Gene-Expression Profiling of Graft-Versus-Host Disease by Oligonucleotide Microarrays.

Graft-versus-host disease (GvHD) is the life-threatening complication of allogeneic hematopoietic stem cells transplantation (allo-HSCT). GvHD is mediated by an immune reaction of donor T lymphocytes against recipient's tissues/cells. Acute GvHD (aGvHD) appearing within the first 100 days post transplantation is the most frequent cause of recipient's death and characterization of biomarkers for early prediction of aGvHD or resistance to corticoid treatment could be of great clinical relevance. Biomarker panels for aGvHD are currently extensively studied by proteomic and gene expression based approaches but so far very few markers were described and validated (Kaiser et al., 2004; Baron et al., 2007; Weissinger et al., 2007; Paczesny et al., 2009).

In this study, we performed microarray gene expression analysis (whole genome Human OneArray, Phalanx) of 43 leukemia patients who received allo-HSCT. Mononuclear cells isolated from peripheral blood samples (Ficoll-Paque) were collected at i) 14 days before transplantation, ii) 20 and iii) 30 days after transplantation and iv) at the time of aGvHD manifestation. We also performed gene-expression analysis for corticoid-resistant vs. corticoid-sensitive aGvHD cases.

The SAM supervised analysis of samples collected at day +20 post transplantation revealed set of differentially expressed genes between groups of patients that developed aGvHD vs. aGvHD-free recipients. Among others, genes CASP1 (encoding caspase 1, protein implicated in apoptosis), HLA-DRA (member of MHC class II family) and LILRA3 (leukocyte immunoglobulin-like receptor, subfamily A member 3) showed the highest difference in expression. Gene expression with regard to corticoid response was analyzed at the time of first aGvHD manifestation. The SAM supervised analysis of gene expression between patients with corticoid-sensitive aGvHD (n=10) or aGvHD resistant to corticoid treatment (n=4) revealed a set of significantly differentially expressed genes including NR4A2 (nuclear receptor subfamily 4; member of the steroid-thyroid hormone-retinoid receptor superfamily), DEDD2 (death effector domain containing 2), TREM1 (triggering receptor expressed on myeloid cells 1), TPK1 (thiamin pyrophosphokinase 1) and HBEGF (heparin-binding EGF-like growth factor).

Oligonucleotide microarrays proved to be a useful tool for expression studies of hematological malignancies and our work shows that they may help to identify markers for early diagnosis/treatment of aGvHD. The limited patients' cohort and their heterogeneity complicate such studies. Our future effort will be focused on experimental group extension, cohort uniformity and verification of the obtained data.

This work is supported by the grant NS9683-4/2008 provided by the IGA MH of the Czech Republic, and MSM0021622430 provided by MEYS of Czech Republic

No relevant conflicts of interest to declare.