Digitally guided microdissection aids somatic mutation detection in difficult to dissect tumors

Abstract

Molecular genetic testing on formalin fixed, paraffin embedded (FFPE) tumors frequently requires dissection of tumor from tissue sections mounted on glass slides. In a process referred to as “manual macrodissection,” the pathologist reviews an H&E stained slide at the light microscope and marks areas for dissection, and then the laboratory performs manual dissection from adjacent sections without the aid of a microscope, using the marked reference H&E slide as a guide. Manual macrodissection may be inadequate for tissue sections with low tumor content. We compared manual macrodissection to a new method, digitally guided microdissection, on a series of 32 FFPE pancreatic cancer samples. KRAS hotspot mutation profiling was performed using the Sequenom MassARRAY system (Agena Bioscience). Digitally guided microdissection was performed on multiple smaller areas of high tumor content selected from within the larger areas marked for manual macrodissection. The KRAS mutant allele fraction and estimated neoplastic cellularity were significantly higher in samples obtained by digitally guided microdissection (p <0.01), and 7 of the 32 samples (22%) showed a detectable mutation only with digitally guided microdissection. DNA quality and yield per cubic millimeter of dissected tissue were similar for both dissection methods. These results indicate a significant improvement in tumor content achievable with digitally guided microdissection.

Relevance of gene mutations and methylation to the growth of pancreatic intraductal papillary mucinous neoplasms based on pyrosequencing

“…The dissected material contained a variety of cell types such as lymphocytes, fibroblasts, acinar cells, besides the target IPMN epithelium. Additionally, the proportion of neoplastic content is different and low in samples 60 , even though their sample sizes are same. However, the study did not compare subtle difference of genetic or epigenetic aberrations between malignant-potential dissected regions, and just compare between malignant-potential and benign dissected regions.…”

Performance of Automated Dissection on Formalin-Fixed Paraffin-Embedded Tissue Sections for the 21-Gene Recurrence Score Assay

“… 25 For example, Geiersbach et al found that the estimated neoplastic cellularity and KRAS mutant allele fraction were significantly higher in samples obtained by mesodissection, and 7 of the 32 samples (22%) showed a detectable mutation only with this technology. 21 Gustafson et al found that the automated dissection of tumor AOIs could detect variants that would otherwise go undetected in 70% of cases, some of which, importantly, were clinically actionable mutations. 25 To date, other complex molecular tests, such as expression analysis, have not been assessed using this system.…”

Neoplastic cell percentage estimation in tissue samples for molecular oncology: recommendations from a modified Delphi study

“…Given its high cost and complexity, laser‐guided microdissection is also not recommended to be used, apart from very rare cases where small areas or individual cells are essential to be analysed. With the current rapid development in digital pathology applications, other digitally guided dissection systems could enter laboratory practice during the next decade …”