| Abstract: |
Efforts to improve graft-versu-tumor (GVT) activity of alloreactive donor T cells are limited by a concomitant rise in graft-versus-host disease (GVHD): We employed experimental allogeneic bone marrow transplantation (allo-BMT) models to assess two novel strategies, in which T lineage cells were genetically engineered to enhance selective effector functions. In addition to the transgenic expression of these molecules on mature donor T cells, we also used genetically engineered precursor T cells as an "off the shelf" adoptive cell therapy.One strategy relies on T cell cytolytic molecule TNF-Related Apoptosis Inducing Ligand (TRAIL), which can induce apoptotic signals through death receptor (DR) 4 and 5 molecules' (only DRS in mice) expressed on target cells. Certain tumor cells are known to express. high levels of surface DR5 making TRAIL an attractive candidate for genetic engineering of donor T cells to enhance GVT activity. Extending previous studies indicating that TRAIL expression in donor T cells is dispensable for the development of GVHD, we evaluated the effect of TRAIL over-expression in donor T cells (mature and precursor) on GVHD and GVT. Mature T cells derived from donor B6 splenocyte's were transduced with a lentiviral TRAIL expression vector. The transduced TRAIL+ T cells were adoptively transferred on day 0 into lethally irradiated CBF1 recipients Of a T cell depleted allo-BMT, bearing LB27.4 tumors, (86 boolean AND CBF1+LB27.4) to assess their GVHD and GVT activity. TRAIL+ T dells displayed significantly enhanced antitumor immunity compared to T cells transduced with a control vector against LB27.4 tumor cell lines in vitro and upon transfer into tumor bearing allo-BMT recipients (p < 0.01)(Fig Interestingly, the recipients treated with TRAIL + T cells had significantly less GVHD. We observed increased DRS expression on host antigen presenting cells (APC) soon after the total body irradiation used in our preparative regimens, suggesting that TRAIL + donor T cells could potently eliminate host APC, resulting in less GVHD. Precursor T cells have the added benefits of reconstituting the T cell compartment without GVHD and "off the shelf" use. We generated TRAIL + precursor T cells from transduced B6 murine hematopoietic stem cells and expanded them using the 0P9-DL1 co-culture system. Adoptive transfer of B6 TRAIL + precursor T cells into syngeneic-transplanted BALB/c mice could reconstitute the T cell compartment with TRAIL expressing T cells. When challenged with localized RENCA tumors, TRAIL + precursor T cells showed enhanced antitumor activity (p < 0.05) compared to mock (GFP)-transduced controls (Fig IB).The second strategy explores the use of promyelocytic leukemia zinc finger (PLZF). PLZF is a member of the BTB-ZF (Broad complex, Tram track, Bric-boolean AND-brac-zinc finger) family Of transcription factors. In NKT and NK cells, it controls key steps in development, activation and effector functions. Its ectopic expression on conventional T cell confers NKT and NK-like properties. Recent studies using transgenic mice have shown that T cells ectopically expressing PLZF spontaneously acquire an effector/memoty phenotype and CD8+ T cells spontaneously express IFN-gamma. Using PLZF + transgenic donor (B6) T cells in A20-tumor challenged 86 BALB/c allo-BMT model, we found that PLZF + T cells have less GVHD activity while GVT activity remained intact (p < 0.001)(Fig 1C). Using adoptive transfer of CFSE-labeled donor T cells we observed that fast proliferating allo-responsive PLZF + T cells died after only a few cell cycles. We then transduced donor T cells with an expression vector and confirmed that transduced PLZF expressing donor T cells have less GVHD activity and intact GVT activity (Fig ID):In conclusion, our data suggest that adoptive therapy, with genetically engineered TRAIL Or PLZF expressing donor T cells exert less GVHD activity while displaying intact or enhanced GVT activity. Furthermore, the "off the shelf" use of genetically enhanced precursor T cells indicates a promising cell therapy strategy to enhance anti-tumor in both autologous and allogeneic BMT patients.[GRAPHICS]. |
