X-exome sequencing of 405 unresolved families identifies seven novel intellectual disability genes.

X-linked intellectual disability (XLID) is a clinically and genetically heterogeneous disorder. During the past two decades in excess of 100 X-chromosome ID genes have been identified. Yet, a large number of families mapping to the X-chromosome remained unresolved suggesting that more XLID genes or loci are yet to be identified. Here, we have investigated 405 unresolved families with XLID. We employed massively parallel sequencing of all X-chromosome exons in the index males. The majority of these males were previously tested negative for copy number variations and for mutations in a subset of known XLID genes by Sanger sequencing. In total, 745 X-chromosomal genes were screened. After stringent filtering, a total of 1297 non-recurrent exonic variants remained for prioritization. Co-segregation analysis of potential clinically relevant changes revealed that 80 families (20%) carried pathogenic variants in established XLID genes. In 19 families, we detected likely causative protein truncating and missense variants in 7 novel and validated XLID genes (CLCN4, CNKSR2, FRMPD4, KLHL15, LAS1L, RLIM and USP27X) and potentially deleterious variants in 2 novel candidate XLID genes (CDK16 and TAF1). We show that the CLCN4 and CNKSR2 variants impair protein functions as indicated by electrophysiological studies and altered differentiation of cultured primary neurons from Clcn4(-/-) mice or after mRNA knock-down. The newly identified and candidate XLID proteins belong to pathways and networks with established roles in cognitive function and intellectual disability in particular. We suggest that systematic sequencing of all X-chromosomal genes in a cohort of patients with genetic evidence for X-chromosome locus involvement may resolve up to 58% of Fragile X-negative cases.

A 1q44 deletion, paternal UPD of chromosome 2 and a deletion due to a complex translocation detected in children with abnormal phenotypes using new SNP array technology.

Children with intellectual disability, dysmorphic features, malformations and/or growth abnormalities frequently display normal karyotypes. Recent studies have shown that genome-wide single nucleotide polymorphism (SNP) arrays can be effective in detecting abnormalities involving copy number variation (CNV), deletions, duplications and loss of heterozygosity (LOH) that routine cytogenetic tests fail to identify. Five patients with various degrees of intellectual disability and/or dysmorphic features and other malformations were whole-genome genotyped using the Human-1 Genotyping BeadChip--Exon-Centrix 100K SNP arrays (Illumina). All patients had undergone routine cytogenetic testing; four patients had normal karyotypes, while one patient had an apparently balanced complex translocation involving chromosomes 1q25, 1q32, 2q23, 7q22 and 16q24. We detected deletions on chromosome 1q44 and 13q31.1 in one patient, and LOH of the entire chromosome 2 in another patient, both with cytogenetically normal karyotypes. The patient with the complex translocation had a deletion on chromosome 7q22.2-22.3, which is in conjunction with one of the translocation breakpoints. Our findings provide further evidence of there being a critical region for the development of microcephaly and corpus callosum abnormalities in children with distal 1q deletions. We have also shown that apparently balanced complex translocations might not be balanced at the DNA level, and we report the fourth case of paternal uniparental disomy of chromosome 2. The results of this study suggest that it may be desirable to investigate idiopathic mental retardation using genome-wide SNP arrays, in conjunction with other cytogenetic and molecular techniques.

Familial 22q11.2 duplication: a three-generation family with a 3-Mb duplication and a familial 1.5-Mb duplication.

We report two familial cases of 22q11.2 duplication detected using multiplex ligation-dependent probe amplification (MLPA). In the first case, eight individuals from a three-generation family were found to carry a 3-Mb 22q11.2 duplication. The individuals carrying the duplication show phenotypic variation. This phenotypic variation includes heart defect (1 in 8 individuals, 1/8), submucous cleft palate (2/8), intellectual disability (2/8), speech delay (2/8), behaviour problems (3/8) and brachydactyly (3/8). In the second case, a 1.5-Mb 22q11.2 duplication was detected in a neonate and her normal mother. The neonate presented with severe laryngomalacia causing intermittent stridor. Cranial ultrasound showed small subependymal cysts bilaterally. There was no heart defect or cleft palate, her chest X ray and renal ultrasound were normal. Review at 2 months of age revealed normal growth and development. Our findings broaden the understanding of 22q11.2 duplication syndrome and demonstrate that MLPA is sensitive for detection and sizing of 22q11.2 microduplications.

Smoking rates and smoking cessation among individuals with multiple sclerosis.

