Common Polymorphic Variants Research Articles

Pharmacogenetic characterization of indacaterol, a novel β2‐adrenoceptor agonist

Indacaterol is a novel beta(2)-adrenoceptor agonist in development for the treatment of chronic obstructive pulmonary disease. The aim of this study was to investigate the comparative pharmacology of indacaterol in recombinant cells expressing the common polymorphic variants of the human beta(2)-adrenoceptor and in human primary airway smooth muscle (ASM) cells. Chinese hamster ovarian-K1 cell lines expressing high and low levels of the common human beta(2)-adrenoceptor variants were generated [Gly16-Glu27-Val34-Thr164(GEVT), RQVT, GQVT] and also the rare GQVI variant. Human primary ASM cells were isolated from explants of trachealis muscle. Adenosine-3',5'-cyclic-monophosphate production was used as an outcome measure. In both the low- and high-expression recombinant GEVT 'wild type' cell lines indacaterol is a high-efficacy agonist. Salmeterol and formoterol were identified as low- and high-efficacy agonists, respectively, and showed similar potencies to indacaterol irrespective of the beta(2)-adrenoceptor genotype. The I164 variant cell line was associated with a reduced capacity to generate adenosine-3',5'-cyclic-monophosphate in response to beta(2)-adrenoceptor agonist. In the human primary ASM cells indacaterol gave a maximal response intermediate between that of salmeterol and formoterol. These data demonstrate that indacaterol is a high-efficacy agonist in recombinant cell systems but acts with lower efficacy in human primary ASM cells. No marked genotype-dependent effects were observed for common variants; however, changes in I164 receptor activity were identified, which were dependent on the level of expression of beta(2)-adrenoceptors.

Promoter and intron 1 polymorphisms of COL1A1 interact to regulate transcription and susceptibility to osteoporosis

Three polymorphisms (-1997G/T; -1663IndelT and +1245G/T) have been identified in the 5' flank of COL1A1 gene that are associated with osteoporosis but the underlying mechanism is unclear. Here we investigated the functional effects of these variants on COL1A1 transcription. Transcription was 2-fold higher with the osteoporosis-associated G-del-T haplotype compared with the common G-Ins-G haplotype. Gel shift assays showed that the region surrounding the -1663IndelT polymorphism recognized a complex of proteins essential for osteoblast differentiation and function including Nmp4 and Osterix, and the osteoporosis-associated -1663delT allele had increased binding affinity for this complex. Chromatin immunoprecipitation assays confirmed that the region flanking -1663insdelT bound a complex of proteins including Osterix and Nmp4 and also showed evidence of recruitment of Nmp4 to the Sp1 binding site in intron 1. Further studies showed that haplotype G-del-T had higher binding affinity for RNA polymerase II, consistent with increased transcription of the G-del-T allele and there was a significant inverse association between carriage of G-del-T and bone mineral density (BMD) in a cohort of 3270 Caucasian women. We conclude that common polymorphic variants in the 5' flank of COLIA1 regulate transcription by affecting DNA-protein interactions and that increased levels of transcription correlate with reduced BMD values in vivo. This is consistent with a model whereby increased COL1A1 transcription predisposes to osteoporosis, probably by increasing production of the alpha 1 chain and disrupting the normal ratio of collagen type 1 alpha 1 and alpha 2 chains.

