Application of specific ELISAs for BMP15 and GDF9 to cumulus cell extracts from infertile women.

Bone morphogenetic protein 15 (BMP15) and growth differentiation factor 9 (GDF9) are oocyte-specific paracrine factors which regulate ovarian cumulus cell (CC) functions. This study aimed to investigate if BMP15 and GDF9 bound to CCs can be characterized, quantified, and show an association with IVF outcomes in infertile women. BMP15 and GDF9 ELISAs were validated and applied to discarded CC extracts. Pooled CCs from individual patients were collected from 120 (cohort 1; BMP15 only) and 81 infertility patients (cohort 2; BMP15 and GDF9) undergoing superovulation. BMP15 and GDF9 levels expressed per CC DNA were correlated with maternal age, clinical and embryology data. Total BMP15 and GDF9 were highly correlated with each other (r = 0.9, p < 0.001). The GDF9:BMP15 ratio was unrelated to oocyte number or age. BMP15/CC DNA and GDF9/CC DNA were unaffected by the type of superovulation and were not related to oocyte/embryo outcomes.

Wnt addiction of genetically defined cancers reversed by PORCN inhibition.

Enhanced sensitivity to Wnts is an emerging hallmark of a subset of cancers, defined in part by mutations regulating the abundance of their receptors. Whether these mutations identify a clinical opportunity is an important question. Inhibition of Wnt secretion by blocking an essential post-translational modification, palmitoleation, provides a useful therapeutic intervention. We developed a novel potent, orally available PORCN inhibitor, ETC-1922159 (henceforth called ETC-159) that blocks the secretion and activity of all Wnts. ETC-159 is remarkably effective in treating RSPO-translocation bearing colorectal cancer (CRC) patient-derived xenografts. This is the first example of effective targeted therapy for this subset of CRC. Consistent with a central role of Wnt signaling in regulation of gene expression, inhibition of PORCN in RSPO3-translocated cancers causes a marked remodeling of the transcriptome, with loss of cell cycle, stem cell and proliferation genes, and an increase in differentiation markers. Inhibition of Wnt signaling by PORCN inhibition holds promise as differentiation therapy in genetically defined human cancers.

Surgical management of vestibular schwannoma: attempted preservation of hearing and facial function.

BACKGROUND: Vestibular schwannomas are benign tumours which usually originate from the vestibular portion of the VIIIth cranial nerve. Treatment options include observation with serial imaging, stereotactic radiation and microsurgical removal. AIM: The goal of surgery was complete eradication of tumour with preservation of hearing and facial nerve function. METHODS: A retrospective review was undertaken of 24 cases of vestibular schwannoma jointly operated upon by a team of neurosurgeons and otologists at the Suez Canal University Hospital, with assessment of VIIth and VIIIth cranial nerve function, tumour size, and extent of growth. All surgery utilised a retromastoid, suboccipital approach. RESULTS: Complete tumour removal was achieved in 19 patients. Anatomical preservation of the facial nerve was possible in 66.6 per cent of patients. Pre-operative, useful hearing was present in four patients, and preserved in 80 per cent. Cerebrospinal fluid leakage was diagnosed in two (8.3 per cent) patients, who responded to conservative therapy. CONCLUSION: The retromastoid, suboccipital surgical approach to the skull base can be safely and successfully achieved using a microsurgical technique, with minimal or no damage to neurovascular structures, even for large tumours.

Whole-exome sequencing and an iPSC-derived cardiomyocyte model provides a powerful platform for gene discovery in left ventricular hypertrophy.

RATIONALE: Left ventricular hypertrophy (LVH) is a heritable predictor of cardiovascular disease, particularly in blacks. OBJECTIVE: Determine the feasibility of combining evidence from two distinct but complementary experimental approaches to identify novel genetic predictors of increased LV mass. METHODS: Whole-exome sequencing (WES) was conducted in seven African-American sibling trios ascertained on high average familial LV mass indexed to height (LVMHT) using Illumina HiSeq technology. Identified missense or nonsense (MS/NS) mutations were examined for association with LVMHT using linear mixed models adjusted for age, sex, body weight, and familial relationship. To functionally assess WES findings, human induced pluripotent stem cell-derived cardiomyocytes (induced pluripotent stem cell-CM) were stimulated to induce hypertrophy; mRNA sequencing (RNA-seq) was used to determine gene expression differences associated with hypertrophy onset. Statistically significant findings under both experimental approaches identified LVH candidate genes. Candidate genes were further prioritized by seven supportive criteria that included additional association tests (two criteria), regional linkage evidence in the larger HyperGEN cohort (one criterion), and publically available gene and variant based annotations (four criteria). RESULTS: WES reads covered 91% of the target capture region (of size 37.2 MB) with an average coverage of 65×. WES identified 31,426 MS/NS mutations among the 21 individuals. A total of 295 MS/NS variants in 265 genes were associated with LVMHT with q-value <0.25. Of the 265 WES genes, 44 were differentially expressed (P < 0.05) in hypertrophied cells. Among the 44 candidate genes identified, 5, including HLA-B, HTT, MTSS1, SLC5A12, and THBS1, met 3 of 7 supporting criteria. THBS1 encodes an adhesive glycoprotein that promotes matrix preservation in pressure-overload LVH. THBS1 gene expression was 34% higher in hypertrophied cells (P = 0.0003) and a predicted conserved and damaging NS variant in exon 13 (A2099G) was significantly associated with LVHMT (P = 4 × 10(-6)). CONCLUSION: Combining evidence from cutting-edge genetic and cellular experiments can enable identification of novel LVH risk loci.

