Correlation between miRseq expression and clinical features
Overview
Introduction

This pipeline uses various statistical tests to identify miRs whose expression levels correlated to selected clinical features.

Summary

Testing the association between 564 genes and 5 clinical features across 442 samples, statistically thresholded by Q value < 0.05, 4 clinical features related to at least one genes.

  • 4 genes correlated to 'Time to Death'.

    • HSA-LET-7G ,  HSA-LET-7B ,  HSA-MIR-628 ,  HSA-MIR-30E

  • 54 genes correlated to 'AGE'.

    • HSA-MIR-424 ,  HSA-MIR-1247 ,  HSA-MIR-337 ,  HSA-MIR-199A-1 ,  HSA-MIR-214 ,  ...

  • 88 genes correlated to 'HISTOLOGICAL.TYPE'.

    • HSA-MIR-9-1 ,  HSA-MIR-9-2 ,  HSA-MIR-34A ,  HSA-MIR-934 ,  HSA-MIR-375 ,  ...

  • 6 genes correlated to 'RADIATIONS.RADIATION.REGIMENINDICATION'.

    • HSA-MIR-128-2 ,  HSA-MIR-3613 ,  HSA-MIR-128-1 ,  HSA-MIR-628 ,  HSA-MIR-181D ,  ...

  • No genes correlated to 'COMPLETENESS.OF.RESECTION'

Results
Overview of the results

Complete statistical result table is provided in Supplement Table 1

Table 1.  Get Full Table This table shows the clinical features, statistical methods used, and the number of genes that are significantly associated with each clinical feature at Q value < 0.05.

Clinical feature Statistical test Significant genes Associated with                 Associated with
Time to Death Cox regression test N=4 shorter survival N=0 longer survival N=4
AGE Spearman correlation test N=54 older N=7 younger N=47
HISTOLOGICAL TYPE ANOVA test N=88        
RADIATIONS RADIATION REGIMENINDICATION t test N=6 yes N=4 no N=2
COMPLETENESS OF RESECTION ANOVA test   N=0        
Clinical variable #1: 'Time to Death'

4 genes related to 'Time to Death'.

Table S1.  Basic characteristics of clinical feature: 'Time to Death'

Time to Death Duration (Months) 0-187.1 (median=15.8)
  censored N = 398
  death N = 39
     
  Significant markers N = 4
  associated with shorter survival 0
  associated with longer survival 4
List of 4 genes significantly associated with 'Time to Death' by Cox regression test

Table S2.  Get Full Table List of 4 genes significantly associated with 'Time to Death' by Cox regression test

HazardRatio Wald_P Q C_index
HSA-LET-7G 0.45 1.275e-05 0.0072 0.291
HSA-LET-7B 0.53 2.604e-05 0.015 0.255
HSA-MIR-628 0.64 4.181e-05 0.023 0.293
HSA-MIR-30E 0.34 6.303e-05 0.035 0.308

Figure S1.  Get High-res Image As an example, this figure shows the association of HSA-LET-7G to 'Time to Death'. four curves present the cumulative survival rates of 4 quartile subsets of patients. P value = 1.27e-05 with univariate Cox regression analysis using continuous log-2 expression values.

Clinical variable #2: 'AGE'

54 genes related to 'AGE'.

Table S3.  Basic characteristics of clinical feature: 'AGE'

AGE Mean (SD) 63.48 (11)
  Significant markers N = 54
  pos. correlated 7
  neg. correlated 47
List of top 10 genes significantly correlated to 'AGE' by Spearman correlation test

Table S4.  Get Full Table List of top 10 genes significantly correlated to 'AGE' by Spearman correlation test

SpearmanCorr corrP Q
HSA-MIR-424 -0.3188 7.082e-12 3.99e-09
HSA-MIR-1247 -0.2843 1.374e-09 7.74e-07
HSA-MIR-337 -0.2722 6.254e-09 3.51e-06
HSA-MIR-199A-1 -0.2647 1.657e-08 9.3e-06
HSA-MIR-214 -0.2622 2.368e-08 1.33e-05
HSA-MIR-199A-2 -0.2617 2.435e-08 1.36e-05
HSA-MIR-409 -0.2616 2.484e-08 1.39e-05
HSA-MIR-34A -0.2605 2.842e-08 1.58e-05
HSA-MIR-493 -0.2591 3.922e-08 2.18e-05
HSA-MIR-199B -0.2552 5.498e-08 3.05e-05

Figure S2.  Get High-res Image As an example, this figure shows the association of HSA-MIR-424 to 'AGE'. P value = 7.08e-12 with Spearman correlation analysis. The straight line presents the best linear regression.

Clinical variable #3: 'HISTOLOGICAL.TYPE'

88 genes related to 'HISTOLOGICAL.TYPE'.

