Correlation between mRNAseq expression and clinical features
Overview
Introduction

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

Summary

Testing the association between 17995 genes and 14 clinical features across 413 samples, statistically thresholded by Q value < 0.05, 13 clinical features related to at least one genes.

  • 127 genes correlated to 'AGE'.

    • ZNF518B|85460 ,  HCG11|493812 ,  C12ORF52|84934 ,  ASB13|79754 ,  RANBP17|64901 ,  ...

  • 39 genes correlated to 'GENDER'.

    • RPS4Y1|6192 ,  ZFY|7544 ,  DDX3Y|8653 ,  UTY|7404 ,  NLGN4Y|22829 ,  ...

  • 4458 genes correlated to 'HISTOLOGICAL.TYPE'.

    • ITGA3|3675 ,  FN1|2335 ,  KCNN4|3783 ,  UNC5CL|222643 ,  TMPRSS4|56649 ,  ...

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

    • C14ORF180|400258 ,  FLJ37543|285668 ,  TAS2R43|259289 ,  ADIPOQ|9370 ,  HPR|3250 ,  ...

  • 6 genes correlated to 'RADIATIONEXPOSURE'.

    • CRNN|49860 ,  C13ORF30|144809 ,  CSNK1A1P|161635 ,  CYP2C9|1559 ,  CYORF15A|246126 ,  ...

  • 8 genes correlated to 'DISTANT.METASTASIS'.

    • C4ORF6|10141 ,  RNF207|388591 ,  CYP21A2|1589 ,  HSPA6|3310 ,  TAF1D|79101 ,  ...

  • 639 genes correlated to 'EXTRATHYROIDAL.EXTENSION'.

    • ASAH2|56624 ,  FAM69C|125704 ,  COL11A1|1301 ,  FAM169A|26049 ,  CYP26A1|1592 ,  ...

  • 1295 genes correlated to 'LYMPH.NODE.METASTASIS'.

    • CREB5|9586 ,  KCNN4|3783 ,  ICAM1|3383 ,  MLEC|9761 ,  FN1|2335 ,  ...

  • 1 gene correlated to 'COMPLETENESS.OF.RESECTION'.

    • CATSPER2P1|440278

  • 673 genes correlated to 'NUMBER.OF.LYMPH.NODES'.

    • CBFB|865 ,  PELI1|57162 ,  FAM60A|58516 ,  ZNF346|23567 ,  CREB5|9586 ,  ...

  • 263 genes correlated to 'NEOPLASM.DISEASESTAGE'.

    • GSPT2|23708 ,  SEMA3G|56920 ,  FAM50B|26240 ,  GJA5|2702 ,  MOGS|7841 ,  ...

  • 1 gene correlated to 'MULTIFOCALITY'.

    • TMTC1|83857

  • 7 genes correlated to 'TUMOR.SIZE'.

    • CLDN2|9075 ,  ZC3H10|84872 ,  C3ORF32|51066 ,  PI16|221476 ,  FOXI2|399823 ,  ...

  • No genes correlated to 'Time to Death'

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=0        
AGE Spearman correlation test N=127 older N=55 younger N=72
GENDER t test N=39 male N=16 female N=23
HISTOLOGICAL TYPE ANOVA test N=4458        
RADIATIONS RADIATION REGIMENINDICATION t test N=26 yes N=18 no N=8
RADIATIONEXPOSURE t test N=6 yes N=2 no N=4
DISTANT METASTASIS ANOVA test N=8        
EXTRATHYROIDAL EXTENSION ANOVA test N=639        
LYMPH NODE METASTASIS ANOVA test N=1295        
COMPLETENESS OF RESECTION ANOVA test N=1        
NUMBER OF LYMPH NODES Spearman correlation test N=673 higher number.of.lymph.nodes N=377 lower number.of.lymph.nodes N=296
NEOPLASM DISEASESTAGE ANOVA test N=263        
MULTIFOCALITY t test N=1 unifocal N=0 multifocal N=1
TUMOR SIZE Spearman correlation test N=7 higher tumor.size N=0 lower tumor.size N=7
Clinical variable #1: 'Time to Death'

No gene related to 'Time to Death'.

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

Time to Death Duration (Months) 0-147.4 (median=9.3)
  censored N = 398
  death N = 10
     
  Significant markers N = 0
Clinical variable #2: 'AGE'

127 genes related to 'AGE'.