A novel vascular endothelial growth factor-directed therapy that selectively activates cytotoxic prodrugs.

We have generated fusion proteins between vascular endothelial growth factor (VEGF) and the bacterial enzyme carboxypeptidase G2 (CPG2) that can activate the prodrug 4-[(2-chloroethyl)(2-mesyloxyethyl)amino]benzoyl-L-glutamic acid (CMDA). Three asparagine residues of CPG2 were mutated to glutamine (CPG2(Q)3) to prevent glycosylation during secretion, and truncations of VEGF(165) were fused to either the C- or N-terminal of CPG2. The K(m) of the fusion proteins (37.5 microM) was similar to that of secreted CPG2(Q)3 (29.5 microM) but greater than that of wild-type CPG2 (8 microM). The affinity of the fusion proteins for VEGF receptor-2 (VEGFR2) (K(d)=0.5-1.1 nM) was similar to that of [(125)I]VEGF (K(d)=0.5 nM) (ELISA) or slightly higher (K(d)=1.3-9.6 nM) (competitive RIA). One protein, VEGF(115)-CPG2(Q)3-H(6), possessed 140% of the enzymic activity of secreted CPG2(Q)3, and had a faster half-maximal binding time for VEGFR2 (77 s), than the other candidates (330 s). In vitro, VEGF(115)-CPG2(Q)3-H(6) targeted CMDA cytotoxicity only towards VEGFR-expressing cells. The plasma half-life of VEGF(115)-CPG2(Q)3-H(6) in vivo was 3 h, comparable to equivalent values observed in ADEPT. We conclude that enzyme prodrug therapy using VEGF as a targeting moiety represents a promising novel antitumour therapy, with VEGF(115)-CPG2(Q)3-H(6) being a lead candidate.

Tumor angiogenesis as a therapeutic target.

Angiogenesis - the formation of new blood vessels within a tumor (or many other tissue types) - has become a hotbed of pharmacological research as well as industrial drug discovery. This is the result of the efforts of a generation of scientists elucidating the complex (patho)physiological, biochemical and molecular events accompanying angiogenesis. It is estimated that >300 drug candidates are currently in various stages of testing, and it is, therefore, impossible to capture all of this in a brief review. Therefore, the emphasis here is on relatively advanced projects that are either in preclinical or clinical development, thus neglecting, to a large extent, the many exciting avenues being pursued in both academic and biotechnology laboratories. Although the potential of the approaches described cannot be overestimated, it is also important to note that there is still no drug on the market that achieves clinical benefit based on a selective modulation or inhibition of angiogenesis.

Strong coherence between solar variability and the monsoon in Oman between 9 and 6 kyr ago.

Variations in the amount of solar radiation reaching the Earth are thought to influence climate, but the extent of this influence on timescales of millennia to decades is unclear. A number of climate records show correlations between solar cycles and climate, but the absolute changes in solar intensity over the range of decades to millennia are small and the influence of solar flux on climate is not well established. The formation of stalagmites in northern Oman has recorded past northward shifts of the intertropical convergence zone, whose northward migration stops near the southern shoreline of Arabia in the present climate. Here we present a high-resolution record of oxygen isotope variations, for the period from 9.6 to 6.1 kyr before present, in a Th-U-dated stalagmite from Oman. The delta18O record from the stalagmite, which serves as a proxy for variations in the tropical circulation and monsoon rainfall, allows us to make a direct comparison of the delta18O record with the Delta14C record from tree rings, which largely reflects changes in solar activity. The excellent correlation between the two records suggests that one of the primary controls on centennial- to decadal-scale changes in tropical rainfall and monsoon intensity during this time are variations in solar radiation.

