Well-differentiated gastroenteropancreatic G3 NET: findings from a large single centre cohort.

Neuroendocrine neoplasms are known to have heterogeneous biological behavior. G3 neuroendocrine tumours (NET G3) are characterized by well-differentiated morphology and Ki67 > 20%. The prognosis of this disease is understood to be intermediate between NET G2 and neuroendocrine carcinoma (NEC). Clinical management of NET G3 is challenging due to limited data to inform treatment strategies. We describe clinical characteristics, treatment, and outcomes in a large single centre cohort of patients with gastroenteropancreatic NET G3. Data was reviewed from 26 cases managed at Queen Elizabeth Hospital, Birmingham, UK, from 2012 to 2019. Most commonly the site of the primary tumour was unknown and majority of cases with identifiable primaries originated in the GI tract. Majority of cases demonstrated somatostatin receptor avidity. Median Ki67 was 30%, and most cases had stage IV disease at diagnosis. Treatment options included surgery, somatostatin analogs (SSA), and chemotherapy with either platinum-based or temozolomide-based regimens. Estimated progression free survival was 4 months following initiation of SSA and 3 months following initiation of chemotherapy. Disease control was observed following treatment in 5/11 patients treated with chemotherapy. Estimated median survival was 19 months; estimated 1 year survival was 60% and estimated 2 year survival was 13%. NET G3 is a heterogeneous group of tumours and patients which commonly have advanced disease at presentation. Prognosis is typically poor, though select cases may respond to treatment with SSA and/or chemotherapy. Further study is needed to compare efficacy of different treatment strategies for this disease.

Familial hypercholesterolaemia workshop for leveraging point-of-care testing and personalised medicine in association with the Lipid and Atherosclerosis Society of Southern Africa.

Familial hypercholesterolaemia (FH) is a common autosomal dominantly inherited disorder in which impaired clearance of plasma low-density lipoprotein cholesterol causes premature atherosclerotic vascular disease and tendon xanthomata. This workshop aimed to consolidate information on the diagnosis and management of FH in South Africa. The genetic causes include mutations in the LDL receptor, apolipoprotein B100 and proprotein convertase subtilisin/kexin type 9 (PCSK9). Additionally, the concatenation of multiple gene variants can result in polygenic FH. Therapeutic measures include a healthy lifestyle, statins and cholesterol-absorption inhibitors that will achieve control of the dyslipidaemia in the majority of cases. The recently introduced monoclonal antibodies to PCSK9 can improve achievement of target concentration in severe cases. FH is present in all sectors of the South African population but there is sparse documentation in the indigenous African populations. FH should be actively sought, diagnosed and treated with judicious pharmacotherapy and screening of relatives.

Subclinical atherosclerosis and its progression are modulated by PLIN2 through a feed-forward loop between LXR and autophagy.

BACKGROUND: Hyperlipidaemia is a major risk factor for cardiovascular disease, and atherosclerosis is the underlying cause of both myocardial infarction and stroke. We have previously shown that the Pro251 variant of perilipin-2 reduces plasma triglycerides and may therefore be beneficial to reduce atherosclerosis development. OBJECTIVE: We sought to delineate putative beneficial effects of the Pro251 variant of perlipin-2 on subclinical atherosclerosis and the mechanism by which it acts. METHODS: A pan-European cohort of high-risk individuals where carotid intima-media thickness has been assessed was adopted. Human primary monocyte-derived macrophages were prepared from whole blood from individuals recruited by perilipin-2 genotype or from buffy coats from the Karolinska University hospital blood central. RESULTS: The Pro251 variant of perilipin-2 is associated with decreased intima-media thickness at baseline and over 30 months of follow-up. Using human primary monocyte-derived macrophages from carriers of the beneficial Pro251 variant, we show that this variant increases autophagy activity, cholesterol efflux and a controlled inflammatory response. Through extensive mechanistic studies, we demonstrate that increase in autophagy activity is accompanied with an increase in liver-X-receptor (LXR) activity and that LXR and autophagy reciprocally activate each other in a feed-forward loop, regulated by CYP27A1 and 27OH-cholesterol. CONCLUSIONS: For the first time, we show that perilipin-2 affects susceptibility to human atherosclerosis through activation of autophagy and stimulation of cholesterol efflux. We demonstrate that perilipin-2 modulates levels of the LXR ligand 27OH-cholesterol and initiates a feed-forward loop where LXR and autophagy reciprocally activate each other; the mechanism by which perilipin-2 exerts its beneficial effects on subclinical atherosclerosis.

