Syk-induced phosphatidylinositol-3-kinase activation in Epstein-Barr virus posttransplant lymphoproliferative disorder.

Posttransplant lymphoproliferative disorder (PTLD)-associated Epstein-Barr virus (EBV)+ B cell lymphomas are serious complications of solid organ and bone marrow transplantation. The EBV protein LMP2a, a B cell receptor (BCR) mimic, provides survival signals to virally infected cells through Syk tyrosine kinase. Therefore, we explored whether Syk inhibition is a viable therapeutic strategy for EBV-associated PTLD. We have shown that R406, the active metabolite of the Syk inhibitor fostamatinib, induces apoptosis and cell cycle arrest while decreasing downstream phosphatidylinositol-3'-kinase (PI3K)/Akt signaling in EBV+ B cell lymphoma PTLD lines in vitro. However, Syk inhibition did not inhibit or delay the in vivo growth of solid tumors established from EBV-infected B cell lines. Instead, we observed tumor growth in adjacent inguinal lymph nodes exclusively in fostamatinib-treated animals. In contrast, direct inhibition of PI3K/Akt significantly reduced tumor burden in a xenogeneic mouse model of PTLD without evidence of tumor growth in adjacent inguinal lymph nodes. Taken together, our data indicate that Syk activates PI3K/Akt signaling which is required for survival of EBV+ B cell lymphomas. PI3K/Akt signaling may be a promising therapeutic target for PTLD, and other EBV-associated malignancies.

Activation of the JAK/STAT pathway in Epstein Barr virus+-associated posttransplant lymphoproliferative disease: role of interferon-gamma.

Epstein Barr virus (EBV) is associated with B-cell lymphomas in posttransplant lymphoproliferative disease (PTLD). Latent membrane protein 1 (LMP1), the major oncogenic protein of EBV, promotes tumorigenesis through activation of NF-kappaB, Erk, p38, JNK and Akt. The Jak/STAT signal transduction pathway is also constitutively active in PTLD-associated EBV(+) B-cell lymphomas. Here we determine the mechanism of Jak/STAT activation in EBV(+) B-cell lymphomas and the role of LMP1 in this process. Immunoprecipitation studies revealed no direct interaction of LMP1 and JAK3, but known associations between JAK3 and common gamma chain, and between LMP1 and TRAF3, were readily detected in EBV(+) B cell lines from patients with PTLD. An inducible LMP1 molecule expressed in EBV(-) BL41 Burkitt's cells demonstrated STAT activation only after prolonged LMP1 signaling. While LMP1 induced IFN-gamma production in BL41 cells, IFN-gamma receptor blockade and IFN-gamma neutralization prior to LMP1 activation markedly decreased STAT1 activation and expression of LMP1-driven IFN-gamma inducible genes. Understanding the mechanisms by which EBV induces cellular signal transduction pathways may facilitate development of new treatments for PTLD.

EBV B lymphoma cell lines from patients with post-transplant lymphoproliferative disease are resistant to TRAIL-induced apoptosis.

Lymphomas associated with post-transplant lymphoproliferative disease (PTLD) represent a significant complication of immunosuppression in transplant recipients. In immunocompetent individuals, EBV-specific cytotoxic T lymphocytes (CTL) prevent the outgrowth of activated B lymphoblasts through apoptosis induction. Soluble versions of TNF-related apoptosis-inducing ligand/Apo2 ligand (TRAIL) can induce apoptosis in numerous tumor cell types. Given the therapeutic potential of TRAIL, we examined the sensitivity of EBV+ spontaneous lymphoblastoid cell lines (SLCL) derived from patients with PTLD to treatment with soluble TRAIL. Despite abundant expression of TRAIL receptors (TRAIL-R), resistance to TRAIL-induced apoptosis was observed in all SLCL examined. This resistance could not be overcome by concomitant treatment with several pharmacological agents. Unlike BJAB positive control cells, for each SLCL tested, cleavage and activation of caspase 8 was inhibited due to failed recruitment of FADD and caspase 8 to TRAIL receptors upon stimulation. Further indicative of a proximal defect, TRAIL receptor aggregation could not be detected on the cell surface of SLCL following ligand engagement. These results suggest that the use of TRAIL for eliminating PTLD-associated tumors may be of limited clinical utility, and illustrate another mechanism by which EBV+ B lymphoma cells can evade tumor surveillance at the level of death receptor signaling.