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==========================================================================
  IOBR v2.2.0.9000  Immuno-Oncology Biological Research 
  For Documentation: https://iobr.github.io/IOBR/
  For Tutorial: https://iobr.github.io/book/
  For Help: https://github.com/IOBR/IOBR/issues

 If you use IOBR in published research, please cite:
 DQ Zeng, YR Fang, ..., GC Yu*, WJ Liao*, 
 Enhancing immuno-oncology investigations through multidimensional decoding 
 of tumor microenvironment with IOBR 2.0. Cell Rep Methods 4, 100910 (2024). 
 &  
 YR Fang, ..., WJ Liao*, DQ Zeng*, 
 Systematic Investigation of Tumor Microenvironment and 
 Antitumor Immunity With IOBR, Med Research (2025). 
 https://onlinelibrary.wiley.com/doi/epdf/10.1002/mdr2.70001 
==========================================================================
IOBRportal database mode: enabled.

Listening on http://127.0.0.1:37443
ℹ Best number of TME clusters: 3
ℹ Cluster distribution:
ℹ Creating heatmap with 10 features
tidyHeatmap says: (once per session) from release 1.7.0 the scaling is set to "none" by default. Please use scale = "row", "column" or "both" to apply scaling
tidyHeatmap says: If you use tidyHeatmap for scientific research, please cite: Mangiola, S. and Papenfuss, A.T., 2020. 'tidyHeatmap: an R package for modular heatmap production based on tidy principles.' Journal of Open Source Software. doi:10.21105/joss.02472.
This message is displayed once per session.
Warning: `when()` was deprecated in purrr 1.0.0.
ℹ Please use `if` instead.
ℹ The deprecated feature was likely used in the tidyHeatmap package.
  Please report the issue at
  <https://github.com/stemangiola/tidyHeatmap/issues>.
Warning: The `size` argument of `element_line()` is deprecated as of ggplot2 3.4.0.
ℹ Please use the `linewidth` argument instead.
ℹ The deprecated feature was likely used in the IOBR package.
  Please report the issue at <https://github.com/IOBR/IOBR/issues>.
Warning: Error in ggsave: could not find function "ggsave"
  96: <Anonymous>
  95: stop
  94: func
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# A tibble: 11 × 5
# Groups:   T_Stage [5]
   T_Stage Cluster  Freq  Prop count
   <chr>   <chr>   <dbl> <dbl> <dbl>
 1 T1      TME1        5  0.56     9
 2 T1      TME2        2  0.22     9
 3 T1      TME3        2  0.22     9
 4 T2      TME1       34  0.81    42
 5 T2      TME2        8  0.19    42
 6 T3      TME1        7  0.88     8
 7 T3      TME3        1  0.12     8
 8 T4      TME1       14  0.78    18
 9 T4      TME2        2  0.11    18
10 T4      TME3        2  0.11    18
11 <NA>    TME1        2  1        2
ℹ Available categories: box, continue2, continue, random, heatmap, heatmap3, tidyheatmap
ℹ Box palettes: nrc, jama, aaas, jco, paired1, paired2, paired3, paired4, accent, set2
'#374E55FF', '#DF8F44FF', '#00A1D5FF', '#B24745FF', '#79AF97FF', '#6A6599FF', '#80796BFF'
ℹ Survival follow-up time range: 4.11 to 153.72 months
ℹ Best cutoff for "EMT3": 0.1
✔ Best cutoff for "EMT3": 0.104
ℹ High EMT3: 38
ℹ Low EMT3: 41
ℹ Maximum follow-up time is 153.7 months; divided into 6 sections
Warning: Using `size` aesthetic for lines was deprecated in ggplot2 3.4.0.
ℹ Please use `linewidth` instead.
ℹ The deprecated feature was likely used in the ggpubr package.
  Please report the issue at <https://github.com/kassambara/ggpubr/issues>.
IOBRportal database mode: enabled.
