Suberoylanilide Hydroxamic Acid Treatment Research Articles

Restoration of Transforming Growth Factor-β Signaling through Receptor RI Induction by Histone Deacetylase Activity Inhibition in Breast Cancer Cells

The loss of transforming growth factor-beta (TGF-beta) response due to the dysregulation of TGF-beta receptors type I (RI) and type II (RII) is well known for its contribution to oncogenesis. Estrogen receptor-expressing breast cancer cells are refractory to TGF-beta-mediated growth control because of the reduced expression of TGF-beta receptors. Although RII is required for the binding of TGF-beta to RI, RI is responsible for directly transducing TGF-beta signals through the Smad protein family. Treatment of estrogen receptor-expressing MCF-7L and ZR75 breast cancer cells with the histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA) led to a dramatic induction of RI. Accumulation of acetylated histones H3 and H4 was observed in the SAHA-treated cells. Chromatin immunoprecipitation analysis followed by PCR with RI promoter-specific primers indicated an accumulation of acetylated histones in chromatin associated with the RI gene, suggesting that histone deacetylation was involved in the transcriptional inactivation of RI. SAHA treatment stimulated RI promoter activity through the inhibition of Sp1/Sp3-associated HDAC activity. Histone acetyltransferase p300 stimulated RI promoter activity, thus further confirming the involvement of HDAC activity in the transcriptional repression of RI. Significantly, SAHA-mediated RI regeneration restored the TGF-beta response in breast cancer cells.

Suberoylanilide hydroxamic acid reduces acute graft-versus-host disease and preserves graft-versus-leukemia effect by inhibiting histone deacetylation

Pro-inflammatory cytokines and the loss of gastrointestinal tract integrity contribute to acute graft-versus-host disease (GVHD) whereas the donor cytotoxic responses are critical for graft-versus-leukemia (GVL) preservation. Suberoylanilide hydroxamic acid (SAHA) is an anti-tumor agent that inhibits the activity of histone deacetylases (HDAC) and reduces the production of proinflammatory cytokines. Using a well characterized allogeneic murine BMT model B6 (H-2b) → B6D2F1 (H-2b/d) we studied the effects of HDAC inhibition by SAHA on acute GVHD. Recipients were transplanted with 2x10 6 donor T and 5x106 BM cells after13 GY TBI. Intra-peritoneal injections of 35 mg/kg/day of SAHA from days +3 to day +7 increased histone H3 acetylation in splenocytes harvested 7 days after BMT, confirming the inhibition of HDAC enzymes. SAHA treatment significantly reduced the serum levels of pro-inflammatory cytokines such as TNF-a, IL-1β and IFN-γ (P < 0.04) in the allogeneic recipients on day +7 after BMT. Intracytoplasmic staining by flow cytometry and RPA analysis of the host splenocytes on day +7 confirmed the decrease in the cytokine protein and mRNA. SAHA significantly improved the survival (P < 0.002) and reduced intestinal damage from GVHD of the allogeneic recipients . However SAHA did not suppress the donor T cell expansion in vivo and the proliferative and cytotoxic responses to host antigens in vitro measured 7 and 14 days after BMT. To test the effect of SAHA on GVL effects, recipients were injected with lethal doses of P815 (H-2d) tumor cells at the time of BMT. SAHA treatment resulted in significantly improved leukemia-free survival after allogeneic BMT (P < 0.05) whereas all the syngeneic BMT recipients of SAHA died of tumor ruling out direct anti-tumor effects of SAHA. Furthermore SAHA increased H3 acetylation in the splenocytes from both the syngeneic and allogeneic leukemic recipients on day +7, confirming that inhibition of HDAC enzymes alone is not sufficient for leukemia free survival in this system. We also tested the effect of SAHA in a second allogeneic BMT model (BALB/c → B6), where it also significantly improved survival (P < 0.001) and preserved GVL effects when recipient mice were injected with lethal doses of EL-4 (H-2b) tumor (P < 0.04). We conclude that HDAC inhibition regulates acute GVHD in these models and suggest that this class of pharmacologic agents may provide a novel strategy to reduce GVHD while maintaining the beneficial GVL effects.

Targeting aberrant transcriptional repression in leukemia: a therapeutic reality?

Targeting aberrant transcriptional repression in leukemia: a therapeutic reality?

Activation of the p21WAF1/CIP1 promoter independent of p53 by the histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) through the Sp1 sites.

Suberoylanilide hydroxamic acid (SAHA) is a novel histone deacetylase inhibitor with high potency in inducing differentiation of cultured murine erythroleukemia cells. We have recently demonstrated that SAHA induces cell cycle arrest and apoptosis in human breast cancer cells, accompanied by up-regulation of the cyclin-dependent kinase inhibitor, p21WAF1/CIP1, via a p53-independent mechanism. In this study, we used p21 gene expression as a model system to elucidate the molecular mechanism(s) underlying SAHA-mediated gene activation. Treatment of human breast cancer cell line MCF7 cells with SAHA induced p21 mRNA as a consequence of an immediate-early gene activation. Moreover, SAHA activated the p21 promoter primarily through two Spl sites located at -82 and -69 relative to the transcription start site. Furthermore, Sp1 and Sp3 proteins were the major factors binding to the Spl site of the p21 promoter. However, SAHA did not alter their DNA binding activities, suggesting that SAHA mediates p21 promoter activity by a mechanism other than altering the DNA binding activities of Sp1 and Sp3. Further studies using the GAL4 luciferase assay system demonstrated that both GAL4-Sp1 and GAL4-Sp3 fusion proteins supported SAHA-mediated gene activation from a promoter driven by five GAL4 DNA binding sites, and that GAL4-Sp3 fusion protein was suppressive in the absence of SAHA treatment. Collectively, our results suggest that SAHA activates the p21 promoter through the Spl sites, and that both Spl and Sp3 proteins can mediate SAHA-induced gene activation.