Table S5.  Basic characteristics of clinical feature: 'HISTOLOGICAL.TYPE'

HISTOLOGICAL.TYPE Labels N
  ENDOMETRIOID ENDOMETRIAL ADENOCARCINOMA 349
  MIXED SEROUS AND ENDOMETRIOID 18
  SEROUS ENDOMETRIAL ADENOCARCINOMA 75
     
  Significant markers N = 88
List of top 10 genes differentially expressed by 'HISTOLOGICAL.TYPE'

Table S6.  Get Full Table List of top 10 genes differentially expressed by 'HISTOLOGICAL.TYPE'

ANOVA_P Q
HSA-MIR-9-1 3.585e-26 2.02e-23
HSA-MIR-9-2 4.153e-26 2.34e-23
HSA-MIR-34A 4.13e-24 2.32e-21
HSA-MIR-934 1.191e-21 6.68e-19
HSA-MIR-375 5.966e-19 3.34e-16
HSA-MIR-195 1.927e-18 1.08e-15
HSA-MIR-497 3.165e-18 1.77e-15
HSA-MIR-548V 6.869e-17 3.83e-14
HSA-MIR-452 3.477e-16 1.93e-13
HSA-MIR-190B 2.88e-15 1.6e-12

Figure S3.  Get High-res Image As an example, this figure shows the association of HSA-MIR-9-1 to 'HISTOLOGICAL.TYPE'. P value = 3.58e-26 with ANOVA analysis.

Clinical variable #4: 'RADIATIONS.RADIATION.REGIMENINDICATION'

6 genes related to 'RADIATIONS.RADIATION.REGIMENINDICATION'.

Table S7.  Basic characteristics of clinical feature: 'RADIATIONS.RADIATION.REGIMENINDICATION'

RADIATIONS.RADIATION.REGIMENINDICATION Labels N
  NO 127
  YES 315
     
  Significant markers N = 6
  Higher in YES 4
  Higher in NO 2
List of 6 genes differentially expressed by 'RADIATIONS.RADIATION.REGIMENINDICATION'

Table S8.  Get Full Table List of 6 genes differentially expressed by 'RADIATIONS.RADIATION.REGIMENINDICATION'

T(pos if higher in 'YES') ttestP Q AUC
HSA-MIR-128-2 5.28 2.37e-07 0.000134 0.6446
HSA-MIR-3613 -5.22 3.512e-07 0.000198 0.6572
HSA-MIR-128-1 5.03 8.438e-07 0.000474 0.6431
HSA-MIR-628 -4.51 9.551e-06 0.00536 0.6223
HSA-MIR-181D 4.21 3.455e-05 0.0193 0.6243
HSA-MIR-660 4.07 6.251e-05 0.0349 0.6145

Figure S4.  Get High-res Image As an example, this figure shows the association of HSA-MIR-128-2 to 'RADIATIONS.RADIATION.REGIMENINDICATION'. P value = 2.37e-07 with T-test analysis.

Clinical variable #5: 'COMPLETENESS.OF.RESECTION'

No gene related to 'COMPLETENESS.OF.RESECTION'.

Table S9.  Basic characteristics of clinical feature: 'COMPLETENESS.OF.RESECTION'

COMPLETENESS.OF.RESECTION Labels N
  R0 302
  R1 24
  R2 14
  RX 24
     
  Significant markers N = 0
Methods & Data
Input

  • Expresson data file = UCEC-TP.miRseq_RPKM_log2.txt

  • Clinical data file = UCEC-TP.clin.merged.picked.txt

  • Number of patients = 442

  • Number of genes = 564

  • Number of clinical features = 5

Survival analysis

For survival clinical features, Wald's test in univariate Cox regression analysis with proportional hazards model (Andersen and Gill 1982) was used to estimate the P values using the 'coxph' function in R. Kaplan-Meier survival curves were plot using the four quartile subgroups of patients based on expression levels

Correlation analysis

For continuous numerical clinical features, Spearman's rank correlation coefficients (Spearman 1904) and two-tailed P values were estimated using 'cor.test' function in R

ANOVA analysis

For multi-class clinical features (ordinal or nominal), one-way analysis of variance (Howell 2002) was applied to compare the log2-expression levels between different clinical classes using 'anova' function in R

Student's t-test analysis

For two-class clinical features, two-tailed Student's t test with unequal variance (Lehmann and Romano 2005) was applied to compare the log2-expression levels between the two clinical classes using 't.test' function in R

Q value calculation

For multiple hypothesis correction, Q value is the False Discovery Rate (FDR) analogue of the P value (Benjamini and Hochberg 1995), defined as the minimum FDR at which the test may be called significant. We used the 'Benjamini and Hochberg' method of 'p.adjust' function in R to convert P values into Q values.

References
[1] Andersen and Gill, Cox's regression model for counting processes, a large sample study, Annals of Statistics 10(4):1100-1120 (1982)
[2] Spearman, C, The proof and measurement of association between two things, Amer. J. Psychol 15:72-101 (1904)
[3] Howell, D, Statistical Methods for Psychology. (5th ed.), Duxbury Press:324-5 (2002)
[4] Lehmann and Romano, Testing Statistical Hypotheses (3E ed.), New York: Springer. ISBN 0387988645 (2005)
[5] Benjamini and Hochberg, Controlling the false discovery rate: a practical and powerful approach to multiple testing, Journal of the Royal Statistical Society Series B 59:289-300 (1995)
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