ℹ Best number of TME clusters: 3
ℹ Cluster distribution:
ℹ Creating heatmap with 2 features
# A tibble: 11 × 5
# Groups:   Cluster [3]
   Cluster Stage      Freq  Prop count
   <chr>   <chr>     <dbl> <dbl> <dbl>
 1 TME1    Stage I       5  0.08    62
 2 TME1    Stage II     29  0.47    62
 3 TME1    Stage III    13  0.21    62
 4 TME1    Stage IV     13  0.21    62
 5 TME1    <NA>          2  0.03    62
 6 TME2    Stage I       2  0.17    12
 7 TME2    Stage II      8  0.67    12
 8 TME2    Stage III     2  0.17    12
 9 TME3    Stage I       2  0.4      5
10 TME3    Stage III     1  0.2      5
11 TME3    Stage IV      2  0.4      5
ℹ Available categories: box, continue2, continue, random, heatmap, heatmap3, tidyheatmap
ℹ Box palettes: nrc, jama, aaas, jco, paired1, paired2, paired3, paired4, accent, set2
'#374E55FF', '#DF8F44FF', '#00A1D5FF', '#B24745FF', '#79AF97FF', '#6A6599FF', '#80796BFF'
ℹ Groups: 4 ("Stage III", "Stage II", "Stage IV", and "Stage I")
ℹ Features: 183
✔ Kruskal-Wallis test complete
ℹ Creating heatmap with 20 features
ℹ Showing top 10 signatures
`height` was translated to `width`.
`height` was translated to `width`.
`height` was translated to `width`.
ℹ Survival follow-up time range: 4.11 to 153.72 months
ℹ Best cutoff for "EMT1": 1.41
✔ Best cutoff for "EMT1": 1.414
ℹ High EMT1: 12
ℹ Low EMT1: 67
ℹ Maximum follow-up time is 153.7 months; divided into 6 sections
ℹ Follow-up time ranges from 4.11 to 153.72 months
  Stage I  Stage II Stage III  Stage IV 
        9        37        16        15 
ℹ Maximum follow-up time is 153.7 months; divided into 6 sections
Registered S3 methods overwritten by 'ggpp':
  method                  from   
  heightDetails.titleGrob ggplot2
  widthDetails.titleGrob  ggplot2
ℹ Time range: 4.11 to 153.72
ℹ Time range: 4.11 to 153.72
ℹ Time range: 4.11 to 153.72
ℹ Input data preview:
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ℹ Calculating spearman correlation (n = 79)
ℹ Exact p-value: 1.3e-01
ℹ Calculating pearson correlation: 6 x 8
ℹ Survival follow-up time range: 4.11 to 153.72 months
ℹ Best cutoff for "EMT1": 1.41
✔ Best cutoff for "EMT1": 1.414
ℹ High EMT1: 12
ℹ Low EMT1: 67
ℹ Maximum follow-up time is 153.7 months; divided into 6 sections
IOBRportal database mode: enabled.
ℹ Best number of TME clusters: 3
ℹ Cluster distribution:
ℹ Survival follow-up time range: 4.11 to 153.72 months
ℹ Best cutoff for "EMT2": 0.33
✔ Best cutoff for "EMT2": 0.334
ℹ High EMT2: 28
ℹ Low EMT2: 51
ℹ Maximum follow-up time is 153.7 months; divided into 6 sections
Ignoring unknown labels:
• colour : "Strata"
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• colour : "Strata"
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• colour : "Strata"
Warning in normalizePath(path) :
  path[1]="C:/Users/qingcongl/Desktop/IOBRshiny_local/IOBR": No such file or directory
Warning: Error in value[[3L]]: Could not find a root 'DESCRIPTION' file that starts with '^Package' in
'C:/Users/qingcongl/Desktop/IOBRshiny_local/IOBR'.
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  file?
  73: <Anonymous>
  72: signalCondition
  71: signal_abort
  70: rlang::abort
  69: cli::cli_abort
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  73: <Anonymous>
  72: signalCondition
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'C:/Users/qingcongl/Desktop/IOBRshiny_local/IOBR'.
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  file?
  73: <Anonymous>
  72: signalCondition
  71: signal_abort
  70: rlang::abort
  69: cli::cli_abort
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IOBRportal database mode: enabled.
