Histone Deacetylase Inhibitor SAHA Reduces Cytokine Storm and Facilitates Induction of Chimerism When Combined with Anti-CD3 mAb in Conditioning of Recipients.

Donor T cell-mediated graft versus leukemia (GVL) and graft versus autoimmunity (GVA) plays a critical role in the treatment of hematological malignancies and refractory autoimmune diseases using allogeneic hematopoietic cell transplantation (HCT). However, graft versus host disease (GVHD) remains a major obstacle in classical HCT, where recipients are usually conditioned with total body irradiation or high dose chemotherapy. We have recently reported a radiation-free and GVHD preventive anti-CD3-conditioning regimen, which allows donor CD8⁺ T cells to facilitate engraftment and mediate GVL without causing GVHD (PNAS, 2007 and J. I. 2007). In order to promote the clinical application of the anti-CD3-conditioning regimen, we need to overcome the cytokine storm that causes flu-like syndrome in patients after anti-CD3 mAb injection and reduce the required donor bone marrow cell dose. Suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, was reported by others to induce apoptosis of human T cell lymphoma cell lines and to reduce serum levels of pro-inflammatory cytokines in HCT recipients. Therefore, we tested whether conditioning with a combination of anti-CD3 and SAHA can reduce the cytokine storm caused by anti-CD3 and reduce the resistance of engraftment mediated by residual host T cells. Accordingly, titrated dose of SAHA (0.125–8 μM) were added to cultures of BALB/c spleen cells stimulated with plate-bound anti-CD3. We found that SAHA augmented apoptosis of anti-CD3 activated T cells in a dose-dependent manner. Although low-dose SAHA (0.125–0.25 μM) did not augment apoptosis, it rendered the residual live T cells partially unresponsive. At the same time, SAHA significantly reduced the pro-inflammatory cytokines (IL-2, IFN-γ, TNF-α, and IL-6) in the culture supernatants. Similarly, in vivo treatment with anti-CD3 and low-dose SAHA (40 μg/g) led to significant reduction of serum levels of those cytokines and partial unresponsiveness of the residual host T cells. Finally, conditioning with combined anti-CD3 and SAHA (40 μg/g) induced complete chimerism without GVHD in 12/12 of old BALB/c as well as in 8/8 of old autoimmune lupus NZB/NZW F1 recipients after two injections of BM cells (2 ×10⁶/g) and CD4⁺ T-depleted spleen cells (4 ×10⁶/g), although conditioning with anti-CD3 alone did not induce any chimerism (0/8). The chimeric NZB/NZW F1 recipients showed complete reversal of autoimmune glomerulonephritis and proteinuria. These results indicate that SAHA can not only reduce cytokine storm but also facilitate engraftment when combined with anti-CD3 for conditioning of recipients. This radiation free and GVHD preventive conditioning regimen may provide a novel approach for clinical HCT.