Genomics and breast cancer: the different levels of inherited susceptibility

Our understanding of inherited breast cancer susceptibility has changed dramatically over the last 5 years, with the discovery of many genes in which mutations influence the risk of developing breast cancer. These fall into three main groups: genes in which mutations confer a high risk of developing cancer, but where such mutations are rare; uncommon mutations in genes conferring a moderate increase in risk (odds ratios of approximately 2–4), and common polymorphic variants which each confer only slight risk alterations (odds ratios rarely above 1.2 for variants conferring increased risk). The discovery of these genes and loci was predicated on linkage analysis (for high-risk genes), screening for mutations in candidate genes selected because they were involved in functional pathways related to BRCA1 and BRCA2 function (moderate risk), and genome-wide association studies (low penetrance polymorphisms). However, with this increased knowledge comes the difficulty of knowing how this information may best be utilized in clinical practice. The review by Ripperger et al1 in this edition of the Journal examines the current knowledge of newly identified moderate and low risk susceptibility genes, classifying the genes involved in inherited breast cancer susceptibility into the categories described above. The tables and diagrams are helpful in indicating the relative frequency and penetrance for breast cancer of genetic variants in these different categories, and give some indication of the proportion of the disease burden associated with mutations in each gene. The discovery of lower penetrance genes2 indicates that inherited breast cancer susceptibility is due to a number of genetic factors, and it may well be that the familial clusters of breast cancer not due to mutations in BRCA1 and BRCA2, originally attributed to BRCA3, are in fact due to a combination of the effects of lower penetrance gene mutations and environmental factors. Healthcare providers will have the dilemma of deciding whether it will be cost-effective to introduce population screening of variants of these low penetrance genes and what level of risk would justify medical intervention. Inheritance of one low penetrance risk-conferring polymorphic allele may only confer a very minor increase in risk of developing cancer, but individuals who have inherited several high-risk polymorphic variants may be at significantly increased risk. It has recently been argued, for instance,3 that individuals who have, say, six ‘high'-risk alleles, may have a sufficiently increased risk to justify initiating mammographic surveillance 10 years before the generally recommended age for women in the general population, and women with six low risk alleles could start screening at a later age.

Effect of Vitamin B12 and Folate on Homocysteine levels in colorectal cancer

Folate and cobalamin (Vitamin B(12)) are two essential micronutrients involved in one-carbon metabolism, which affects heart disease, neural tube defects and cancer. Methylenetetrahydrofolate reductase, the key enzyme involved in one carbon metabolism produces methyl tetrahydrofolate from methylene tetrahydrofolate, which in turn donates methyl group to homocysteine to generate methionine. There exist two common low function polymorphic variants of the methylenetetrahydrofolate reductase gene involving nucleotides 677 C→T and 1298 A→C, which are associated with hyperhomocysteinemia. These polymorphisms are also linked with increased risk for certain cancers such as breast cancer and at the same time providing a protective effect on colorectal cancer. In this case control study, we have evaluated levels of folic acid, vitamin B(12) and homocysteine in patients with colorectal cancer. Folate and homocysteine levels did not differ significantly between the two groups; however an increasing trend was noticed with increase in homocysteine levels. Vitamin B(12) levels were increased in cases compared to control group.

Multivariate permutation analysis associates multiple polymorphisms with subphenotypes of major depression.

Unipolar major depressive disorder (MDD) is a prevalent, disabling condition with multiple genetic and environmental factors impacting disease risk. The diagnosis of MDD relies on a cumulative measure derived from multiple trait dimensions and alone is limited in elucidating MDD genetic determinants. We and others have proposed that MDD may be better dissected using paradigms that assess how specific genes associate with component features of MDD. This within-disease design requires both a well-phenotyped cohort and a robust statistical approach that retains power with multiple tests of genetic association. In the present study, common polymorphic variants of genes related to central monoaminergic and cholinergic pathways that previous studies align with functional change in vitro or depression associations in vivo were genotyped in 110 individuals with unipolar MDD. Subphenotypic characteristics were examined using responses to individual items assessed with the Structured Clinical Interview for Diagnostic and Statistical Manual of Mental Disorders (DSM IV), the 17-item Hamilton Rating Scale for Depression (HAM-D) and the NEO Five Factor Inventory. Multivariate Permutation Testing (MPT) was used to infer genotype-phenotype relationships underlying dimensional findings within clinical categories. MPT analyses show significant associations of the norepinephrine transporter (NET, SLC6A2) -182 T/C (rs2242446) with recurrent depression [odds ratio, OR = 4.15 (1.91-9.02)], NET -3081 A/T (rs28386840) with increase in appetite [OR = 3.58 (1.53-8.39)] and the presynaptic choline transporter (CHT, SLC5A7) Ile89Val (rs1013940) with HAM-D-17 total score {i.e. overall depression severity [OR = 2.74 (1.05-7.18)]}. These relationships illustrate an approach to the elucidation of gene influences on trait components of MDD and with replication, may help identify MDD subpopulations that can benefit from more targeted pharmacotherapy.

