The genomes of most, if not all, flowering plants have undergone whole genome duplication events during their evolution. The impact of such polyploidy events is poorly understood, as is the fate of most duplicated genes. We sequenced an approximately 1 million-bp region in soybean (Glycine max) centered on the Rpg1-b disease resistance gene and compared this region with a region duplicated 10 to 14 million years ago. These two regions were also compared with homologous regions in several related legume species (a second soybean genotype, Glycine tomentella, Phaseolus vulgaris, and Medicago truncatula), which enabled us to determine how each of the duplicated regions (homoeologues) in soybean has changed following polyploidy. The biggest change was in retroelement content, with homoeologue 2 having expanded to 3-fold the size of homoeologue 1. Despite this accumulation of retroelements, over 77% of the duplicated low-copy genes have been retained in the same order and appear to be functional. This finding contrasts with recent analyses of the maize (Zea mays) genome, in which only about one-third of duplicated genes appear to have been retained over a similar time period. Fluorescent in situ hybridization revealed that the homoeologue 2 region is located very near a centromere. Thus, pericentromeric localization, per se, does not result in a high rate of gene inactivation, despite greatly accelerated retrotransposon accumulation. In contrast to low-copy genes, nucleotide-binding-leucine-rich repeat disease resistance gene clusters have undergone dramatic species/homoeologuespecific duplications and losses, with some evidence for partitioning of subfamilies between homoeologues.The comparative approach to studying genes and genomes is a powerful method for addressing both

Metastatic melanoma is the most aggressive form of this cancer. It is important to understand factors that increase or decrease metastatic activity in order to more effectively research and implement treatments for melanoma. Increased cell invasion through the extracellular matrix is required for metastasis and is enhanced by matrix metalloproteinases (MMPs). Tissue inhibitor of metalloproteinases 3 (TIMP3) inhibits MMP activity. It was previously shown by our group that miR-21, a potential regulator of TIMP3, is over-expressed in cutaneous melanoma. It was therefore hypothesized that increased levels of miR-21 expression would lead to decreased expression of TIMP3 and thereby enhance the invasiveness of melanoma cells. miR-21 over-expression in the melanoma cell lines WM1552c, WM793b, A375 and MEL 39 was accomplished via transfection with pre-miR-21. Immunoblot analysis of miR-21-overexpressing cell lines revealed reduced expression of TIMP3 as compared to controls. This in turn led to a significant increase in the invasiveness of the radial growth phase cell line WM1552c and the vertical growth phase cell line WM793b (p < 0.05), but not in the metastatic cell lines A375 or MEL 39. The proliferation and migration of miR-21 over-expressing cell lines was not affected. Reduced expression of TIMP3 was achieved by siRNA knockdown and significantly enhanced invasion of melanoma cell lines, mimicking the effects of miR-21 over-expression. Treatment of tumor cells with a linked nucleic acid antagomir to miR-21 inhibited tumor growth and increased tumor expression of TIMP3 in vivo in 01B74 Athymic NCr-nu/nu mice. Intra-tumoral injections of anti-miR-21 produced similar effects. This data shows that increased expression of miR-21 enhanced the invasive potential of melanoma cell lines through TIMP3 inhibition. Therefore, inhibition of miR-21 in melanoma may reduce melanoma invasiveness.

Retrotransposons and their remnants often constitute more than 50% of higher plant genomes. Although extensively studied in monocot crops such as maize (Zea mays) and rice (Oryza sativa), the impact of retrotransposons on dicot crop genomes is not well documented. Here, we present an analysis of retrotransposons in soybean (Glycine max). Analysis of approximately 3.7 megabases (Mb) of genomic sequence, including 0.87 Mb of pericentromeric sequence, uncovered 45 intact long terminal repeat (LTR)-retrotransposons. The ratio of intact elements to solo LTRs was 8:1, one of the highest reported to date in plants, suggesting that removal of retrotransposons by homologous recombination between LTRs is occurring more slowly in soybean than in previously characterized plant species. Analysis of paired LTR sequences uncovered a low frequency of deletions relative to base substitutions, indicating that removal of retrotransposon sequences by illegitimate recombination is also operating more slowly. Significantly, we identified three subfamilies of nonautonomous elements that have replicated in the recent past, suggesting that retrotransposition can be catalyzed in trans by autonomous elements elsewhere in the genome. Analysis of 1.6 Mb of sequence from Glycine tomentella, a wild perennial relative of soybean, uncovered 23 intact retroelements, two of which had accumulated no mutations in their LTRs, indicating very recent insertion. A similar pattern was found in 0.94 Mb of sequence from Phaseolus vulgaris (common bean). Thus, autonomous and nonautonomous retrotransposons appear to be both abundant and active in Glycine and Phaseolus. The impact of nonautonomous retrotransposon replication on genome size appears to be much greater than previously appreciated.Transposable elements are abundant components of plant genomes. They are typically divided into two groups based on their mechanism of transposition. Class I transposons transpose via an RNA intermediate and must therefore use reverse transcriptase (RT) during the replication process. Class II transposons do not have an RNA intermediate and usually use a cut-andpaste mechanism for transposition . Elements of both classes have had major impacts on genome structure, apparently not only promoting mutations of genes and affecting gene regulatory sequences but also playing a substantial role in the creation of new genes by "exon-shuffling" and retrotransposition (Jin and

