GLI2-Mediated Melanoma Invasion and Metastasis

Background The transforming growth factor-β (TGF-β) pathway, which has both tumor suppressor and pro-oncogenic activities, is often constitutively active in melanoma and is a marker of poor prognosis. Recently, we identified GLI2, a mediator of the hedgehog pathway, as a transcriptional target of TGF-β signaling. Methods We used real-time reverse transcription-polymerase chain reaction (RT-PCR) and western blotting to determine GLI2 expression in human melanoma cell lines and subsequently classified them as GLI2high or as GLI2low according to their relative GLI2 mRNA and protein expression levels. GLI2 expression was reduced in a GLI2high cell line with lentiviral expression of short hairpin RNA targeting GLI2. We assessed the role of GLI2 in melanoma cell invasiveness in Matrigel assays. We measured secretion of matrix metalloproteinase (MMP)-2 and MMP-9 by gelatin zymography and expression of E-cadherin by western blotting and RT-PCR. The role of GLI2 in development of bone metastases was determined following intracardiac injection of melanoma cells in immunocompromised mice (n = 5-13). Human melanoma samples (n = 79) at various stages of disease progression were analyzed for GLI2 and E-cadherin expression by immunohistochemistry, in situ hybridization, or RT-PCR. All statistical tests were two-sided. Results Among melanoma cell lines, increased GLI2 expression was associated with loss of E-cadherin expression and with increased capacity to invade Matrigel and to form bone metastases in mice (mean osteolytic tumor area: GLI2high vs GLI2low, 2.81 vs 0.93 mm2, difference = 1.88 mm2, 95% confidence interval [CI] = 1.16 to 2.60, P < .001). Reduction of GLI2 expression in melanoma cells that had expressed high levels of GLI2 substantially inhibited both basal and TGF-β-induced cell migration, invasion (mean number of Matrigel invading cells: shGLI2 vs shCtrl (control), 52.6 vs 100, difference = 47.4, 95% CI = 37.0 to 57.8, P = .024; for shGLI2 + TGF-β vs shCtrl + TGF-β, 31.0 vs 161.9, difference = −130.9, 95% CI = −96.2 to −165.5, P = .002), and MMP secretion in vitro and the development of experimental bone metastases in mice. Within human melanoma lesions, GLI2 expression was heterogeneous, associated with tumor regions in which E-cadherin was lost and increased in the most aggressive tumors. Conclusion GLI2 was directly involved in driving melanoma invasion and metastasis in this preclinical stud

Nutrition, diet and immunosenescence

Ageing is characterized by immunosenescence and the progressive decline in immunity in association with an increased frequency of infections and chronic disease. This complex process affects both the innate and adaptive immune systems with a progressive decline in most immune cell populations and defects in activation resulting in loss of function. Although host genetics and environmental factors, such as stress, exercise and diet can impact on the onset or course of immunosenescence, the mechanisms involved are largely unknown. This review focusses on identifying the most significant aspects of immunosenescence and on the evidence that nutritional intervention might delay this process, and consequently improve the quality of life of the elderly

Novel avian paramyxovirus isolated from gulls in Caspian seashore in Kazakhstan.

Three isolates APMV/gull/Kazakhstan/5976/2014, APMV/gull/Kazakhstan/ 5977/2014 and APMV/gull/Kazakhstan/5979/2014, were obtained from independent samples during annual surveillance for avian influenza and paramyxoviruses in wild birds from the Caspian Sea coast in Western Kazakhstan, and were initially identified as putative paramyxoviruses on the basis of electron microscopy. Hemagglutination Inhibition Assays with antisera to nine known APMV serotypes (APMV1-9) indicated no relation to any of them. Next generation sequencing of whole genome sequences indicated the three isolates were genetically identical, and had a nucleotide structure typical for all APMVs, consisting of six genes 3'-NP-P-M-F-HN-L-5'. Phylogenetic analyses, and assessment of amino acid identities, suggested the most closely related lineages to be APMV-2, 8, 10 and 15, but the novel isolate had less than 64% identity to them and all other known avian paramyxoviruses. This value was above levels considered to generally define other APMV serotypes. Estimates of the evolutionary divergence of the nucleotide sequences of the genomes of APMVs have shown that novel Kazakhstan APMV strain was closest to APMV-2, APMV-8, APMV-10 and APMV-15, with calculated distance values of 2.057, 2.058, 2.026 and 2.286 respectively, which is above values considered to differentiate other serotypes (observed minimum was 1.108 between APMV-1 and recently isolated APMV/UPO216/Korea). Together, the data suggest that isolate APMV/gull/Kazakhstan/5976/2014 and other two should be considered as the first representative of a novel APMV-20 group, and is the first time that avian paramyxoviruses have been found infecting members of the gull family, extending the known taxonomic host range

