoma bodies, and pleomorphic features were frequently observed (in 34.2%, 39.5%, and 39.5% of tumors, respectively).
Conclusions:
In-depth knowledge of the advantages and disadvantages of the different RET testing methodologies could help clinical and molecular tumor boards implement and maintain sensible algorithms for the rapid and effective detection of RET fusions in patients with NSCLC. The likelihood of RET false-negative results with both FISH and RT-PCR reinforces the need for upfront NGS in patients with NSCLC.
Insights
Accurate detection of RET fusions in non-small cell lung cancer (NSCLC) is crucial for targeted therapy. This study highlights limitations in fluorescence in situ hybridization (FISH) and reverse transcriptase-polymerase chain reaction (RT-PCR) assays, reinforcing the need for upfront next-generation sequencing (NGS).
Area of Science:
- Oncology
- Molecular Diagnostics
- Genomics
Background:
- RET inhibitors offer significant response rates in non-small cell lung cancer (NSCLC).
- Accurate identification of RET fusions, a key predictive biomarker, remains challenging.
- Current guidelines recommend next-generation sequencing (NGS) or alternative methods like fluorescence in situ hybridization (FISH) and reverse transcriptase-polymerase chain reaction (RT-PCR).
Purpose of the Study:
- To evaluate the real-world performance of various RET fusion detection assays in a multicenter cohort of NSCLC patients.
- To compare the sensitivity and specificity of NGS, FISH, and RT-PCR for identifying RET fusions.
Main Methods:
- A cohort of 38 RET fusion-positive NSCLC specimens was analyzed.
- Specimens were tested using RNA-based NGS (reference standard), two different RET FISH assays, and one RT-PCR assay.
- Common RET fusion partners and specific fusion events were identified.
Main Results:
- Next-generation sequencing (NGS) identified a broader range of RET fusions compared to FISH and RT-PCR.
- Three RET fusions were missed by RT-PCR, and two were missed by FISH, despite being positive by NGS.
- Histopathological features such as signet ring cells and psammoma bodies were frequently observed in these tumors.
Conclusions:
- Single-analyte assays like FISH and RT-PCR have limitations and can yield false-negative results for RET fusions.
- Understanding the performance characteristics of different RET testing methodologies is essential for developing effective diagnostic algorithms.
- Upfront NGS is reinforced as the preferred method for comprehensive RET fusion detection in NSCLC patients to avoid diagnostic gaps.
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