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We develop statistical methods for tackling two important problems in genetic association studies. First, we propose a Bayesian approach to overcome the winner's curse in genetic studies. Second, we consider a Bayesian latent variable model for analyzing longitudinal family data with pleiotropic phenotypes. Winner's curse in genetic association studies refers to the estimation bias of the reported odds ratios (OR) for an associated genetic variant from the initial discovery samples. It is a consequence of the sequential procedure in which the estimated effect of an associated genetic marker must first pass a stringent significance threshold. We propose a hierarchical Bayes method in which a spike-and-slab prior is used to account for the possibility that the significant test result may be due to chance. We examine the robustness of the method using different priors corresponding to different degrees of confidence in the testing results and propose a Bayesian model averaging procedure to combine estimates produced by different models. The Bayesian estimators yield smaller variance compared to the conditional likelihood estimator and outperform the latter in the low power studies. We investigate the performance of the method with simulations and applications to four real data examples. Pleiotropy occurs when a single genetic factor influences multiple quantitative or qualitative phenotypes, and it is present in many genetic studies of complex human traits. The longitudinal family studies combine the features of longitudinal studies in individuals and cross-sectional studies in families. Therefore, they provide more information about the genetic and environmental factors associated with the trait of interest. We propose a Bayesian latent variable modeling approach to model multiple phenotypes simultaneously in order to detect the pleiotropic effect and allow for longitudinal and/or family data. An efficient MCMC algorithm is developed to obtain the posterior samples by using hierarchical centering and parameter expansion techniques. We apply spike and slab prior methods to test whether the phenotypes are significantly associated with the latent disease status. We compute Bayes factors using path sampling and discuss their application in testing the significance of factor loadings and the indirect fixed effects. We examine the performance of our methods via extensive simulations and apply them to the blood pressure data from a genetic study of type 1 diabetes (T1D) complications.

We develop statistical methods for tackling two important problems in genetic association studies. First, we propose a Bayesian approach to overcome the winner's curse in genetic studies. Second, we consider a Bayesian latent variable model for analyzing longitudinal family data with pleiotropic phenotypes. Winner's curse in genetic association studies refers to the estimation bias of the reported odds ratios (OR) for an associated genetic variant from the initial discovery samples. It is a consequence of the sequential procedure in which the estimated effect of an associated genetic marker must first pass a stringent significance threshold. We propose a hierarchical Bayes method in which a spike-and-slab prior is used to account for the possibility that the significant test result may be due to chance. We examine the robustness of the method using different priors corresponding to different degrees of confidence in the testing results and propose a Bayesian model averaging procedure to combine estimates produced by different models. The Bayesian estimators yield smaller variance compared to the conditional likelihood estimator and outperform the latter in the low power studies. We investigate the performance of the method with simulations and applications to four real data examples. Pleiotropy occurs when a single genetic factor influences multiple quantitative or qualitative phenotypes, and it is present in many genetic studies of complex human traits. The longitudinal family studies combine the features of longitudinal studies in individuals and cross-sectional studies in families. Therefore, they provide more information about the genetic and environmental factors associated with the trait of interest. We propose a Bayesian latent variable modeling approach to model multiple phenotypes simultaneously in order to detect the pleiotropic effect and allow for longitudinal and/or family data. An efficient MCMC algorithm is developed to obtain the posterior samples by using hierarchical centering and parameter expansion techniques. We apply spike and slab prior methods to test whether the phenotypes are significantly associated with the latent disease status. We compute Bayes factors using path sampling and discuss their application in testing the significance of factor loadings and the indirect fixed effects. We examine the performance of our methods via extensive simulations and apply them to the blood pressure data from a genetic study of type 1 diabetes (T1D) complications.