Table S2.  Basic characteristics of clinical feature: 'AGE'

AGE Mean (SD) 46.6 (16)
  Significant markers N = 127
  pos. correlated 55
  neg. correlated 72
List of top 10 genes significantly correlated to 'AGE' by Spearman correlation test

Table S3.  Get Full Table List of top 10 genes significantly correlated to 'AGE' by Spearman correlation test

SpearmanCorr corrP Q
ZNF518B|85460 -0.3422 8.722e-13 1.57e-08
HCG11|493812 -0.3173 4.093e-11 7.36e-07
C12ORF52|84934 0.3169 4.337e-11 7.8e-07
ASB13|79754 0.3115 9.585e-11 1.72e-06
RANBP17|64901 -0.3114 1.14e-10 2.05e-06
STL|7955 -0.3084 1.502e-10 2.7e-06
IL20RA|53832 0.3084 1.665e-10 2.99e-06
MSL3L2|151507 -0.3052 2.381e-10 4.28e-06
EPN3|55040 0.297 7.373e-10 1.33e-05
GPI|2821 0.2903 1.825e-09 3.28e-05

Figure S1.  Get High-res Image As an example, this figure shows the association of ZNF518B|85460 to 'AGE'. P value = 8.72e-13 with Spearman correlation analysis. The straight line presents the best linear regression.

Clinical variable #3: 'GENDER'

39 genes related to 'GENDER'.

Table S4.  Basic characteristics of clinical feature: 'GENDER'

GENDER Labels N
  FEMALE 307
  MALE 106
     
  Significant markers N = 39
  Higher in MALE 16
  Higher in FEMALE 23
List of top 10 genes differentially expressed by 'GENDER'

Table S5.  Get Full Table List of top 10 genes differentially expressed by 'GENDER'

T(pos if higher in 'MALE') ttestP Q AUC
RPS4Y1|6192 83.65 1.028e-170 1.85e-166 1
ZFY|7544 84 3.646e-166 6.56e-162 1
DDX3Y|8653 87.01 1.572e-157 2.83e-153 1
UTY|7404 83.08 2.656e-132 4.78e-128 1
NLGN4Y|22829 60.52 2.266e-131 4.08e-127 1
KDM5D|8284 78.03 8.45e-123 1.52e-118 1
PRKY|5616 37.47 3.752e-112 6.75e-108 0.9924
USP9Y|8287 66.01 4.011e-109 7.21e-105 0.9999
XIST|7503 -42.46 5.508e-72 9.91e-68 0.9928
CYORF15A|246126 53.1 4.894e-58 8.8e-54 1

Figure S2.  Get High-res Image As an example, this figure shows the association of RPS4Y1|6192 to 'GENDER'. P value = 1.03e-170 with T-test analysis.

Clinical variable #4: 'HISTOLOGICAL.TYPE'

4458 genes related to 'HISTOLOGICAL.TYPE'.

Table S6.  Basic characteristics of clinical feature: 'HISTOLOGICAL.TYPE'

HISTOLOGICAL.TYPE Labels N
  OTHER SPECIFY 7
  THYROID PAPILLARY CARCINOMA - CLASSICAL/USUAL 286
  THYROID PAPILLARY CARCINOMA - FOLLICULAR (>= 99% FOLLICULAR PATTERNED) 86
  THYROID PAPILLARY CARCINOMA - TALL CELL (>= 50% TALL CELL FEATURES) 34
     
  Significant markers N = 4458
List of top 10 genes differentially expressed by 'HISTOLOGICAL.TYPE'

Table S7.  Get Full Table List of top 10 genes differentially expressed by 'HISTOLOGICAL.TYPE'

ANOVA_P Q
ITGA3|3675 2.257e-37 4.06e-33
FN1|2335 2.785e-37 5.01e-33
KCNN4|3783 7.66e-35 1.38e-30
UNC5CL|222643 9.357e-35 1.68e-30
TMPRSS4|56649 1.292e-34 2.32e-30
MUC1|4582 1.912e-34 3.44e-30
SFTPB|6439 2.687e-34 4.83e-30
KRT19|3880 4.991e-34 8.98e-30
TM7SF4|81501 5.856e-34 1.05e-29
SERPINA1|5265 2.073e-33 3.73e-29

Figure S3.  Get High-res Image As an example, this figure shows the association of ITGA3|3675 to 'HISTOLOGICAL.TYPE'. P value = 2.26e-37 with ANOVA analysis.