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 3985 genes without length information
ℹ Found 1679 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54658 unique genes
✔ Applied log2 transformation
ℹ When yinter = -3
ℹ Potential outliers: 
ℹ Calculating signature scores using ssGSEA method
ℹ Log2 transformation not necessary (data appears to already be log-scaled)
ℹ Calculating scores for 166 signature(s)
ℹ GSVA version 2.4.9
ℹ Searching for rows with constant values
ℹ Calculating ssGSEA scores for 165 gene sets
ℹ Calculating ranks
ℹ Calculating rank weights
ℹ Normalizing ssGSEA scores
✔ Calculations finished
ℹ Best number of TME clusters: 4
ℹ Cluster distribution:
ℹ Creating heatmap with 173 features
Warning in grid.Call.graphics(C_text, as.graphicsAnnot(x$label), x$x, x$y,  :
  conversion failure on 'IFNγ_iCAF' in 'mbcsToSbcs': for γ (U+03B3)
Warning in grid.Call.graphics(C_text, as.graphicsAnnot(x$label), x$x, x$y,  :
  conversion failure on 'TGFβ_myCAF' in 'mbcsToSbcs': for β (U+03B2)
# A tibble: 3 × 5
# Groups:   ARID1A [2]
  ARID1A    PIK3CA     Freq  Prop count
  <chr>     <chr>     <dbl> <dbl> <dbl>
1 mutatant  mutatant      4  1        4
2 wild_type mutatant      1  0.17     6
3 wild_type wild_type     5  0.83     6
ℹ Available categories: box, continue2, continue, random, heatmap, heatmap3, tidyheatmap
ℹ Box palettes: nrc, jama, aaas, jco, paired1, paired2, paired3, paired4, accent, set2
'#374E55FF', '#DF8F44FF', '#00A1D5FF', '#B24745FF', '#79AF97FF', '#6A6599FF', '#80796BFF'
ℹ Available categories: box, continue2, continue, random, heatmap, heatmap3, tidyheatmap
ℹ Random palettes: 1 (palette1), 2 (palette2), 3 (palette3), 4 (palette4)
ℹ Available categories: box, continue2, continue, random, heatmap, heatmap3, tidyheatmap
ℹ Random palettes: 1 (palette1), 2 (palette2), 3 (palette3), 4 (palette4)
ℹ Groups: "High" and "Low"
ℹ Features: 171
✔ Wilcoxon test complete
ℹ Groups: 4 ("TME2", "TME4", "TME1", and "TME3")
ℹ Features: 171
✔ Kruskal-Wallis test complete
ℹ Creating heatmap with 20 features
ℹ Showing top 10 signatures
`height` was translated to `width`.
`height` was translated to `width`.
ℹ Survival follow-up time range: 0.47 to 28.97 months
ℹ Best cutoff for "CD_8_T_effector": -1.28
✔ Best cutoff for "CD_8_T_effector": -1.285
ℹ High CD_8_T_effector: 8
ℹ Low CD_8_T_effector: 1
ℹ Maximum follow-up time is 29 months; divided into 6 sections
Ignoring unknown labels:
• colour : "Strata"
Ignoring unknown labels:
• colour : "Strata"
Ignoring unknown labels:
• colour : "Strata"
Ignoring unknown labels:
• colour : "Strata"
Ignoring unknown labels:
• colour : "Strata"
Ignoring unknown labels:
• colour : "Strata"
ℹ Follow-up time ranges from 0.47 to 28.97 months
TME1 TME2 TME3 TME4 
   2    2    2    3 
ℹ Maximum follow-up time is 29 months; divided into 6 sections
Ignoring unknown labels:
• colour : "Strata"
ℹ Time range: 0.47 to 28.97
ℹ Input data preview:
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Setting levels: control = 0, case = 1
Setting direction: controls > cases
ℹ Input data preview:
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
ℹ Input data preview:
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls < cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
ℹ Input data preview:
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls > cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
Setting levels: control = 0, case = 1
Setting direction: controls < cases
Warning in ci.auc.smooth.roc(smooth.roc, ...) :
  NA value(s) produced during bootstrap were ignored.
