ALK gene rearrangements have been described in 3–5% of lung adenocarcinomas (ADC), and the presence of dysregulated ALK has been described in at least 18 cancer types (reviewed in 1). Since lung cancer is a common type of cancer, an estimated 70,000 cases of non-small cell lung cancer (NSCLC) which have ALK genomic alterations occur world-wide each year, making it the most common ALK-positive type of cancer (2–6). In the majority of NSCLC cases, ALK rearrangements are not found with other oncogenic mutations, such as mutations in epidermal growth factor receptor (EGFR) or KRAS (7–9). Currently, there are at least 22 known different partners that form fusion proteins with ALK (10), which in the past has posed a challenge in establishing a diagnostic method broad enough to detect all ALK fusions.
Molecular diagnostic testing for ALK is recommended to guide NSCLC therapy, and is essential to select patients for treatment with ALK tyrosine kinase inhibitors (TKIs) (11–12, Figure 1). Screening for ALK fusions in NSCLC is important, as “ALK-positive” tumors harboring a rearranged ALK gene/fusion protein are highly sensitive to ALK TKI therapies. However, these treatments should be limited to patients whose tumors contain this abnormality as detected by a sensitive and accurate diagnostic assay. For many years, fluorescent in situ hybridization (FISH) was considered the only validated diagnostic assay for detection of ALK aberrations. Yet several disadvantages to this method have been described (13–15), such as the high cost per ALK-positive patient, need for specialized expertise, long turnaround times and complex results interpretation (16). The FISH readout can be challenging, as splitting patterns can be subtle, particularly in cases with small intrachromosomal deletions and inversions. As a result, interpretation can vary significantly between laboratories.
Figure 1. Example of diagnostic testing algorithm for NSCLC patients to determine proper course of treatment. (click on image to enlarge)
Considering the limitations of ALK FISH testing, an optimal technique for large-scale screening in a clinical setting is still being determined (23) and other diagnostic assays are still being explored (24). Immunohistochemistry (IHC) is an alternative method for which commercial tests are available, and is faster and less expensive than FISH, widely adopted in pathology labs and is able to detect ALK rearrangements independent of the fusion partners. However, the sensitivity and reproducibility of this method, two critical aspects when performing ALK detection, still require optimization (17) and the suitability of this method has recently been questioned (20) due to poor concordance with FISH results. Reverse transcription-polymerase chain reaction (RT-PCR) of mRNA is a sensitive and specific alternative to FISH and IHC (18–19), and has been recommended in many studies (18, 14, 23) due to improved sensitivity, reliability, and results interpretation versus FISH and IHC. However, classical methods of RT-PCR also have disadvantages, such as requiring multiple primer sets to detect all fusion variants, and being prone to RNA degradation issues.
Awlays abreast of biomarker testing and innovation, several years ago QIAGEN identified a need for an improved method for ALK fusion testing (see webinar here). To address these challenges for ALK detection in patients, QIAGEN is preparing to launch a new CE-IVD kit for ALK fusion detection, the therascreen ALK RGQ RT-PCR Kit (therascreen ALK test), in Spring 2017. QIAGEN’s therascreen ALK test is based on the one-step RT-PCR method, and uses Scorpions technology to detect expression of RNA transcripts encoding the ALK tyrosine kinase domain. This approach ensures broad detection of all ALK fusions with just one RT-PCR assay, since the ALK tyrosine kinase domain is conserved in all ALK fusions. QIAGEN’s simplified Sample to Insight workflow enables quick turnaround time for faster results versus other standard methods, and reduced laboratory costs. Less hands-on time is required, due to the ready-to-use kit with pre-prepared solutions and one-step RT-PCR procedure. A built in control monitors and detects RNA degradation, ensuring a low failure rate. In addition, the therascreen ALK test is highly sensitive and provides precise ALK fusion detection from even low amounts of starting material, enabling patients for whom even minimal biopsy was obtained to be tested for ALK. Results interpretation for the therascreen ALK test is simplified and made objective, thanks to QIAGEN’s unique Rotor-Gene AssayManager results interpretation software. In addition, the same workflow and Rotor-Gene Q MDx 5plex HRM instrument can be used to test both ALK fusions and EGFR mutations from the same patient tissue sample when the therascreen ALK test is used together with the therascreen EGFR RGQ PCR Kit. This helps to save precious patient sample, as well as valuable lab resources.
Marchetti et al. (21) performed a technical validation to evaluate a new algorithm for quick and easy ALK fusion detection using QIAGEN’s ALK RT-PCR test to establish the ALK status in a large cohort of lung ADC patients (read the Oncology Solutions review here). The results of the technical validation study suggest that the QIAGEN’s ALK RT-PCR test could be useful in clinical practice as a complementary assay in multi-test diagnostic algorithms or even as a standalone or screening test for the selection of patients to be treated with ALK inhibitors.
In addition, Liu et al. (22) recently performed a clinical validation study of QIAGEN’s ALK RT-PCR test for detection of ALK aberrations in NSCLC, with the goal to identify an efficient and effective approach for testing genetic alterations in both EGFR and ALK from small biopsies and cytology specimens. Their study included a retrospective review of cases previously tested by FISH, which they retested using QIAGEN’s ALK RT-PCR test for identification of ALK fusions. They reported 100% sensitivity and specificity of QIAGEN’s ALK RT-PCR test to detect ALK fusions compared to FISH, and concluded that the assay allows rapid identification of ALK alterations, can be performed in conjunction with EGFR testing, and does not require use of additional valuable tumor tissue.
QIAGEN continues to expand its portfolio of innovative Sample to Insight technologies for oncology biomarker detection to help care givers more accurately and efficiently identify the right treatment for the right patient. QIAGEN’s oncology solutions include convenient biomarker testing, reliable, efficient and cost effective workflows, and secure systems with experienced services and support. Find out more about QIAGEN’s partnerships and oncology companion diagnostics solutions in the US and oncology solutions available throughout Europe.
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