Search

A high proportion of coronavirus disease 2019 (COVID-19) survivors may develop long-term cognitive impairment. We aimed to develop a multivariate causal model exposing the links between COVID-19-associated biomarkers, illness-related variables, and their effects on cognitive performance.In this prospective study, we assess the potential drivers for the development of cognitive impairment in patients with severe COVID-19 pneumonia aged ≥ 18 years at 6-month follow-up after hospital discharge, using the Montreal Cognitive Assessment (MoCA). Patients with pre-existing cognitive impairment were excluded. Laboratory results at hospital admission were clustered by principal component analysis (PCA) and included in a path analysis model evaluating the causal relationship between age, comorbidities, hypoxemia, invasive mechanical ventilation (IMV) requirement, in-hospital delirium, and cognitive performance.We studied 92 patients: 54 (58.7%) men and 38 (41.3%) women, with median age of 50 years (interquartile range 42-55), among whom 50 (54.4%) tested positive for cognitive impairment at 6-month follow-up. Path analysis revealed a direct link between the thrombo-inflammatory component of PCA (C-reactive protein, fibrinogen, and neutrophils) and hypoxemia severity at hospital admission. Our model showed that low PaOIn this study, biomarkers of thrombo-inflammation in COVID-19 and low PaO

Latin America continues to be severely underrepresented in genomics research, and fine-scale genetic histories as well as complex trait architectures remain hidden due to the lack of Big Data. To fill this gap, the Mexican Biobank project genotyped 1.8 million markers in 6,057 individuals from 32 states and 898 sampling localities across Mexico with linked complex trait and disease information creating a valuable nationwide genotype-phenotype database. Through a suite of state-of-the-art methods for ancestry deconvolution and inference of identity-by-descent (IBD) segments, we inferred detailed ancestral histories for the last 200 generations in different Mesoamerican regions, unraveling native and colonial/post-colonial demographic dynamics. We observed large variations in runs of homozygosity (ROH) among genomic regions with different ancestral origins reflecting their demographic histories, which also affect the distribution of rare deleterious variants across Mexico. We analyzed a range of biomedical complex traits and identified significant genetic and environmental factors explaining their variation, such as ROH found to be significant predictors for trait variation in BMI and triglycerides.

Background and aims Both insulin resistance (IR) and visceral adipose tissue (VAT) are related cardiometabolic risk factors; nevertheless, their joint effect on endothelial functionality is controversial. This study aims to evaluate the joint effect of IR and VAT on endothelial functionality using the pulse-waveform analysis and explore the mediating role of VAT on the effect of IR on arterial pressure, arterial stiffness and incident arterial hypertension. Methods and results We measured VAT (n = 586) using two methods (dual-energy X-ray absorptiometry and a clinical surrogate), arterial stiffness (with pulse-waveform velocity), and IR (using three methods: HOMA2-IR (n = 586), a frequently sampled intravenous glucose tolerance test (n = 131) and euglycemic hyperinsulinemic clamping (n = 97)) to confirm the mediator effect of IR on VAT. The incidence of arterial hypertension attributable to the mediating effect of IR related to VAT was evaluated using a prospective cohort (n = 6850). Adjusted linear regression models, causal mediation analysis, and Cox-proportional hazard risk regression models were performed to test our objective. IR and VAT led to increased arterial stiffness and increased blood pressure; the combination of both further worsened vascular parameters. Nearly, 57% (ΔE→MY 95% CI: 31.7–100.0) of the effect of IR on altered pulse-wave velocity (PWV) analysis was mediated through VAT. Moreover, VAT acts as a mediator of the effect of IR on increased mean arterial pressure (ΔE→MY 35.7%, 95% CI: 23.8–59) and increased hypertension risk (ΔE→MY 69.1%, 95% CI: 46.1–78.8). Conclusion VAT acts as a mediator of IR in promoting arterial stiffness and arterial hypertension. Both phenomena should be targeted to ameliorate the cardiometabolic risk.