L’approche GWAS est un outil essentiel pour la découverte de gènes associés aux maladies, mais elle pose des problèmes de puissance statistique quand il est impossible d’échantillonner génétiquement des dizaines de milliers de sujets. Les résultats présentés ici—ALDsuite, un programme en utilisant une correction nouvelle et efficace pour le déséquilibre de liaison (DL) ancestrale de la population locale, en permettant l'utilisation de marqueurs denses dans le MALD, et la démonstration que la méthode simpleM fournit une correction optimale pour les comparaisons multiples dans le GWAS—réaffirment la valeur de l'analyse en composantes principales (APC) pour capturer l’essence de la complexité des systèmes de grande dimension. L’APC est déjà la norme pour corriger la structure de la population dans le GWAS; mes résultats indiquent qu’elle est aussi une stratégie générale pour faire face à la forte dimensionnalité des données génomiques d'association. / GWAS is an essential tool for disease gene discovery, but has severe problems of statistical power when it is impractical to genetically sample tens of thousands of subjects. The results presented here—a novel, effective correction for local ancestral population LD allowing use of dense markers in MALD using the ALDsuite and the demonstration that the simpleM method provides an optimum Bonferroni correction for multiple comparisons in GWAS, reiterate the value of PCA for capturing the essential part of the complexity of high- dimensional systems. PCA is already standard for correcting for population substructure in GWAS; my results point to it’s broader applicability as a general strategy for dealing with the high dimensionality of genomic association data.

A pigmentação humana é uma característica variável e complexa determinada por fatores genéticos e hormonais, exposição à radiação ultravioleta, idade, doenças, entre outros. Alguns polimorfismos em genes de pigmentação têm sido associados com a diversidade fenotípica de cor da pele, cabelo e olhos e em populações homogêneas. A técnica denominada Fenotipagem Forense pelo DNA (FDP) vem beneficiando a ciência forense em vários países e auxiliando investigações criminais por ser capaz de sugerir, com boa precisão, os possíveis fenótipos para as características externamente visíveis (EVCs) em amostras de origem desconhecida. No presente trabalho foram avaliadas as associações entre os SNPs presentes nos genes SLC24A5 (rs1426654; rs16960620; rs2555364), TYR (rs1126809) e ASIP (rs6058017) com cor de pele, cabelo e olhos em indivíduos da população brasileira para apontar o possível uso desses marcadores na prática forense em populações miscigenadas. Os voluntários responderam um questionário no qual fizeram a autodeclaração dessas características e estes dados foram usados para as comparações entre genótipos e fenótipos. Os resultados mostraram que para os SNPs rs2555364 e rs1426654 o alelo ancestral esteve associado com as características cor de pele negra, cabelos castanhos ou pretos e olhos castanhos. Além disso, o alelo ancestral do SNP rs6058017 foi significativamente associado com cor de pele negra e olhos castanhos. Inversamente, os alelos variantes destes SNPs são correlacionados com características de pigmentação clara para as EVCs avaliadas, corroborando os estudos prévios realizados em diferentes populações. Esses resultados mostram que a informação molecular pode ser útil para a inferência de EVCs, e a técnica de FDP é uma importante ferramenta para estudos forenses em amostra brasileira / Human pigmentation is a variable and complex trait determined by genetic and hormonal factors, exposure to ultraviolet radiation, age, diseases, among others. Some polymorphisms in pigmentation genes have been associated with the phenotypic diversity of skin, hair and eyes color in homogeneous populations. Forensic DNA Phenotyping (FDP) is benefiting forensic science in several countries, helping in criminal investigations due to its ability to suggest, with good accuracy, the possible phenotypes for externally visible characteristics (EVCs) in samples of unknown origin. Herein, we evaluated the associations between the SNPs present in the genes SLC24A5 (rs1426654; rs16960620; rs2555364), TYR (rs1126809) and ASIP (rs6058017) with skin, hair and eyes color in individuals of the Brazilian population in order to point out the possible use of these markers in forensic practice in admixed populations. The volunteers answered a questionnaire in which they self reported these characteristics for comparison between genotypes and phenotypes. The results showed that for the SNPs rs2555364 and rs1426654 the ancestral allele was associated with characteristics of black skin color, brown or black hair and brown eyes. In addition, the ancestral allele of the SNP rs6058017 was significantly associated with black skin color and brown eyes. Inversely, the variant alleles of these SNPs are correlated with fair pigmentation characteristics for the evaluated EVCs, corroborating the previous studies performed in different populations. These results show that molecular information may be useful for the inference of EVCs, and the FDP technique is an important tool for forensic studies in a Brazilian sample

Introduction: The Restless Legs Syndrome (RLS) is a frequent neurological disorder with a prevalence ranging from 5 - 10%. RLS is characterized by an urge to move the lower extremities during the night, thus RLS causes sleep disturbance. It presents as both idiopathic and secondary form. Idiopathic RLS is associated with common genetic variants in MEIS1, BTBD9, PTPRD and MAP2K5/SCOR1. Recently, multiple sclerosis (MS) was identified as a common cause for secondary RLS, the prevalence of RLS in patients with MS ranges from 13.3 to 37.5%. The aim of our study was to analyse the clinical and genetic aspects of this disorder, especially in patients with multiple sclerosis. In the clinical part, we evaluated the prevalence of RLS among Czech patients with MS and we compared the extent of brain damage between patients with and without RLS using magnetic resonance imaging (MRI). In the genetic part, we further analysed the impact of known genetic variants (MEIS1, BTBD9, MAP2K5/SCOR1, PTPRD) for RLS in other European populations and in patients with MS. Methods: Clinical part: Each patient with MS underwent a semi-structured interview. A patient was considered to be affected by RLS if he/she met all four standard criteria at life- long interval. Lesion load (LL - T2), brain atrophy - T1 and brain...