Coronary heart disease mortality in treated familial hypercholesterolaemia: Update of the UK Simon Broome FH register.

BACKGROUND AND AIMS: Patients with familial hypercholesterolaemia (FH) have an elevated risk of coronary heart disease (CHD). Here we compare changes in CHD mortality in patients with heterozygous (FH) pre 1992, before lipid-lowering therapy with statins was used routinely, and in the periods 1992-2008 and 2008-2016. METHODS: 1903 Definite (DFH) and 1650 Possible (PFH) patients (51% women) aged 20-79 years, recruited from 21 lipid clinics in the United Kingdom and followed prospectively between 1980 and 2016 for 67,060 person-years. The CHD standardised mortality ratio (SMR) compared to the population in England and Wales was calculated (with 95% Confidence intervals). RESULTS: There were 585 deaths, including 252 from CHD. Overall, the observed 2.4-fold excess coronary mortality for treated DFH post-1991 was significantly higher than the 1.78 excess for PFH (35% 95% CI 3%-76%). In patients with DFH and established coronary disease, there was a significant excess coronary mortality in all time periods, but in men it was reduced from a 4.83-fold excess (2.32-8.89) pre-1992 to 4.66 (3.46-6.14) in 1992-2008 and 2.51 (1.01-5.17) post-2008, while in women the corresponding values were 7.23 (2.65-15.73), 4.42 (2.70-6.82) and 6.34 (2.06-14.81). Primary prevention in men with DFH resulted in a progressive reduction in coronary mortality over the three time-periods, with no excess mortality evident post-2008 (0.89 (0.29-2.08)), although in women the excess persisted (post-2008 3.65 (1.75-6.72)). CONCLUSIONS: The results confirm the benefit of statin treatment in reducing CHD mortality, but suggest that FH patients with pre-existing CHD and women with FH may not be treated adequately.

The Genetic Spectrum of Familial Hypercholesterolemia (FH) in the Iranian Population.

Familial hypercholesterolemia (FH) is an autosomal dominant disorder associated with premature cardiovascular disease (CVD). Mutations in the LDLR, APOB, and PCSK9 genes are known to cause FH. In this study, we analysed the genetic spectrum of the disease in subjects from the Iranian population with a clinical diagnosis of FH. Samples were collected from 16 children and family members from five different cities of Iran. Probands were screened for mutations in the LDLR, APOB, and PCSK9 genes using next generation sequencing, with results confirmed by Sanger sequencing. The likely pathology of identified variants was examined using in silico tools. Of the probands, 14 had a clinical diagnosis of homozygous FH and two of heterozygous FH. No mutations were found in either APOB or PCSK9, but nine probands were homozygous for seven different LDLR mutations, with p.(Trp577Arg) occurring in three and p.Val806Glyfs*11 occurring in two patients. Two mutations were novel: p.(Leu479Gln) and p.(Glu668*). Seven probands with a clinical diagnosis of FH were mutation negative. This pilot study, integrating clinical and molecular-based techniques, begins to elucidate the FH heterogeneity and the mutation spectrum in the Iranian population. Such information is important for future disease management and cost savings.

Analysis of mutations causing familial hypercholesterolaemia in black South African patients of different ancestr.

