The novel combination of sirolimus and bortezomib prevents graft-versus-host disease but maintains the graft-versus-leukemia effect after allogeneic transplantation.

BACKGROUND: We have previously shown that bortezomib induces a depletion of alloreactive T cells and allows the expansion of T cells with suppressive properties. In the current study, we analyzed the potential synergistic effect of bortezomib in conjunction with sirolimus in order to reduce-graft-versus-host disease without hampering graft-versus-leukemia effect in the allogeneic transplant setting. DESIGN AND METHODS: We evaluated the effect of sirolimus, bortezomib or the combination of both in the proliferation and activation of in vitro stimulated T lymphocytes. Pathways involved in this synergy were also analyzed using Western blot assays. Finally, BALB/c mice receiving C57BL/6 allogeneic donor bone marrow with splenocytes were used to measure in vivo the effect of this novel combination on the risk of graft-versus-host disease. RESULTS: The combination of both drugs synergistically inhibited both activation and proliferation of stimulated T cells. Also, the production of Th1 cytokines (IFN γ, IL-2 and TNF) was significantly inhibited. This effect was due, at least in part, to the inhibition of Erk and Akt phosphorylation. In vivo, the combination reduced the risk of graft-versus-host disease without hampering graft-versus-leukemia effect, as shown in mice receiving graft-versus-host disease prophylaxis with sirolimus plus bortezomib being infused with tumor WEHI cells plus C57BL/6 donor BM and splenocytes. CONCLUSIONS: The current study reveals a synergistic effect of the combination sirolimus and bortezomib to prevent graft-versus-host disease while maintaining the graft-versus-leukemia effect.

Limbus damage in ocular graft-versus-host disease.

Graft-versus-host disease (GVHD) is the major cause of morbidity and mortality among patients undergoing allogeneic hematopoietic stem cell transplantation. Although animal models have been clearly established for the study of skin, liver, and gut, currently there is no equivalent experiemental model for analyzing ocular involvement, which is rather common, especially among patients diagnosed with chronic GVHD. In the current study we have developed a murine model of ocular GVHD and, for the first time, we describe the histopathologic features involving cornea and limbus, which could play a role in the physiopathology of the disease at the ocular level. Our results represent a major finding that allows us to define a model for evaluating new therapeutic strategies for treating ocular GVHD prior to their use in clinical setting.

Isolation and characterization of mesenchymal stromal cells from human degenerated nucleus pulposus: comparison with bone marrow mesenchymal stromal cells from the same subjects.

STUDY DESIGN: To identify mesenchymal stromal cells (MSC) from degenerate human nucleus pulposus (NP) and compare them with bone marrow (BM) MSC. OBJECTIVE: To test whether MSC obtained from NP and BM from the same subjects share similar biologic characteristics. SUMMARY OF BACKGROUND DATA: Recent studies have proposed biologic strategies for the treatment of intervertebral disc degeneration, including cell therapy. Bone marrow (BM) MSC could be an attractive approach to restore disc function, and there is evidence that NP may contain MSC-like cells. METHODS: Tissue samples were obtained from degenerate lumbar NP and from iliac crest of the same 16 patients with degenerative disc diseases, undergoing discectomy and fusion procedures. MSC isolated from both sources were compared regarding their expansion time, immunophenotype, differentiation ability, and molecular analysis. RESULTS: In all cases, MSC from NP were isolated and expanded. They fulfil nearly all morphological, inmunophenotypical, and differentiation criteria described by the International Society of Cell Therapy for MSC, with the exception that NP-MSC are not able to differentiate into adipocytes. Slight differences were observed with BM-MSC from the same subjects. CONCLUSION: The NP contains mesenchymal stem cells. These cells were quite similar to mesenchymal stem cells from BM, with the exception of their adipogenic differentiation ability. These findings suggest that we may treat intervertebral disc degeneration by cell therapy (MSC from BM) and by stimulating endogenous MSC from NP.

Immunomodulatory effect of 5-azacytidine (5-azaC): potential role in the transplantation setting.

Cytokine genes are targets of multiple epigenetic mechanisms in T lymphocytes. 5-azacytidine (5-azaC) is a nucleoside-based DNA methyltransferase inhibitor that induces demethylation and gene reactivation. In the current study, we analyzed the effect of 5-azaC in T-cell function and observed that 5-azaC inhibits T-cell proliferation and activation, blocking cell cycle in the G(0) to G(1) phase and decreasing the production of proinflammatory cytokines such as tumor necrosis factor-alpha and interferon-gamma. This effect was not attributable to a proapoptotic effect of the drug but to the down-regulation of genes involved in T-cell cycle progression and activation such as CCNG2, MTCP1, CD58, and ADK and up-regulation of genes that induce cell-growth arrest, such as DCUN1D2, U2AF2, GADD45B, or p53. A longer exposure to the drug leads to demethylation of FOXP3 promoter, overexpression of FOXP3, and expansion of regulatory T cells. Finally, the administration of 5-azaC after transplantation prevented the development of graft-versus-host disease, leading to a significant increase in survival in a fully mismatched bone marrow transplantation mouse model. In conclusion, the current study shows the effect of 5-azaC in T lymphocytes and illustrates its role in the allogeneic transplantation setting as an immunomodulatory drug, describing new pathways that must be explored to prevent graft-versus-host disease.