Background: In addition to patient-related systemic factors directing the immune response, the pathomechanisms of appendicitis (AP) might also include insufficient drainage leading to inflammation caused by decreased peristalsis. Genetic predisposition accounts for 30%–50% of AP. M. Hirschsprung (HSCR), also characterized by disturbed peristalsis, is associated with variants in the RET proto-oncogene. We thus hypothesized that RET variants contribute to the etiology of AP. Methods: DNA from paraffin-embedded appendices and clinical data of 264 children were analyzed for the RET c.135A>G variant (rs1800858, NC_000010.11:g.43100520A>G). In 46 patients with gangrenous or perforated AP (GAP), peripheral blood DNA was used for RET sequencing. Results: Germline mutations were found in 13% of GAP, whereas no RET mutations were found in controls besides the benign variant p.Tyr791Phe (NC_000010.11:g.43118460A>T). In GAP, the polymorphic G-allele in rs2435352 (NC_000010.11:g.43105241A>G) in intron 4 was underrepresented (p = 0.0317). Conclusion: Our results suggest an impact of the RET proto-oncogene in the etiology of AP. Mutations were similar to patients with HSCR but no clinical features of HSCR were observed. The pathological phenotypes in both populations might thus represent a multigenic etiology including RET germline mutations with phenotypic heterogeneity and incomplete penetrance.

Coronary artery disease (CAD), Hypertension (Ht) and Type 2 Diabetes Mellitus (T2DM) are all global health problems. This is particularly evident amongst South Asian population groups. The conventional risk factors do not fully explain the higher prevalence of these diseases among South Asians. The endothelial Nitric Oxide Synthase (eNOS) gene is responsible for the production of Nitric Oxide (NO), which may contribute to the physiology of all three disease states. Endothelial dysfunction (which is characterised by a reduction in basal NO) has been shown to be present in, or prior to all three diseases. Numerous variations exist within the eNOS gene, of these variations three have been shown to have a possible functional effect. The first is the Glu298Asp polymorphism within the exon region of the gene, resulting in an amino acid substitution of Glutamate (Glu) to Aspartate (Asp). The second, known as the T-786C polymorphism, is a thymine to cytosine mutation at position -786 in the promoter region. Finally a VNTR polymorphism in Intron 4 causes either a 4 27bp repeat or a 5 27bp repeat. It is hypothesised that these variations could have an effect on the ability of eNOS to produce NO and thus may increase the risk or contribute to the development of the diseases. Previous studies on these variants have shown conflicting results and further studies are warranted to understand and confirm the role of eNOS gene polymorphisms in cardio-metabolic diseases. There is very limited research into the distributions of these genetic variants and their interaction in diseases processes in North Indian populations. Objectives: 1. To analyse through a case control study three different polymorphisms of the eNOS gene for possible association with Coronary Artery Disease (CAD), Hypertension (Ht) and Type 2 Diabetes Mellitus (T2DM) in North Indian population groups. 2. To statistically evaluate descriptive statistics including; age, gender, smoking, dietary behaviours and lipid parameters for possible influence on disease and potential interaction with genetic polymorphisms. 3. To evaluate linkage disequilibrium between the three eNOS variants and carryout haplotype analysis to work out haplotype risk in different diseases. 4. To analyse through a case control study the deletion variant of the Angiotensin-converting enzyme (ACE) gene for possible association with Coronary Artery Disease (CAD), Hypertension (Ht) and Type 2 Diabetes Mellitus (T2DM) in North Indian population groups. 