BACKGROUND: Familial hypercholesterolaemia (FH) is usually caused by mutations in three genes (LDLR, APOB and PCSK9). OBJECTIVE: To identify the spectrum of FH-causing mutations in black South African (SA) patients. METHODS: DNA samples of 16 unrelated South African FH patients with elevated low-density lipoprotein cholesterol levels, tendon xanthomas and corneal arcus (3 clinically homozygous FH and 13 heterozygous FH) of ethnic African origin were screened for mutations in the LDLR (coding region, promoter and intron/exon boundaries), APOB (part of exon 26) and PCSK9 genes (exon 7), using high-resolution melting. RESULTS: Eight LDLR mutations were identified, for an overall detection rate of 8/19 predicted FH-causing alleles (42.1%). The previously reported six base pair deletion p.(D47_G48del) was found in two patients, and two novel variants (c.1187-25T>C and c.1664T>G p.(L555R)) were found, both predicted to be pathogenic using in silico web-based predictive algorithms. No pathogenic variants in APOB or PCSK9 were found. CONCLUSIONS: These findings contribute to the knowledge of allelic heterogeneity in the spectrum of FH-causing mutations in black SA patients, signifying their ancestral diversity. The relatively low overall detection rate may reflect locus heterogeneity of the FH phenotype in black SA FH patients.

Would raising the total cholesterol diagnostic cut-off from 7.5 mmol/L to 9.3 mmol/L improve detection rate of patients with monogenic familial hypercholesterolaemia?

A previous report suggested that 88% of individuals in the general population with total cholesterol (TC) > 9.3 mmol/L have familial hypercholesterolaemia (FH). We tested this hypothesis in a cohort of 4896 UK civil servants, mean (SD) age 44 (±6) years, using next generation sequencing to achieve a comprehensive genetic diagnosis. 25 (0.5%) participants (mean age 49.2 years) had baseline TC > 9.3 mmol/L, and overall we found an FH-causing mutation in the LDLR gene in seven (28%) subjects. The detection rate increased to 39% by excluding eight participants with triglyceride levels over 2.3 mmol/L, and reached 75% in those with TC > 10.4 mmol/L. By extrapolation, the detection rate would be ∼25% by including all participants with TC > 8.6 mmol/L (2.5 standard deviations from the mean). Based on the 1/500 FH frequency, 30% of all FH-cases in this cohort would be missed using the 9.3 mmol/L cut-off. Given that an overall detection rate of 25% is considered economically acceptable, these data suggest that a diagnostic TC cut-off of 8.6 mmol/L, rather than 9.3 mmol/L would be clinically useful for FH in the general population.

Upregulation of gingival tissue miR-200b in obese periodontitis subjects.

Increased local immune and inflammatory responses in obese individuals with periodontitis may explain the aggressive clinical presentation and altered treatment response when compared to that of normal weight subjects. Our goal was to identify any differences in microRNA (miRNA) expression profiles of gingival tissue in periodontitis when obesity is present, which may suggest novel molecular pathways that this miRNA network may affect. Total RNA was extracted from gingival tissue biopsies collected from normal weight and obese individuals with periodontitis; miRNA expression profiling was performed with Affymetrix GeneChip miRNA 3.0 arrays; and results were validated with quantitative reverse transcription polymerase chain reaction (qRT-PCR). In silico identification of previously confirmed miRNA gene targets was conducted through miRTarBase and miRWalk databases, and pathway enrichment analysis identified enriched miRNA gene sets. Expression of selected genes in the same biopsy samples was tested with qRT-PCR. The gingival tissue miRNA profile of obese patients, compared to that of normal weight patients, showed 13 upregulated and 22 downregulated miRNAs, among which miR-200b was validated by qRT-PCR to be significantly increased in obesity. Functional analysis of 51 experimentally validated miR-200b gene targets identified enrichment of genes involved in cell motility, differentiation, DNA binding, response to stimulus, and vasculature development pathways not previously identified in the obesity-specific disease profile. Furthermore, the expression of the miR-200b gene targets ZEB1/2, GATA2, and KDR was confirmed by qRT-PCR as being lower in obese patients with periodontitis versus normal weight patients, suggesting a role of miR-200b in regulation of a set of gene targets and biological pathways relevant to wound healing and angiogenesis. Functional studies to explore the role of miR-200b in the above processes may offer new insights on putative therapeutic targets for this group of patients.

Telomere shortening over 6 years is associated with increased subclinical carotid vascular damage and worse cardiovascular prognosis in the general population.

