Formalin fixation on HER-2 and PD-L1 expression in gastric cancer: A pilot analysis using the same surgical specimens with different fixation times.
Author(s): Kai K, Yoda Y, Kawaguchi A, Minesaki A, Iwasaki H, Aishima S, Noshiro H
Publication: World J Clin Cases, 2019, Vol. 7, Page 419-430
PubMed ID: 30842953 PubMed Review Paper? No
Purpose of Paper
This paper sought to identify potential differences in programmed death-ligand 1 (PD-L1) and human epidermal growth factor receptor 2 (HER-2) immunohistochemical staining among gastric tumor specimens that were fixed in 10% formalin or 10% neutral buffered formalin (NBF) for 6 h, 24 h, 48 h, 1 week, or >3 months. HER-2 dual ISH for copy number determination was also performed in a subset of HER-2 immunopositive specimens.
Conclusion of Paper
No significant differences in the percentage of immunopositive cases were observed among gastric tumor specimens fixed in 10% formalin or 10% NBF and stained for HER-2 (81.8 vs 60%) or PD-L1 (63.4 vs. 60%). Effects associated with time in formalin were limited to a loss of PD-L1 staining among the majority of specimens fixed for 3-28 months, as no differences were present for PD-L1 or HER-2 (by either IHC or DISH) among specimens fixed for 6, 24, or 48 h, irrespective of the threshold of positivity. When HER-2 immunohistochemistry and DISH results were compared, two out of the five immunopositive cases displayed discrepancies among some of the fixation timepoints, which the authors postulated were the result of tissue heterogeneity as no trend was apparent.
Studies
-
Study Purpose
This study explored potential differences in PD-L1 and HER-2 immunohistochemical staining among 32 gastric tumor specimens procured by gastrectomy that were divided and fixed for 6 h, 24 h, 48 h, 1 week, or >3 months in either 10% formalin (22 cases) or 10% NBF (10 cases). Formalin-fixed paraffin-embedded (FFPE) sections (3 µm) from each block were stained for HER-2 with an autostainer and the HercepTest II Kit. DISH analysis of a subset of HER-2 positive cases was performed with a Ventana INFORM Dual ISH HER-2 Kit and presented as HER-2/CEP17 ratio. PD-L1 immunohistochemical analysis was performed on tissue microarrays assembled with 2.0 mm cores and stained using an autostainer and the PD-L1 IHC 22C3 pharmDx Kit. Scoring was completed by two pathologists based on the percentage of immunopositive cells for PD-L1 and the 0, 1+, 2+, 3+ scoring system for HER-2.
Summary of Findings:
Gastric tumor specimens fixed in 10% formalin or 10% NBF did not differ significantly in the percentage of HER-2 positive (81.8 vs 60%) or PD-L1 positive (63.4 vs. 60%) cases. No significant differences in immunostaining for HER-2 or PD-L1 were observed among specimens that were fixed for 6, 24, or 48 h compared to specimens fixed for 1 week in either 10% formalin or 10% NBF. Adjusting the threshold of positivity for PD-L1 to 1%, 10%, or 50% failed to reveal any significant differences among gastric tumor specimens fixed for 6 h to 1 week. Using a PD-L1 cut-off of 50%, immunopositive staining was lost when a subset of specimens was fixed for 3-28 months in either 10% formalin or 10% NBF in four of the five cases examined.
Similarly, no significant difference was observed in HER-2/CEP17 ratio determined by DISH among specimens fixed for 6, 24, or 48 h compared to those fixed for 1 week for either fixative. When HER-2 immunohistochemistry and DISH results were compared, two out of the five immunopositive cases displayed discrepancies among some of the fixation timepoints, which the authors postulated were the result of tissue heterogeneity as no trend was apparent.
Biospecimens
Preservative Types
- Formalin
Diagnoses:
- Neoplastic - Carcinoma
Platform:
Analyte Technology Platform Protein Immunohistochemistry DNA In situ hybridization Protein Tissue microarray Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Biospecimen Preservation Time in fixative 6 h
24 h
48 h
1 week
3-28 months
Biospecimen Preservation Type of fixation/preservation Formalin (buffered)
Formalin (unbuffered)
In situ hybridization Specific Targeted nucleic acid HER-2/CEP17
Tissue microarray Specific Targeted peptide/protein PD-L1
Immunohistochemistry Specific Targeted peptide/protein PD-L1
HER-2