br Materials and methods br Results

Materials and methods
Results
Discussion In this study, we investigated two novel candidate therapeutics, the related CRM1-inhibiting SINEs KPT-185 and KPT-276, in treatment of NHL. The results showed that KPT-185 induced growth inhibition, cell-cycle arrest, and apoptosis in tumor faah inhibitors in vitro. And we observed similar antitumor activity of KPT-276 in an NOD/SCID mouse model of NHL formed by Jeko-1 cells. KPT-276 administered orally at a dose of 100mg/kg three times a week exhibited marked activity against established Jeko-1 tumor xenografts in vivo. Under these conditions, KPT-276 was well tolerated, with no serious resulting weight loss or side effects similar to those of chemotherapy. In general, the management of NHL is remaining a big challenge, especially for the T-cell lymphomas. Compared with the more common aggressive B-cell lymphomas, more patients with T-cell lymphomas will be refractory to initial therapy and those who achieve responses often experience with shorter progression-free survival [33]. The current therapeutic strategies for T cell lymphomas remain poorly defined, which are commonly extrapolated from treatment paradigms of B-cell lymphomas and generally have not been studied in a prospective manner. To improve the outcome of patients with T cell lymphomas, it is imperative that new therapeutic options be explored [34]. Herein, we tested the effect of KPT-185 on the growth of seven types of cell lines, including four B cell lymphomas and three T-cell lymphomas. Notably, the three T cell lines (Hut102, Jurkat and Hut78) were more sensitive than the B cell lines to the KPT-185. Therefore, our data demonstrated that KPT-185 may be a new promising agent for the treatment of T cell lymphomas. Recent studies suggested that CRM1 overexpression is associated with progression and mortality of several human cancers and that CRM1 is an oncogene [3]. In the present study, we observed reduction of CRM1 protein expression treated with KPT-185 in NHL cell lines, which exhibited a correlation with the efficacy of KPT-185 in vitro. For previous studies demonstrated that knockdown of CRM1 gene expression could result in inhibition of tumor cell growth [28], we considered that the decreased CRM1 expression accompanying treatment with KPT-185 may be a reason for this SINE\'s therapeutic efficacy. Furthermore, CRM1 is known to export primarily p53+/+ from the nucleus to the cytoplasm of cancer cells via its C-terminal NES, allowing for efficient degradation of p53 by the proteasome [9]. Also, researchers have reported that p53 was considered to be a critical mediator of CRM1 inhibitor-induced cell apoptosis [19], [25]. Interestingly, in this present study, we noticed that treatment with KPT-185 decreased the expression of mutant p53 in Mino, Jurkat, and RL cells lines but increased the expression of p53+/+ in Granta519 cells. Given the different functions of various p53 genotypes in tumor growth and survival, these results indicated that p53 may be a mediator of SINE-induced antitumor activity. Another CRM1-dependent process affected by treatment with KPT-185 was the reduction of NF-κB expression in Jeko-1, Hut102, Mino, RL, Jurkat and Hut78 cells. NF-κB plays important roles in various types of cancer cells and is an important target for anticancer therapy [35]. Therefore, downregulated expression of NF-κB may be a reasonable explanation for the antilymphoma activity of KPT-185. Notably, we made the novel observation that expression of the growth-promoting protein survivin was greatly downregulated in the cells with KPT-185 treatment. Furthermore, the enhanced nuclear staining of survivin in Jeko-1 tumor sections visualized by immunohistochemistry indicated that survivin was trapped within the nucleus by KPT-276 treatment. As an inhibitor member of the apoptosis protein family, survivin exhibits antiapoptotic characteristics. Authors have reported elevated survivin expression to confer radiation or drug resistance, whereas artificial knockdown of survivin expression increases the susceptibility of cells to apoptosis [36], [37], [38]. Furthermore, earlier studies have demonstrated that survivin is actively and rapidly exported into the cytoplasm, and it contains a canonical NES domain. Exclusion of survivin from the nucleus is mediated by the classic nuclear export mechanism involving the interaction between the transport receptor CRM1 and Ran-guanosine triphosphate [12], [39], [40]. Recent reports also demonstrated that preferentially cytoplasmic survivin is “cytoprotective” in tumor cells, whereas nuclear survivin may be indicative of “impaired survivin function.” [13], [14], [41] Therefore, patients with cancer having predominantly nuclear survivin in tumor cells may have reduced tumor-protective survivin activity, which may ultimately translate into a favorable prognosis. Therefore, targeting the survivin/CRM1 interface may be a powerful approach to NHL therapy. Our present results indicated an important antitumor mechanism of SINEs based on expression and cellular localization of survivin.