Association between the BRCA2N372H variant and male breast cancer risk: a population-based case-control study in Tuscany, Central Italy

BackgroundMale breast cancer (MBC) is a rare disease and little is known about its aetiology. Germ-line mutations of BRCA2 and, at lower frequency, of BRCA1 are implicated in a relatively small proportion of MBC cases. Common polymorphic variants in BRCA1 and BRCA2 genes may represent breast cancer (BC) susceptibility alleles and could be associated with a modestly increased risk of MBC at population level. Considering the relevant role of BRCA2 in MBC, we investigated whether the BRCA2 N372H variant, representing the only common non-synonymous polymorphism in BRCA2, might modulate the risk of BC in male populations.MethodsA case-control study was performed comparing a population-based series of 99 MBC cases, characterized for BRCA1 and BRCA2 mutations, with 261 male population controls, all residing in Tuscany, Central Italy. All MBC cases and controls were genotyped for the BRCA2 N372H allele by TaqMan allelic discrimination assays. To evaluate the genotype specific risk of the BRCA2 N372H variant, MBC carriers of germ-line BRCA1/2 mutations were excluded from the analyses.ResultsNo association emerged in univariate and age-adjusted analyses. Age-specific analyses suggested an increased risk for the HH homozygous genotype in subjects younger than 60 years. A statistically significant interaction emerged between this genotype and age (p = 0.032). When analyses were restricted to MBC cases enrolled in the first 4 years following diagnosis, a recessive model showed a significantly increased risk of MBC in HH subjects younger than 60 years (OR = 5.63; 95% CI = 1.70;18.61).ConclusionOverall, our findings, although based on a relatively small series, suggest that the BRCA2 HH homozygous genotype might be positively associated with an increased risk of MBC in men younger than 60 years.

Common variants in the ATM, BRCA1, BRCA2, CHEK2 and TP53 cancer susceptibility genes are unlikely to increase breast cancer risk

IntroductionCertain rare, familial mutations in the ATM, BRCA1, BRCA2, CHEK2 or TP53 genes increase susceptibility to breast cancer but it has not, until now, been clear whether common polymorphic variants in the same genes also increase risk.MethodsWe have attempted a comprehensive, single nucleotide polymorphism (SNP)- and haplotype-tagging association study on each of these five genes in up to 4,474 breast cancer cases from the British, East Anglian SEARCH study and 4,560 controls from the EPIC-Norfolk study, using a two-stage study design. Nine tag SNPs were genotyped in ATM, together with five in BRCA1, sixteen in BRCA2, ten in CHEK2 and five in TP53, with the aim of tagging all other known, common variants. SNPs generating the common amino acid substitutions were specifically forced into the tagging set for each gene.ResultsNo significant breast cancer associations were detected with any individual or combination of tag SNPs.ConclusionIt is unlikely that there are any other common variants in these genes conferring measurably increased risks of breast cancer in our study population.

The Genetics of Low-Density Lipoprotein Receptor-Related Protein 5 in Bone: A Story of Extremes

A few years ago, human genetic studies provided compelling evidence that the low-density lipoprotein receptor-related protein 5 (LRP5) is involved in the regulation of bone homeostasis because pathogenic LRP5 mutations were found in monogenic conditions with abnormal bone density. On the one hand, the osteoporosis pseudoglioma syndrome results from loss of function of LRP5, whereas on the other hand, gain-of-function mutations in LRP5 cause conditions with an increased bone density. On the molecular level, these types of mutations result in disturbed (respectively, decreased and increased) canonical Wnt signaling, an important metabolic pathway in osteoblasts during embryonic and postnatal osteogenesis. This signaling cascade is activated by binding of Wnt ligand to the Frizzled/LRP5 receptor complex. In addition to the involvement of LRP5 in conditions with extreme bone phenotypes, the genetic profile of this gene has also been shown to contribute to the determination of bone density in the general population. Quite a number of studies already demonstrated that common polymorphic variants in LRP5 are associated with bone mineral density and consequently osteoporosis, a multifactorial trait with low bone mass and porous bone structure. These genetic studies together with results obtained from in vitro and in vivo studies emphasize the importance of LRP5 and canonical Wnt signaling in the regulation of bone homeostasis. Therefore, unraveling the exact mechanisms of this signaling cascade has become an important area in bone research. This review focuses on the genetics of LRP5 and summarizes the findings on monogenic bone conditions as well as the current knowledge of its involvement in the pathogenesis of osteoporosis.