PURPOSE: Adults with physical disabilities tend to smoke at higher rates than smokers in the general population. No study to date, however, has assessed smoking prevalence and cessation among individuals with multiple sclerosis (MS). Such information is critically needed because smoking is more deleterious for individuals with MS than for smokers without MS and increases MS risk. METHOD: Questionnaires were sent to 700 National Multiple Sclerosis Society Rhode Island Chapter members. RESULTS: Based on a 50% response rate, results demonstrated a 15.2% current smoker prevalence rate, which is lower than USA and Rhode Island general adult population averages. Individuals who smoked, however, tended to be heavy smokers, consuming 20 - 30 cigarettes daily, and had been smoking 10 years or longer. Smokers varied in their interest in quitting but seemed confident in their ability to do so. Respondents reported that it was difficult to quit because smoking was pleasurable; smoking was helpful in coping with boredom and with having MS; withdrawal symptoms were unpleasant; and treatment for tobacco dependence was expensive. CONCLUSIONS: Efficacious smoking cessation interventions for smokers with MS should be developed to address a critical health need for a population of highly nicotine-dependent smokers who face numerous obstacles to quitting.

Autosomal dominant congenital non-progressive ataxia overlaps with the SCA15 locus.

BACKGROUND: Most patients with pure nonprogressive congenital cerebellar ataxia have a sporadic form of unknown heredity and etiology. Several small families have been reported with a dominantly inherited nonprogressive congenital ataxia (NPCA). METHODS: The authors ascertained and clinically characterized a four-generation pedigree segregating an autosomal dominant type of congenital nonprogressive cerebellar ataxia associated with cognitive impairment. Following the exclusion of several SCA localizations (SCA-1, 2, 3, 4, 5, 6, 7, 8, 10, 12, 17, IOSCA, and DRPLA), a genome-wide linkage study was performed. RESULTS: Examination of the family showed that all affected members had gait ataxia and cognitive disability with variable features of dysarthria, dysmetria, dysdiadochokinesia, nystagmus, dystonic movements, and cerebellar hypoplasia on imaging. Clinical signs of pyramidal tract dysfunction and sensory changes were absent. A genome-wide search in this family detected linkage to chromosome 3p with a maximum two-point lod score of 4.26 at D3S3630. This localization to the pter is distal to D3S1304, as defined by a recombination event. This overlaps with the SCA15 locus, with the critical overlapping region between the microsatellite markers, D3S1304 and D3S1620 (approximately 8 cM). CONCLUSION: Autosomal dominant congenital nonprogressive cerebellar ataxia with or without cerebellar hypoplasia overlaps with the SCA15 locus on chromosome 3pter.

Growth hormone receptor antagonist administration inhibits growth of human colorectal carcinoma in nude mice.

Increasing evidence has accumulated in support of the hypothesis that growth hormone (GH) and insulin-like growth factors (IGFs) play a role in carcinogenesis. In order to test this hypothesis, female nude mice were xenografted with two different human colorectal cancer cell lines (COLO 205 and HT-29) and randomized to receive placebo or a GH receptor antagonist (GHRA) (B2036-PEG) every second day for 16 days. The tumour volume was measured in each animal throughout the study and by the end of the experiment the tumour weights were recorded. After 16 days of therapy in nude mice with the COLO 205 colorectal cancer, GHRA treatment caused a 39% reduction in tumour volume (p < 0.02) and a 44% reduction in tumour weight (p < 0.01). GHRA treatment equally reduced circulating IGF-I and IGFBP-3 levels, while apoptosis was increased in the treatment group. Expression of IGF-I, IGF-II and the corresponding receptors in COLO 205 tumours was also decreased by the treatment. GHRA had no effect on the growth of the HT-29 colorectal cancer despite pronounced reduction in serum IGF-I. The present study thereby demonstrates a central role for the GH/IGF system in the pathogenesis of some colorectal cancers and suggests that specific GHR blockade may present a new concept in the treatment of colorectal cancer.

Examining the effects of tobacco treatment policies on smoking rates and smoking related deaths using the SimSmoke computer simulation model.

