comparison of hepatocellular carcinoma mirna expression profiling as evaluated by next generation...
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Comparison of Hepatocellular Carcinoma miRNA Expression Profiling as Evaluated by Next Generation Sequencing and Microarray
PLoS ONE 9(9): e106314. doi:10.1371/journal.pone.0106314
Yoshiki Murakami1, Toshihito Tanahashi2,3, Rina Okada4, Hidenori Toyoda5, Takashi Kumada5, Masaru Enomoto1, Akihiro Tamori1, Norifumi Kawada1, Y-h Taguchi6, Takeshi Azuma4
1 Department of Hepatology, Osaka City University Graduate School of Medicine, Osaka, Japan2 Department of Medical Pharmaceutics, Kobe Pharmaceutical University, Kobe, Japan, 3 Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan4 Division of Gastroenterology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Japan5 Department of Gastroenterology, Ogaki Municipal Hospital, Ogaki, Japan6 Department of Physics, Chuo University, Tokyo 112-8551, Japan
Purpose: How can we measure microRNA expression using MiSeq + miRDeep2?
MiSeq: Desktop sequencerMiRDeep2: easy to use pipeline for miRNA-seq
We would like to measure miRNA expression as easy & precisely as possible ….
(without any complicated or sophisticated or advanced procedures)
How accurate are those outcomes?
Materials:14 HCC (liver cancer) + 6 Normal tissue (same patients)
MiRNA-seq : 14 HCC + 6 Normal tissueMicroarray : 14 HCC
Questions:How coincident between miRNA-seq and microarray?
Can miRNA-seq diagnose HCC vs normal tissue?
Are novel miRNAs detected?
How coincident between miRNA-seq and microarray?
log10(NGS)
log1
0(m
icro
arra
y)Corr. Coef. = 0.6059
Microarray:Agilent microRNA GprocessedSignal + mean=0 + SD=1
miRDeep2:Bowtie + hg19 + miRBase 18 + default settings
Essentially, no normalization, but they are correlated very well.
Te
ch
ni c
al re
plic
ate
s
An
oth
er p
atie
nt
Independent of technical replicates / patients, miRNA-seq and microarray are well correlated without any sophisticated/complicated/advanced normalizations
Next: Consistency of differential miRNA expression between miRNA-seq and microarray
log10(NGS)
log1
0(m
icro
arra
y)K-177 vs CU-087
Corr. Coef. = 0.5555
Com
pari
sons
bet
wee
n te
chni
cal r
eplic
ates
An
oth
er p
atie
nt
Independent of technical replicates / patients, miRNA-seq and microarray of differential expression were well correlated without any sophisticated/complicated/advanced normalizations
Comparisons between technical replicates of miRNA-seq
0.986
0.984 0.986
Comparisons between technical replicates of miRNA-seq of differential expression
0.913
0.929 0.907
0.926 0.852 0.865
0.834 0.925 0.843 0.899
0.853 0.830 0.843 0.925 0.896
0.918 0.841 0.856 0.922 0.818 0.850
0.830 0.919 0.842 0.845 0.922 0.847 0.899
0.836 0.815 0.914 0.844 0.818 0.922 0.921 0.899
Independent of technical replicates / patients, reproducibilities of miRNA-seq of expression and differential expression were well without any sophisticated/complicated/advanced normalizations
MiRNA-seq expression/differential expression does not require any sophisticated/complicated/advanced normalizations, possibly because of the fact that limited number of miRNAs are generally expressive.
Can miRNA-seq diagnose HCC ?
11 miRNAs were selected using PCA based unsupervised FE
miR-10a-5p, miR-122-5p, miR-146b-5p, miR-148a-3p, miR-192-5p, miR-22-3p, miR-26a-5p, and miR-27b-3p, miR-10b-5p, miR-143-3p, and miR-21-5p
True Tumor ControlPredicted Tumor 12 0 Control 2 6
M features
N sample
Q
=+ noise
M' N✕matrix
Q N matrix✕
N(≪M)samples
M'(≪M)✕Q matrix
M features
M features embedded
PC1
PC1
M'
N(≪M)samples
Individual miRNAs are not always expressed between tumor and control
Combinatorial expression is important !
Discovery of Novel miRNAs
Conclusions
※Miseq+miRDeep2 detect miRNA expression and differential miRNA expression coincident with microarray
・Miseq+miRDeep2 give reproducible miRNA expression and differential miRNA expression
・miRNA expression measured by Miseq+miRDeep2 can diagnose between HCC and normal control
・Miseq+miRDeep2 can identify novel miRNAs