Obesity predisposes individuals to multiple cardiometabolic disorders, including type 2 diabetes (T2D). As body mass index (BMI) cannot reliably differentiate fat from lean mass, the metabolically detrimental abdominal obesity has been estimated using waist-hip ratio (WHR). Waist-hip ratio adjusted for body mass index (WHRadjBMI) in turn is a well-established sex-specific marker for abdominal fat and adiposity, and a predictor of adverse metabolic outcomes, such as T2D. However, the underlying genes and regulatory mechanisms orchestrating the sex differences in obesity and body fat distribution in humans are not well understood.We searched for genetic master regulators of WHRadjBMI by employing integrative genomics approaches on human subcutaneous adipose RNA sequencing (RNA-seq) data (n ~ 1400) and WHRadjBMI GWAS data (n ~ 700,000) from the WHRadjBMI GWAS cohorts and the UK Biobank (UKB), using co-expression network, transcriptome-wide association study (TWAS), and polygenic risk score (PRS) approaches. Finally, we functionally verified our genomic results using gene knockdown experiments in a human primary cell type that is critical for adipose tissue function.Here, we identified an adipose gene co-expression network that contains 35 obesity GWAS genes and explains a significant amount of polygenic risk for abdominal obesity and T2D in the UKB (n = 392,551) in a sex-dependent way. We showed that this network is preserved in the adipose tissue data from the Finnish Kuopio Obesity Study and Mexican Obesity Study. The network is controlled by a novel adipose master transcription factor (TF), TBX15, a WHRadjBMI GWAS gene that regulates the network in trans. Knockdown of TBX15 in human primary preadipocytes resulted in changes in expression of 130 network genes, including the key adipose TFs, PPARG and KLF15, which were significantly impacted (FDR0.05), thus functionally verifying the trans regulatory effect of TBX15 on the WHRadjBMI co-expression network.Our study discovers a novel key function for the TBX15 TF in trans regulating an adipose co-expression network of 347 adipose, mitochondrial, and metabolically important genes, including PPARG, KLF15, PPARA, ADIPOQ, and 35 obesity GWAS genes. Thus, based on our converging genomic, transcriptional, and functional evidence, we interpret the role of TBX15 to be a main transcriptional regulator in the adipose tissue and discover its importance in human abdominal obesity.

Background Obesity predisposes individuals to multiple cardiometabolic disorders, including type 2 diabetes (T2D). As body mass index (BMI) cannot reliably differentiate fat from lean mass, the metabolically detrimental abdominal obesity has been estimated using waist-hip ratio (WHR). Waist-hip ratio adjusted for body mass index (WHRadjBMI) in turn is a well-established sex-specific marker for abdominal fat and adiposity, and a predictor of adverse metabolic outcomes, such as T2D. However, the underlying genes and regulatory mechanisms orchestrating the sex differences in obesity and body fat distribution in humans are not well understood. Methods We searched for genetic master regulators of WHRadjBMI by employing integrative genomics approaches on human subcutaneous adipose RNA sequencing (RNA-seq) data (n ~ 1400) and WHRadjBMI GWAS data (n ~ 700,000) from the WHRadjBMI GWAS cohorts and the UK Biobank (UKB), using co-expression network, transcriptome-wide association study (TWAS), and polygenic risk score (PRS) approaches. Finally, we functionally verified our genomic results using gene knockdown experiments in a human primary cell type that is critical for adipose tissue function. Results Here, we identified an adipose gene co-expression network that contains 35 obesity GWAS genes and explains a significant amount of polygenic risk for abdominal obesity and T2D in the UKB (n = 392,551) in a sex-dependent way. We showed that this network is preserved in the adipose tissue data from the Finnish Kuopio Obesity Study and Mexican Obesity Study. The network is controlled by a novel adipose master transcription factor (TF), TBX15, a WHRadjBMI GWAS gene that regulates the network in trans. Knockdown of TBX15 in human primary preadipocytes resulted in changes in expression of 130 network genes, including the key adipose TFs, PPARG and KLF15, which were significantly impacted (FDR < 0.05), thus functionally verifying the trans regulatory effect of TBX15 on the WHRadjBMI co-expression network. Conclusions Our study discovers a novel key function for the TBX15 TF in trans regulating an adipose co-expression network of 347 adipose, mitochondrial, and metabolically important genes, including PPARG, KLF15, PPARA, ADIPOQ, and 35 obesity GWAS genes. Thus, based on our converging genomic, transcriptional, and functional evidence, we interpret the role of TBX15 to be a main transcriptional regulator in the adipose tissue and discover its importance in human abdominal obesity.