5. To determine a possible interactive effect of the eNOS polymorphisms with the ACE polymorphism. Subjects and Methods: The Glu298Asp and Intron 4 variants were genotyped using a PCR-RFLP technique, the T-786C variant was genotyped using a real time-PCR technique. The ACE deletion variant was also genotyped using a standard PCR technique. The genotyping was undertaken in a total of 457 CAD patients and 220 matched controls from Lucknow, Uttar Pradesh in North India, 319 T2DM patients and 307 matched controls from Punjab, North India and 210 Ht and 162 matched controls, also from Punjab, North India. Results: CAD: The Glu298Asp was significantly associated with CAD among smokers (TT+GT vs. GG OR=2.84 (CI: 1.61-5.0), p<0.001). The Intron 4 variant was also significantly associated with CAD in a smoking dependent manner (4aa+4ab vs. 4bb OR=0.56 (CI: 0.33-0.96). The T-786C variant showed no overall influence on CAD risk. There was also evidence for both synergistic and haplotypic effects of the eNOS gene on CAD status (haplotype G-C-4b OR=4.76 (CI: 1.43-15.78), p<0.001). The ACE genetic variant was confirmed to be a strong independent risk factor for CAD under a dominant model (OR=2.18 (CI: 1.46-3.25), p<0.001). There was no evidence for an interactive effect between the ACE deletion and any of the three eNOS variants incorporated in the current study. Ht: The Glu298Asp variant was not shown to increase Ht risk, with a reduced risk association found under a recessive model (OR=0.316 (CI:0.089-1.116)), p=0.061). The T-786C variant s role in disease remained unclear with the findings showing a non significant increased risk. The Intron 4 variant was also shown to increase Ht risk, in a non significant manner. Sufficiently powered studies would be required to clarify these possible associations. The combined analysis, using logistic regression and haplotype analysis revealed no significant associations, but there was a possible protective effect of the T-C-4b haplotype (OR=0.46 (CI: 0.21-1.01), p=0.054). The ACE gene variant was confirmed to be a strong independent risk factor for Ht under a recessive model (OR=1.81 (CI: 1.20-2.74), p=0.01). Again there was no evidence for an interactive effect between the ACE deletion and any of the three eNOS variants in hypertension. T2DM: The Glu298Asp variant was found to be associated with T2DM under a dominant model, the protective effect remained significant following adjustment for conventional risk factors and other gene variants (OR=0.407 (CI: 0.231-0.717), p=0.002). The T-786C variant showed no overall influence on T2DM risk. The Intron 4 variant also found no overall influence. Haplotype analysis found the T-T-4b was found to be significantly protective for T2DM (OR=0.41 (CI: 0.26-0.65), p=0.0002). Finally the ACE gene variant was confirmed to be a risk factor for T2DM under a dominant model (OR=2.62 (CI: 1.51-4.54), p=0.001). Overall Conclusions: To conclude, this study successfully identified the frequency of three eNOS gene variants and the ACE deletion variant in three complex diseases within north Indian populations. There is a clear role of the eNOS gene in all three diseases and consequently the genetic variants have susceptible/protective associations. The association with disease was found to be present at an individual level, in association with risk factors and at a haplotypic level. These findings warrant further studies to confirm and untangle the genetics of complex diseases and genetic risk profiles calculations which will contribute to the field of medical genomics/personalised medicare and interventions among North Indian populations.

