Supernumerary marker chromosomes (SMCs) in Turner syndrome are mostly derived from the Y chromosome

DNA and FISH (fluorescence in situ hybridization) analysis were carried out in 12 patients with stigmata of Turner syndrome to determine whether the Supernumerary M arker C hromosome (SMC) found cytogenetically in each of these patients was derived from the Y chromosome. The presence of a Y chromosome in these patients may predispose them to develop gonadoblastoma. PCR-Southern blot analysis, followed by FISH, was used to detect the presence of Y chromosome material. The S ex determining R egion Y (SRY), T estis S pecific P rotein Y -encoded (TSPY) and Y -chromosome R NA R ecognition M otif (YRRM) genes, which map at Yp11.31, Yp11.1–11.2 and Yp11.2/Yq11.21–11.23, respectively, were selected as markers, because they span the whole Y chromosome, and more importantly, they are considered to be involved in the development of gonadoblastoma. It was shown that in 12 patients, all of whom had an SMC, the SMC of 11 was derived from the Y chromosome. Furthermore, the presence of the SRY, TSPY and YRRM gene sequences was determined and FISH analysis confirmed the Y origin of the SMCs. The methodology described in this report is a rapid, reliable and sensitive approach which may be easily applied to determine the Y origin of an SMC carried in Turner syndrome. The identification of an SMC is important for the clinical management and prognostic counseling of these patients with Turner syndrome.     

Detection and incidence of cryptic Y chromosome sequences in Turner syndrome patients

The presence of Y chromosome sequences in Turner syndrome (TS) patients may predispose them to gonadoblastoma formation with an estimated risk of 15–25%. The aim of this study was to determine the presence and the incidence of cryptic Y chromosome material in the genome of TS patients. The methodology involved a combination of polymerase chain reaction (PCR) and nested PCR followed by Southern blot analysis of three genes—the sex determining region Y (SRY), testis specific protein Y encoded (TSPY) and RNA binding motif protein (RBM) (previously designated as YRRM) and nine additional STSs spanning all seven intervals of the Y chromosome. The methodology has a high sensitivity as it detects one 46, XY cell among 10⁵ 46, XX cells. Reliability was ensured by taking several precautions to avoid false positive results. We report the results of screening 50 TS patients and the identification of cryptic Y chromosome material in 12 (24%) of them. Karyotypes were divided in four groups: 5 (23.8%) patients out of the 21 TS patients which have the 45, X karyotype (group A) also have cryptic Y sequences; none (0%) of the 7 patients who have karyotypes with anomalies on one of the X chromosomes have Y mosaicism (group B); 1 (6.3%) of the 16 patients with a mosaic karyotype have Y material (group C); and 6 (100%) out of 6 patients with a supernumerary marker chromosome (SMC) have Y chromosome sequences (group D). Nine of the 12 patients positive for cryptic Y material were recalled for a repeat study. Following new DNA extraction, molecular analysis was repeated and, in conjunction with fluorescent in situ hybridization (FISH) analysis using the Y centromeric specific probe Yc-2, confirmed the initial positive DNA findings. This study used a reliable and sensitive methodology to identify the presence of Y chromosome material in TS patients thus providing not only a better estimate of a patient's risk in developing either gonadoblastoma or another form of gonadal tumor but also the overall incidence of cryptic Y mosaicism.     

Fast Eating Is Associated with Increased BMI among High-School Students

Fast self-reported eating rate (SRER) has been associated with increased adiposity in children and adults. No studies have been conducted among high-school students, and SRER has not been validated vs. objective eating rate (OBER) in such populations. The objectives were to investigate (among high-school student populations) the association between OBER and BMI z-scores (BMIz), the validity of SRER vs. OBER, and potential differences in BMIz between SRER categories. Three studies were conducted. Study 1 included 116 Swedish students (mean ± SD age: 16.5 ± 0.8, 59% females) who were eating school lunch. Food intake and meal duration were objectively recorded, and OBER was calculated. Additionally, students provided SRER. Study 2 included students (n = 50, mean ± SD age: 16.7 ± 0.6, 58% females) from Study 1 who ate another objectively recorded school lunch. Study 3 included 1832 high-school students (mean ± SD age: 15.8 ± 0.9, 51% females) from Sweden (n = 748) and Greece (n = 1084) who provided SRER. In Study 1, students with BMIz ≥ 0 had faster OBER vs. students with BMIz < 0 (mean difference: +7.7 g/min or +27%, p = 0.012), while students with fast SRER had higher OBER vs. students with slow SRER (mean difference: +13.7 g/min or +56%, p = 0.001). However, there was “minimal” agreement between SRER and OBER categories (κ = 0.31, p < 0.001). In Study 2, OBER during lunch 1 had a “large” correlation with OBER during lunch 2 (r = 0.75, p < 0.001). In Study 3, fast SRER students had higher BMIz vs. slow SRER students (mean difference: 0.37, p < 0.001). Similar observations were found among both Swedish and Greek students. For the first time in high-school students, we confirm the association between fast eating and increased adiposity. Our validation analysis suggests that SRER could be used as a proxy for OBER in studies with large sample sizes on a group level. With smaller samples, OBER should be used instead. To assess eating rate on an individual level, OBER can be used while SRER should be avoided.     

Randomized trial of amifostine in locally advanced non-small-cell lung cancer patients receiving chemotherapy and hyperfractionated radiation: radiation therapy oncology group trial 98-01.