ℹ Computing spearman correlation for 172 features
✔ Correlation analysis complete
ℹ Computing pearson partial correlation for
✔ Partial correlation analysis complete
ℹ Calculating spearman correlation (n = 10)
ℹ Exact p-value: 6e-02
`geom_smooth()` using formula = 'y ~ x'
`geom_smooth()` using formula = 'y ~ x'
ℹ Calculating spearman correlation (n = 10)
ℹ Exact p-value: 6e-02
ℹ Groups: "Diffuse", "Intestinal", and "Mixed"
`geom_smooth()` using formula = 'y ~ x'
`geom_smooth()` using formula = 'y ~ x'
ℹ Calculating pearson correlation: 3 x 11
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 3985 genes without length information
ℹ Found 1679 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54658 unique genes
✔ Applied log2 transformation
User session ended. Running garbage collection...
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 3985 genes without length information
ℹ Found 1679 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54658 unique genes
✔ Applied log2 transformation
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 3985 genes without length information
ℹ Found 1679 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54658 unique genes
✔ Applied log2 transformation
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 3985 genes without length information
ℹ Found 1679 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54658 unique genes
✔ Applied log2 transformation
ℹ Creating heatmap with 10 features
ℹ Creating heatmap with 10 features
ℹ Creating heatmap with 10 features
ℹ Trying mirror 1/4: <https://github.com>
trying URL 'https://github.com/IOBR/IOBR/releases/download/data-v1.0/anno_hug133plus2.rda'
Warning in utils::download.file(url, cache_file, mode = "wb", quiet = !verbose) :
  URL 'https://github.com/IOBR/IOBR/releases/download/data-v1.0/anno_hug133plus2.rda': Timeout of 60 seconds was reached
! Mirror 1 failed: cannot open URL 'https://github.com/IOBR/IOBR/releases/download/data-v1.0/anno_hug133plus2.rda'
ℹ Trying mirror 2/4: <https://ghproxy.vip/https://github.com>
trying URL 'https://ghproxy.vip/https://github.com/IOBR/IOBR/releases/download/data-v1.0/anno_hug133plus2.rda'
Content type 'application/octet-stream' length 208824 bytes (203 KB)
==================================================
downloaded 203 KB

✔ Download complete: "anno_hug133plus2"
ℹ Row number of original eset: 54675
✔ 83% of probes in expression set were annotated
ℹ Found 23366 duplicate symbols, using "mean" method
ℹ Row number after filtering duplicated gene symbol: 21752
ℹ Log2 transformation not necessary (data appears to already be log-scaled)
ℹ Loading cached data: "anno_hug133plus2"
ℹ Row number of original eset: 54675
✔ 83% of probes in expression set were annotated
ℹ Found 23366 duplicate symbols, using "mean" method
ℹ Row number after filtering duplicated gene symbol: 21752
ℹ Log2 transformation not necessary (data appears to already be log-scaled)
ℹ When yinter = -3
ℹ Potential outliers: "GSM1523817", "GSM1523858", "GSM1523984", "GSM1523988", and "GSM1524030"
ℹ Calculating signature scores using ssGSEA method
ℹ Log2 transformation not necessary (data appears to already be log-scaled)
ℹ Calculating scores for 279 signature(s)
ℹ GSVA version 2.4.9
ℹ Searching for rows with constant values
ℹ Calculating ssGSEA scores for 279 gene sets
ℹ Calculating ranks
ℹ Calculating rank weights
ℹ Normalizing ssGSEA scores
✔ Calculations finished
ℹ Best number of TME clusters: 4
ℹ Cluster distribution:
User session ended. Running garbage collection...
ℹ Loading cached data: "anno_hug133plus2"
ℹ Row number of original eset: 54675
✔ 83% of probes in expression set were annotated
ℹ Found 23366 duplicate symbols, using "mean" method
ℹ Row number after filtering duplicated gene symbol: 21752
ℹ Log2 transformation not necessary (data appears to already be log-scaled)
IOBRportal database mode: enabled.
User session ended. Running garbage collection...
IOBRportal database mode: enabled.
User session ended. Running garbage collection...
Warning: closing unused connection 3 ()
ℹ Loading cached data: "anno_grch38"
ℹ Using local annotation (anno_grch38) for TPM conversion
! Omitting 153 genes without length information
ℹ Found 1680 duplicate symbols. Using "mean" for ranking.
✔ Reduced to 54767 unique genes
✔ Applied log2 transformation
User session ended. Running garbage collection...


Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Closed TCGA pool.
Closed OtherCohort pool.
Closed Immunotherapy pool.
Closed CancerCohort pool.
Execution halted