Although recent studies have shown that adenosine-to-inosine (A-to-I) RNA editing occurs in microRNAs, its effects on tumor growth and metastasis are not well understood. We present evidence of CREB-mediated low expression of ADAR1 in metastatic melanoma cell lines and tumor specimens. Re-expression of ADAR1 resulted in the suppression of melanoma growth and metastasis in vivo. Consequently, we identified 3 miRs undergoing A-to-I editing in the low-metastatic melanoma but not in highly metastatic cell lines. One of these miRs, miR-455-5p has two A-to-I RNA editing sites. The biological function of edited miR-455-5p is different from the unedited form as it recognizes different set of genes. Indeed, w.t. miR-455-5p promotes melanoma metastasis via inhibition of the tumor suppressor gene CPEB1. Moreover, w.t. miR-455 enhances melanoma growth and metastasis in vivo while the edited form inhibits these features. These results demonstrate a previously unrecognized role of RNA editing in melanoma progression.

Summary Malignant melanoma is one of the most aggressive cancers and can disseminate from a relatively small primary tumor and metastasize to multiple sites, including the lung, liver, brain, bone, and lymph nodes. Elucidating the molecular and genetic changes that take place during the metastatic process has led to a better understanding of why melanoma is so metastatic. Herein, we describe the unique features that distinguish melanoma from other solid tumors and contribute to the malignant phenotype of melanoma cells. For example, although melanoma cells are highly antigenic, they are extremely efficient at evading host immune response. Melanoma cells share numerous cell surface molecules with vascular cells, are highly angiogenic, are mesenchymal in nature, and possess a higher degree of ‘stemness’ than do other solid tumors. Finally, analysis of melanoma mutations has revealed that the gene expression profile of malignant melanoma is different from that of other cancers. Elucidating these molecular and genetic processes in highly metastatic melanoma can lead to the development of improved treatment and individualized therapy options.

BackgroundThe pharmaceutical and biotechnology industries depend on findings from academic investigators prior to initiating programs to develop new diagnostic and therapeutic agents to benefit cancer patients. The success of these programs depends on the validity of published findings. This validity, represented by the reproducibility of published findings, has come into question recently as investigators from companies have raised the issue of poor reproducibility of published results from academic laboratories. Furthermore, retraction rates in high impact journals are climbing.Methods and FindingsTo examine a microcosm of the academic experience with data reproducibility, we surveyed the faculty and trainees at MD Anderson Cancer Center using an anonymous computerized questionnaire; we sought to ascertain the frequency and potential causes of non-reproducible data. We found that ∼50% of respondents had experienced at least one episode of the inability to reproduce published data; many who pursued this issue with the original authors were never able to identify the reason for the lack of reproducibility; some were even met with a less than “collegial” interaction.ConclusionsThese results suggest that the problem of data reproducibility is real. Biomedical science needs to establish processes to decrease the problem and adjudicate discrepancies in findings when they are discovered.

Melanoma is the deadliest form of skin cancer in which patients with metastatic disease have a five year survival-rate of less than 10%. Recently the over expression of a beta galactoside binding protein, galectin-3 (LGALS3), has been correlated with metastatic melanoma in patients. We have previously shown that silencing galectin-3 in metastatic melanoma cells reduces tumor growth and metastasis. Gene expression profiling identified the pro-tumorigenic gene autotaxin (ENPP2) to be down regulated after silencing galectin-3. Here we report that galectin-3 regulates autotaxin expression at the transcriptional level by modulating the expression of the transcription factor NFAT1 (NFATC2). Silencing galectin-3 reduced NFAT1 protein expression which resulted in decreased autotaxin expression and activity. Reexpression of autotaxin in galectin-3 silenced melanoma cells rescues angiogenesis, tumor growth and metastasis in vivo. Silencing NFAT1 expression in metastatic melanoma cells inhibited tumor growth and metastatic capabilities in vivo. Our data elucidate a previously unidentified mechanism by which galectin-3 regulates autotaxin, and assign a novel role for NFAT1 during melanoma progression.

