AGR2 protein expression in colorectal tumour epithelialcompartment

We read with great interest the manuscript by Tian et al published in Gut. Tian et al report that in colorectal cancer (CRC) at stages III/IV, the presence of extracellular AGR2 in the tumour environment is due to its secretion by tumourassociated neutrophils (TANs). This, in turn, promotes tumour cells migration and invasion. Although these findings are original and exciting, it is essential to put them in the context of the biology of anterior gradient (AGR) proteins. AGR2 is the most studied member of the AGR family, which contains three proteins and belongs to the superfamily of protein disulfide isomerases. AGR13 exhibit all the features of endoplasmic reticulum (ER) resident proteins by containing a signal peptide and an ER retention motif. AGR2 and AGR3 can be secreted in the extracellular milieu to trigger cell migration, and for AGR2 only, epithelialtomesenchymal transition, chemoattraction of monocytes and myofibroblast activation. 6 Moreover, AGR2 is strongly expressed in endodermderived organs (lung stomach, colon prostate, intestine) and as such, is almost exclusively expressed in mucosal epithelial cells. Its overexpression has been associated with tumour aggressiveness in various cancers. Tian et al claim that in CRC, AGR2 is only produced by tumourinfiltrating neutrophils (TANs) which impacts on the migration of CRC cells through the activation of signalling pathways depending on CD98hccCT and Rho GTPases. Although this result is interesting, it is surprising that no expression of AGR2 is detected in tumour epithelial cells. We aimed at further documenting the expression levels of AGR2 protein in TANinfiltrated CRC. To do so, 21 CRC samples from our European institutions and covering 3 aetiologies including microsatellite stable (MSS) and mismatch repair and instable (MSI) adenocarcinomas (ADK) and CRC from patients with inflammatory bowel disease (IBD) associated with primary sclerosing cholangitis (IBD+PSC) or not (IBDCRC) at stages III/IV. These samples were processed for immunohistochemistry with antiAGR2 (antiAGR2 (M03) 1C3, Abnova, 1/800) and antiMPO antibody as a neutrophil marker (antiMPO (MAB3174), R&D System, 1/1000) and the images quantified (figure 1A,B). We also show data from the Human Protein Atlas (figure 1C,D). Mutually exclusive staining of AGR2 and MPO in CRC stages III/IV from various aetiologies were found thereby indicating that if TAN do express AGR2, it is marginal (figure 1A). Riener et al showed that AGR2 expression is lost or decreased in the majority of leftsided CRC, and is significantly associated with reduced overall patient survival suggesting that AGR2 might be a tumour suppressor. However, these results neither did correspond to the Tian et al article nor to their previous report in which results in discrepancies were speculated to come from variabilities in subjects or experimentalists. Hence one should rely on large international cohorts of patients with CRC to link AGR2 expression to specific aetiologies. Recent results from our laboratories did not confirm AGR2 loss of expression Letter