High-Throughput Multiplex Immunohistochemical Imaging of the Tumor and Its Microenvironment
0
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
Purpose The aim of this study was to develop a formalin-fixed paraffin-embedded (FFPE) tissue based multiplex immunochemistry (mIHC) method for high-throughput comprehensive tissue imaging and demonstrate its feasibility, validity, and usefulness. Materials and Methods The mIHC protocol was developed and tested on tissue microarray slides made from archived gastric cancer (GC) tissue samples. On a single FFPE slide, cyclic immunochemistry for multiple markers of immune cells and cytokeratin for tumor cells was performed; hematoxylin staining was used for demarcation of nuclei. Whole slides were digitally scanned after each cycle. For interpretation of mIHC results, we performed computer-assisted image analysis using publicly available software. Results Using mIHC, we were able to characterize the tumor microenvironment (TME) of GCs with accurate visualization of various immune cells harboring complex immunophenotypes. Spatial information regarding intratumoral and peritumoral TME could be demonstrated by digital segmentation of image guided by cytokeratin staining results. We further extended the application of mIHC by showing that subcellular localization of molecules can be achieved by image analysis of mIHC results. Conclusion We developed a robust method for high-throughput multiplex imaging of FFPE tissue slides. The feasibility and adaptability of mIHC suggest that it is an efficient method for in situ single- cell characterization and analysis.
Abstract
Purpose The aim of this study was to develop a formalin-fixed paraffin-embedded (FFPE) tissue based multiplex immunochemistry (mIHC) method for high-throughput comprehensive tissue imaging and demonstrate its feasibility, validity, and usefulness. Materials and Methods The mIHC protocol was developed and tested on tissue microarray slides made from archived gastric cancer (GC) tissue samples. On a single FFPE slide, cyclic immunochemistry for multiple markers of immune cells and cytokeratin for tumor cells was performed; hematoxylin staining was used for demarcation of nuclei. Whole slides were digitally scanned after each cycle. For interpretation of mIHC results, we performed computer-assisted image analysis using publicly available software. Results Using mIHC, we were able to characterize the tumor microenvironment (TME) of GCs with accurate visualization of various immune cells harboring complex immunophenotypes. Spatial information regarding intratumoral and peritumoral TME could be demonstrated by digital segmentation of image guided by cytokeratin staining results. We further extended the application of mIHC by showing that subcellular localization of molecules can be achieved by image analysis of mIHC results. Conclusion We developed a robust method for high-throughput multiplex imaging of FFPE tissue slides. The feasibility and adaptability of mIHC suggest that it is an efficient method for in situ single- cell characterization and analysis.
참고문헌 (26)
- Hanahan D, "Hallmarks of cancer : the next generation", Cell, 144, pp.646~674 (2011)
- Hodi FS, "Improved survival with ipilimumab in patients with metastatic melanoma", N Engl J Med, 363, pp.711~723 (2010)
- Ansell SM, "PD-1 blockade with nivolumab in relapsed or refractory Hodgkin's lymphoma", N Engl J Med, 372, pp.311~319 (2015)
- Garon EB, "Pembrolizumab for the treatment of non-small-cell lung cancer", N Engl J Med, 372, pp.2018~2028 (2015)
- Mlecnik B, "Comprehensive intrametastatic immune quantification and major impact of immunoscore on survival", J Natl Cancer Inst, 110, pp.97~108 (2018)
- Tsujikawa T, "Quantitative multiplex immunohistochemistry reveals myeloid-inflamed tumor-immune complexity associated with poor prognosis", Cell Rep, 19, pp.203~217 (2017)
- Giesen C, "Highly multiplexed imaging of tumor tissues with subcellular resolution by mass cytometry", Nat Methods, 11, pp.417~422 (2014)
- Jerby-Arnon L, "A cancer cell program promotes T cell exclusion and resistance to checkpoint blockade", Cell, 175, pp.984~997.e24 (2018)
- Kyu Sang Lee, "Stromal Expression of MicroRNA-21 in Advanced Colorectal Cancer Patients with Distant Metastases", Journal of Pathology and Translational Medicine, 50(4), pp.270~277 (2016), 대한병리학회[DOI]
- Jones TR, "CellProfiler Analyst : data exploration and analysis software for complex image-based screens", BMC Bioinformatics, 9, pp.482~ (2008)
- Amir el AD, "viSNE enables visualization of high dimensional single-cell data and reveals phenotypic heterogeneity of leukemia", Nat Biotechnol, 31, pp.545~552 (2013)
- Umar A, "Revised Bethesda guidelines for hereditary nonpolyposis colorectal cancer(Lynch syndrome)and microsatellite instability", J Natl Cancer Inst, 96, pp.261~268 (2004)
- Natsume A, "Chromatin regulator PRC2 is a key regulator of epigenetic plasticity in glioblastoma", Cancer Res, 73, pp.4559~4570 (2013)
- Su IH, "Polycomb group protein ezh2 controls actin polymerization and cell signaling", Cell, 121, pp.425~436 (2005)
- Bataille F, "Multiparameter immunofluorescence on paraffin-embedded tissue sections", Appl Immunohistochem Mol Morphol, 14, pp.225~228 (2006)
- Viegas MS, "An improved and cost-effective methodology for the reduction of autofluorescence in direct immunofluorescence studies on formalinfixed paraffin-embedded tissues", Eur J Histochem, 51, pp.59~66 (2007)
- Robertson D, "Multiple immunofluorescence labelling of formalin-fixed paraffinembedded(FFPE)tissue", BMC Cell Biol, 9, pp.13~ (2008)
- Cancer Genome Atlas Research Network, "Comprehensive molecular characterization of gastric adenocarcinoma", Nature, 513, pp.202~209 (2014)
- Sotiriou C, "Breast cancer classification and prognosis based on gene expression profiles from a population-based study", Proc Natl Acad Sci U S A, 100, pp.10393~10398 (2003)
- Qiu P, "Extracting a cellular hierarchy from high-dimensional cytometry data with SPADE", Nat Biotechnol, 29, pp.886~891 (2011)
- Bruggner RV, "Automated identification of stratifying signatures in cellular subpopulations", Proc Natl Acad Sci U S A, 111, pp.E2770~7 (2014)
- Taylor CR, "Quantification of immunohistochemistry : issues concerning methods, utility and semiquantitative assessment II", Histopathology, 49, pp.411~424 (2006)
- Mengel M, "Inter-laboratory and inter-observer reproducibility of immunohistochemical assessment of the Ki-67 labelling index in a large multi-centre trial", J Pathol, 198, pp.292~299 (2002)
- Howat WJ, "Antibody validation of immunohistochemistry for biomarker discovery : recommendations of a consortium of academic and pharmaceutical based histopathology researchers", Methods, 70, pp.34~38 (2014)
- O'Hurley G, "Garbage in, garbage out : a critical evaluation of strategies used for validation of immunohistochemical biomarkers", Mol Oncol, 8, pp.783~798 (2014)
- Remark R, "In-depth tissue profiling using multiplexed immunohistochemical consecutive staining on single slide", Sci Immunol., 1, pp.aaf6925~ (2016)