Tumour tissue sampling for lung cancer management in the era of personalised therapy: what is good enough for molecular testing?

Abstract

In the era of personalised cancer therapy, the demand for molecular profiling of the patient's tumour is steadily increasing. In advanced nonsmall cell lung cancer (NSCLC) patients, testing for epidermal growth factor receptor (EGFR) mutations and anaplastic lymphoma kinase (ALK) gene rearrangements has become an essential component of clinical practice to select patients who are most likely to benefit from EGFR and ALK tyrosine kinase inhibitors, respectively. Furthermore, obtaining tissue specimens from recurrent or metastatic tumours or from patients who develop resistance to initial effective therapies are essential for our understanding of the molecular basis of tumour progression and development of drug resistance. Therefore, the sampling of tumour tissue that is representative and is adequate in quantity and quality for pathological diagnosis and genomic profiling is crucial. In this review, we will discuss factors that should be considered in obtaining and processing biopsy specimens to enable routine molecular analysis in NSCLC patients. @ERSpublications Personalised therapy: adequate tumour tissue sampling for pathological diagnosis and molecular profiling is critical

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

“…The use of less invasive procedures results in smaller amount of content in cytology samples, which may cause false-negative results [28]. To overcome the limitations of small sample sizes, cytological rapid on-site evaluations (C-ROSE) should be applied during sampling to assess the adequateness of the biopsies [45].…”

“…Depending on the techniques utilized for sampling, tissue biopsies range from large surgical resections to small biopsies by endoscopy or by image-guided transthoracic core-needle biopsy [28].…”

“…This underlines the need for caution in distinguishing sequence artifacts from the naturally occurring T790M mutation. To minimize the adverse effects of formalin fixation in FFPE samples, the following measures should be adopted: (1) preanalysis of the tumor cell ratios to ensure adequate amounts of amplifiable DNA templates and performance of manual enrichments in cases with abundant nonspecific tumor contents; (2) fixation of samples in cold 10% neutral formalin (at 4 °C) in the shortest possible times, typically 8-18 h for large surgical resections, and 6-12 h for small biopsies [28]; (3) use of heat treatment during DNA extraction, usually at >90 °C, to remove formalin-induced cross-links and to facilitate tissue digestion with proteinase K [40]; (4) pretreatment of DNA extracted by uracil-DNA glycosylase in order to prevent modified bases [41]; (5) use of short amplicons and application of unique molecular tagging combined with bidirectional strand sequencing of DNAs to distinguish sequence artifacts from naturally occurring mutations [42]; (6) performance of amplifications of FFPE samples in duplicates to ensure accurate results [12]; and (7) utilization of specific DNA polymerases that do not read through uracil residues and abasic sites and use of high-fidelity DNA polymerases to decrease polymeraseinduced errors, when using PCR-based methods [38].…”

Improving Molecular Testing and Personalized Medicine in Non-Small-Cell Lung Cancer in Ontario

“…In addition to determining malignancy, lung origin, and pathologic subtype, pathologists evaluate diagnostic specimens to ensure that samples meet the criteria for molecular analysis. Whether cytology or biopsy specimens are obtained, tumour cellularity is the key determinant of the likelihood of successful molecular testing [45][46][47][48] ; both techniques can potentially yield adequate material for diagnostic molecular tests 49 . With respect to cytology specimens, preparation of a cell block is still preferred 24,50 , but other specimens are also suitable for analysis 27 .…”

Ready or Not? Lung Cancer Diagnosis in 2015

“…Bronchoalveolar lavage and bronchial brushings offer even poorer yields, with diagnostic sensitivity of 43% and 54% respectively. Those poor yields pose major challenges in the diagnosis of lung cancer, because, on top of necessary material for diagnosis, standard molecular testing currently requires 100-300 cells to perform EGFR genotyping, plus additional material to test for ALK rearrangement and to perform relevant ihc staining for subtyping 22,23 .…”