Cool glacial temperatures and changes in moisture source recorded in oman groundwaters

Concentrations of atmospheric noble gases (neon, argon, krypton, and xenon) dissolved in groundwaters from northern Oman indicate that the average ground temperature during the Late Pleistocene (15,000 to 24,000 years before present) was 6.5 degrees +/- 0.6 degrees C lower than that of today. Stable oxygen and hydrogen isotopic groundwater data show that the origin of atmospheric water vapor changed from a primarily southern, Indian Ocean source during the Late Pleistocene to a dominantly northern, Mediterranean source today. The reduced northern water vapor source is consistent with a drier Last Glacial Maximum through much of northern Africa and Arabia.

Permanent I-125 brain stem implants in children.

Between 1988 and 1997, 28 children have had iodine-125 implants for CNS tumors performed in our institution. Ten had stereotactic implantation in the brain stem region, and nine had the diagnosis of brain stem glioma (8 diffuse pontine, 1 midbrain tumor). Their ages ranged from 1.8 to 12 years. All patients had histological confirmation of malignancy (7 high-grade glioma, 2 low-grade glioma, 1 PNET). Diffuse pontine glioma patients received external beam radiation (50 Gy) followed by a fractionated stereotactic boost of 3 Gyx4 fractions. After 4-6 weeks, patients were reevaluated for stereotactic interstitial I-125 therapy. The planned implant dose was 82.9 Gy to the enhancing tumor (4 cGy per h). Preliminary results indicated that no surgical complications were associated with the catheter placement. Four patients have died (7-9 months from diagnosis) and four patients remain alive (5-38 months from diagnosis, median 10 months). Two autopsies confirmed the presence of progressive glioblastoma multiforme and intralesional necrosis. In one patient who received an implant alone for midbrain LGA, necrosis without tumor was found on biopsy after 36 months. He was successfully treated with hyperbaric oxygen therapy. The implementation of permanent I-125 implants appears to have a role in the management of pediatric CNS malignancy. This study confirms the results of previous reports regarding the safety of stereotactic interstitial brachytherapy in the brain stem. Tumor control for patients with high-grade brain stem glioma remains poor even with high focal radiation doses.

Interactive image-guided surgical resection of intracranial arteriovenous malformations.

Surgical excision is the only treatment method that immediately prevents increased morbidity or mortality as a result of hemorrhage from arteriovenous malformations (AVMs). For those lesions located deep within the cerebral hemispheres or near eloquent areas, conventional surgical resection may be associated with an unacceptable degree of morbidity and mortality. Herein we report our experience in the resection of these lesions using interactive image guidance. There were five women and five men in the patient group. Their age ranged from 16 to 73 years (mean = 41). Clinical presentation included hemorrhage (n = 7), headaches (n = 2), and seizures (n = 1). All lesions were classified using the Spetzler-Martin grading system as follows: grade I (n = 4), grade II (n = 5), and grade III (n = 1). The locations of the lesions were supratentorial (9) and infratentorial (1). Surgical planning was carried out using the Neurological Surgery Planning System software developed at Wayne State University. An infrared-based system was used to locate and define the lesion intraoperatively. For those lesions located near or within eloquent areas, an awake craniotomy with functional mapping was carried out. Clinical follow-up ranged from 3 to 62 months (mean = 34). Complete surgical excision was achieved in all patients, which was demonstrated postoperatively by digital substraction angiography. The preoperative neurological status remained unchanged in seven patients and improved in three. There was no associated morbidity and mortality with this technique. Image-guided surgical resection of arteriovenous malformations represents a valuable technique, especially in small deep-seated lesions and in those near eloquent areas.

Markers of tumor angiogenesis and proteolysis independently define high- and low-risk subsets of node-negative breast cancer patients.

PURPOSE: To compare the prognostic impact of tumor angiogenesis factors (vascular endothelial growth factor [VEGF], angiogenin, and basic fibroblast growth factor [bFGF]), tumor proteolysis factors (urokinase-type plasminogen activator [uPA] and plasminogen activator inhibitor-1 [PAI-1]), and conventional tumor markers (stage, grade, and steroid receptors) in early breast cancer. PATIENTS AND METHODS: In the primary clinical study, tumor angiogenesis and other factors were detected in frozen biopsies from 305 primary breast tumors. VEGF expression was assessed by chemiluminescence immunosorbent assay (ICMA); angiogenin, bFGF, uPA, and PAI-1 by enzyme-linked immunosorbent assay (ELISA); and steroid receptors (estrogen receptor [ER] and progesterone receptor [PgR]) by enzyme immunoassay (EIA). In the validating clinical study, another set of 190 node-negative primary breast tumor samples were collected at a separate institution. RESULTS: Univariate analysis of the primary study showed that VEGF levels were positively correlated with recurrence (P < .001). Angiogenin levels were positively correlated with disease relapse (P < .005) for the overall collective group, but not within the node-negative subset. No significant correlations were found between tumor bFGF levels and patient survival. In multivariate regression analysis, the only independent predictors of relapse-free survival (RFS) were VEGF, uPA, and lymph node status. In the validation set, the distribution of VEGF and uPA values were similar to those in the primary study; low expression of both VEGF and uPA identified patients with a < or = 20% likelihood of recurrence within 7 years. CONCLUSION: Separate primary and validating clinical studies concur that tumor VEGF level is the most important prognostic parameter among several markers of tumor angiogenesis and proteolysis.

