Performance of Idylla KRAS assay on extracted DNA and de-stained cytology smears: Can we rescue small sample?

In the era of precision medicine, molecular profiling of tumors provides important prognostic and predictive information to guide therapeutic decisions. A large fraction of patients with solid tumor malignancies presents with advanced-stage disease, in which cytology specimens or small biopsies are often the only source of material available for molecular analysis, such as next generation sequencing (NGS) testing. This problem is especially prominent in lung cancer patients, approximately 70 % of whom are diagnosed with advanced, stage IV disease [1]. In many of these patients, cellblocks (CB) are a convenient material for molecular testing due to their similarity to formalin-fixed, paraffin-embedded (FFPE) tissue blocks. However, the quantity not sufficient (QNS) rate of CB for molecular testing varies, with reports up to 64 % depending on the institution [2], [3]. Thus, there is a growing interest in validating cytology specimens other than CB for routine molecular testing. Studies have shown that direct smears, residual liquid-based cytology solution, and supernatant from fine needle aspiration can yield substantial amounts of DNA for mutation analysis on different platforms, including the Biocartis Idylla, which is a fully automated PCR system [4], [5], [6], [7], [8].

Molecular testing for advanced stage cancer is restricted not only by scant tissue specimens, but also by the lack of suitable of testing platforms within institutions. Many small community hospitals lack the infrastructure to implement and support NGS testing. In these settings, the Biocartis Idylla system provides a relatively simple option to test key, actionable biomarkers, such as EGFR, KRAS, BRAF, NRAS, MSI, and gene fusion events while minimizing the technical skills and requirements needed for NGS testing.

The Idylla system is fully automated and designed for FFPE tissue sections. As designed by the manufacturer, Idylla analyzes 5 μm to 10 μm thick FFPE tissue sections ranging from 25 mm2 to 50 mm2 and containing at least 10 % neoplastic nuclei.

Even in labs equipped with NGS testing, the Idylla system is often used to “rescue” test for cases with insufficient DNA for NGS study [9]. Depending on the NGS methodology and size of the gene panel, NGS may require up to 100 ng of DNA, whereas Idylla requires only a single FFPE tissue section yielding <10 ng of DNA [10], [11], [12]. Additionally, the Idylla system provides 2–3-h turnaround times making it very useful when results are needed urgently, compared to 7–21 days for NGS. The Idylla system therefore complements NGS testing to optimize a laboratory's overall molecular testing workflow [7].

Although Idylla requires less DNA than NGS, the manufacturer has not defined the minimum quantity of DNA from sources other than FFPE. Previous research focusing on EGFR showed that molecular testing with the Idylla system was feasible using small amounts of extracted DNA and cytology smears [7], [13]. The present study aims to assess the performance of Idylla for KRAS using scant amounts of DNA and cytology smears. KRAS is one of the most frequently mutated genes in pancreatic (95 %), colorectal (45 %) and lung (35 %) adenocarcinomas. Furthermore, KRAS G12C occurs frequently in lung adenocarcinomas (13 %), colorectal cancer (1–3 %), and other cancers [14], [15], [16]. Based on recent clinical trials of KRAS G12C targeted inhibition in patients with advanced disease [17], the FDA has approved sotorasib in KRAS G12C NSCLC. To expand therapeutic opportunities for these types of patients, we evaluated the Idylla KRAS assay using either purified FFPE DNA or tissue obtained directly from the cytology smears in order to rescue QNS samples for NGS testing.

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