Possibility of Cell Block Specimens from Overnight-Stored Bile for Next-Generation Sequencing of Cholangiocarcinoma.

The identification of anticancer therapies using next-generation sequencing (NGS) is necessary for the treatment of cholangiocarcinoma. NGS can be easily performed when cell blocks (CB) are obtained from bile stored overnight. We compared NGS results of paired CB and surgically resected specimens (SRS) from the same cholangiocarcinoma cases. Of the prospectively collected 64 bile CBs from 2018 to 2023, NGS was performed for three cases of cholangiocarcinoma that could be compared with the SRS results. The median numbers of DNA and RNA reads were 95,077,806 [CB] vs. 93,161,788 [SRS] and 22,101,328 [CB] vs. 24,806,180 [SRS], respectively. We evaluated 588 genes and found that almost all genetic alterations were attributed to single-nucleotide variants, insertions/deletions, and multi-nucleotide variants. The coverage rate of variants in SRS by those found in CB was 97.9-99.2%, and the coverage rate of SRS genes by CB genes was 99.6-99.7%. The NGS results of CB fully covered the variants and genetic alterations observed in paired SRS samples. As bile CB is easy to prepare in general hospitals, our results suggest the potential use of bile CB as a novel method for NGS-based evaluation of cholangiocarcinoma.

Formalin-fixed paraffin-embedded sample conditions for deep next generation sequencing.

INTRODUCTION: Precision medicine is only possible in oncology practice if targetable genes in fragmented DNA, such as DNA from formalin-fixed paraffin-embedded (FFPE) samples, can be sequenced using next generation sequencing (NGS). The aim of this study was to examine the quality and quantity of DNA from FFPE cancerous tissue samples from surgically resected and biopsy specimens. METHODS: DNA was extracted from unstained FFPE tissue sections prepared from surgically resected specimens of breast, colorectal and gastric cancer, and biopsy specimens of breast cancer. A total quantity of DNA ≥60 ng from a sample was considered adequate for NGS. The DNA quality was assessed by Q-ratios, with a Q-ratio >0.1 considered sufficient for NGS. RESULTS: The Q-ratio for DNA from FFPE tissue processed with neutral-buffered formalin was significantly better than that processed with unbuffered formalin. All Q-ratios for DNA from breast, colorectal and gastric cancer samples indicated DNA levels sufficient for NGS. DNA extracted from gastric cancer FFPE samples prepared within the last 7 years is suitable for NGS analysis, whereas those older than 7 years may not be suitable. Our data suggested that adequate amounts of DNA can be extracted from FFPE samples, not only of surgically resected tissue but also of biopsy specimens. CONCLUSIONS: The type of formalin used for fixation and the time since FFPE sample preparation affect DNA quality. Sufficient amounts of DNA can be extracted from FFPE samples of both surgically resected and biopsy tissue, thus expanding the potential diagnostic uses of NGS in a clinical setting.

Plastic-Embedded, Surgically Resected Specimens for Teaching Surgery / 医学教育

I have used plastic-embedded, surgically resected specimens to teach surgery. These specimens were obtained from more than 100 cases of important surgical diseases, such as gastric cancer, goiter, and inflammatory diseases. They are more easily carried than are formalin-preserved specimens. After I teach my students each disease, I show the corresponding specimen, explain the pathologic changes, and discuss the surgical procedure with which they were obtained. I then let the students handle and examine the specimens. Observation of actual specimens will help students clearly understand diseases and give stronger impressions than can words, illustrations, or photographs.