Through these our recent reports, we confirmed that TNIK can be a potential target for inducing apoptosis activity of KY-05009 and dovitinib in cancer cells. In the present study, we investigated the Prochloraz manganese level of TNIK expression in human MM cells from patients and the apoptosis-inducing effect of KY-05009 and dovitinib in the IL-6-dependent MM RPMI8226 cell line. and induces apoptosis [7, 11]. A few reports have exhibited the expression of TNIK and cancer cell proliferation in several types of cancer, but the relevance of TNIK Prochloraz manganese to hematological malignancies, especially MM, has not been sufficiently described [6C11]. In our previous studies, Rabbit Polyclonal to GRP94 we investigated the apoptosis-inducing effect of tyrosine kinase inhibitor dovitinib and its inhibition of TNIK kinase activity and endogenous Wnt signaling in human MM cells . TNIK is usually highly expressed in MM cells compared to normal peripheral blood mononuclear cells (PBMCs), and inhibition of TNIK expression by siRNA induces cell death. KY-05009 and dovitinib have a high affinity for the ATP binding site in TNIK and inhibit the protein expression of TNIK and transcriptional activity of Wnt target genes [11, 12]. Through these our recent reports, we confirmed that TNIK can be a potential target for inducing apoptosis activity of KY-05009 and dovitinib in cancer cells. In the present study, we investigated the level of TNIK expression in human MM cells from patients and the apoptosis-inducing effect of KY-05009 and dovitinib in the IL-6-dependent MM RPMI8226 cell line. IL-6 enhanced cell proliferation, mRNA Prochloraz manganese and protein expression, and the transcriptional activity of Wnt target genes. KY-05009 exerted synergistic anti-proliferative effects with dovitinib and brought on caspase-dependent apoptosis in RPMI8226 cells. We hypothesize that a possible mode of action of KY-05009 and dovitinib is usually a high affinity for TNIK and subsequent inhibition of kinase activity [11, 12]. This inhibitory effect against TNIK may suppress the proliferation of RPMI8226 cells. Thus, our results suggest that TNIK could be an anti-cancer target for the investigation of treating MM by inhibiting Wnt signaling-mediated MM cell proliferation. RESULTS IL-6 stimulates the proliferation of RPMI8226 cells IL-6 has been identified as a major growth factor for myeloma cell proliferation and [13C16]. In particular, paracrine regulation of IL-6 stimulates myeloma cell proliferation in patients . To confirm the effect of IL-6 around the production of cytokines in MM cells, we analyzed the expression of cytokines and whether IL-6 treatment induces paracrine effects of other cytokines, such as IL-1, IL-2, IL-4, IL-5, and tumor necrosis factor (TNF)-, on cultured supernatant or protein expression in cell lysates (Physique ?(Figure1A).1A). Serum-starved RPMI8226 cells were treated with recombinant human IL-6 in serum-free medium for 72 h and the culture supernatants and cell lysates isolated to analyze secreted factors and their influence on protein expression in MM cells. A human cytokine array showed that migration inhibitor factor (MIF), an inflammatory mediator, was constitutively produced regardless of IL-6 treatment. IL-6 was only expressed in the supernatant in response to IL-6 treatment (Physique ?(Physique1A,1A, left). We also observed that IL-8, an activator of osteoclast differentiation and bone resorption in MM, was expressed in both untreated controls and the IL-6-treated group, but IL-6 only increased in the IL-6-treated lysate group (Physique ?(Physique1A,1A, right). Next, we assessed the Prochloraz manganese stimulatory effect of IL-6 around the proliferation of MM cells. RPMI8226 cells were incubated with recombinant human IL-6 for 24 to 72 h. As shown in Figure ?Physique1B,1B, IL-6 stimulated the proliferation of MM cells in a dose- and time-dependent manner. These results support an increased level of IL-6 in cultured supernatants and cell lysates correlating with MM cell proliferation. Open in a separate window Physique 1 IL-6 activates MM cell proliferation(A) RPMI8226 cells were treated with recombinant human IL-6 for 72 h. After incubation, cytokine expression in cell supernatants and lysates was analyzed by human cytokine array. The expression of IL-6 was normalized by the density of control spots. (B) Cell viability of RPMI8226 cells Prochloraz manganese after treatment with IL-6 (0-100 ng/mL) in serum-free medium for 24-72 h. Data are presented as meanSD. The experiments were performed in triplicate. *< 0.01, *< 0.001 versus control. IL-6 activates TNIK expression and the transcriptional activity of Wnt signaling Our previous studies exhibited the association between canonical Wnt signaling and the survival of MM cells [17, 18]. In addition, targeting of the constitutively active -catenin/TCF4 transcriptional complex has been identified as a potent therapeutic strategy.
Through these our recent reports, we confirmed that TNIK can be a potential target for inducing apoptosis activity of KY-05009 and dovitinib in cancer cells
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