OBJECTIVES: To develop a simulation model to predict the effects of different smoking treatment policies on quit rates, smoking rates, and smoking attributable deaths. METHODS: We first develop a decision theoretic model of quitting behaviour, which incorporates the decision to quit and the choice of treatment. A model of policies to cover the costs of different combinations of treatments and to require health care provider intervention is then incorporated into the quit model. The policy model allows for the smoker to substitute between treatments and for policies to reduce treatment effectiveness. The SimSmoke computer simulation model is then used to examine policy effects on smoking rates and smoking attributable deaths. RESULTS: The model of quit behaviour predicts a population quit rate of 4.3% in 1993, which subsequently falls and then increases in recent years to 4.5%. The policy model suggests a 25% increase in quit rates from a policy that mandates brief interventions and the coverage of all proven treatments. Smaller effects are predicted from policies that mandate more restricted coverage of treatments, especially those limited to behavioural treatment. These policies translate into small reductions in the smoking rate at first, but increase to as much as a 5% reduction in smoking rates. They also lead to substantial savings in lives. CONCLUSIONS: Tobacco treatment policies, especially those with broad and flexible coverage, have the potential to increase smoking cessation substantially and decrease smoking rates in the short term, with fairly immediate reductions in deaths.

Long-term treatment of acromegaly with pegvisomant, a growth hormone receptor antagonist.

BACKGROUND: Pegvisomant is a new growth hormone receptor antagonist that improves symptoms and normalises insulin-like growth factor-1 (IGF-1) in a high proportion of patients with acromegaly treated for up to 12 weeks. We assessed the effects of pegvisomant in 160 patients with acromegaly treated for an average of 425 days. METHODS: Treatment efficacy was assessed by measuring changes in tumour volume by magnetic resonance imaging, and serum growth hormone and IGF-1 concentrations in 152 patients who received pegvisomant by daily subcutaneous injection for up to 18 months. The safety analysis included 160 patients some of whom received weekly injections and are excluded from the efficacy analysis. FINDINGS: Mean serum IGF-1 concentrations fell by at least 50%: 467 mg/L (SE 24), 526 mg/L (29), and 523 mg/L (40) in patients treated for 6, 12 and 18 months, respectively (p<0.001), whereas growth hormone increased by 12.5 mg/L (2.1), 12.5 mg/L (3.0), and 14.2 mg/L (5.7) (p<0.001). Of the patients treated for 12 months or more, 87 of 90 (97%) achieved a normal serum IGF-1 concentration. In patients withdrawn from pegvisomant (n=45), serum growth hormone concentrations were 8.0 mg/L (2.5) at baseline, rose to 15.2 mg/L (2.4) on drug, and fell back within 30 days of withdrawal to 8.3 mg/L (2.7). Antibodies to growth hormone were detected in 27 (16.9%) of patients, but no tachyphylaxis was seen. Serum insulin and glucose concentrations were significantly decreased (p<0.05). Two patients experienced progressive growth of their pituitary tumours, and two other patients had increased alanine and asparate aminotransferase concentrations requiring withdrawal from treatment. Mean pituitary tumour volume in 131 patients followed for a mean of 11.46 months (0.70) decreased by 0.033 cm(3) (0.057; p=0.353). INTERPRETATION: Pegvisomant is an effective medical treatment for acromegaly.

Effect of clean indoor air laws on smokers: the clean air module of the SimSmoke computer simulation model.

OBJECTIVES: To develop a simulation model to examine the effects of clean indoor air laws on prevalence rates and smoking attributable deaths. METHODS: Based on empirical and theoretical research, the effects of clean air laws are modelled by type of law. The model considers clean air laws at the state levels between 1993 and 2000, and projects the number of smokers and smoking attributable deaths in the USA under different scenarios from 2000 onward. RESULTS: The model predicts that comprehensive clean air laws have the potential to reduce substantially the number of smokers and smoking attributable deaths, and these effects are predicted to grow over time. The predicted impact of new worksite laws are reduced when previously implemented private and public worksite restrictions are taken into account. CONCLUSIONS: Clean indoor air laws have the ability to reduce smoking rates substantially and save lives, but their impact is likely to depend on their comprehensiveness and prior private worksite restrictions in place.

Smoking treatment interventions and policies to promote their use: a critical review.

Despite evidence regarding the effectiveness and cost-effectiveness of smoking cessation treatments, the involvement of health care workers in providing advice and the use of treatment is limited. Barriers to treatment access have been suggested as part of the reason physicians do not routinely provide brief interventions and smokers do not use proven cessation methods. The purpose of this paper is to review tobacco treatments and policy research, and highlight where more research is warranted to develop more effective policies at the private and public levels. Special attention is devoted to the recently released Guidelines (Fiore et al., Treating Tobacco Use and Dependence. Clinical Practice Guideline. Rockville, MD: USDHHS, 2000). We also consider the policy issues faced by private and public insurers, such as cost-effectiveness, the different forms that policies could take, and potential problems in implementing those policies. The efficacy of brief interventions and cessation treatments has been well studied. Less understood, however, is their impact in population-based investigations. In particular, more information is needed on the prevalence of their use and on how treatment use depends on prior smoking, treatment history, and demographic subgroup. Public and private health policies designed to increase access to cessation treatments and information dissemination through brief interventions by health care providers also have the potential to increase cessation. Further research on the impact of these interventions on use and quit rates, particularly over longer periods of time, and how these effects depend on the form of the intervention, is warranted.