Large-Scale Population-Based Study Shows No Association Between Common Polymorphisms of the TGFB1 Gene and BMD in Women

The TGFB1 gene is a strong functional candidate for regulating genetic susceptibility to osteoporosis. We studied five common polymorphisms of TGFB1 in relation to osteoporosis-related phenotypes in a population-based cohort of 2975 British women, but found no significant association with bone mass, bone loss, bone markers, or fracture. The gene encoding TGFB1 is a strong functional candidate for genetic susceptibility to osteoporosis. Several polymorphisms have been identified in TGFB1, and previous work has suggested that allelic variants of TGFB1 may regulate BMD and susceptibility to osteoporotic fracture. We studied the relationship between common polymorphisms of TGFB1 and several osteoporosis-related phenotypes including BMD at the lumbar spine and femoral neck, measured by DXA; bone loss over a 6-year period; biochemical markers of bone turnover (urinary free deoxypyridinoline and free pyridinoline/creatinine ratio and serum N-terminal propeptide of type 1 collagen), and fractures in a population-based study of 2975 women from the United Kingdom. Participants were genotyped for single nucleotide polymorphisms (SNPs) in the TGFB1 promoter (G-800A; rs1800468; C-509T; rs1800469), exon 1 (T29C; rs1982073 and G74C; rs1982073); and exon 5 (C788T; rs1800471) on PCR-generated fragments of genomic DNA. Haplotypes were constructed from genotype data using the PHASE software program, and genotypes and haplotypes were related to the phenotypes of interest using general linear model ANOVA, with correction for confounding factors including age, height, weight, menopausal status, hormone replacement therapy (HRT) use, physical activity score, and dietary calcium intake. The polymorphisms were in strong linkage disequilibrium, and four common haplotypes accounted for >95% of alleles at the locus. There was no association between individual SNPs and BMD, bone loss, or biochemical markers of bone turnover. Haplotype analysis showed a nominally significant association with femoral neck BMD (p = 0.042) and with incident osteoporotic fracture (p = 0.013), but these were not significant after correcting for multiple testing. Common polymorphic variants of the TGFB1 gene did not influence BMD or bone loss in this population.

Streamlined assessment of gene variants by high resolution melt profiling utilizing the ornithine transcarbamylase gene as a model system

Ornithine transcarbamylase (OTC) deficiency is an X-linked, semidominant genetic disorder and the most prevalent inherited defect of the urea cycle. Molecular genetic testing of the OTC gene is critically important for clinical diagnosis, carrier testing, and prenatal diagnosis. Private mutations are observed throughout the OTC gene with more than 340 reported disease-causing mutations. High-resolution melt profiling was adapted to perform homogeneous analysis of the 10 coding regions and their intronic flanks in a 96-well plate format. The 10 DNA fragments ranging from 146 bp to 266 bp are amplified in a PCR run. A common analysis condition simultaneously generates melting profiles from all 10 fragments. To streamline analysis, deviant profiles resulting from common polymorphic variants are triaged using redundant assessment with melt profile controls in selected whole-exon assays and a separate multiplex genotyping assay. The test is further streamlined by recovering dye-stained amplification product from the melt profiling plate to serve as DNA sequencing template. Described herein is the comprehensive analysis of the OTC gene in 23 OTC-deficient patients. This system provides a rapid means to localize sequence variants, markedly reducing the need for DNA sequencing, and is applicable to other genes and disorders.

Metabolic endophenotype and related genotypes are associated with oxidative stress in children with autism

Autism is a behaviorally defined neurodevelopmental disorder usually diagnosed in early childhood that is characterized by impairment in reciprocal communication and speech, repetitive behaviors, and social withdrawal. Although both genetic and environmental factors are thought to be involved, none have been reproducibly identified. The metabolic phenotype of an individual reflects the influence of endogenous and exogenous factors on genotype. As such, it provides a window through which the interactive impact of genes and environment may be viewed and relevant susceptibility factors identified. Although abnormal methionine metabolism has been associated with other neurologic disorders, these pathways and related polymorphisms have not been evaluated in autistic children. Plasma levels of metabolites in methionine transmethylation and transsulfuration pathways were measured in 80 autistic and 73 control children. In addition, common polymorphic variants known to modulate these metabolic pathways were evaluated in 360 autistic children and 205 controls. The metabolic results indicated that plasma methionine and the ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH), an indicator of methylation capacity, were significantly decreased in the autistic children relative to age-matched controls. In addition, plasma levels of cysteine, glutathione, and the ratio of reduced to oxidized glutathione, an indication of antioxidant capacity and redox homeostasis, were significantly decreased. Differences in allele frequency and/or significant gene-gene interactions were found for relevant genes encoding the reduced folate carrier (RFC 80G > A), transcobalamin II (TCN2 776G > C), catechol-O-methyltransferase (COMT 472G > A), methylenetetrahydrofolate reductase (MTHFR 677C > T and 1298A > C), and glutathione-S-transferase (GST M1). We propose that an increased vulnerability to oxidative stress (endogenous or environmental) may contribute to the development and clinical manifestations of autism.

