Chronic graft versus host disease (cGVHD) is the major cause of late morbidity and mortality after allogeneic hematopoietic cell transplantation. Little therapeutic progress has been made due to inadequate animal models. We (Blazar lab) developed a new cGVHD model resulting in broad multi-organ injury and advanced the field by making the important observation that IgG2c deposited in tissues causes cGVHD. Germinal centers (GCs) are sites where B cells proliferate, differentiate, undergo somatic mutation to produce plasma cells that secrete high amounts of immunoglobulin. GC formation is supported by subsets of T cells, Tfollicular helper (TFH) cells, and suppressed by Tfollicular regulatory (TFR) cells. Our central hypothesis is that cGVHD is modulated by the balance between GC facilitating TFH and suppressive TFR cells that can controlled by altering intracellular signalling and shifting from a dependency upon glycolysis, that favour TFH cells, to fatty acid oxidation (FAO) that favors TFR over TFH cells. TFH, TFR and GC B cells metabolic requirements have not been defined in scleroderma cGVHD models that have a distinct pathophysiology and represent a large unmet medical need will be pursued by the MacDonald lab.
TCR/CD28 and IL-2R signalling via the phosphatidylinositol 3-kinase (PI3K) pathway are essential for T cell activation. Resting and activated Tnaive, Teffectors (Teffs), Tmemory and Tregs use distinct energy pathways. In acute GVHD, rapidly dividing Teffs use aerobic glycolysis and oxidative phosphorylation. Thus, we will determine metabolism pathways in cGVHD, which should be under lower metabolic stress than acute GVHD. We then will pursue perturbations favoring TFRs vs TFHs and GC B cells. The lipid phosphatase PTEN (phosphatase and tensin homolog on chromosome 10) is the primary PI3K inhibitor. CD28 costimulation is required for T cells to increase glucose uptake and the glycolytic rate in response to activation. Tregs and Teffector cells have different metabolic needs, that correlate with their proliferative rates and survival. We have seen during acute GVHD, oxygen consumption rates by Seahorse, lactate, glucose transporter GLUT1, mitochondrial membrane potential and mass as well as superoxide production are significantly higher and pyruvate lower than syngenic BMT controls. In contrast, Tregs express low GLUT1 levels, and high levels of phospho-AMP-activated kinase that inhibit mTOR complex 1. Our data from mice that globally overexpress PTEN suggests the metabolic signature of Tregs may be a result of elevated PTEN activity. Furthermore, our evidence shows that PTEN-deleted Tregs use glycolysis vs. oxidative phosphorylation for energy demands, which may lead to loss of regulatory capacity and conversion into pathogenic Teffs. This may be important for lineage stability; blockade of glycolysis in activated Teffs promotes Treg induction.
Additional preliminary data demonstrate that (exogenous) oral administration of short chain fatty acids can be used to treat cGVHD. Additionally, pathogenic Th17 cells rely on ACC1-mediated de novo fatty acid synthesis and CD8+ T cells rely on de novo lipogenesis for proliferation and survival. ACC1 inhibition favors induction of Tregs by preventing acetyl-CoA conversion to malonyl-CoA so that cells cannot produce lipids needed for rapidly proliferating cells and without malonyl-CoA, increased FAO occurs, favoring Tregs. Conversely, pathogenic Th17 cells rely on ACC1-mediated de novo fatty acid synthesis and CD8+ T cells rely on de novo lipogenesis for proliferation and survival. Our preliminary data with ACC1 knockout T cells and a small molecule ACC1 inhibitor has demonstrated that cGVHD can be prevented and treated, respectively. Lastly, metformin, a drug in widespread use world wide to reduce insulin resistance and treat metabolic syndrome, has been effective in treating cGVHD. Metformin favors FAO. Confirmatory studies are needed as is direct evidence that FAO is favored in TFR cells under these conditions. We propose that elevated glycolysis leads to loss of Tregs, while FAO favors Tregs and can be used to treat cGVHD.
Hypothesis: CD28 and PI3K downstream signaling pathways regulate cGVHD
We will test the hypotheses that (A) signals from CD28 and receptors that activate PI3K modulate cGVHD via TFH/TFR balance and (B) that favouring lipid oxidation (vs glycolysis) in cGVHD mice will restore TFH/TFR balance. Innovation: Metabolic profiling of cGVHD has not been undertaken and no one has treated cGVHD by altering metabolism. Also novel are our small molecule approaches to alter signalling via PTEN, PI3K in Tregs.
To apply for this PhD project; please contact the Project Supervisor/s: Dr Kelli MacDonald | Kelli.MacDonald@qimrberghofer.edu.au; Professor Bruce Blazar | firstname.lastname@example.org.