This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.ABSTRACTHodgkin's disease and Non-Hodgkin's lymphomas (NHL) are associated with Epstein-Barr virus (EBV). Our group has successfully generated EBV-specific CTL (EBV-specific CTL) in patients with EBV-positive Hodgkin's disease.1 After infusion these CTL home to the tumor sites and persist in the circulation for up to 9 months. However although transient clinical benefits were seen, only two complete responses were achieved with CTL therapy alone. At present the anti-lymphoma activity of EBV-specific CTL is limited by several factors, including 1) EBV-specific CTL lines generated by standards methods are dominated by T-cell clones not reactive to the subdominant EBV proteins LMP1 and LMP2 expressed in lymphoma, and 2) infused EBV-specific CTL did not significantly expand in vivo after adoptive transfer. We are currently conducting 2 RAC and FDA approved Phase I clinical trials as an initial step to overcome both limitations. The majority of EBV-positive lymphomas in immunocompetent patients express a limited subset of EBV antigens, EBNA1, LMP1, and LMP2. In the first clinical trial we are generating CTL against the subdominant LMP2 (LMP2-specific CTL) in an effort to increase the tumor-specificity of the infused CTL lines. One strategy to enhance the in vivo expansion of T cells is to reduce the lymphocyte pool prior to T-cell infusion. This should result in homeostatic proliferation of the infused T cells to restore the lymphoid compartment. In the second clinical trial we are therefore evaluating a pair of monoclonal antibodies, targeting the CD45 antigen (CD45 MAbs), to lymphodeplete patients prior to adoptive transfer of LMP2-specific CTL. The preliminary results of both studies are encouraging. Infusion of LMP2-specific CTL was safe and resulted in 4 complete responses in 8 patients treated with active disease. The use of CD45 MAbs was safe, resulted in transient lymphodepletion, and enhanced CTL expansion in comparison to giving CTL alone. Although these results are encouraging we hypothesize that broadening the tumor-specificity of CTL will further increase anti-tumor effect of infused CTL. Since CTL escape mutants have been described for tumors as well as viruses and the expression of LMP1 and LMP2 in tumors is heterogeneous, we propose in this Phase I clinical the infusion of LMP1- and LMP2-specific CTL after lymphodepletion to 1) ensure that good CTL epitopes are available regardless of the patient's HLA type and 2) generate the broadest CTL response possible against the malignant lymphoma cells.I. HYPOTHESIS We hypothesize that a) the anti-tumor activity of CTL infused into patients with EBV-positive Hodgkin's disease and Non-Hodgkin's Lymphoma (NHL) can be increased by broadening the specificity of infused T cells to the EBV antigens LMP1 and LMP2.b) the anti-tumor activity of CTL infused into patients with EBV-positive Hodgkin's disease and NHL can be increased by prior depletion of the T cell compartment to allow for expansion of infused tumor-specific T cells. c) that CD45 monoclonal antibodies (Mabs) will safely and substantially deplete normal T cells, and their short half-life will allow infusion of LMP1- and LMP2-specific CTL shortly after CD45 MAb administration, enabling their expansion prior to the recovery of endogenous T cells. II.
SPECIFIC AIMS Aim 1) To perform a phase I study of LMP1- and LMP2-specific CTL infusion after administration of CD45 MAbs in patients with Hodgkin's disease and NHL. Primary clinical endpoints of this study will be safety and hematopoietic and immune reconstitution. In our previous studies in animal and humans, lytic CD45 MAbs safely depleted lymphocytes of all lineages, but spared hematopoietic progenitor cells, which have a low level of target antigen expression. The antibodies have a half-life of circa 8 hours and therefore cause only transient neutropenia with no measurable stem cell toxicity. The degree of T cell depletion will be determined by total and differential T cell counts and phenotyping. The clinical implications of transient lymphopenia in these patients will be monitored with emphasis on the occurrence of potential side effects such as infection.
Aim 2) To analyze the effect of CD45 MAb-mediated host T cell depletion on the expansion, persistence and anti-tumor effector function of subsequently infused LMP1- and LMP2-specific CTL. We will use multimer staining and functional ELISPOT assays to monitor the expansion and persistence of adoptive transferred LMP1- and LMP2-specific CTL. Anti-tumor effects will be evaluated by measuring cell-free EBV-DNA in plasma, tumor size on imaging and clinical outcome.
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