A framework for evaluating and improving clean indoor air laws.

Recent attention has focused on clean indoor air laws as part of a comprehensive tobacco control strategy. Clean air laws have the potential to reduce smoking behaviors as well as second-hand smoke. Many states do not have these laws but most firms now have policies restricting smoking indoors. The authors developed a framework intended to guide in the evaluation and improvement of clean air laws. This article examines the relationship between public laws and private restrictions and how smoking restrictions are likely to affect smoking behavior throughout the population. The article also discusses the information needed to better understand clean air laws and improve future public policy.

Cancer and the potential place for growth hormone receptor antagonist therapy.

Pegvisomant is a recombinant protein, structurally similar to natural human growth hormone (GH), which is capable of binding to the GH receptor as a competitive antagonist. As well as being evaluated for the treatment of acromegaly, pegvisomant is being investigated as a possible antineoplastic agent, initially in mice. So far, in vitro efficacy against meningioma and in vivo efficacy against colon and breast cancer cell lines have been examined.

Growth hormone and insulin-like growth factor-I: effects on the growth of glioma cell lines.

Growth hormone (GH) and insulin-like growth factor-I (IGF-I) are known to be mitogens for many types of neoplasms. To investigate their role in tumors of glial origin, in vitro and in vivo experiments were performed with a panel of immortalized glioma cell lines (D54, SNB-19, U87, U251 and U373). Initial analysis for mRNA expression demonstrated the following: GH receptor (5/5 cell lines positive), IGF-I (0/5), IGF-II (0/5), IGF-I receptor (5/5), IGF-II receptor (2/5). Thus, each cell line expressed the necessary receptors to respond to GH and the IGFs but there was no autocrine IGF production by the tumors themselves. IGF-I stimulated mitogenesis as measured by [(3)H]thymidine uptake experiments in U251 and U373 cells. However, when these two IGF-responsive cell lines were xenografted into mice, tumor development and growth rates were not significantly different in GH-deficient animals (despite having IGF-I serum concentrations only 31% of normal). Because our studies were performed in immunocompromised animals, GH or IGF effects on immune surveillance, known to be important from some syngeneic glioma models, would not be likely to contribute to our findings. Nevertheless, these studies are important because they demonstrate that the growth of glioma cell lines in an in vivo environment can remain robust in a GH/IGF-I-deficient setting, even if in vitro experiments indicate that IGF-I is mitogenic.

Effect of policies directed at youth access to smoking: results from the SimSmoke computer simulation model.

OBJECTIVES: To develop a simulation model to predict the effects of youth access policies on retail compliance, smoking rates, and smoking attributable deaths. METHODS: A model of youth access policies is developed based on empirical research and a theory of perceived risk. The model incorporates substitution into other sources as retail sales are restricted, and is used to project the number of smokers and smoking related deaths. Various policies to limit youth access to cigarettes are evaluated, and we explore how efficient policies may be developed. RESULTS: The model predicts that a well designed and comprehensive policy that includes sufficient compliance checks, penalties, and community involvement has the potential to reduce the number of young smokers. Because smoking related deaths occur later in life, the effects on health are largely delayed. CONCLUSIONS: A well designed youth access policy has the ability to affect youth smoking rates in the short term, and will lead to savings in lives in future years. The ability of retail oriented policies to reduce youth smoking, however, is limited. Other tobacco control policies, including those directed at non-retail sources of cigarettes, are also needed.

Corticosteroid-induced inhibition of adrenal androgen production in selected patients with prostate cancer.