Erythrocytes infected by mature forms of the Plasmodium falciparum parasite adhere to other components of the vascular space, a behavior considered critical to the pathogenesis of severe malaria. Adhesion is mediated by the P. falciparum erythrocyte membrane protein 1 (PfEMP1), a highly variant antigen expressed by the parasite and subject to switching during the course of an infection. The host ligands of PfEMP1 include CD36, ICAM-1 and the ABO antigens. By employing a series of population- and family-based association studies from multiple African populations, we examined whether variation in the genes underlying these molecules affects susceptibility to severe malaria. Our results suggest that a common frameshift mutation in the ABO glycosyltransferase, responsible for blood group O, is associated with protection from severe malarial phenotypes (P=2x10⁻⁷), particularly severe malarial anaemia. However, we found no significant disease associations with variation in either the ICAM1 or CD36 genes. We focused on two particular functional polymorphisms, the missense ICAM-1Kilifi and the CD36 nonsense mutation T1264G. We genotyped both markers in around 10,000 individuals, but neither demonstrated an association with severe malarial phenotypes. Malaria has been a profound selection pressure shaping human genetic diversity. The last decade has seen the development of several haplotype-based methods to detect signatures of recent positive evolutionary selection. These techniques are potentially invaluable tools in our hunt for genetic variants that protect from life threatening malaria. We used simulations and empirical data from the International HapMap Project to demonstrate the validity of searching for long regions of haplotype homozygosity, as an approach to finding alleles undergoing selective sweeps. We analysed genetic data from a range of populations, particularly those utilized by HapMap, to investigate whether our candidate genes were associated with signals of recent positive selection. We characterized the distribution of a selection event associated with the CD36 1264G allele, focused in Central-West Africa, and demonstrated a novel signal of low population differentiation at the ABO gene, suggestive of longstanding balancing selection. Our work confirms that variation in the host ligands of PfEMP1 modulates severe malaria susceptibility, and highlights the value of using signals of selection, along with functional experiments and genetic association studies, to dissect the biology of severe malaria.

INTRODUÇÃO: A persistência de um microambiente cronicamente inflamado no estômago tem sido descrita como um componente crítico tanto para a iniciação, quanto para a progressão tumoral. Além disso, variações genéticas tem demonstrado influenciar na variabilidade interindividual da resposta inflamatória. Desta forma, esse estudo teve como objetivo investigar a associação de polimorfismos em genes relacionados à resposta inflamatória com o risco para o desenvolvimento do câncer gástrico, com suas variáveis anatomopatológicas e com a sobrevida global e livre de doença em uma amostra da população Brasileira. MÉTODOS: Dezesseis variantes genéticas selecionadas em onze genes (COX-2, OGG1, TNFB, TNFA, HSPA1L, HSPA1B, VEGFA, IL17F, LGALS3, PHB e TP53) foram genotipadas em 262 indivíduos controles e 178 pacientes diagnosticados com câncer gástrico. As análises de associações genéticas foram realizadas em diferentes modelos (Genótipos, Alelos, Dominante e Recessivo) considerando a amostra total de casos (N=178) e estratificada somente para os casos com o subtipo histológico difuso de Lauren (N=112). Também foi investigado o desequilíbrio de ligação entre os polimorfismos e as análises de associação com os haplótipos formados foram realizadas por meio dos softwares Haploview e PLINK. RESULTADOS: No estudo caso-controle, indivíduos portadores do alelo Pro do polimorfismo rs1042522 (TP53) apresentaram risco cerca de duas vezes maior em desenvolver o câncer gástrico em análise multivariada, sendo esse risco ainda maior quando considerado somente os casos com o subtipo difuso. Por outro lado, a presença do alelo A do polimorfismo rs699947 (VEGFA) foi associada com uma proteção para o câncer gástrico. Em relação às variáveis anatomopatológicas, os polimorfismos rs689466 (COX-2); rs1052133 (OGG1); rs699947, rs833061 e rs2010963 (VEGFA); rs4644 (LGALS3) e rs1042522 (TP53) foram significativamente associados com características de pior progressão da doença, enquanto que rs5275 (COX-2); rs2227956 (HSPA1L) e rs3025039 (VEGFA) foram