PMC:3342329 / 19094-26766 JSON TXT

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{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/3342329","sourcedb":"PMC","sourceid":"3342329","source_url":"http://www.ncbi.nlm.nih.gov/pmc/3342329","text":"Discussion\nIn the present study, we found that tumor weight and volume were much smaller in the CCR5−/− mice compared to the CCR5+/+ mice inoculated with B16 melanoma cells (Figure 1A and 1B). The tumor growth inhibition in the CCR5−/− mice was associated with the inhibition of constitutively activated NF-κB and elevation of IL-1Ra in the tumor tissue. This anti-tumor activity was also associated with decreased expression of NF-κB target anti-apoptotic, cell proliferative, and tumor promoting genes (Bcl-2 and C-IAP1) but with increased expression of their target apoptotic genes; cleaved caspase-3 and 9, and Bax (Figure 3A-D). In the tumor and the spleen, the number of cytotoxic CD8+ T cells, CD57+ natural killer cells, and the levels of IL-1Ra were increased (Figure 4 and 5). These findings suggest that melanoma growth was inhibited in the CCR5−/− mice, and that the elevation of IL-1Ra, accompanied with decreased NF-κB, is significant in the inhibition of melanoma growth in CCR5−/− mice.\nNF-κB regulates the expression of over 200 genes that control the immune system, cancer cell growth and inflammation [39]. Because of its abilities to induce the expression of a large array of inflammatory mediators and its roles as core transcription factors in diverse immune responses, NF-κB has been recognized as a major factor responsible for cytokine-associated cancer development or anti-tumor immunity. In the melanoma tumor tissues of the CCR5−/− mice, the expression of NF-κB target cell death genes (Bax, caspase-3 and 9) and the DNA binding activity of NF-κB, were significantly inhibited, but the expression of cell death inhibitory NF-κB target genes, such as Bcl-2 and cIAP were enhanced. In addition, CCR5−/− prevented the phosphorylation of IκB, accompanied with the inhibition of the p50 and p65 translocation into the nucleus (Figure 2A-C). Many tumors, including melanoma, have increased levels of NF-κB [40], which is likely acting as a survival factor for melanoma growth. Thus, the inhibitions of NF-κB activity and the expression of target genes are critical in the inhibition of tumor growth in CCR5−/− mice. Although the mechanism is not clear as to how CCR5−/− downregulates NF-κB, it is noteworthy that NF-κB is activated or inactivated by many cytokines.\nNumerous studies showed that cytokine affect tumor growth by regulating NF-κB. Inflammatory cytokines, including IL-1, IL-6 and IL-8, activate NF-κB pathways in tumor cells [41]. The interleukin 10 (IL-10) is well known as an anti-tumor factor as well as an anti-inflammatory factor [42], [43]. IL-10 inhibits constitutively activated NF-κB both in vitro [44] and in vivo [45]. Inhibition of NF-κB by the IL-10 contributes to the anti-tumor effects through the activation of caspase-3 and apoptosis in Colon26-bearing mice [43]. Similarly, several studies have demonstrated the ability of IL-1Ra to abrogate the proinflammatory effects of IL-1 [46]. It has been reported that IL-1 induces the activation of NF-κB in several cancers, and increases cell cycle progression, and the suppression of apoptotic cell death leading to tumor promotion. The results of various studies over the past 10 years indicate that the major function of IL-1Ra is to regulate the pleiotropic effects of IL-1 by competitively blocking its functions [47]. The ability of IL-1 to induce its own antagonist is of interest in understanding the regulation and dysregulation of IL-1 signaling in normal and diseased tissue. Up-regulation of the receptor antagonist represents a mechanism for turning off IL-1 signaling and creating a temporary state of refractoriness. The elevated expression of IL-1Ra, without any change of IL-1 in tumors, suggests that the IL-1-signaling system may be dysregulated by IL-1Ra in tumors. IL-1Ra has been shown to inhibit tumor progression in prostate, breast, and skin cancer [14], [15], [16]. Intracellular IL-1Ra (icIL-1Ra) inhibits IL-6 and IL-8 production in Caco-2 cells by blocking NF-κB pathways [37]. We found that the levels of IL-1Ra were significantly elevated in the tumor tissues of CCR5−/− mice inoculated with B16 melanoma cells. Taken together, these data indicate IL-1Ra could be critically involved in the tumor growth inhibition of CCR5−/− mice through the inactivation of NF-κB in the present study.\nCCR5 increases tumor growth in a local model and inhibits the efficacy of a dendrite cell vaccine in CCR5+/+ mice compared with CCR5−/− mice [10]. Cheng J and Sung RS examined the effect of CCR5 on the immune response to adenovirus vectors and graft function in an islet transplant model. They found that CCR5 absence does not prevent the local immune response to adenovirus transduction [48]. Additionally, Tan MC et al. suggested that disruption of CCR5/CCL5 signaling, either by reducing CCL5 production by tumor cells or by systemic administration of a CCR5 inhibitor (TAK-779), reduced Treg (regulatory T cells) migration to tumors so that tumors are smaller in control mice [9]. These data support the important premise that inhibition of tumor growth in CCR5−/− mice could be related with the inhibition of immune escape. Similar to this finding, we found that the CCR5+/+ mice injected with melanoma cells apparently have a lower number of cytotoxic T cells (CD8+positive T Lymphocytes cells) and natural killer cells (CD57 positive NK cells), as well as dendrite cells, in spleen tissues compared to those without melanoma cells. But the lymphocytes of the CCR5−/− mice did not decrease when injected with melanoma cells. These differences of the immune response between the CCR5+/+ mice and the CCR5−/− mice, involving tumor immune tolerance, may be connected with the inhibition of cancer progression. Taken together, in the cytokine array experiment, the level of MIP-1α (which has a potent activity of T cell and B cell migration) was increased in the CCR5−/− mice compared with those in the CCR5+/+ mice. Thus, relatively increased tumor associated cytotoxic lymphocytes could play an important role in inhibiting the tumor growth in CCR5−/− mice.\nActivation of NF-κB could modulate subcellular localization of NK cells and pathogenesis [49]. Inhibition of NF-κB activity prevented NK cell depletion, and thus increased anti-tumor activity by decreasing IL-6 production [50]. Thus, the decreased NF-κB and elevated IL-1Ra could also be associated in the infiltration of cytotoxic lymphocytes into tumor, resulting in tumor growth inhibition. In contrast, IL-23, a tumor growth promoting cytokine, increased NF-κB and immune cell infiltration in oral tumor [51]. These data indicate that NF-κB could be involved in the cytokine mediated anti-tumor activities of immune cells. It is also noteworthy that IL-1Ra inhibited melanoma tumor growth by increasing the number of myeloid suppressor cells in tumor [25]. The 3-methylcholatrene-induced tumor incidence was reduced in IL-1α knockout mice, but increased in IL-1Ra mice with concomitant maturation of NK cells and anti-tumor immunity [52]. These data indicate that the decrease of NF-κB, and thus increase of IL-1Ra could be significant in tumor growth inhibition of CCR5−/− mice.\nOur data thus show significant clinical implications of CCR5, because CCR5 is a G protein–coupled receptor and it is amenable to use as a small molecule for the inhibition of CCR5. Several such compounds have already been shown to be safe in clinical trials for use in certain diseases such as HIV [53]. Thus, we believe CCR5 is a cancer target that warrants continued investigation. In summary, this study showed that melanoma cells growth was inhibited via modulation of NF-κB/IL-1Ra pathways in CCR5−/− 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