Functional mechanisms underlying pleiotropic risk alleles at the 19p13.1 breast-ovarian cancer susceptibility locus

A locus at 19p13 is associated with breast cancer (BC) and ovarian cancer (OC) risk. Here we analyse 438 SNPs in this region in 46,451 BC and 15,438 OC cases, 15,252 BRCA1 mutation carriers and 73,444 controls and identify 13 candidate causal SNPs associated with serous OC (P=9.2 × 10-20), ER-negative BC (P=1.1 × 10-13), BRCA1-associated BC (P=7.7 × 10-16) and triple negative BC (P-diff=2 × 10-5). Genotype-gene expression associations are identified for candidate target genes ANKLE1 (P=2 × 10-3) and ABHD8 (P<2 × 10-3). Chromosome conformation capture identifies interactions between four candidate SNPs and ABHD8, and luciferase assays indicate six risk alleles increased transactivation of the ADHD8 promoter. Targeted deletion of a region containing risk SNP rs56069439 in a putative enhancer induces ANKLE1 downregulation; and mRNA stability assays indicate functional effects for an ANKLE1 3′-UTR SNP. Altogether, these data suggest that multiple SNPs at 19p13 regulate ABHD8 and perhaps ANKLE1 expression, and indicate common mechanisms underlying breast and ovarian cancer risk

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P &lt; 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

The FANCM:p.Arg658* truncating variant is associated with risk of triple-negative breast cancer

Abstract: Breast cancer is a common disease partially caused by genetic risk factors. Germline pathogenic variants in DNA repair genes BRCA1, BRCA2, PALB2, ATM, and CHEK2 are associated with breast cancer risk. FANCM, which encodes for a DNA translocase, has been proposed as a breast cancer predisposition gene, with greater effects for the ER-negative and triple-negative breast cancer (TNBC) subtypes. We tested the three recurrent protein-truncating variants FANCM:p.Arg658*, p.Gln1701*, and p.Arg1931* for association with breast cancer risk in 67,112 cases, 53,766 controls, and 26,662 carriers of pathogenic variants of BRCA1 or BRCA2. These three variants were also studied functionally by measuring survival and chromosome fragility in FANCM−/− patient-derived immortalized fibroblasts treated with diepoxybutane or olaparib. We observed that FANCM:p.Arg658* was associated with increased risk of ER-negative disease and TNBC (OR = 2.44, P = 0.034 and OR = 3.79; P = 0.009, respectively). In a country-restricted analysis, we confirmed the associations detected for FANCM:p.Arg658* and found that also FANCM:p.Arg1931* was associated with ER-negative breast cancer risk (OR = 1.96; P = 0.006). The functional results indicated that all three variants were deleterious affecting cell survival and chromosome stability with FANCM:p.Arg658* causing more severe phenotypes. In conclusion, we confirmed that the two rare FANCM deleterious variants p.Arg658* and p.Arg1931* are risk factors for ER-negative and TNBC subtypes. Overall our data suggest that the effect of truncating variants on breast cancer risk may depend on their position in the gene. Cell sensitivity to olaparib exposure, identifies a possible therapeutic option to treat FANCM-associated tumors