PURPOSE: To test the ability of the cytoprotectant, amifostine, to reduce chemoradiotherapy-induced esophagitis and evaluate its influence on quality of life (QOL) and swallowing symptoms. PATIENTS AND METHODS: A total of 243 patients with stage II to IIIA/B non-small-cell lung cancer received induction paclitaxel 225 mg/m(2) intravenously (IV) days 1 and 22 and carboplatin area under the curve (AUC) days 1 and 22, followed by concurrent weekly paclitaxel (50 mg/m(2) IV) and carboplatin (AUC 2), and hyperfractionated radiation therapy (69.6 Gy at 1.2 Gy bid). Patients were randomly assigned at registration to amifostine (AM) 500 mg IV four times per week or no AM during chemoradiotherapy. Beyond standard toxicity end points, physician dysphagia logs (PDLs), daily patient swallowing diaries, and QOL (EORTC QLQ-C30/LC-13) were also collected. Swallowing AUC analyses were calculated from patient diaries and PDLs. RESULTS: A total of 120 patients were randomly assigned to receive AM, and 122, to receive no AM (one patient was ineligible); 72% received AM per protocol or with a minor deviation. AM was associated with higher rates of acute nausea (P = .03), vomiting (P = .007), cardiovascular toxicity (P = .0001), and infection or febrile neutropenia (P = .03). The rate of >/= grade 3 esophagitis was 30% with AM versus 34% without AM (P = .9). Patient diaries demonstrated lower swallowing dysfunction AUC with amifostine (z test P = .025). QOL was not significantly different between the two arms, except for pain, which showed more clinically meaningful improvement and less deterioration at 6 weeks follow-up (v pretreatment) in the AM arm (P = .003). The median survival rates for both arms were comparable (AM, 17.3 v no AM, 17.9 months; P = .87). CONCLUSION: AM did not significantly reduce esophagitis >/= grade 3 in patients receiving hyperfractionated radiation and chemotherapy. However, patient self-assessments suggested a possible advantage to AM that is being explored with modified dosing route strategies.     

Reconstruction of an active SOCS3-based E3 ubiquitin ligase complex in vitro: identification of the active components and JAK2 and gp130 as substrates

Abstract

SOCS3 (suppressor of cytokine signaling 3) inhibits the intracellular signaling cascade initiated by exposure of cells to cytokines. SOCS3 regulates signaling via two distinct mechanisms: directly inhibiting the catalytic activity of Janus kinases (JAKs) that initiate the intracellular signaling cascade and catalysing the ubiquitination of signaling components by recruiting components of an E3 ubiquitin ligase complex. Here we investigate the latter mode-of-action biochemically by reconstructing a SOCS3-based E3 ubiquitin ligase complex in vitro using fully purified, recombinant components and examining its ability to promote the ubiquitination of molecules involved in the cytokine signaling cascade. We show that SOCS3 is an active substrate recruitment module for a Cullin5-based E3 ligase and have defined the core protein components required for ubiquitination. SOCS3-induced polyubiquitination was rapid and could proceed through a number of different ubiquitin lysines. SOCS3 catalyzed the ubiquitination of both the IL-6 receptor common chain (gp130) and JAK2.     

Difference in outcomes after antibody‐mediated rejection between abo‐incompatible and positive cross‐match transplantations

Graft survival seems to be worse in positive cross‐match (HLAi) than in ABO‐incompatible (ABOi) transplantation. However, it is not entirely clear why these differences exist. Sixty‐nine ABOi, 27 HLAi and 10 combined ABOi+HLAi patients were included in this retrospective study, to determine whether the frequency, severity and the outcome of active antibody‐mediated rejection (AMR) were different. Five‐year death‐censored graft survival was better in ABOi than in HLAi and ABOi+HLAi patients (99%, 69% and 64%, respectively, P = 0.0002). Features of AMR were found in 38%, 95% and 100% of ABOi, HLAi and ABOi+HLAi patients that had a biopsy, respectively (P = 0.0001 and P = 0.001). After active AMR, a declining eGFR and graft loss were observed more frequently in HLAi and HLAi+ABOi than in ABOi patients. The poorer prognosis after AMR in HLAi and ABOi+HLAi transplantations was not explained by a higher severity of histological lesions or by a less aggressive treatment. In conclusion, ABOi transplantation offers better results than HLAi transplantation, partly because AMR occurs less frequently but also because outcome after AMR is distinctly better. HLAi and combined ABOi+HLAi transplantations appear to have the same outcome, suggesting there is no synergistic effect between anti‐A/B and anti‐HLA antibodies.     

Discovery of an orally active small-molecule irreversible inhibitor of protein disulfide isomerase for ovarian cancer treatment

Protein disulfide isomerase (PDI), an endoplasmic reticulum chaperone protein, catalyzes disulfide bond breakage, formation, and rearrangement. The effect of PDI inhibition on ovarian cancer progression is not yet clear, and there is a need for potent, selective, and safe small-molecule inhibitors of PDI. Here, we report a class of propynoic acid carbamoyl methyl amides (PACMAs) that are active against a panel of human ovarian cancer cell lines. Using fluorescent derivatives, 2D gel electrophoresis, and MS, we established that PACMA 31, one of the most active analogs, acts as an irreversible small-molecule inhibitor of PDI, forming a covalent bond with the active site cysteines of PDI. We also showed that PDI activity is essential for the survival and proliferation of human ovarian cancer cells. In vivo, PACMA 31 showed tumor targeting ability and significantly suppressed ovarian tumor growth without causing toxicity to normal tissues. These irreversible small-molecule PDI inhibitors represent an important approach for the development of targeted anticancer agents for ovarian cancer therapy, and they can also serve as useful probes for investigating the biology of PDI-implicated pathways.