INTRODUCTION: Leucocyte telomere length (LTL) is an important determinant of telomere function and cellular replicative capacity. The aim of the present study was to examine prospectively the associations between telomere shortening (TS) and both the progression of atherosclerosis and the incidence of cardiovascular events (CVEs). MATERIALS AND METHODS: Leucocyte telomere length was measured by quantitative polymerase chain reaction to determine the ratio of telomere length to single-copy gene (T/S) in 768 subjects (462 female and 306 male) enrolled in a large general population survey [the Progressione della Lesione Intimale Carotidea (PLIC study)]. Common carotid artery intima-media thickness was determined at baseline and after 6 years of follow-up, and the associations between TS and the progression of atherosclerosis and incidence of CVEs were evaluated. RESULTS: Mean LTL was 1.25 ± 0.92 T/S (median 1.14) at baseline and 0.70 ± 0.37 T/S (median 0.70) after 6 years of follow-up. Median 6-year LTL change was -0.46 T/S [interquartile range (IQR) -0.57 to 1.06], equating to -0.078 T/S [IQR(-0.092 to 0.176)] per year. Of note, telomere lengthening occurred in 30.4% of subjects. After adjustment for classical cardiovascular disease (CVD) risk factors (age, gender, smoking, physical activity, alcohol consumption, systolic blood pressure, glucose levels, lipid profile and therapies), TS was associated with incident subclinical carotid vascular damage [hazard ratio (HR) 5.19, 95% confidence interval (CI) 1.20-22.4, P = 0.028]. Finally, subjects in whom LTL shortened over time showed an increased risk of incident CVE, compared to those in whom LTL lengthened (HR 1.69, CI 1.02-2.78, P = 0.041). CONCLUSION: These data indicate that TS is associated with increased risk of subclinical carotid vascular damage and increased incidence of CVEs beyond CVD risk factors in the general population, whereas LTL lengthening is protective.

Functional analysis of TCF7L2 genetic variants associated with type 2 diabetes.

BACKGROUND AND AIMS: Common non-coding variations within the TCF7L2 locus have a strong influence on type 2 diabetes (T2D) susceptibility through uncharacterised mechanisms. An islet-specific functional polymorphism has been identified, although this does not explain the association between genotype and gene expression in other cell types. This study sought to identify these other functional TCF7L2 variants. METHODS AND RESULTS: Alternative splicing and gene expression from TCF7L2 was examined from peripheral blood mononuclear cells from 100 healthy Caucasians using two T2D-associated SNPs, rs7903146 and rs12255372. Electrophoretic mobility shift assays and luciferase reporter assays were performed with these SNPs and those in strong LD to determine potential SNP functionality. Individuals homozygous for rs7903146 and rs12255372 T2D risk alleles (TT/TT) expressed 2.6-fold greater levels of TCF7L2 mRNA compared to individuals homozygous for the non-risk alleles (CC/GG, p = 0.006), although differentially spliced TCF7L2 transcripts did not differ by T2D risk-associated genotype. From SNPs identified to be in strong LD with the T2D-associated SNPs, rs7903146 and rs12255372, five (rs4132670, rs4506565, rs7903146, rs7901695, rs17747324) demonstrated allele-specific binding in electrophoretic mobility shift assays (EMSA). In luciferase reporter assays, rs4132670 exhibited 1.3-fold higher levels of enhancer activity in the Huh7 cell line (p = 3.8 × 10(-5)) and 2-fold higher levels in a WiDr colon carcinoma cell line (p = 0.008). CONCLUSIONS: These results suggest that rs4132670, located in a region of chromatin accessibility, is a non-tissue-specific candidate functional SNP that has the potential to play a role in TCF7L2 gene expression and T2D risk.

LDL oxidative modification and carotid atherosclerosis: results of a multicenter study.

