Transcriptome profiling of archived sectioned formalin-fixed paraffin-embedded (AS-FFPE) tissue for disease classification.
BACKGROUND
Archived tissues from previously completed prospective trials represent invaluable resource for biomarker development. However, such specimens are often stored as sections on glass slides, in which RNA is severely degraded due to prolonged air exposure. We evaluated whether a proportion of archived sectioned formalin-fixed paraffin-embedded (AS-FFPE) tissues yield transcriptome profiles comparable to freshly cut (FC) FFPE tissues, which can be used for retrospective class prediction analysis.


METHODS
Genome-wide transcriptome profiles of 6 to 7-year-old AS-FFPE tissue sections (generated from 5 to 16-year-old blocks) of 83 hepatocellular carcinoma (HCC) and 47 liver cirrhosis samples were generated by using whole-genome DASL assay (Illumina) and digital transcript counting (nCounter) assay (NanoString), and gene signature-based prediction of HCC subclasses and prognosis was compared with previously generated FC-FFPE profiles from the same tissue blocks.


RESULTS
RNA quality and assay reproducibility of AS-FFPE RNA were comparable to intermediate to poor quality FC-FFPE samples (RNA Integrity Number: up to 2.50, R-square for technical replicates: up to 0.93). Analyzable transcriptome profiles were obtained in 64 (77%) HCC and 36 (77%) cirrhosis samples. Statistically more confident predictions based on random resampling-based method (nearest template prediction) were obtained in 37 (58%) HCC and 13 (36%) cirrhosis samples. Predictions made in FC-FFPE profiles were reproduced in 36 (97%) HCC and 11 (85%) cirrhosis AS-FFPE profiles. nCounter assay was tested in 24 cirrhosis samples, which yielded confident prediction in 15 samples (63%), of which 10 samples (67%) showed concordant predictions with FC-FFPE profiles.


CONCLUSIONS
AS-FFPE tissues yielded poorer quality RNA and transcriptome profiles compared to FC-FFPE tissues. Statistically more confident class prediction was feasible in 37 of 83 HCC samples and 13 of 47 cirrhosis samples. These results suggest that AS-FFPE tissues can be regarded as a resource for retrospective transcriptome-based class prediction analysis when they are the only available materials.
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Kensuke
K
Kojima
Illumina, Inc., San Diego, California, United States of America.
Craig
C
April
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Claudia
C
Canasto-Chibuque
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Xintong
X
Chen
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Manjeet
M
Deshmukh
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Anu
A
Venkatesh
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Poh Seng
PS
Tan
Department of Hepatology, Toranomon Hospital, Tokyo, Japan.
Masahiro
M
Kobayashi
Department of Hepatology, Toranomon Hospital, Tokyo, Japan.
Hiromitsu
H
Kumada
Illumina, Inc., San Diego, California, United States of America.
Jian-Bing
JB
Fan
Liver Cancer Program, Tisch Cancer Institute, Division of Liver Diseases, Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, United States of America.
Yujin
Y
Hoshida
D006528:Carcinoma, Hepatocellular / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism; D005404:Fixatives / Q000737:chemistry; D005557:Formaldehyde / Q000737:chemistry; D020869:Gene Expression Profiling; D006801:Humans; D008103:Liver Cirrhosis / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism; D008113:Liver Neoplasms / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism; D016612:Paraffin Embedding; D011379:Prognosis; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016707:Tissue Fixation; D059467:Transcriptome
D005404:Fixatives; D012333:RNA, Messenger; D005557:Formaldehyde