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Updated: Oct 27, 2026
Heterogeneity Mapping of Protein Expression in Tumors using Quantitative Immunofluorescence
Published on: October 25, 2011
Integrated histopathological, immunohistochemical, and molecular profiling reveals hidden heterogeneity within
Yi-Chen Yeh1, Min-Shu Hsieh2, Lei-Chi Wang1
1Department of Pathology and Laboratory Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; School of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan.
Objectives:
Thymoma classification is based primarily on histologic features, but the extent to which histologic subtypes reflect underlying molecular diversity remains incompletely defined. Molecular characterization is further complicated by abundant intratumoral lymphocytes, which can reduce tumor purity and limit mutation detection using conventional sequencing approaches.
Methods:
In this study, we performed an integrated histopathological, immunohistochemical, and molecular analysis of 138 surgically resected thymomas. Immunohistochemical staining for ASS1, SATB1, BCL2, and CD20 was evaluated, and high-sensitivity molecular barcoding-based targeted sequencing was used to improve detection of low-variant allele frequency mutations. Epithelial-rich tumors were further evaluated by whole-exome sequencing and TCGA multi-omics analysis.
Results:
GTF2I p.L424H mutations were highly prevalent in type A (28/33, 84.8 %) and AB thymomas (50/51, 98.0 %). Among type B thymomas, GTF2I mutations were confined to type B1 tumors (5/16, 31.3 %) and absent in B2 and B3 tumors. GTF2I-mutated B1 thymomas showed low or absent ASS1 expression, resembling type A/AB thymomas, whereas GTF2I-wild-type B1 thymomas showed higher ASS1 expression, similar to other type B thymomas. Whole-exome sequencing revealed molecular heterogeneity among GTF2I-wild-type type A thymomas, with a subset showing high copy number alteration burden and immunophenotypic overlap with type B3 thymomas. TCGA multi-omics analysis further identified a rare GTF2I-mutated type B3 thymoma with type A-like molecular features despite typical B3 morphology.
Conclusion:
Our findings demonstrate that thymoma subtypes harbor molecular heterogeneity not fully captured by conventional histologic classification. Integrating histopathological, immunohistochemical, and molecular features may provide a more biologically informative framework for thymoma stratification.
