In this study we evaluated Treg as measured by the CD4+
phenotype for its value as a cellular biomarker at GVHD onset. Because Tregs rely on exogenous IL-2 in order to function (17
), situations characterized by impaired IL-2 production might alter Treg frequency. Calcineurin inhibitors, are frequently administered to allogeneic BMT patients as in this study, and have been shown to impair the expansion and suppressive function of Tregs in experimental models (12
). Treg frequency at day + 30 after autologous and after allogeneic BMT in patients without GVHD were similar, suggesting that allogeneic transplant recipients receiving tacrolimus have similar Treg recovery to autologous transplant recipients without tacrolimus. However, absolute Tregs were less in allogeneic BMTs patients with no GVHD compared to autologous BMT patients, an effect attributed to lower absolute lymphocyte counts. This supports a study in which low dose cyclosporine had no effect on CD4+
cell frequencies at day +30 following T cell depleted BMT (10
Previous studies measuring Tregs following BMT have not analyzed their frequency at time of GVHD onset. We observed that Treg frequencies measured within 24hrs of GVHD diagnosis were significantly less than allogeneic patients without GVHD, and correlated inversely with GVHD severity. This association supports an earlier report from Johns Hopkins University that demonstrated a correlation of FOXP3 mRNA with GVHD severity when using samples at GVHD onset (26
). This finding has not been consistently confirmed in other studies (10
). As a single biomarker, Treg frequency at GVHD onset had only modest diagnostic value (AUC = 0.69), but the inverse relationship between Treg frequency and GVHD severity demonstrates its utility. It should also be noted that we measured Tregs in the peripheral blood, not in the target tissue, where they presumably exert their greatest effect. Interestingly, in a small cohort of patients we observed an overall decrease in Tregs prior to clinical onset of GVHD, suggesting that a decline in Treg frequency may herald onset of the disease. Although these analyses are correlative in nature, this observation indicates that systematic measurements of Treg frequencies may be able to predict the occurrence of GVHD and extends the findings of a previous study in which the frequency of CD4+
T cells decreased during the initial phases of GVHD (27
Importantly, Treg frequency measured at GVHD onset may have prognostic value, which is a major feature of clinically relevant biomarkers (28
). In our study, Treg frequency of less than 0.5% correlated with poor outcomes (maximum overall GVHD grade, NRM, OS). This finding suggests that prospective measurement of Treg frequency at GVHD onset may improve standard clinical GVHD grading and further risk stratify groups of patients. Identification of patients at high risk for severe GVHD early in their transplant course may impact clinical decisions to include more stringent monitoring and/or intense treatment.
Consistent with its prognostic value, Treg frequency at GVHD onset correlated with eventual treatment response at four weeks. It is not known whether altering immunosupression in patients with low Treg frequencies might subsequently improve outcomes through the in vivo
expansion of Tregs. Recently, treatments such as rapamycin and extracorporeal photophoresis have been shown to increase Tregs and protect from experimental GVHD (29
). Prospective studies of Treg expansion for the treatment and prevention of GVHD in high risk patients are therefore warranted.