Identification and assessment of molecular biomarkers in haematopoietic stem cell transplantation /

PhD ThesisHaematopoietic stem cell transplantation (HSCT) has become a central treatment modality in the management of various hematologic malignancies, but it is not without treatment sequelae. The major complication of HSCT is acute or chronic graft-versus host disease (GvHD). GvHD is an immunologically mediated disease that contributes substantially to transplant-related morbidity and mortality. One reason for the lack of progress in the treatment of acute GvHD (aGvHD) is the lack of reliable biomarkers. There is a need to develop diagnostic tools that can identify patients who are at higher risk of aGvHD progression following allogeneic HSCT and predict GvHD occurrence before clinical symptoms manifest. During the past decade, many reports have identified genetic variants such as single nucleotide polymorphisms (SNPs) that influence the risk of aGvHD after allogeneic HCT. In addition, since miRNAs are key regulators of gene expression, miRNA-related SNPs including SNPs in miRNA genes and target sites may function as regulatory SNPs through modifying miRNA regulation to affect the phenotypes and disease susceptibility. Firstly, this study investigated the impact of rs2910164 and rs2431697 in miR-146a, rs3027898 in IRAK1 and rs10511792 in MICA for their association with HSCT outcome and showed that there was a significant association between carrying the C variant in rs2910164 in miR-146a and an increased non relapse mortality (NRM) post-HSCT. For rs2431697 in miR-146a, the presence of the T allele was associated with a trend towards an increased NRM in patients post-HSCT. In the case of rs3027898 in IRAK1, the C allele was associated with a decreased risk of relapse in patients which was more apparent when patients were homozygous for the C allele. For rs1051792 in MICA, this study showed that the MICA-129 Met variant was significantly associated with low overall survival (OVS) post-HSCT, which was more apparent in the group of patients receiving non-TCD treatment. This study also revealed that the presence of the MICA-129 Met allele in patients was significantly associated with an increased risk of relapse and the presence of the MICA-129 Val variant in patients was significantly associated with an increased risk of developing aGVHD post-HSCT. Investigation of gene expression and the protein levels of MICA in the GI tract showed that there was a significant association between decreased expression of MICA and aGvHD which was observed again in the case of MICA protein levels, where high levels of MICA protein were observed in patients with no active GIGvHD. Assessment of the levels of iii soluble MICA in sera of patients post HSCT showed a significant association between high levels of soluble MICA and aGvHD post-HSCT. Alongside MICA, this study investigated the mRNA and protein levels of a panel of genes (C1QTNF7, LGALS7, ANP32A, HTRA1, PIK3AP1, PSTPIPI, MSR1 and CXCL9) in RNA from blood samples and patient sera at different time points pre and post-HSCT. This study showed that there was a significant downregulation in the expression levels of MSR1 and ANP32A in aGvHD patients post-HSCT while a significant upregulation in the expression levels of CXCL9 was observed in aGvHD patients. Investigation of the association between the levels of proteins and the incidence of aGvHD showed that there was a significant association between upregulated protein levels of LGALS7 and aGvHD. Finally, a microRNA profiling in GI samples taken from aGvHD patients was performed aiming for the identification of miRNAs associated with the incidence of aGvHD in the GI tract after HSCT. This study identified 4 miRs that were dysregulated in patients in association with aGvHD, and a validation study was carried out for hsa-miR-34a-5p which expression was shown to be significantly decreased in patients with aGvHD (1-4) compared those patients with no aGvHD