# Comparative spatial whole transcriptome analysis of matched frozen and formalin-fixed paraffin-embedded colorectal cancer tissues

## TL;DR

This study compares gene expression in colorectal cancer tissues preserved using different methods, showing how each method affects spatial transcriptomic data quality.

## Contribution

The study provides a direct comparison of spatial transcriptomic profiles from FFPE, FF, and SF CRC tissues, highlighting their relative strengths.

## Key findings

- FFPE tissues provided better spatial resolution of cellular morphology and gene expression.
- FF and SF tissues showed higher transcript abundance and CRC gene expression.
- All tissue types shared 132 differentially expressed genes linked to CRC pathways.

## Abstract

Spatial transcriptomic technology enables resolving a tumor microenvironment heterogeneity of colorectal cancer (CRC) tissue samples. However, there is limited evidence regarding the relative strengths and weaknesses of different CRC sample types and preservation methods, including formalin-fixed paraffin-embedded (FFPE), fresh frozen (FF) and snap frozen (SF). Here, we examine gene expression profiles in 36 tissue areas, FFPE, FF and SF samples, derived from a patient with stage IV CRC, using spatial analysis of the whole transcriptome. The tissue samples obtained from surgical resection of the cancer were subjected to a detailed histological evaluation and spatial transcriptomics analysis to compare the quality of the sample between different preservation methods. Spatial transcriptomic analysis demonstrated that FFPE tissue samples offered improved resolution of cellular morphology in the tumor microenvironment, facilitating greater detail of gene expression and cell types. In contrast, gene expression analysis of FF and SF tissues embedded in optimal cutting temperature compound revealed a high level of CRC gene expression. FFPE, FF and SF samples shared a significant overlap among the top 300 genes that were strongly associated with key biological processes in CRC. In summary, the spatial analysis of the whole transcriptome across FFPE, FF and SF CRC tissues revealed intratumoral heterogeneity and highlighted key transcriptomic signatures, suggesting the specific value of different tissue preservation methods in the profiling of CRC.

•Spatial transcriptomics applied to matched FFPE, FF, and SF CRC tissues.•FFPE preserved spatial context; FF/SF retained higher transcript abundance.•Shared 132 DEGs across all tissue types linked to key CRC pathways.•FFPE enables reliable spatial profiles and preserves transcriptomic signatures.

Spatial transcriptomics applied to matched FFPE, FF, and SF CRC tissues.

FFPE preserved spatial context; FF/SF retained higher transcript abundance.

Shared 132 DEGs across all tissue types linked to key CRC pathways.

FFPE enables reliable spatial profiles and preserves transcriptomic signatures.