Prostaglandin E₂ (PGE₂) Alters the Molecular and Functional Properties of Umbilical Cord Blood T Cells Via Modulating Wnt/β-Catenin Signaling

Abstract 226

Umbilical cord blood transplantation (UCBT) has extended the availability of hematopoietic stem cell transplantation (HSCT) to patients without compatible adult stem cell donors. However, prolonged time to engraftment, delayed immunologic reconstitution and late memory T cell skewing compromise the favorable outcome of UCBT. Studies in zebrafish and mouse models have shown that the prostaglandin compound, 16,16 dimethyl prostaglandin E₂ (PGE₂) increases HSC number, homing and engraftment by modifying the Wnt signaling cascade through cAMP/PKA-mediated stabilization of β-catenin and expression of Wnt target genes. We performed a Phase Ib clinical trial of double UCBT (dUCBT), using one untreated and one ex vivo PGE₂-treated UCB unit, to determine safety of this uproach. 12 subjects with hematologic malignancies were enrolled (median age 58.1 years (19.8–66.5)). The median time to engraftment was 17.5 days, superior to a 21 day median for a historic control group (n=53; p=0.045). The PGE₂-UCB was the dominant source of hematopoiesis in 10 of 12 subjects (p=0.039) with full T cell chimerism as early as 13 days after dUCBT, suggesting that PGE₂ regulated the outcome of UCBT by T cell-dependent mechanisms. Here we examined whether PGE₂ might alter the properties of UCB T cells thereby affecting T cell reconstitution after UCBT. We determined that EP2 and EP4 receptors were constitutively expressed in UCB T cells and mediated increase of intracellular cAMP in response to PGE₂. PGE₂ modulated the Wnt/β-catenin pathway in UCB T cells as determined by upregulation of β-catenin and expression of Wnt target genes including Lef1, Tcf7 and Runx1. Similarly to pharmacologic activation of Wnt/β-catenin signaling by the Gsk3β inhibitor TWS119, PGE₂ inhibited proliferation of UCB T cells in response to stimulation via TCR/CD3 and CD28. Under these conditions, a significant proportion of T cells expressed naïve immunophenotypic features with high levels of CD45RA, CD62L and CD127. To examine whether these PGE₂-mediated effects on UCB T cells might have in vivo implications we assessed reconstitution of T lymphocytes in PGE₂-UCBT recipients and compared the pattern of recovery with that of dUCBT recipients without PGE₂ treatment. The numbers of total CD3⁺ T lymphocytes, CD4⁺ and CD8⁺ T subsets remained significantly lower (p=0.036) in PGE₂-UCBT recipients during the first year after UCBT. In contrast, T cell receptor excisions circles (TRECs), a marker of naïve post-thymic T cells and indicator of thymic function, were substantially higher in PGE₂-UCBT recipients and never reached the nadir observed in dUCBT recipients without PGE₂, who had values below the limit of detection through 100 days. Importantly, PGE₂-UCBT recipients displayed potent antiviral immunity as determined by the reduced incidence of CMV viremia and no cases of EBV-mediated posttransplant lymphoproliferative disorder (PTLD), which caused significant morbidity and mortality in dUCBT recipients without PGE₂. In mouse models Wnt/β-catenin signaling promotes the generation of a central memory/stem cell memory T cell population with naïve immunophenotypic features but potent immune function. To examine whether such PGE₂-mediated Wnt/β-catenin imprinting might be evident in PGE₂-UCBT recipients, we examined expression of the Wnt/β-catenin target Eomes, a transcription factor that links the long-term memory CD8⁺ T cells to effector potency and protective immunity. Real time qPCR revealed that expression of Eomes was significantly elevated in the PBMCs of PGE₂-UCBT recipients compared to control UCBT recipients. Consistent with the role of Wnt/β-catenin to maintain a central memory/stem cell memory phenotype in CD8⁺ cells, there was an increased fraction of CD62L⁺ cells within the CD8⁺ populations in recipients of PGE₂-UCBT. Our in vitro and in vivo findings indicate that PGE₂-UCB treatment improves HSC engraftment while preserving a naïve T cell compartment and favoring the generation of long-lived memory CD8⁺ cells via Wnt-mediated gene programming. This novel treatment might significantly improve the outcome of UCBT where delayed engraftment and impaired immunity are serious causes of morbidity and mortality.