Breast Cancer Risk is not Associated with Polymorphic Forms of Xeroderma Pigmentosum Genes in a Cohort of Women from Washington County, Maryland

The genes mutated in the cancer-prone syndrome, xeroderma pigmentosum (XP genes), have been well studied both biochemically and mechanistically. These genes are important components of the DNA nucleotide excision repair (NER) pathway, which protects against environmentally-induced cancers. XP genes are also downstream of the hereditary breast cancer syndrome gene, BRCA1, suggesting that XP genes may be important to hereditary forms of breast cancer as well. Although mutated XP genes are rare, polymorphic forms with potential functional deficiencies are common, and could pose a significant cancer risk in the general population. This study tested the hypothesis that common polymorphic variants of XP genes were associated with the risk of breast cancer among a population of women in Washington County, Maryland. Five single nucleotide polymorphisms (SNPs) among four XP genes (XPC, XPD, XPF and XPG) were genotyped from DNA samples collected at baseline, and then analyzed by conditional logistic regression for association with the incidence of breast cancer. 321 cases were individually matched to 321 controls, by age and menopausal status. No significant associations were found between breast cancer risk and any of the XP genotypes. Odds ratios for all genotypes ranged from 0.61 to 1.14, and none were statistically significant. Adjustment and stratification for family history of breast cancer did not alter the findings. These results suggest that polymorphisms of XP genes are not likely to be significant risk factors for women within the general population. This study did not address, however, risks for subpopulations of women with high exposures to DNA damaging agents.

Iron and infection: effects of host iron status and the iron-regulatory genes haptoglobin and NRAMP1 (SLC11A1) on host–pathogen interactions in tuberculosis and HIV

There are many lines of evidence illustrating that iron plays a pivotal role in modulating the battle for survival between mammalian hosts and their pathogens. Each displays considerable genetic investment in a wide range of mechanisms for acquiring and maintaining iron. These competitive mechanisms are highly complex, existing within an interacting matrix of absorption, transport, storage and detoxification systems, each of which are iron-responsive and thus able to adapt to the different phases of infection. Considerable genetic polymorphism in some of these systems, with signals of geographic selection in the hosts, and niche selection in the pathogens, indicates that they are critical for species survival. In this review we briefly summarize the role of iron in host immune function before reviewing the available evidence that iron modulates susceptibility and disease outcomes in HIV and TB (tuberculosis). We then examine the putative role of iron-related host genes by focussing on two candidate genes, haptoglobin and NRAMP1, for which there are common polymorphic variants in humans with strong evidence of functionally distinct biochemical phenotypes that would be predicted to influence the course of HIV and TB infections. Finally, we examine the limited evidence so far available that nutrient-gene interactions are likely to influence the way in which gene variants can protect against infection. We conclude that there is a wealth of evidence associating alterations in iron balance and in iron-regulatory systems with disease progression, but that many issues related to the direction of causality, mechanisms of action and sensitivity to pharmacological intervention remain to be elucidated. Since iron is probably the most widely prescribed compound throughout the world, used in both preventative and treatment regimens, a deeper understanding of the host-pathogen interactions relating to iron constitutes an important area for both basic and clinical research.