OBJECTIVE: Serum LDL conjugated diene concentration is a marker of oxidative modification of LDL. We investigated the relationship between LDL conjugated dienes and cross-sectional subclinical atherosclerosis assessed by carotid IMT in high-risk subjects of a multicenter study. METHODS: Serum LDL conjugated dienes and ultrasonographically assessed carotid intima-media thickness (IMT(mean), IMT(max) and IMT(mean-max)) were available for 553 subjects from Finland, France, Italy, the Netherlands, and Sweden. RESULTS: In multivariate regression analysis, gender (p < 0.001), age (p < 0.001), systolic blood pressure (IMT(mean), p = 0.01; IMT(mean-max), p = 0.05) and serum LDL conjugated dienes (p = 0.02 for both IMT(mean) and IMT(mean-max)) were the strongest determinants of IMT variation, adjusted for study center, ultrasound videotape reader and serum LDL cholesterol. Pack-years of smoking, added into the regression model, did not destroy the significant association between increased serum LDL conjugated dienes and IMT. Ratio of LDL conjugated dienes to LDL particle cholesterol was higher in subjects of Northern recruiting centers than of Southern centers (r = 0.39, p < 0.0001). CONCLUSIONS: There was a cross-sectional association between in vivo increased LDL oxidative modification and subclinical atherosclerosis after adjustment for traditional risk factors. The subjects in Northern countries of Europe had more oxidatively modified lipids per cholesterol in LDL particle than subjects in Southern countries.

Genetic analysis of NR0B1 in congenital adrenal hypoplasia patients: identification of a rare regulatory variant resulting in congenital adrenal hypoplasia and hypogonadal hypogonadism without testicular carcinoma in situ.

There have been few testicular histology reports of adult patients with congenital adrenal hypoplasia/hypogonadal hypogonadism (AHC/HH), but Leydig cell hyperplasia has been observed, an indicator of the possibility of malignant transformation. We aimed to define the basis of AHC/HH in 4 pedigrees of different ethnic backgrounds. One patient was elected to have testicular biopsy which was examined for evidence of carcinoma in situ (CIS). NR0B1 mutation analysis was performed by sequence analysis. NR0B1 expression was investigated by RT-PCR. Testicular biopsy sections were stained with HE or immunostained for OCT3/4, an established marker of CIS. We identified NR0B1 variants in the 4 AHC pedigrees: pedigree 1 (United Arab Emirates), c.1130A>G predicting p.(Glu377Gly); pedigree 2 (English Caucasian), c.327C>A predicting p.(Cys109*); pedigree 3 (Oman), a 6-bp deletion of a direct repeat, c.857_862delTGGTGC predicting p.(Leu286_Val287del); pedigree 4 (English Caucasian), c.1168+1G>A, a regulatory variant within the NR0B1 splice donor site. This last male patient, aged 30 years, presented with evidence of HH but incomplete gonadotrophin deficiency, following an earlier diagnosis of Addison's disease at 3 years. Hormonal therapy induced virilisation. Testicular biopsy was performed. The c.1168+1G>A variant abrogated normal splicing of testicular mRNA. Histological examination showed poorly organised testicular architecture and absence of spermatozoa. Morphological analyses and the absence of immunohistochemical staining for OCT3/4 excluded the presence of malignant germ cell cancer and its precursor lesion, CIS. These studies add to the knowledge of the types and ethnic diversity of NR0B1 mutations and their associated phenotypes, and provide insight into the assessment and interpretation of testicular histology in AHC and HH.

Effects of genetic variation on chromatin structure and the transcriptional machinery: analysis of the IL6 gene locus.

The presence of functional regulatory polymorphism at the interleukin 6 (IL6) locus is uncertain, with many conflicting in vitro findings. To examine the in vivo effect of the three putative functional IL6 promoter variants, -174G>C, -572G>C and -6331T>C, two complementary techniques, allele-specific chromatin immunoprecipitation and allele-specific formaldehyde-assisted isolation of regulatory elements, were carried out using unrelated lymphoblast cell lines of known genotype. There were no allele-specific effects for all three single-nucleotide polymorphisms (SNPs) under basal conditions. Upon IL-1ß stimulation, however, allele-specific effects were seen for the -6331 allele, which showed both increased RNA polymerase II loading (56%, P=0.001) and increased open chromatin (59%, P=0.004) for the T allele, which is in line with previous reports of this SNP and the effects from acute inflammation. These studies highlight the importance of examining chromatin under different environmental conditions when studying the functionality of regulatory polymorphisms.

Review article: pancreatic renin-angiotensin systems in health and disease.