Clinical variable #5: 'RADIATIONS.RADIATION.REGIMENINDICATION'

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

Table S8.  Basic characteristics of clinical feature: 'RADIATIONS.RADIATION.REGIMENINDICATION'

RADIATIONS.RADIATION.REGIMENINDICATION Labels N
  NO 13
  YES 400
     
  Significant markers N = 26
  Higher in YES 18
  Higher in NO 8
List of top 10 genes differentially expressed by 'RADIATIONS.RADIATION.REGIMENINDICATION'

Table S9.  Get Full Table List of top 10 genes differentially expressed by 'RADIATIONS.RADIATION.REGIMENINDICATION'

T(pos if higher in 'YES') ttestP Q AUC
C14ORF180|400258 12.31 6.753e-14 1.21e-09 0.8788
FLJ37543|285668 14.21 2.111e-13 3.79e-09 0.8845
TAS2R43|259289 9.59 1.583e-11 2.84e-07 0.7814
ADIPOQ|9370 10.64 3.358e-11 6.03e-07 0.9115
HPR|3250 10.64 1.738e-09 3.12e-05 0.9232
LOC440173|440173 8.99 1.531e-08 0.000275 0.8827
ENPP3|5169 9 2.594e-08 0.000465 0.8582
CXORF48|54967 8.73 2.205e-07 0.00396 0.8335
C6ORF203|51250 8.56 3.197e-07 0.00574 0.876
HCFC1|3054 -8.69 3.584e-07 0.00643 0.9002

Figure S4.  Get High-res Image As an example, this figure shows the association of C14ORF180|400258 to 'RADIATIONS.RADIATION.REGIMENINDICATION'. P value = 6.75e-14 with T-test analysis.

Clinical variable #6: 'RADIATIONEXPOSURE'

6 genes related to 'RADIATIONEXPOSURE'.

Table S10.  Basic characteristics of clinical feature: 'RADIATIONEXPOSURE'

RADIATIONEXPOSURE Labels N
  NO 354
  YES 16
     
  Significant markers N = 6
  Higher in YES 2
  Higher in NO 4
List of 6 genes differentially expressed by 'RADIATIONEXPOSURE'

Table S11.  Get Full Table List of 6 genes differentially expressed by 'RADIATIONEXPOSURE'

T(pos if higher in 'YES') ttestP Q AUC
CRNN|49860 -12.13 7.086e-21 1.27e-16 0.927
C13ORF30|144809 -11.05 2.252e-19 4.05e-15 0.8687
CSNK1A1P|161635 -7.71 2.612e-11 4.7e-07 0.8219
CYP2C9|1559 -7.55 8.719e-11 1.57e-06 0.7341
CYORF15A|246126 6.56 1.31e-09 2.35e-05 0.606
EIF1AY|9086 5.43 3.311e-07 0.00595 0.7776

Figure S5.  Get High-res Image As an example, this figure shows the association of CRNN|49860 to 'RADIATIONEXPOSURE'. P value = 7.09e-21 with T-test analysis.

Clinical variable #7: 'DISTANT.METASTASIS'

8 genes related to 'DISTANT.METASTASIS'.

Table S12.  Basic characteristics of clinical feature: 'DISTANT.METASTASIS'

DISTANT.METASTASIS Labels N
  M0 231
  M1 8
  MX 173
     
  Significant markers N = 8
List of 8 genes differentially expressed by 'DISTANT.METASTASIS'

Table S13.  Get Full Table List of 8 genes differentially expressed by 'DISTANT.METASTASIS'

ANOVA_P Q
C4ORF6|10141 8.042e-08 0.00145
RNF207|388591 8.664e-08 0.00156
CYP21A2|1589 1.104e-07 0.00199
HSPA6|3310 5.558e-07 0.01
TAF1D|79101 9.498e-07 0.0171
SNORA8|654320 1.436e-06 0.0258
ADAMTS10|81794 2.04e-06 0.0367
PPP1R13L|10848 2.142e-06 0.0385

Figure S6.  Get High-res Image As an example, this figure shows the association of C4ORF6|10141 to 'DISTANT.METASTASIS'. P value = 8.04e-08 with ANOVA analysis.

Clinical variable #8: 'EXTRATHYROIDAL.EXTENSION'

639 genes related to 'EXTRATHYROIDAL.EXTENSION'.

