Novel Mediators of Disease Pathogenesis
Biological Phenotyping and Insights into Disease Pathogenesis
The CVBRU has funded a PhD student, Ms Amy Shepherd, to work with Dr Lawrie. Ms Shepherd commenced her MRes project in October 2008 and the PhD programme a year later on the project “Characterising T-cell phenotypes and investigating Heat Shock Protein interaction in Pulmonary Arterial Hypertension.” Below is an update on the progress made.
T-cell Phenotyping of Patient Samples: Ms Shepherd has established a 12-colour flow cytometry protocol to phenotype lymphocyte populations in patients with PH, including basic CD4/CD8, regulatory T-cells and activation state. She has compared 4 different criteria on a cohort of around 25 patient samples obtained via the CVBRU.
The team has found interesting and novel differences between classification of PH in terms of T-cell profile and activation. This work has been submitted in abstract form to the British Thoracic Society Winter Meeting. The current focus for this work is to expand the disease and control groups (healthy and systemic sclerosis) whilst examining recruited patients at follow-up to determine the effect of treatment/disease progression on their T-cell profile.
Ms Shepherd is also establishing a functional assay to assess the anti-proliferative capacity of T-regs isolated from patients.
HSP70 Expression in Serum of Patients with PH: HSP70 can be expressed and secreted from endothelial cells. High expression of HSP70 has been reported in the serum of patients with systemic hypertension. We are now assaying serum from all CVBRU PH patients; including the 25 that Ms Shepherd has T-cell profiles on, for HSP70 levels, to correlate this with T-cell profiles and potential clinical phenotype data.
TNF Related Apoptosis-Inducing Ligand (TRAIL)
Work from Dr Lawrie has shown that TRAIL protein is unregulated in end stage PH patients. Following a British Heart Foundation Clinical Research Training Fellowship awarded to Dr Abdul Hameed, this has now been validated in two rodent modesl of PH. Dr Hameed has demonstrated that either genetic deletion of TRAIL in a murine model, or treatment with an anti-TRAIL antibody in a rat model, prevents development of PH. Furthermore, administration of recombinant TRAIL to the TRAIL knockout mouse restores development of PH.
This data suggests that TRAIL is required for disease progression in both models. We are currently assessing the ability of anti-TRAIL treatment to stabilise/reverse disease pathogenesis. Full analysis of this date is ongoing but the signs are very encouraging that TRAIL may be a potential new target in PH.
Osteoprotegerin (OPG):
Published work from Dr Lawrie in preclinical models and CVBRU patients has indicated that OPG protein is unregulated in end stage PH and is downstream of a number of key pathways in PH. This data and others suggest OPG may be a novel target for the treatment of PH.
The molecular structure and signalling pathways related to OPG and vascular cells are not fully understood. To enable further therapeutic targeting of this pathway the CVBRU is providing support through the appointment of a post-doctoral structural biologist to solve the crystal structure of OPG as a preliminary to rational drug design of OPG antagonists.
Interleukin-1 (IL-1):
Previous studies have demonstrated that IL-1 receptor antagonist (Il-1Ra) can prevent PH in a fatal rat model. We have demonstrated that administering IL-1Ra for four weeks normalises both haemodynamics and pulmonary vascular remodelling in a murine PAH model. This avenue is highly promising for translation since a number of drugs are already on the market to target IL-1. We thus plan to undertake pilot studies with IL-1Ra in patients with PH.