Fetal Programming of Polycystic Ovary Syndrome by Androgen Excess: Evidence from Experimental, Clinical, and Genetic Association Studies

Polycystic ovary syndrome (PCOS) is a common endocrine disorder of premenopausal women, characterized by hyperandrogenism, polycystic ovaries, and chronic anovulation along with insulin resistance and abdominal obesity as frequent metabolic traits. Although PCOS manifests clinically during adolescence, emerging data suggest that the natural history of PCOS may originate in intrauterine life. Evidence from experimental, clinical, and genetic research supporting the hypothesis for the fetal origins of PCOS has been analyzed. Female primates, exposed in utero to androgen excess, exhibit the phenotypic features of PCOS during adult life. Clinical observations also support a potential fetal origin of PCOS. Women with fetal androgen excess disorders, including congenital 21-hydroxylase deficiency and congenital adrenal virilizing tumors, develop features characteristic of PCOS during adulthood despite the normalization of androgen excess after birth. The potential mechanisms of fetal androgen excess leading to a PCOS phenotype in humans are not clearly understood. However, maternal and/or fetal hyperandrogenism can provide a plausible mechanism for fetal programing of PCOS, and this, in part, may be genetically determined. Thus, genetic association studies have indicated that common polymorphic variants of genes determining androgen activity or genes that influence the availability of androgens to target tissues are associated with PCOS and increased androgen levels. These genomic variants may provide the genetic link to prenatal androgenization in human PCOS. Prenatal androgenization of the female fetus induced by genetic and environmental factors, or the interaction of both, may program differentiating target tissues toward the development of PCOS phenotype in adult life.

Coumarins are competitive inhibitors of cytochrome P450 1B1, with equal potency for allelic variants

Coumarins are naturally occurring chemicals with potential as chemopreventive agents, several with known action on the cytochrome P450 1A family. We examined whether cytochrome P450 1B1 (CYP1B1) was inhibited by coumarins, whether such inhibition was competitive, and whether inhibition varied between common polymorphic variants of this enzyme. We tested the inhibition properties of four coumarins, bergamottin, isopimpinellin, isoimperatorin, and imperatorin in an assay for oxidation of (-)benzo[a]pyrene-7R-trans-7,8-dihyrodiol (B[a]P-7,8-diol) by CYP1B1 using yeast-microsome expressed enzymes. These assays were performed with wild-type enzyme and five single-amino acid polymorphic variants. All four coumarins are competitive inhibitors of CYP1B1, with Ki values equal to 587, 11, 6 and 1 muM respectively. Inhibition parameters were consistent between five haplotypes of CYP1B1, three representing common haplotypes in Asians, African-Americans and European-Americans, and two with baseline kinetic parameters previously shown to be potentially different from wild-type. Coumarins are capable of inhibiting carcinogen activation by CYP1B1 with varying potencies, and their efficacy as chemopreventive agents is not likely to be affected by polymorphism in this enzyme.

Human ATP:Cob(I)alamin Adenosyltransferase and Its Interaction with Methionine Synthase Reductase

The final step in the conversion of vitamin B(12) into coenzyme B(12) (adenosylcobalamin, AdoCbl) is catalyzed by ATP:cob(I)alamin adenosyltransferase (ATR). Prior studies identified the human ATR and showed that defects in its encoding gene underlie cblB methylmalonic aciduria. Here two common polymorphic variants of the ATR that are found in normal individuals are expressed in Escherichia coli, purified, and partially characterized. The specific activities of ATR variants 239K and 239M were 220 and 190 nmol min(-1) mg(-1), and their K(m) values were 6.3 and 6.9 mum for ATP and 1.2 and 1.6 mum for cob(I)alamin, respectively. These values are similar to those obtained for previously studied bacterial ATRs indicating that both human variants have sufficient activity to mediate AdoCbl synthesis in vivo. Investigations also showed that purified recombinant human methionine synthase reductase (MSR) in combination with purified ATR can convert cob(II)alamin to AdoCbl in vitro. In this system, MSR reduced cob(II)alamin to cob(I)alamin that was adenosylated to AdoCbl by ATR. The optimal stoichiometry for this reaction was approximately 4 MSR/ATR and results indicated that MSR and ATR physically interacted in such a way that the highly reactive reaction intermediate [cob(I)alamin] was sequestered. The finding that MSR reduced cob(II)alamin to cob(I)alamin for AdoCbl synthesis (in conjunction with the prior finding that MSR reduced cob(II)alamin for the activation of methionine synthase) indicates a dual physiological role for MSR.