Interactive image guided surgery of the pineal region.

A wide variety of lesions occur in the pineal region. Of these, only 25% may benefit from surgical resection, being benign or radioresistant in nature. In the remaining 75% (malignant or radiosensitive), a conservative approach is preferred. Surgical resection of lesions arising from the pineal region has been associated with high morbidity and mortality, due to the complex microanatomy of this area. Recently computer-assisted surgery has emerged as an ideal method to safely approach many intracranial lesions. In the present study we discuss our experience in interactive image-guided surgical management of pineal mass lesions, in terms of patient selection, methodology, and surgical morbidity and mortality. From July 1992 to December 1996, 15 patients underwent interactive image guided surgical procedures. There were eight men and seven women. Age ranged from 5 to 79 years (mean 30). Preoperatively all patients underwent neuroimaging studies that included computed tomography and magnetic resonance imaging under stereotactic conditions. Surgical planning was carried out using the Neurological Surgical Planning System software developed at Wayne State University. In patients with a preoperative diagnosis of a malignant or radiosensitive lesion (n = 10), an interactive image-guided stereotactic biopsy was considered, whereas for those with benign or radioresistant lesions (n = 5) surgical excision using an infrared based system was carried out. Histological diagnosis was obtained in all patients. There was no morbidity and mortality associated with computer-assisted procedures for biopsies the diagnostic yield was 100%. For patients that underwent interactive image-guided surgery, gross total removal was accomplished in 3 and in 1 patient a subtotal resection greater than 90% was achieved. Interactive image-guided biopsy represents a safe and accurate method in the diagnosis of malignant and/or radiosensitive pineal lesions. Using this technique surgeons can simulate preoperatively the surgical trajectory, thus avoiding damage to critical and vascular structures. For patients with benign or radioresistant lesions, computer-assisted surgical excision is recommended.

Radiosurgical treatment of meningiomas.

From January 1992 to November 1996, 17 patients with the diagnosis of intracranial meningioma underwent radiosurgical treatment. Of these, 7 patients were treated using a Linac-based system (group 1), and 10 using the Leksell Gamma Knife unit (group 2). The follow-up ranged between 12 and 48 (median 33) months for group 1 and between 1 and 11 (median 5) months for group 2, consisting of clinical and MRI assessments every 3 months during the first year, and every 6 months thereafter. There were 14 women and 3 men. The mean age was 42 years. Prior to radiosurgery, 15 patients underwent surgical procedures. Histological diagnosis was consistent with benign meningioma, except in 2 patients (malignant meningioma). In 15 patients with benign meningiomas there was no evidence of tumor growth as demonstrated by clinical and radiological evaluation, in 2 patients with a malignant histological type there was tumor progression.

Interactive image-guided resection of cerebral cavernous malformations.

Between July 1992, and February 1997, 15 patients with cavernous malformations underwent interactive image-guided resection of their lesions. There were eight women and seven men in the group, ranging in age from 6 years to 62 years (mean 34 years). Clinical presentations included seizures (n = 7), headache (n = 4), and hemorrhage (n = 4). Prior conventional subtotal resection had been performed in one patient, and a history of prior hemorrhage was found for two patients. Diagnosis was made using magnetic resonance imaging and digital substraction angiography. Locations of the lesions were temporal (n = 9), frontal (n = 3), thalamus (n = 1), basal ganglia (n = 1), and pons (n = 1). Size ranged from 9 to 20 mm (mean 12 mm). For those lesions located near or within eloquent areas (n = 7), an awake craniotomy with functional cortical and subcortical mapping was performed. An infrared system was used intraoperatively to confirm the location and the extent of the resection of these lesions in real time. In 1996 we started using a robotic microscope to aid in localization and resection. Clinical follow-up ranged from 2 to 54 months (mean 27 months). In all 15 patients, complete surgical resection was achieved as demonstrated by postoperative magnetic resonance imaging studies. Two patients had postoperative transient neurological deficits (13.3%) that cleared over a 6-month period. One of them had a lesion in the pons, with multiple cranial nerve deficits postoperatively that gradually improved. There was no associated mortality. Histological diagnosis was consistent with cavernous angioma in all cases. Clinical follow-up revealed that 13 patients experienced complete recovery from preoperative symptoms and two patients with seizures showed marked improvement. We conclude that interactive image-guided surgery for deep-seated cavernous malformations represents a very accurate and safe approach.

