RNA-Seq Reveals Differences in Expressed Tumor Mutation Burden in Colorectal and Endometrial Cancers with and without...

Area of Science:

• Oncology
• Genomics
• Biomarker Discovery

Background:

• Tumor mutation burden (TMB) is a key biomarker for predicting immunotherapy response.
• Accurate TMB measurement is crucial, especially in formalin-fixed paraffin-embedded (FFPE) tissues.

Purpose of the Study:

• To develop and validate a method for measuring expressed TMB (eTMB) using RNA sequencing in FFPE samples.
• To assess the correlation between eTMB and microsatellite instability (MSI) status in colorectal and endometrial cancers.

Main Methods:

• RNA sequencing was performed on FFPE tumor and normal tissue samples from 58 patients (46 MSI-H, 12 MSS).
• A novel method was developed to accurately measure eTMB by removing FFPE-derived artifacts using mutation signatures.
• DNA TMB was compared with eTMB using whole-exome sequencing in a subset of patients.

Main Results:

• A significant difference in median eTMB was observed between MSI-H (27.3 mut/Mb) and MSS (6.7 mut/Mb) tumors (P < 10⁻⁸).
• MSI-H tumors with DNA mismatch-repair mutations showed higher eTMB (28.1 mut/Mb) than those with MLH1 hypermethylation (17.5 mut/Mb) (P = 0.037).
• MSI status, tumor type, and age were significantly associated with eTMB; DNA TMB correlated well with eTMB (Spearman r = 0.83).

Conclusions:

• RNA sequencing provides a reliable method for measuring eTMB in FFPE tumor specimens.
• eTMB measurement can effectively differentiate MSI-H from MSS tumors, supporting its utility as a biomarker.
• This approach has implications for patient stratification and treatment selection in immunotherapy.