Constitutive activation of signal transducers and activators of transcription predicts vorinostat resistance in cutaneous T-cell lymphoma.

Vorinostat is a histone deacetylase inhibitor that induces differentiation, growth arrest, and/or apoptosis of malignant cells both in vitro and in vivo and has shown clinical responses in approximately 30% of patients with advanced mycosis fungoides and Sézary syndrome cutaneous T-cell lymphoma (CTCL). The purpose of this study was to identify biomarkers predictive of vorinostat response in CTCL using preclinical model systems and to assess these biomarkers in clinical samples. The signal transducer and activator of transcription (STAT) signaling pathway was evaluated. The data indicate that persistent activation of STAT1, STAT3, and STAT5 correlate with resistance to vorinostat in lymphoma cell lines. Simultaneous treatment with a pan-Janus-activated kinase inhibitor resulted in synergistic antiproliferative effect and down-regulation of the expression of several antiapoptotic genes. Immunohistochemical analysis of STAT1 and phosphorylated tyrosine STAT3 (pSTAT3) in skin biopsies obtained from CTCL patients enrolled in the vorinostat phase IIb trial showed that nuclear accumulation of STAT1 and high levels of nuclear pSTAT3 in malignant T cells correlate with a lack of clinical response. These results suggest that deregulation of STAT activity plays a role in vorinostat resistance in CTCL, and strategies that block this pathway may improve vorinostat response. Furthermore, these findings may be of prognostic value in predicting the response of CTCL patients to vorinostat.

Regulation of c-Jun N-terminal kinase and p38 kinase pathways in endothelial cells.

The rapid and transient induction of E-selectin gene expression by inflammatory tumor necrosis factor (TNF)-alpha in endothelial cells is mediated by signaling pathways which involve c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) kinase pathways. To explore this regulation, we first observed that in the continuous presence of cytokine TNF, activation of JNK-1 in both nuclear and cytoplasmic compartments peaked at 15-30 min, with activity returning to uninduced levels by 60 min. Phosphorylation of both the p38 kinase and its molecular target, the nuclear transcription factor, activating transcription factor-2, were transient after TNF-alpha or interleukin (IL)-1beta induction. However, cycloheximide treatment prolonged the TNF-alpha-induced JNK-1 kinase activity beyond 60 min, suggesting that protein synthesis is required to limit this signaling cascade. We investigated the possible role of the dual-specificity phosphatases MAPK phosphatase (MKP)-1 and MKP-2 in limiting cytokine-induced MAPK signaling. Maximum induction of MKP-1 mRNA and nuclear protein levels by TNF-alpha or IL-1beta were noted at 60 min and their expression correlated with the termination of JNK kinase activity, whereas nuclear levels of MKP-2 were not significantly affected by treatment with TNF-alpha or IL-1beta. Transient overexpression of MKP-1 demonstrated significant specific inhibition of E-selectin promoter activity consistent with a regulatory role for dual-specificity phosphatases. Inhibition of MKP-1 expression through the use of small interfering RNAs prolonged the cytokine-induced p38 and JNK kinase phosphorylation. Our results suggest that endogenous inhibitors of the MAPK cascade, such as the dual-specificity phosphatases like MKP-1 may be important for the postinduction repression of MAPK activity and E-selectin transcription in endothelial cells. Thus, these inhibitors may play an important role in limiting the inflammatory effects of TNF-alpha and IL-1beta.