OBJECTIVE: To determine the effect of administration of corticosteroids on adrenal androgen production and the serologic markers of prostate cancer. METHODS: Six patients with prostate cancer who had a serum testosterone concentration that exceeded 20 ng/dL despite treatment with medical or surgical castration were treated with dexamethasone. All patients were asymptomatic, but four were demonstrating progressive increases in serum prostate-specific antigen (PSA) concentrations. Dexamethasone, 1 mg at bedtime, was given initially and then increased to 1 mg twice daily if serum testosterone concentrations remained > or =10 ng/dL. The effect of treatment on PSA concentration was monitored. RESULTS: The mean testosterone concentration (and standard error of the mean) was 47.5 +/- 7.9 ng/dL before administration of dexamethasone; this decreased to 5.2 +/- 3.0 ng/dL during therapy (P = 0.002). The effect was rapid (overnight) and sustainable (for 6 months). Although the duration of follow-up is limited, PSA concentrations generally stabilized (23.5 +/- 6.1 ng/mL at baseline in comparison with 15.6 +/- 1.1 ng/mL approximately 2 months after initiation of dexamethasone therapy; P = 0.24). Two patients required 1 mg of dexamethasone twice daily to suppress serum testosterone levels to <10 ng/dL. CONCLUSION: Administration of corticosteroids in a manner opposing the normal circadian glucocorticoid production effectively and rapidly decreases adrenal androgen production in patients with prostate cancer treated with orchiectomy or luteinizing hormone-releasing hormone agonists. This reduction of androgen production was generally associated with a decrease or stabilization of PSA concentrations in all patients with increased PSA levels. Overnight dexamethasone suppression testing is useful in determining the minimal effective dose.

Antitumor activity of the growth hormone receptor antagonist pegvisomant against human meningiomas in nude mice.

OBJECT: The authors have previously demonstrated that modulation of the growth hormone (GH)/insulin-like growth factor-I (IGF-I) axis can significantly affect meningioma growth in vitro. These studies were performed to evaluate the efficacy of GH receptor blockade in vivo. METHODS: Primary cultures from 15 meningioma tumors obtained in humans were xenografted into athymic mice. Approximately 1.5 million cells from each of the 15 tumors were implanted into the flanks of two female mice, one pair for each tumor. One animal from each of the 15 pairs was then treated with the GH receptor antagonist pegvisomant and the other with vehicle alone for 8 weeks. The tumor volume was measured using digital calipers three times per week. The mean tumor volume at the initiation of injections was 284 +/- 18.8 mm3 in the vehicle group and 291.1 +/- 20 mm3 in the pegvisomant group. After 8 weeks of treatment, the mean volume of tumors in the pegvisomant group was 198.3 +/- 18.9 mm3 compared with 350.1 +/- 23.5 mm3 for the vehicle group (p < 0.001). The serum IGF-I concentration in the vehicle group was 319 +/- 12.9 microg/L compared with 257 +/- 9.7 in the pegvisomant group (p < 0.02). A small but significant decrease was observed in circulating IGF binding protein (IGFBP)-3 levels, whereas slight increases occurred with respect to serum IGFBP-1 and IGFBP-4 levels. In the placebo group the tumor weight was 0.092 +/- 0.01 g compared with 0.057 +/- 0.01 g in the pegvisomant group (p < 0.02). The IGF-I and IGF-II concentrations were measured in the tumors by using a tissue extraction method. These human-specific immunoassays demonstrated that there was no autocrine production of IGF-I in any of the tumors, either in the pegvisomant or vehicle group. The IGF-I levels were highly variable (0-38.2 ng/g tissue) and did not differ significantly between treatment groups. CONCLUSIONS: In an in vivo tumor model, downregulation of the GH/IGF-I axis significantly reduces meningioma growth and, in some instances, causes tumor regression. Because the concentrations of IGF-II in tumor did not vary with pegvisomant treatment and there was no autocrine IGF-I production by the tumors, the mechanism of the antitumor effect is most likely a decrease of IGF-I in the circulation and/or surrounding host tissues. Because the authors have previously demonstrated that the GH receptor is ubiquitously expressed in meningiomas, direct blockade of the GH receptor on the tumors may also be contributing to inhibitory actions.

A computer simulation model of mass media interventions directed at tobacco use.

OBJECTIVES: The goal of this study was to develop a simulation model to examine the effects of tobacco control mass media interventions on smoking rates and smoking-attributable deaths. METHODS: The model projects the number of smokers and smoking-related deaths. Based on empirical and theoretical research, the effects of media interventions, varying in magnitude and duration, directed at all smokers and directed specifically at youth under age 18 are modeled. RESULTS: The model predicts that sustained media interventions of sufficient magnitude and duration directed at all smokers have the potential to substantially reduce the number of smokers and premature deaths, with the effects growing over time. For the same expenditures, youth interventions would appear to have smaller and more delayed effects. CONCLUSIONS: Media interventions, particularly those targeted at the general population and of sufficient scale and duration, have the ability to substantially reduce smoking rates and save lives, but their effects are likely to depend on how they are implemented.