BACKGROUND: In addition to the circulating (endocrine) renin-angiotensin system (RAS), local renin-angiotensin systems are now known to exist in diverse cells and tissues. Amongst these, pancreatic renin-angiotensin systems have recently been identified and may play roles in the physiological regulation of pancreatic function, as well as being implicated in the pathogenesis of pancreatic diseases including diabetes, pancreatitis and pancreatic cancer. AIM: To review and summarise current knowledge of pancreatic renin-angiotensin systems. METHODS: We performed an extensive PubMed, Medline and online review of all relevant literature. RESULTS: Pancreatic RAS appear to play various roles in the regulation of pancreatic physiology and pathophysiology. Ang II may play a role in the development of pancreatic ductal adenocarcinoma, via stimulation of angiogenesis and prevention of chemotherapy toxicity, as well as in the initiation and propagation of acute pancreatitis (AP); whereas, RAS antagonism is capable of preventing new-onset diabetes and improving glycaemic control in diabetic patients. Current evidence for the roles of pancreatic RAS is largely based upon cell and animal models, whilst definitive evidence from human studies remains lacking. CONCLUSIONS: The therapeutic potential for RAS antagonism, using cheap and widely available agents, and may be untapped and such roles are worthy of active investigation in diverse pancreatic disease states.

Probabilistic cost-effectiveness analysis of cascade screening for familial hypercholesterolaemia using alternative diagnostic and identification strategies.

OBJECTIVE: To estimate the probabilistic cost-effectiveness of cascade screening methods in familial hypercholesterolaemia (FH) from the UK NHS perspective. DESIGN: Economic evaluation (cost utility analysis) comparing four cascade screening strategies for FH: Using low-density lipoprotein (LDL) cholesterol measurements to diagnose affected relatives (cholesterol method); cascading only in patients with a causative mutation identified and using DNA tests to diagnose relatives (DNA method); DNA testing combined with LDL-cholesterol testing in families with no mutation identified, only in patients with clinically defined 'definite' FH (DNA+DFH method); DNA testing combined with LDL-cholesterol testing in no-mutation families of both 'definite' and 'probable' FH patients (DNA+DFH+PFH). A probabilistic model was constructed to estimate the treatment benefit from statins, with all diagnosed individuals receiving high-intensity statin treatment. POPULATION: A cohort of 1000 people suspected of having FH aged 50 years for index cases and 30 years for relatives, followed for a lifetime. MAIN OUTCOMES: Costs, quality-adjusted life-years (QALY) and incremental cost-effectiveness ratios (ICER). RESULTS: The DNA+DFH+PFH method was the most cost-effective cascade screening strategy. The ICER was estimated at £3666/QALY. Using this strategy, of the tested relatives 30.6% will be true positives, 6.3% false positives, 61.9% true negatives and 1.1% false negatives. Probabilistic sensitivity analysis showed that this approach is 100% cost-effective using the conventional benchmark for cost-effective treatments in the NHS of between £20,000 and £30,000 per QALY gained. CONCLUSION: Cascade testing of relatives of patients with DFH and PFH is cost-effective when using a combination of DNA testing for known family mutations and LDL-cholesterol levels in the remaining families. The approach is more cost-effective than current primary prevention screening strategies.

Association between the rs4880 superoxide dismutase 2 (C>T) gene variant and coronary heart disease in diabetes mellitus.

Mitochondrial superoxide dismutase 2 (SOD2) is an endogenous anti-oxidant enzyme. The rs4880 gene variant results in a C>T substitution, influencing SOD enzymatic activity. This variant has been associated with micro- and macro-vascular complications in diabetes mellitus. Our aim was to examine the association between this variant and coronary heart disease (CHD) risk in a cross-sectional sample of subjects with diabetes. 776 Caucasian subjects with diabetes were genotyped. CHD risk, oxidised-LDL and plasma total anti-oxidant status (TAOS) were analysed in relation to genotype. In females, the TT genotype was associated with CHD (CC/CT/TT: No CHD vs. CHD: 22.4/56.0/21.6% vs. 12.0/50.0/38.0%, p=0.03; for CC/CT vs. TT, p=0.01). The odds ratio for CHD associated with the TT genotype compared to CC/CT was 2.22 [95%CI: 1.17-4.24], p=0.01. The TT genotype was also associated with significantly lower plasma TAOS. In males, no association was observed between genotype and CHD risk, but CHD was significantly associated with age, lower HDL, higher triglycerides, higher BMI and cigarette smoking. The TT genotype of this variant is associated with increased CHD risk and lower plasma anti-oxidant defences in females with diabetes. This modest genotype-effect is not apparent in males where traditional risk factors may play a greater role.