associados às variáveis de melhor progressão da doença, na amostra total de casos. Também foi observado que o polimorfismo rs909253 (TNFB) foi capaz de predizer uma melhor progressão da doença quando considerado somente os casos com subtipo difuso. Além disso, em relação ao impacto sobre a sobrevida, rs909253 (TNFB) foi associado a um melhor prognóstico quando analisadas ambas as curvas de sobrevida global e livre de doença, enquanto que portadores do alelo His do polimorfismo rs4644 (LGALS3) apresentaram um pior prognóstico com menor tempo de sobrevida livre de doença. Por fim, nas análises de associação com os haplótipos, identificamos que o haplótipo CTC (formado por rs699947, rs833061 e rs2010963 do gene VEGFA), demonstrou ser um fator de maior suscetibilidade ao câncer gástrico. Também foram observadas associações entre o haplótipo GG (TNFB/TNFA) e invasão perineural; haplótipo ACG (VEGFA) e invasão sanguínea; haplótipo CTC (VEGFA) e invasão para outros órgãos; haplótipo GT (rs689466 e rs5275 do gene COX-2) e subtipo histológico intestinal. CONCLUSÕES: Os resultados desse estudo nos ajudaram a esclarecer o potencial papel desses polimorfismos em genes envolvidos com a modulação da resposta inflamatória na patogênese do câncer gástrico, indicando que variantes genéticas do hospedeiro atuam conjuntamente com outros fatores, influenciando na suscetibilidade, progressão e prognóstico dessa doença / INTRODUCTION: The chronic inflammatory microenvironment in the stomach has been described as a critical component for both tumor initiation and progression. Furthermore, genetic variants have shown to influence the interindividual variations in the inflammatory response. Therefore, we aimed to investigate whether polymorphisms in inflammatory response related-genes were associated with risk for gastric tumor development, clinical outcomes, overall and disease free survival of this disease in a Brazilian population sample. METHODS: Sixteen selected genetic variants in eleven genes (COX-2, OGG1, TNFB, TNFA, HSPA1L, HSPA1B, VEGFA, IL17F, LGALS3, PHB and TP53) were genotyped in 262 control individuals and 178 gastric cancer patients. Genetic association analyses were investigated in different models (Genotype, Allele, Dominant and Recessive) in both total sample (N=178) and stratified for the diffuse histological subtype based on Lauren´s classification (N=112). We also calculated the linkage disequilibrium among the polymorphisms and the haplotype associations were carried out using Haploview and PLINK softwares. RESULTS: In the case-control study, rs1042522 (TP53) Pro allele carriers presented about 2-fold higher risk for developing gastric cancer in a multivariate analysis and this association was even stronger when analyzing only cases with the diffuse subtype. On the other hand, the presence of A allele of rs699947 (VEGFA) was associated with a protection for developing gastric cancer. About the significant associations detected with the clinicopathological features, we found that rs689466 (COX-2); rs1052133 (OGG1); rs699947, rs833061 and rs2010963 (VEGFA); rs4644 (LGALS3) and rs1042522 (TP53) were able to predict outcomes associated with a worse progression of the disease while rs5275 (COX-2); rs2227956 (HSPA1L) and rs3025039 (VEGFA) were associated with better outcomes in the total sample. We also observed that the polymorphism rs909253 (TNFB) was able to predict a better outcome only for the individuals diagnosed with the diffuse subtype. Additionally, regarding the impact on the survival curves, rs909253 (TNFB) was associated with a better prognosis when analyzing both the overall and disease-free survivals while rs4644 (LGALS3) His allele carriers presented a worse prognosis with shorter disease-free survival. Finally, concerning the haplotype associations, we found that CTC haplotype (composed by rs699947, rs833061 and rs2010963 of VEGFA) showed an association with gastric malignancy. We also observed associations between GG haplotype (TNFB/TNFA) and perineural invasion; ACG haplotype (VEGFA) and venous vascular invasion; CTC haplotype (VEGFA) and invasion to other organs and GT haplotype (rs689466 and rs5275 of COX-2 gene) and the intestinal histologic subtype. CONCLUSIONS: These results helped us to clarify the potential role of these polymorphisms in genes involved in the modulation of the inflammatory response in the pathogenesis of gastric malignancy, highlighting that the host genetic variants act together with other factors to influence in the susceptibility, progression and prognosis of gastric cancer