Cellular and in vivo characterization of the MCR rat mammary tumor model.

The slowly growing, transplantable MCR-83 rat mammary tumor is estrogen-dependent and non-metastasizing. A rapidly growing, estrogen-independent, metastasizing subline (MCR-86) was subsequently isolated in vivo. We have established and characterized cell lines from both MCR rat mammary tumors. MCR cell lines and tumors were studied in vivo and in vitro. Analysis of DNA from tumors and cell lines showed that mutations had not occurred in codons 12, 13 and 61 of the Ha-ras and Ki-ras genes. Additionally, dominant transforming activity could not be detected by DNA transfection using NIH 3T3 focus-forming assay. No gene amplification was detected for either the EGF-receptor or c-erbB-2 genes. Differences in the tyrosine phosphorylation patterns were found between the 2 MCR cell lines. Addition of serum to starved cells resulted in the tyrosine phosphorylation of a 120-kDa protein, which was elevated in the MCR-86. The lack of ras activation in the MCR tumors differentiates this model from the widely studied, chemically induced rodent mammary tumors. In addition, the differences in the cellular phosphotyrosine patterns between MCR-83 and MCR-86 suggests the occurrence of alterations in signalling pathways that involve tyrosine protein kinases.

Obesity among secondary school students in Bahrain.

The aim of this study is to estimate the prevalence of obesity and factors associated with it in Bahraini secondary school students. A cross-sectional study involving a sample of 825 students (417 boys and 408 girls) aged 15 to 21 years was obtained from secondary schools. Obesity was determined using body mass index (BMI = Wt/Ht2). The findings revealed that 15.6% of boys and 17.4% of girls were either overweight or obese (BMI > or = 25). Family size, parents education, and family history of obesity were significantly associated with obesity among boys, while family history was the only socio-economic factors statistically associated with obesity among girls. Meal patterns such as eating between meals, number of meals per day, and method of eating were not associated with obesity in students. Boys who ate alone were 3 times more likely to be obese than those who ate with family members (odd ratio = 3.4). Measures to prevent and control obesity among children are suggested.

New S-adenosylmethionine decarboxylase inhibitors with potent antitumor activity.

Methylglyoxal bis(guanylhydrazone) (MGBG) has been studied clinically as an antitumor and antileukemic agent and is recognized as a potent but nonspecific inhibitor of the key polyamine biosynthetic enzyme, S-adenosylmethionine decarboxylase (SAMDC). A series of four SAMDC inhibitors with structural features similar to MGBG have been found to have improved potency and specificity toward the target enzyme, SAMDC. Relative to MGBG, the new derivatives were much more effective in inhibiting partially purified preparations of SAMDC (50% inhibitory concentration, 10 to 100 nM), much less effective at inhibiting diamine oxidase, and inactive toward ornithine decarboxylase. The inhibitors varied relative to MGBG in their ability to compete with spermidine for uptake, with two being similar and two being less effective. Against L1210 leukemic cells and T24 bladder carcinoma cells, the compounds were slightly less effective than MGBG at inhibiting cell growth, with 50% inhibitory concentration values of 1 to 10 microM as compared with 0.5 and 1.1 microM, respectively, for MGBG. Under 50% growth-inhibitory conditions, the inhibitors decreased SAMDC activity, increased ornithine decarboxylase activity and putrescine pools, and markedly depleted spermidine and spermine pools of L1210 cells. At the same time, mitochondrial integrity as assessed by whole-cell pyruvate oxidation and mitochondrial DNA content was not affected as it was with MGBG. At doses less than one tenth that of the maximally tolerated dose, all of the new inhibitors strongly suppressed the growth of B16 melanoma in vivo with minimal weight loss or toxicity. At doses less than one sixth the maximally tolerated dose, they effectively inhibited the growth of T24 human bladder carcinoma xenografts. In these same systems, MGBG showed only marginal antitumor activity. These studies identify two potent and efficacious inhibitors of SAMDC as potential antitumor agents and reaffirm the importance of SAMDC as a target in anticancer drug discovery.