Determining crystal structures through crowdsourcing and coursework

We show here that computer game players can build high-quality crystal structures. Introduction of a new feature into the computer game Foldit allows players to build and real-space refine structures into electron density maps. To assess the usefulness of this feature, we held a crystallographic model-building competition between trained crystallographers, undergraduate students, Foldit players and automatic model-building algorithms. After removal of disordered residues, a team of Foldit players achieved the most accurate structure. Analysing the target protein of the competition, YPL067C, uncovered a new family of histidine triad proteins apparently involved in the prevention of amyloid toxicity. From this study, we conclude that crystallographers can utilize crowdsourcing to interpret electron density information and to produce structure solutions of the highest quality

The predictive ability of the 313 variant–based polygenic risk score for contralateral breast cancer risk prediction in women of European ancestry with a heterozygous BRCA1 or BRCA2 pathogenic variant

Abstract: Purpose: To evaluate the association between a previously published 313 variant–based breast cancer (BC) polygenic risk score (PRS313) and contralateral breast cancer (CBC) risk, in BRCA1 and BRCA2 pathogenic variant heterozygotes. Methods: We included women of European ancestry with a prevalent first primary invasive BC (BRCA1 = 6,591 with 1,402 prevalent CBC cases; BRCA2 = 4,208 with 647 prevalent CBC cases) from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA), a large international retrospective series. Cox regression analysis was performed to assess the association between overall and ER-specific PRS313 and CBC risk. Results: For BRCA1 heterozygotes the estrogen receptor (ER)-negative PRS313 showed the largest association with CBC risk, hazard ratio (HR) per SD = 1.12, 95% confidence interval (CI) (1.06–1.18), C-index = 0.53; for BRCA2 heterozygotes, this was the ER-positive PRS313, HR = 1.15, 95% CI (1.07–1.25), C-index = 0.57. Adjusting for family history, age at diagnosis, treatment, or pathological characteristics for the first BC did not change association effect sizes. For women developing first BC < age 40 years, the cumulative PRS313 5th and 95th percentile 10-year CBC risks were 22% and 32% for BRCA1 and 13% and 23% for BRCA2 heterozygotes, respectively. Conclusion: The PRS313 can be used to refine individual CBC risks for BRCA1/2 heterozygotes of European ancestry, however the PRS313 needs to be considered in the context of a multifactorial risk model to evaluate whether it might influence clinical decision-making

A case-only study to identify genetic modifiers of breast cancer risk for BRCA1/BRCA2 mutation carriers

Abstract: Breast cancer (BC) risk for BRCA1 and BRCA2 mutation carriers varies by genetic and familial factors. About 50 common variants have been shown to modify BC risk for mutation carriers. All but three, were identified in general population studies. Other mutation carrier-specific susceptibility variants may exist but studies of mutation carriers have so far been underpowered. We conduct a novel case-only genome-wide association study comparing genotype frequencies between 60,212 general population BC cases and 13,007 cases with BRCA1 or BRCA2 mutations. We identify robust novel associations for 2 variants with BC for BRCA1 and 3 for BRCA2 mutation carriers, P < 10−8, at 5 loci, which are not associated with risk in the general population. They include rs60882887 at 11p11.2 where MADD, SP11 and EIF1, genes previously implicated in BC biology, are predicted as potential targets. These findings will contribute towards customising BC polygenic risk scores for BRCA1 and BRCA2 mutation carriers

Functional mechanisms underlying pleiotropic risk alleles at the 19p13.1 breast-ovarian cancer susceptibility locus

A locus at 19p13 is associated with breast cancer (BC) and ovarian cancer (OC) risk. Here we analyse 438 SNPs in this region in 46,451 BC and 15,438 OC cases, 15,252 BRCA1 mutation carriers and 73,444 controls and identify 13 candidate causal SNPs associated with serous OC (P = 9.2 x 10(-20)), ER-negative BC (P = 1.1 x 10(-13)), BRCA1-associated BC (P = 7.7 x 10(-16)) and triple negative BC (P-diff = 2 x 10(-5)). Genotype-gene expression associations are identified for candidate target genes ANKLE1 (P = 2 x 10(-3)) and ABHD8 (PPeer reviewe