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German Team Debuts ALK Testing Method Similar to Pfizer Team's but with Potential for Broader Appeal

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A group of researchers from Germany's Robert Bosch Hospital and other associated institutions have described a PCR-based method they have developed to detect ALK fusions that takes advantage of some aspects of a method published earlier this year by Pfizer researchers, but which they say could be more readily adoptable by a wider variety of testing labs.

The team published an explanation of the method and reported on its performance on formalin fixed tissue samples in a study in the Journal of Thoracic Oncology this month.

Like a method published last month by researchers from Pfizer and South Korea's Samsung Medical Center, the German team's method differs from other PCR-based ALK testing approaches, which target specific ALK fusion transcripts, and instead relies on measurement of overall differences in expression of the 3' end and 5' end of the ALK encoding kinase domain to indicate the presence of ALK fusions.

But according to the German group, by developing the method using quantitative RT-PCR, rather than the NanoString platform used by the Pfizer team, they have made their technique more readily adoptable by diagnostic labs, the vast majority of which are accustomed to standard PCR techniques, but do not have NanoString or other specialized commercial instruments.

Claudia Kalla, the study's senior author, told PGx Reporter in an email that the group was impressed with the results the method was able to generate in its initial study, and is moving on to a follow-up project to test the method prospectively.

Based on the success of this prospective study, the team will decide whether to adopt the method for clinical ALK testing, she wrote.

Kalla's group conducts ALK testing as members of The Robert Bosch Hospital's department of clinical pathology. Prior to developing this method she said the team has used breakapart fluorescent in situ hybridization probes to diagnose ALK rearrangements.

In the US, the only test for detecting ALK fusions that has been approved by the US Food and Drug Administration is Abbott's breakapart FISH assay, which is used to help determine whether patients are likely to respond to Pfizers ALK inhibitor Xalkori (crizotinib).

However, issues have been raised in regard to the suitability of FISH as a method for widespread screening of the NSCLC population to identify those with ALK, or other mutations. For instance, a 2012 study by University of Colorado researchers that found that broadly testing all advanced NSCLC patients in order to identify the small subset of ALK-positive individuals who should be treated with Xalkori did not meet a cost-effectiveness bar of less than $100,000 per quality-adjusted life year gained. In addition, other researchers from the University of Colorado and Tel Aviv University have published research suggesting that FISH may miss some rare ALK fusions that don't fit the FDA-approved test's inclusion criteria.

"Usually, breakapart FISH probes [work] very well to detect specific translocations … however, FISH analysis of ALK rearrangements is unusually complicated," Kalla wrote.

"In search for a reliable and easy-to-perform approach we were thinking about the phenomenon of unbalanced ALK transcript expression as a result of ALK translocation and decided to develop a respective method. Our main object was to develop a method that can cope with poor-quality RNA isolated from routine specimens" such as formalin-fixed paraffin-embedded tumor samples, she explained.

The resulting achievement was an assay relying on qRT-PCR and utilizing small amplicons designed to overcome the technical problems associated with amplifying FFPE RNA.

Instead of targeting known and specific ALK fusions, the group's method relies on measuring unbalanced expression of the 3' and 5' ends of the ALK gene. According to Kalla, one advantage of this is that it allows the assay to detect rearrangements independently of what the fusion partner is.

Additionally, the method allows the detection of full-length ALK transcripts. Although the role of these transcripts in lung cancer has not been established, it plays a role in other disease like neuroblastoma and sarcomas, she wrote.

In the study, Kalla and her colleagues tested the method on 652 non-small cell lung cancer specimens, comparing the results to FISH and immunohistochemistry measurements.

According to the authors, their method accurately typed 97 percent of 182 tumors compared to FISH and IHC results, and also suggested the presence of rearrangements in three tumor samples that had insufficient or ambiguous FISH results.

Kalla said the team estimates that the cost of the group's method should be at least half that of FISH.

Abbott Molecular has previously told PGx Reporter that Medicare reimbursement for manually running its ALK test ranges from $329 per test to $571 per test, with average reimbursement at $440 per test.

Moreover, Kalla wrote, "besides saving material costs and time, the strong point of the qRT-PCR technique is that it opens up the possibility of optimized tissue processing: Using the extraction method mentioned in the manuscript both RNA and DNA can be isolated simultaneously from the same tumor cells, thus expression analysis can be combined with DNA mutation analysis … which allows for a comprehensive analysis of even small samples."

Kalla said the researchers have not had discussions about the commercial potential of the method, but if the results of their follow up prospective analyses hold up, they would potentially adopt the method for their own clinical testing purposes.

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