The HPSE Gene Insulator-A Novel Regulatory Element That Affects Heparanase Expression, Stem Cell Mobilization, and the Risk of Acute Graft versus Host Disease.

The HPSE gene encodes heparanase (HPSE), a key player in cancer, inflammation, and autoimmunity. We have previously identified a strong HPSE gene enhancer involved in self-regulation of heparanase by negative feedback exerted in a functional rs4693608 single-nucleotide polymorphism (SNP) dependent manner. In the present study, we analyzed the HPSE gene insulator region, located in intron 9 and containing rs4426765, rs28649799, and rs4364254 SNPs. Our results indicate that this region exhibits HPSE regulatory activity. SNP substitutions lead to modulation of a unique DNA-protein complex that affects insulator activity. Analysis of interactions between enhancer and insulator SNPs revealed that rs4693608 has a major effect on HPSE expression and the risk of post-transplantation acute graft versus host disease (GVHD). The C alleles of insulator SNPs rs4364254 and rs4426765 modify the activity of the HPSE enhancer, resulting in altered HPSE expression and increased risk of acute GVHD. Moreover, rs4426765 correlated with HPSE expression in activated mononuclear cells, as well as with CD3 levels and lymphocyte counts following G-CSF mobilization. rs4363084 and rs28649799 were found to be associated with CD34+ levels. Our study provides new insight into the mechanism of HPSE gene regulation and its impact on normal and pathological processes in the hematopoietic system.

CMV Seropositive Status Increases Heparanase SNPs Regulatory Activity, Risk of Acute GVHD and Yield of CD34+ Cell Mobilization.

Heparanase is an endo-ß-glucuronidase that is best known for its pro-cancerous effects but is also implicated in the pathogenesis of various viruses. Activation of heparanase is a common strategy to increase viral spread and trigger the subsequent inflammatory cascade. Using a Single Nucleotide Polymorphisms (SNP)-associated approach we identified enhancer and insulator regions that regulate HPSE expression. Although a role for heparanase in viral infection has been noticed, the impact of HPSE functional SNPs has not been determined. We investigated the effect of cytomegalovirus (CMV) serostatus on the involvement of HPSE enhancer and insulator functional SNPs in the risk of acute graft versus host disease (GVHD) and granulocyte-colony stimulating factor related CD34+ mobilization. A significant correlation between the C alleles of insulator rs4364254 and rs4426765 and CMV seropositivity was found in healthy donors and patients with hematological malignancies. The risk of developing acute GVHD after hematopoietic stem cell transplantation was identified only in CMV-seropositive patients. A significant correlation between the enhancer rs4693608 and insulator rs28649799 and CD34+ cell mobilization was demonstrated in the CMV-seropositive donors. It is thus conceivable that latent CMV infection modulates heparanase regulatory regions and enhances the effect of functional SNPs on heparanase function in normal and pathological processes.

Modification of heparanase gene expression in response to conditioning and LPS treatment: strong correlation to rs4693608 SNP.

Heparanase is an endo-ß-glucuronidase that specifically cleaves the saccharide chains of HSPGs, important structural and functional components of the ECM. Cleavage of HS leads to loss of the structural integrity of the ECM and release of HS-bound cytokines, chemokines, and bioactive angiogenic- and growth-promoting factors. Our previous study revealed a highly significant correlation of HPSE gene SNPs rs4693608 and rs4364254 and their combination with the risk of developing GVHD. We now demonstrate that HPSE is up-regulated in response to pretransplantation conditioning, followed by a gradual decrease thereafter. Expression of heparanase correlated with the rs4693608 HPSE SNP before and after conditioning. Moreover, a positive correlation was found between recipient and donor rs4693608 SNP discrepancy and the time of neutrophil and platelet recovery. Similarly, the discrepancy in rs4693608 HPSE SNP between recipients and donors was found to be a more significant factor for the risk of aGVHD than patient genotype. The rs4693608 SNP also affected HPSE gene expression in LPS-treated MNCs from PB and CB. Possessors of the AA genotype exhibited up-regulation of heparanase with a high ratio in the LPS-treated MNCs, whereas individuals with genotype GG showed down-regulation or no effect on HPSE gene expression. HPSE up-regulation was mediated by TLR4. The study emphasizes the importance of rs4693608 SNP for HPSE gene expression in activated MNCs, indicating a role in allogeneic stem cell transplantation, including postconditioning, engraftment, and GVHD.