Table S14.  Basic characteristics of clinical feature: 'EXTRATHYROIDAL.EXTENSION'

EXTRATHYROIDAL.EXTENSION Labels N
  MINIMAL (T3) 110
  MODERATE/ADVANCED (T4A) 13
  NONE 270
  VERY ADVANCED (T4B) 1
     
  Significant markers N = 639
List of top 10 genes differentially expressed by 'EXTRATHYROIDAL.EXTENSION'

Table S15.  Get Full Table List of top 10 genes differentially expressed by 'EXTRATHYROIDAL.EXTENSION'

ANOVA_P Q
ASAH2|56624 4.689e-15 8.44e-11
FAM69C|125704 6.159e-14 1.11e-09
COL11A1|1301 5.794e-13 1.04e-08
FAM169A|26049 6.843e-13 1.23e-08
CYP26A1|1592 8.149e-13 1.47e-08
COL1A1|1277 1.103e-12 1.98e-08
FOXJ1|2302 1.186e-12 2.13e-08
SRPX2|27286 1.375e-12 2.47e-08
SLC25A42|284439 2.654e-12 4.77e-08
ZNF540|163255 3.772e-12 6.79e-08

Figure S7.  Get High-res Image As an example, this figure shows the association of ASAH2|56624 to 'EXTRATHYROIDAL.EXTENSION'. P value = 4.69e-15 with ANOVA analysis.

Clinical variable #9: 'LYMPH.NODE.METASTASIS'

1295 genes related to 'LYMPH.NODE.METASTASIS'.

Table S16.  Basic characteristics of clinical feature: 'LYMPH.NODE.METASTASIS'

LYMPH.NODE.METASTASIS Labels N
  N0 191
  N1 48
  N1A 77
  N1B 59
  NX 38
     
  Significant markers N = 1295
List of top 10 genes differentially expressed by 'LYMPH.NODE.METASTASIS'

Table S17.  Get Full Table List of top 10 genes differentially expressed by 'LYMPH.NODE.METASTASIS'

ANOVA_P Q
CREB5|9586 2.403e-15 4.32e-11
KCNN4|3783 2.297e-14 4.13e-10
ICAM1|3383 6.153e-14 1.11e-09
MLEC|9761 6.576e-14 1.18e-09
FN1|2335 7.421e-14 1.34e-09
CTSC|1075 9e-14 1.62e-09
SFTPB|6439 1.351e-13 2.43e-09
WFS1|7466 1.452e-13 2.61e-09
TMEM117|84216 2.118e-13 3.81e-09
PROS1|5627 2.435e-13 4.38e-09

Figure S8.  Get High-res Image As an example, this figure shows the association of CREB5|9586 to 'LYMPH.NODE.METASTASIS'. P value = 2.4e-15 with ANOVA analysis.

Clinical variable #10: 'COMPLETENESS.OF.RESECTION'

One gene related to 'COMPLETENESS.OF.RESECTION'.

Table S18.  Basic characteristics of clinical feature: 'COMPLETENESS.OF.RESECTION'

COMPLETENESS.OF.RESECTION Labels N
  R0 325
  R1 35
  R2 2
  RX 26
     
  Significant markers N = 1
List of one gene differentially expressed by 'COMPLETENESS.OF.RESECTION'

Table S19.  Get Full Table List of one gene differentially expressed by 'COMPLETENESS.OF.RESECTION'

ANOVA_P Q
CATSPER2P1|440278 3.665e-09 6.6e-05

Figure S9.  Get High-res Image As an example, this figure shows the association of CATSPER2P1|440278 to 'COMPLETENESS.OF.RESECTION'. P value = 3.66e-09 with ANOVA analysis.

Clinical variable #11: 'NUMBER.OF.LYMPH.NODES'

673 genes related to 'NUMBER.OF.LYMPH.NODES'.

Table S20.  Basic characteristics of clinical feature: 'NUMBER.OF.LYMPH.NODES'

NUMBER.OF.LYMPH.NODES Mean (SD) 3.57 (6.3)
  Significant markers N = 673
  pos. correlated 377
  neg. correlated 296
List of top 10 genes significantly correlated to 'NUMBER.OF.LYMPH.NODES' by Spearman correlation test

Table S21.  Get Full Table List of top 10 genes significantly correlated to 'NUMBER.OF.LYMPH.NODES' by Spearman correlation test

SpearmanCorr corrP Q
CBFB|865 0.3887 6.519e-13 1.17e-08
PELI1|57162 0.3859 9.801e-13 1.76e-08
FAM60A|58516 0.3824 1.635e-12 2.94e-08
ZNF346|23567 -0.3766 3.725e-12 6.7e-08
CREB5|9586 0.3751 4.644e-12 8.35e-08
CSGALNACT2|55454 0.3725 6.661e-12 1.2e-07
CCDC109B|55013 0.3692 1.043e-11 1.88e-07
TAGLN2|8407 0.3679 1.25e-11 2.25e-07
S100A10|6281 0.3613 3.056e-11 5.5e-07
TMEM117|84216 0.3592 4.067e-11 7.31e-07

Figure S10.  Get High-res Image As an example, this figure shows the association of CBFB|865 to 'NUMBER.OF.LYMPH.NODES'. P value = 6.52e-13 with Spearman correlation analysis. The straight line presents the best linear regression.

