THBS4, a novel stromal molecule of diffuse-type gastric adenocarcinomas, identified by transcriptome-wide expression profiling.

Gastric adenocarcinomas can be divided into two major histological types, the diffuse and intestinal type (Laurén classification). Since they diverge in many clinical and molecular characteristics, it is widely accepted that they represent distinct disease entities that may benefit from different therapeutic approaches. Gene expression profiling studies have identified numerous genes that are differentially expressed between them. However, none of these studies covered the whole transcriptome and the published gene lists reveal little overlap, raising the need for further, more comprehensive analyses. Here, we present the first transcriptome-wide expression profiling study comparing the two types (diffuse n=19, intestinal n=24), which identified >1000 genes that are differentially expressed. Among them, thrombospondin 4 (THBS4) showed the strongest correlation to histological type, with vast overexpression in the diffuse type. Quantitative real-time PCR validated this strong overexpression and revealed that intestinal tumors generally lack THBS4 expression. Immunohistochemistry demonstrated THBS4 overexpression on the protein level (n=10) and localized THBS4 to the stromal aspect. Its expression was primarily observed within the extracellular matrix surrounding the tumor cells, with the highest intensities found in regions of high tumor cell density and invasion. Intestinal tumors and matched non-neoplastic gastric epithelium and stroma did not feature any relevant THBS4 expression in a preliminary selection of analyzed cases (n=5). Immunohistochemical colocalization and in vitro studies revealed that THBS4 is expressed and secreted by cancer-associated fibroblasts. Furthermore, we show that THBS4 transcription in fibroblasts is stimulated by tumor cells. This study is the first to identify THBS4 as a powerful marker for diffuse-type gastric adenocarcinomas and to provide an initial characterization of its expression in the course of this disease.