Cost-effectiveness analysis of the use of a high-intensity statin compared to a low-intensity statin in the management of patients with familial hypercholesterolaemia.

OBJECTIVES: To estimate, using probabilistic decision-analytic modelling techniques, the cost effectiveness of treating familial hypercholesterolaemia (FH) patients with high-intensity statins compared to treatment with low-intensity statins. For the purpose of this economic analysis, and based on their known differences, statins were categorised as high intensity if they produce greater LDL-cholesterol reductions than simvastatin 40 mg (e.g., simvastatin 80 mg and appropriate doses of atorvastatin and rosuvastatin or combination of statins + ezetimibe). METHODS: A lifetime Markov model was developed to estimate the incremental cost per quality adjusted life year (QALY) of treating a hypothetical cohort of 1000 FH patients aged between 20 and 70 years. Baseline coronary heart disease risks reported in the NICE TA 94 on statins, and age-adjusted risk of cardiovascular disease reported in the FH population, were used to populate the model. A meta-analysis estimate of the reduction in cardiovascular events from using high-intensity compared with low-intensity statins was obtained from published trials. Results were interpreted using a cost-effectiveness threshold of pound20 000/QALY. RESULTS: Fewer cardiovascular events and deaths were predicted to occur in the group treated with higher-intensity statins, and the incremental cost-effectiveness ratio (ICER) was estimated at pound11 103/QALY. The ICER remained below the pound20 000 threshold for 20-39-year-olds and 40-59-year-olds, but rose above this threshold in individuals aged over 60 years. One-way sensitivity analysis showed that results were most sensitive to variation in treatment effect on mortality and the cost of high-intensity statins. CONCLUSIONS: Modelling demonstrates that high-intensity statins are cost-effective for the treatment of younger FH patients. If, as is likely, the relative price of high-intensity statins fall in the future as they come off patent, then their cost effectiveness will improve further.

Development of a high-resolution melting method for mutation detection in familial hypercholesterolaemia patients.

AIMS: Current screening methods, such as single strand conformational polymorphism (SSCP) and denaturing high performance liquid chromatography (dHPLC) that are used for detecting mutations in familial hypercholesterolaemia (FH) subjects are time consuming, costly and only 80-90% sensitive. Here we have tested high-resolution melt (HRM) analysis for mutation detection using the Rotor-Gene(6000) realtime rotary analyser. Methods and subjects Polymerase chain reaction and melt conditions (HRM) for 23 fragments of the LDL-receptor gene, a region of exon 26 in the APOB gene (including p.R3527Q) and exon 7 of the PCSK9 gene (including p.D374Y) were optimized. Two double stranded DNA saturating dyes, LC-Green and Syto9, were compared for sensitivity. Eighty-two samples with known mutations were used as positive controls. Twenty-eight Greek FH heterozygous patients and two homozygous patients from the UK and Croatia were screened. RESULTS: HRM was able to identify all the positive control mutations tested, with similar results with either dye. Eight different variations were found in 17 of the 28 Greek FH patients for an overall detection rate of 61%: c.41delT (1), p.W165X (1), p.C173R (3), p.S286R (2), p.V429M (4), p.G549D (4), p.V613I (1), and a previously unreported mutation p.F694V (1) which is predicted to be FH-causing by functional algorithms. Mutations were found in both the homozygous patients; p.Q92X (Croatia) and p.Y489C (UK); both patients were homozygous for their respective mutations. CONCLUSIONS: HRM is a sensitive, robust technique that could significantly reduce the time and cost of screening for mutations in a clinical setting.