There were errors published in J. Cell Sci. 122, 1842-1851.In Fig. 3, panels A and C (rather than A and B) were mistakenly labeled as wild type and panels B and D (rather than C and D) were mistakenly labeled as β8 -/-. The correct figure is shown below:In Fig. 5, the units on the y axis for panel C should be in micrometers (μm), not millimeters (mm). The correct figure is shown below:The authors apologise for these mistakes.

Nuclear factor of activated T cell (NFAT1, NFATC2) is a transcription factor that binds and positively regulates interleukin-2 expression during T cell activation. NFAT1 has important roles in both innate and adaptive immune responses, but its involvement in cancer is not completely understood. We previously demonstrated that NFAT1 contributes to melanoma growth and metastasis by regulating the autotaxin gene (Enpp2). Here, we report a strong correlation between NFAT1 expression and metastatic potential in melanoma cell lines and tumor specimens. To elucidate the mechanisms underlying NFAT1 overexpression during melanoma progression, we conducted a microarray on a highly metastatic melanoma cell line in which NFAT1 expression was stably silenced. We identified and validated two downstream targets of NFAT1, IL-8 and MMP-3. Accordingly, NFAT1 depletion in metastatic melanoma cell lines was associated with reduced IL-8 and MMP-3 expression, whereas NFAT1 overexpression in a weakly metastatic cell line induced expression of these targets. Restoration of NFAT1 expression recovered IL-8 and MMP-3 expression levels back to baseline, indicating that both are direct targets of NFAT1. Moreover, in vivo studies demonstrated that NFAT1 and MMP-3 promoted melanoma tumor growth and lung metastasis. Collectively, our findings assign a new role for NFAT1 in melanoma progression, underscoring the multifaceted functions that immunomodulatory factors may acquire in an unpredictable tumor microenvironment.

The effects of AURKA overexpression associated with poor clinical outcomes have been attributed to increased cell cycle progression and the development of genomic instability with aneuploidy. We used RNA interference to examine the effects of AURKA overexpression in human bladder cancer cells. Knockdown had minimal effects on cell proliferation but blocked tumor cell invasion. Whole genome mRNA expression profiling identified nicotinamide N-methyltransferase (NNMT) as a downstream target that was repressed by AURKA. Chromatin immunoprecipitation and NNMT promoter luciferase assays revealed that AURKA’s effects on NNMT were caused by PAX3-mediated transcriptional repression and overexpression of NNMT blocked tumor cell invasion in vitro. Overexpression of AURKA and activation of its downstream pathway was enriched in the basal subtype in primary human tumors and was associated with poor clinical outcomes. We also show that the FISH test for the AURKA gene copy number in urine yielded a specificity of 79.7% (95% confidence interval [CI] = 74.2% to 84.1%), and a sensitivity of 79.6% (95% CI = 74.2% to 84.1%) with an AUC of 0.901 (95% CI = 0.872 to 0.928; P < 0.001). These results implicate AURKA as an effective biomarker for bladder cancer detection as well as therapeutic target especially for its basal type.

The progression of melanoma toward the metastatic phenotype occurs in a defined stepwise manner. While many molecular changes take place early in melanoma development, progression toward the malignant phenotype, most notably during the transition from the radial growth phase (RGP) to the vertical growth phase (VGP) involves deregulated expression of several transcription factors. For example, the switch from RGP to VGP is associated with the loss of the transcription factor AP2α and gain of transcriptional activity of cAMP-responsive element binding protein. Together with the upregulation of microphthalmia-associated transcription factor, activating transcription factor 2, nuclear factor kappa B, and other transcription factors, these changes lead to dysregulated expression or function of important cellular adhesion molecules, matrix degrading enzymes, survival factors, as well as other factors leading to metastatic melanoma. Additionally, recent evidence suggests that microRNAs and RNA editing machinery influence the expression of transcription factors or are regulated themselves by transcription factors. Many of the downstream signaling molecules regulated by transcription factors, such as protease activated receptor-1, interleukin-8, and MCAM/MUC18 represent new treatment prospects.