Clinical variable #12: 'NEOPLASM.DISEASESTAGE'

263 genes related to 'NEOPLASM.DISEASESTAGE'.

Table S22.  Basic characteristics of clinical feature: 'NEOPLASM.DISEASESTAGE'

NEOPLASM.DISEASESTAGE Labels N
  STAGE I 232
  STAGE II 46
  STAGE III 90
  STAGE IV 2
  STAGE IVA 35
  STAGE IVC 5
     
  Significant markers N = 263
List of top 10 genes differentially expressed by 'NEOPLASM.DISEASESTAGE'

Table S23.  Get Full Table List of top 10 genes differentially expressed by 'NEOPLASM.DISEASESTAGE'

ANOVA_P Q
GSPT2|23708 7.713e-17 1.39e-12
SEMA3G|56920 1.011e-13 1.82e-09
FAM50B|26240 1.409e-13 2.53e-09
GJA5|2702 1.119e-11 2.01e-07
MOGS|7841 1.653e-11 2.97e-07
CYP26A1|1592 2.764e-11 4.97e-07
HARS2|23438 1.316e-10 2.37e-06
ZNF518B|85460 4.636e-10 8.34e-06
PECI|10455 5.139e-10 9.24e-06
MKKS|8195 1.377e-09 2.48e-05

Figure S11.  Get High-res Image As an example, this figure shows the association of GSPT2|23708 to 'NEOPLASM.DISEASESTAGE'. P value = 7.71e-17 with ANOVA analysis.

Clinical variable #13: 'MULTIFOCALITY'

One gene related to 'MULTIFOCALITY'.

Table S24.  Basic characteristics of clinical feature: 'MULTIFOCALITY'

MULTIFOCALITY Labels N
  MULTIFOCAL 177
  UNIFOCAL 227
     
  Significant markers N = 1
  Higher in UNIFOCAL 0
  Higher in MULTIFOCAL 1
List of one gene differentially expressed by 'MULTIFOCALITY'

Table S25.  Get Full Table List of one gene differentially expressed by 'MULTIFOCALITY'

T(pos if higher in 'UNIFOCAL') ttestP Q AUC
TMTC1|83857 -4.77 2.571e-06 0.0463 0.6369

Figure S12.  Get High-res Image As an example, this figure shows the association of TMTC1|83857 to 'MULTIFOCALITY'. P value = 2.57e-06 with T-test analysis.

Clinical variable #14: 'TUMOR.SIZE'

7 genes related to 'TUMOR.SIZE'.

Table S26.  Basic characteristics of clinical feature: 'TUMOR.SIZE'

TUMOR.SIZE Mean (SD) 2.92 (1.6)
  Significant markers N = 7
  pos. correlated 0
  neg. correlated 7
List of 7 genes significantly correlated to 'TUMOR.SIZE' by Spearman correlation test

Table S27.  Get Full Table List of 7 genes significantly correlated to 'TUMOR.SIZE' by Spearman correlation test

SpearmanCorr corrP Q
CLDN2|9075 -0.2914 1.014e-07 0.00183
ZC3H10|84872 -0.279 3.198e-07 0.00575
C3ORF32|51066 -0.2746 6.844e-07 0.0123
PI16|221476 -0.2718 8.584e-07 0.0154
FOXI2|399823 -0.2644 1.501e-06 0.027
LOC151162|151162 -0.2628 1.551e-06 0.0279
SEC22A|26984 -0.2571 2.648e-06 0.0476

Figure S13.  Get High-res Image As an example, this figure shows the association of CLDN2|9075 to 'TUMOR.SIZE'. P value = 1.01e-07 with Spearman correlation analysis. The straight line presents the best linear regression.

Methods & Data
Input

  • Expresson data file = THCA-TP.uncv2.mRNAseq_RSEM_normalized_log2.txt

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

  • Number of patients = 413

  • Number of genes = 17995

  • Number of clinical features = 14

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

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

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

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] Lehmann and Romano, Testing Statistical Hypotheses (3E ed.), New York: Springer. ISBN 0387988645 (2005)
[4] Howell, D, Statistical Methods for Psychology. (5th ed.), Duxbury Press:324-5 (2002)
[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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