Increased VFA/SFA is associated with decreased PFS and OS in patients with metastatic melanoma treated with AA therapy, indicating body fat distribution is an important prognostic factor.

In patients with metastatic melanoma and RECIST SD on initial post-therapy CT images, a model incorporating CT texture analysis of target lesions, tumor size changes, and baseline LDH levels was highly accurate in predicting OS.

Sorafenib is an oral multi-kinase inhibitor that was originally developed as a Raf kinase inhibitor. We hypothesized that sorafenib would also have inhibitory effects upon cytokine signaling pathways in immune cells. Peripheral blood mononuclear cells (PBMCs) from normal donors were treated with varying concentrations of sorafenib, and stimulated with IFN-α or IL-2. Phosphorylation of STAT1 and STAT5 was measured by flow cytometry and confirmed by immunoblot analysis. Changes in IFN-α- and IL-2 stimulated gene expression were measured by qPCR, and changes in cytokine production were evaluated by ELISA. Cryopreserved PBMCs were obtained from cancer patients before and after receiving 400 mg of sorafenib twice daily. Patient PBMCs were thawed, stimulated with IL-2 or IFN-α, and evaluated for phosphorylation of STAT1 and STAT5. Pre-treatment of PBMCs with 10 μM sorafenib decreased STAT1 and STAT5 phosphorylation after treatment with IFN-α or IL-2. This inhibitory effect was observed in PBMCs from healthy donors over a range of concentrations of sorafenib (5-20 μM), IL-2 (2-24 nM), and IFN-α (101- 106 U/mL). This effect was observed in immune cell subsets including T cells, B cells, NK cells, regulatory T cells and myeloid-derived suppressor cells. Pre-treatment with sorafenib also inhibited PBMC expression of IFN-α- and IL-2-regulated genes and inhibited NK cell production of IFN-γ, RANTES, MIP1-α, and MIG in response to IFN-α stimulation. PBMCs from patients receiving sorafenib therapy showed decreased responsiveness to IL-2 and IFN-α treatment. Sorafenib is a Raf kinase inhibitor that could have off-target effects on cytokine-induced signal transduction in immune effector cells.

miR expression profiles can be measured in indeterminate Spitz tumors and correlate with markers of malignant potential.

Purpose To predict survival in patients with metastatic melanoma by evaluating a combination of serum lactate dehydrogenase (LDH) level and initial computed tomographic (CT) findings of tumor devascularization after antiangiogenic therapy. Materials and Methods Consent was waived for this institutional review board–approved, retrospective, secondary analysis. Forty-four patients with metastatic melanoma received bevacizumab therapy in a randomized prospective phase II trial. Target lesions on the initial posttherapy CT images were evaluated by using Response Evaluation Criteria in Solid Tumors, the Choi criteria, and Morphology, Attenuation, Size, and Structure (MASS) criteria. Cox proportional hazards models were used to assess the association of baseline clinical variables including serum LDH and imaging findings with progression-free and overall survival. The receiver operating characteristic curve with area under the curve (AUC) was used to evaluate accuracy. Results In multivariate analysis, a high baseline serum LDH level was associated with decreased progression-free survival (hazard ratio = 1.29 for each increase of 100 IU/L; P = .002) and overall survival (hazard ratio = 1.44 for each increase of 100 IU/L; P = .001). Evaluation with MASS criteria of the first CT examination after therapy strongly predicted progression-free (P < .001) and overall (P < .001) survival. Baseline serum LDH level was moderately accurate for predicting progression-free survival at 9 months (AUC = 0.793) and overall survival at 18 months (AUC = 0.689). The combination of baseline serum LDH levels and evaluation with MASS criteria at the first CT examination after therapy had significantly higher accuracy for predicting progression-free survival at 9 months (AUC = 0.969) and overall survival at 18 months (AUC = 0.813) than did baseline serum LDH levels alone for prediction of progression-free survival (P = .020). Conclusion A combination of baseline serum LDH levels and evaluation with MASS criteria at the first CT examination after bevacizumab therapy had the highest accuracy for predicting survival in patients with metastatic melanoma.