Multiplatform comparison of molecular oncology tests performed on cytology specimens and formalin‐fixed, paraffin‐embedded tissue

Abstract

BACKGROUND: Molecular oncology testing is important for patient management, and requests for the molecular analysis of cytology specimens are increasingly being made. Formalin-fixed, paraffin-embedded (FFPE) cell blocks of such specimens have been routinely used for molecular diagnosis. However, the inability to assess cellularity before cell block preparation is a pitfall of their use. In this study, various cytologic preparations were tested with several molecular test platforms, and the results were compared with paired FFPE tissue. METHODS: Seventy-seven cytology cases, including fine-needle aspiration smears, touch preparations, and SurePath thin-layer preparations, were selected from the archives.Areas of interest were removed from the slide with a matrix capture solution. DNA extracted from the cells was evaluated by mutation analysis for BRAF, epidermal growth factor receptor (EGFR), RAS, and a 50-gene panel with various testing platforms (single-nucleotide primer extension assay, Sanger sequencing, and next-generation sequencing). Thirty-eight tumors with available FFPE tissue were tested in parallel. RESULTS: The average DNA concentration was 299 ng/mL for the cytology specimens and 171 ng/mg for the paired FFPE tissue. Point mutations and large deletions were detected in BRAF, KRAS, NRAS, HRAS, and EGFR genes. In comparison with FFPE tissue, 5-to 8-fold less input DNA was needed when cytology preparations were used. The concordance between cytology specimens and FFPE tissue was 100%. CON-CLUSIONS: Cytologic preparations were found to be a reliable source for molecular oncology testing. DNA derived from cytology specimens performed well on multiple platforms, and 100% concordance was observed between cytology specimens and FFPE tissue. Cancer (Cancer Cytopathol) 2015;123:30-9.

Molecular Testing of Non–Small Cell Lung Carcinoma Diagnosed by Endobronchial Ultrasound–Guided Transbronchial Fine-Needle Aspiration: The Cleveland Clinic Experience

“…Given the diversity of cytopathologic specimen types and the different methods of fixation and processing available, options for procuring DNA from cytologic specimens for molecular analysis vary; material can be successfully obtained from cell blocks, smears, cytospins, and liquid-based preparations. 7,9,11,12,14,[19][20][21][22][23][24][25] Most commonly, cell blocks are used for molecular testing on cytopathologic specimens. 21 The cell block closely approximates surgical pathology material; tissue fragments from cytopathologic specimens are embedded in paraffin and are sectioned akin to surgical specimens.…”

“…The successful use of smeared slides to acquire DNA for EGFR testing has been described in several publications. 9,11,28 Smears are decoverslipped, and tumor cells are removed from areas of the slide with a high tumor fraction by scraping with a scalpel or dissecting needle.…”

“…Cytology specimens provide highquality DNA for molecular testing 7,8 and are comparable to, if not superior to, formalin-fixed, paraffin-embedded surgical pathology specimens, for the evaluation of clinically relevant mutations. [8][9][10][11] Although cell blocks are frequently used for molecular testing of non-small cell lung carcinomas, at our institution they were suboptimal for analysis in a high proportion of cases. An alternate strategy for EGFR and ALK testing was therefore developed, using material from the liquid-based cytology sample for EGFR mutation testing by nextgeneration sequencing (NGS), and the ThinPrep slide for fluorescence in situ hybridization (FISH) testing for ALK gene rearrangements.…”

Techniques for maximizing the performance of molecular pathology testing: responsibilities of all pathologists

“…Cytological materials that were obtained with easily reproducible and comfortable methods constitute a valuable resource for diagnosis and treatment studies in routine pathological process. Studies have shown that cytological specimens used in molecular studies show up to 100% compatibility with histological specimens (12). Especially in the majority of non-small cell lung carcinomas, the diagnosis is provided with cytological material and small biopsies.…”

Sample types applied for molecular diagnosis of therapeutic management of advanced non-small cell lung cancer in the precision medicine

“…According to statistics, the diagnosis and molecular profiling of 25%-30% NSCLCs are made by small biopsies, and up to 40% by cytology biopsies [15]. The success rates of both small and cytology biopsies range between 83% and 100% in published studies [16][17][18][19][20][21]. It is known that 90% of patients with early stage NSCLCs who receive surgical resections or have locally advanced NSCLCs and receive definitive chemoradiation, eventually relapse [22].…”