SNPs, Haplotypes, and Cancer: Applications in Molecular Epidemiology

SNPs, Haplotypes, and Cancer: Applications in Molecular Epidemiology

Common genetic variants that relate to disorders of lipid transport in Spanish subjects with premature coronary artery disease

Fifteen common polymorphic variants at six loci (apolipoproteins AI, B, CIII and E, hepatic lipase and lipoprotein lipase) involved in plasma lipid transport have been studied in 210 northern Spanish men, of whom 98 had proven coronary artery disease. The other 112 men were clinically free from coronary artery disease and acted as controls. The genotypes were investigated for relationships with plasma lipid and lipoprotein levels, as well as for the presence of coronary artery disease. As expected, the mean levels of plasma triacylglycerols (triglycerides) and lipoprotein (a) and the number of smokers were significantly higher in the disease group, and high-density lipoprotein (HDL)-cholesterol was significantly lower. Surprisingly, plasma cholesterol and low-density lipoprotein cholesterol were not different between the two groups. With regard to the common mutations, plasma triacylglycerol levels were related to the HindIII variants of lipoprotein lipase (P<0.05), to the apolipoprotein CIII variant (C3175G in exon 4) and to the apolipoprotein AI XmnI polymorphisms (P<0.05 and P<0.02 respectively). The apolipoprotein E variants were related to plasma cholesterol (P<0.05), HDL-cholesterol (P<0.02), plasma triacylglycerols (P<0.05) and the triacylglycerol/HDL ratio (P<0.01). Only the three-codon insertion/deletion variants of the apolipoprotein B signal peptide region discriminated between the two groups with or without arterial disease (P=0.02). The possible functional effects of these common mutations are discussed.

Genetic restriction of HIV-1 pathogenesis to AIDS by promoter alleles of IL10.

IL10 is a powerful TH-2 cell cytokine produced by lymphoid cells that limits HIV-1 replication in vivo, ostensibly by inhibiting macrophage/monocyte and T-cell lymphocyte replication and secretion of inflammatory cytokines (IL1, TNFalpha, IL6, IL8, and IL12). A genetic epidemiological scan of patients enrolled in AIDS cohorts for candidate gene-linked short tandem repeat polymorphisms revealed significant genotype associations for HIV-1 infection and progression to AIDS with markers adjacent to and tracking (by linkage disequilibrium) common single nucleotide polymorphic variants in the IL10 promoter region. Individuals carrying the IL10-5'-592A (IL10-5'A) promoter allele possibly were at increased risk for HIV-1 infection, and once infected they progressed to AIDS more rapidly than homozygotes for the alternative IL10-5'-592 C/C (IL10-+/+) genotype, particularly in the later stages of HIV-1 infection. An estimated 25-30% of long-term nonprogressors (who avoid clinical AIDS for 10 or more years after HIV-1 infection) can be attributed to their IL10-+/+ promoter genotype. Alternative IL10 promoter alleles are functionally distinct in relative IL10 production, in retention of an avian erythroblastosis virus transcription factor recognition sequence and in binding to specific putative nuclear transcription factors, suggesting a potential mechanism whereby IL10-5'A down-regulation of inhibitory IL10 facilitates HIV-1 replication in vivo, accelerating the onset of AIDS.

A single-nucleotide polymorphic variant of the RET proto-oncogene is underrepresented in sporadic Hirschsprung disease.

Hirschsprung disease (HSCR) is an inherited disorder characterised by absence of intrinsic ganglion cells in the distal gastrointestinal tract. Different susceptibility genes, involved in either the Ret-tyrosine kinase or the endothelin signalling pathways, contribute to HSCR phenotype. Interestingly, alterations of these genes are detected in only 30-50% of all HSCR patients, suggesting the involvement of modifier genes and/or additional genetic or environmental risk factors. In complex disorders common polymorphic variants can be associated with the disease phenotype, thus modifying the risk of recurrence. To investigate whether sequence variants of the RET proto-oncogene may be associated with the development of the HSCR phenotype, we analysed 92 Italian patients for the 2508C > T synonymous substitution in exon 14 (S836S) finding that the T allele is clearly less frequent than in control individuals (Fisher exact test P = 0.0002). On the other hand, this RET variant allele is overrepresented in patients affected with medullary thyroid carcinoma. Assuming a direct effect of this single-nucleotide polymorphism in predisposing to RET associated pathologies, we have performed functional tests which excluded any possible involvement of the C and T alleles in DNA-protein binding, transcript stability and RNA splicing and editing.