Comparison of Methods for Testing Mismatch Repair Status in Endometrial Cancer.

Approximately 20-30% of endometrial carcinomas (EC) are characterized by mismatch repair (MMR) deficiency (dMMR) or microsatellite instability (MSI), and their testing has become part of the routine diagnosis. The aim of this study was to establish and compare the MMR status using various approaches. Immunohistochemistry (IHC), PCR-based MSI, and the detection of defects in the four key MMR genes (MLH1, PMS2, MSH2, and MSH6) via methylation-specific multiplex ligation-dependent probe amplification (MLPA) and targeted next-generation sequencing (NGS) were performed. MSH3 expression was also evaluated. A set of 126 early-stage EC samples were analyzed, 53.2% of which were dMMR and 46.8% of which were proficient MMR (pMMR) as determined using IHC, whereas 69.3% were classified as microsatellite stable, while 8.8% and 21.9% were classified MSI-low (MSI-L) and MSI-high (MSI-H), respectively. In total, 44.3% of the samples showed genetic or epigenetic alterations in one or more genes; MLH1 promoter methylation was the most common event. Although acceptable concordance was observed, there were overall discrepancies between the three testing approaches, mainly associated with the dMMR group. IHC had a better correlation with MMR genomic status than the MSI status determined using PCR. Further studies are needed to establish solid conclusions regarding the best MMR assessment technique for EC.

EBI2 regulates CXCL13-mediated responses by heterodimerization with CXCR5.

B-cell movement into lymphoid follicles depends on the expression of the chemokine receptor CXCR5 and the recently reported Epstein-Barr virus-induced receptor 2 (EBI2). In cooperation with CXCR5, EBI2 helps to position activated B cells in the follicle, although the mechanism is poorly understood. Using human HEK293T cells and fluorescence resonance energy transfer (FRET) techniques, we demonstrate that CXCR5 and EBI2 form homo- and heterodimers. EBI2 expression modulated CXCR5 homodimeric complexes, as indicated by the FRET(50) value (CXCR5 homodimer, 0.9851±0.0784; CXCR5 homodimer+EBI2, 1.7320±0.4905; P<0.05). HEK293T cells expressing CXCR5/EBI2 and primary activated murine B cells both down-modulated CXCR5-mediated responses, such as Ca(2+) flux, cell migration, and MAPK activation; this modulation did not occur when primary B cells were obtained from EBI2(-/-) mice. The mechanism involves a reduction in binding affinity of the ligand (CXCL13) for CXCR5 (K(D): 5.05×10(-8) M for CXCR5 alone vs. 1.49×10(-7) M for CXCR5/EBI2) and in the efficacy (E(max)) of G-protein activation in CXCR5/EBI2-coexpressing cells (42.33±4.3%; P<0.05). These findings identify CXCR5/EBI2 heterodimers as functional units that contribute to the plasticity of CXCL13-mediated B-cell responses.