Non-myeloablative bone marrow transplantation for Mucopolysaccharide diseases

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Abstract

The Mucopolysaccharide (MPS) diseases are a group of lysosomal storage disorders, caused by a lack of the enzymes required for catabolism of glycosaminoglycans (GAGs), leading to severe neurological decline, skeletal deformities, organomegaly, cardiac and respiratory compromise, and premature death. The severe form of MPS I, Hurler syndrome, can be successfully treated using haematopoietic stem cell transplantation (HSCT), but the risks associated with myeloablation and immune suppression limit the broader application of HSCT to attenuated diseases. Successful engraftment in MPS I has been difficult to achieve, and requires fully myeloablative conditioning, whilst reduced intensity conditioning is a risk factor for graft rejection. Non-myeloablative conditioning generating reliable graft acceptance and high donor chimerism could increase safety and applicability of HSCT in genetic disease, therefore the aim of this research was to identify such a regimen in a clinically relevant mouse model of HSCT.Conditioning regimens developed in existing mouse models of HSCT have had limited clinical success, and often require clinically unachievable high cell doses or less stringent strain combinations to overcome allogeneic transplant rejection. To improve clinical relevance we used CBA donors and C57BL/6 recipients, which require full myeloablation with busulfan and immune suppression using non-depleting anti-CD4 and anti-CD8 monoclonal antibodies for engraftment of low cell doses across a major histocompatibility complex barrier. In syngeneic transplant donor chimerism was improved by generating a greater ratio of donor:recipient haematopoietic cells in the bone marrow initially, therefore we tested granulocyte colony stimulating factor (G-CSF), high cell dose and stem cell niche disruption and compared this to anti-CD40L costimulatory blockade in allogeneic transplant performed with a reduced dose of busulfan that was insufficient for graft acceptance. Despite improvements in initial engraftment with some of these treatments, only combined signal 1 and 2 T cell blockade were effective in reducing the dose of busulfan required for long-term graft acceptance. Early detection of MPS is important in treatment success; good disease biomarkers are vital, and biomarkers suitable for monitoring treatment outcome in MPS are lacking. We evaluated serum heparin cofactor II-thrombin (HCII-T) complex for MPS. We determined optimal sample collection and storage conditions, assay limitations and developed measurement in dried blood spots. Dermatan sulphate has a greater effect on in vivo HCII-T complex formation than heparan sulphate, thus in the MPS mouse models HCII-T is a reliable biomarker for MPS I, but not MPS IIIA or IIIB. HCII-T is greatly elevated in MPS I, II and VI patients, who all store dermatan sulphate, but it is also elevated by a small but significant amount in MPS III patients, who store heparan sulphate. HCII-T was also measured longitudinally in MPS I, II and VI patients, compared to an existing clinical biomarker, and validated against clinical outcomes to show that it is a good biomarker of short-term treatment outcomes and responds rapidly to perturbations in treatment.Finally, we determined whether an engraftment defect was observed in the MPS I mouse model, and show that this is present following both syngeneic and allogeneic HSCT. The effect of enzyme replacement therapy (ERT) and anti-inflammatory treatment prior to allogeneic HSCT was investigated, and initial results suggest that ERT, but not ibuprofen, may improve HSCT outcome.Overall, a clinically relevant mouse model of allogeneic HSCT has been developed and used to determine a non-myeloablative conditioning regimen that generates high levels of donor chimerism with a minimal dose of busulfan and blockade of both signal 1 and 2 of T cell activation. The conditions required to observe an engraftment defect in MPS I mice have also been defined, and preliminary studies have suggested that ERT, but not anti-inflammatory treatment, may overcome the engraftment defect in MPS I. Alongside this work, the HCII-T biomarker has been evaluated in MPS mouse models and patients, determining that it correlates well with short-term treatment outcomes. The techniques and models developed here will provide an excellent basis for further work in developing non-myeloablative conditioning for bone marrow transplant in MPS I.

Keyword(s)

Biomarkers; Bone marrow transplant; Haematopoietic stem cell transplant; Mucopolysaccharidosis; Reduced intensity conditioning

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