Hypertension is associated with insulin resistance (IR), metabolic syndrome (MS), and arterial stiffness. Non–insulin‐based IR indexes were developed as tools for metabolic screening. Here, we aimed to evaluate the novel non–insulin‐based Metabolic Score for IR (METS‐IR) index for the prediction of incident hypertension and arterial stiffness evaluated using pulse wave velocity (PWV) analysis, compared with other non–insulin‐based IR indexes. We evaluated two populations, a cross‐sectional evaluation of high‐risk individuals (n = 305) with a wide range of metabolic comorbidities and dyslipidemia in whom PWV measurement was performed and a 3‐year prospective cohort of normotensive individuals (N = 6850). We observed a positive correlation between METS‐IR and PWV in the cross‐sectional cohort, which was higher compared with other non–insulin‐based fasting IR indexes; furthermore, PWV values >75th percentile were associated with the upper tercile of METS‐IR values. In the prospective cohort, we observed an increased risk for incident hypertension for the upper METS‐IR tercile (METS‐IR ≥ 46.42; HR: 1.81, 95% CI: 1.41‐2.34), adjusted for known cardiovascular risk factors, and observed that METS‐IR had greater increases in the predictive capacity for hypertension along with SBP and the Framingham Hypertension Risk Prediction Model compared with other non–insulin‐based IR indexes. Therefore, METS‐IR is a novel non–insulin‐based IR index which correlates with arterial stiffness and is a predictor of incident hypertension, complementary to previously validated risk prediction models.

Objective A haplotype at chromosome 17p13 that reduces expression and function of the solute carrier transporter SLC16A11 is associated with increased risk for type 2 diabetes in Mexicans. We aim to investigate the detailed metabolic profile of SLC16A11 risk haplotype carriers to identify potential physiological mechanisms explaining the increased type 2 diabetes risk. Design Cross-sectional study. Methods We evaluated carriers (n = 72) and non-carriers (n = 75) of the SLC16A11 risk haplotype, with or without type 2 diabetes. An independent sample of 1069 subjects was used to replicate biochemical findings. The evaluation included euglycemic–hyperinsulinemic clamp, frequently sampled intravenous glucose tolerance test (FSIVGTT), dual-energy X-ray absorptiometry (DXA), MRI and spectroscopy and subcutaneous abdominal adipose tissue biopsies. Results Fat-free mass (FFM)-adjusted M value was lower in carriers of the SLC16A11 risk haplotype after adjusting for age and type 2 diabetes status (β = −0.164, P = 0.04). Subjects with type 2 diabetes and the risk haplotype demonstrated an increase of 8.76 U/L in alanine aminotransferase (ALT) (P = 0.02) and of 7.34 U/L in gamma-glutamyltransferase (GGT) (P = 0.05) compared with non-carriers and after adjusting for gender, age and ancestry. Among women with the risk haplotype and normal BMI, the adipocyte size was higher (P < 0.001). Conclusions Individuals carrying the SLC16A11 risk haplotype exhibited decreased insulin action. Higher serum ALT and GGT levels were found in carriers with type 2 diabetes, and larger adipocytes in subcutaneous fat in the size distribution in carrier women with normal weight.

The purpose of this perspective is to analyze the use of precision medicine and its potential for the next few decades with a special focus on the low- and middle-income countries, using diabetes as a paradigm. Precision medicine has improved the diagnosis, prevention, treatment, and prognosis of several malignant neoplasia. Furthermore, this approach is useful in the management of monogenic diabetes. However, its impact in the current practice for the majority of the cases of diabetes is quite limited. Precision medicine has not fulfilled the expectations because it implies a long-term process composed by several feedback loops, and a number of internal and external validations and calibrations to target specific populations. If we want to obtain the expected benefits, the academic community and science agencies should work together to create the budgets and infrastructure that warrant the transfer of knowledge to the whole of society.

Background Familial hypertriglyceridemia (FHTG) is a partially characterized primary dyslipidemia which is frequently confused with other forms hypertriglyceridemia. The aim of this work is to search for specific features that can help physicians recognize this disease. Methods This study included 84 FHTG cases, 728 subjects with common mild-to-moderate hypertriglyceridemia (CHTG) and 609 normotriglyceridemic controls. All subjects underwent genetic, clinical and biochemical assessments. A set of 53 single nucleotide polymorphisms (SNPs) previously associated with triglycerides levels, as well as 37 rare variants within the five main genes associated with hypertriglyceridemia (i.e. LPL, APOC2, APOA5, LMF1 and GPIHBP1) were analyzed. A panel of endocrine regulatory proteins associated with triglycerides homeostasis were compared between the FHTG and CHTG groups. Results Apolipoprotein B, fibroblast growth factor 21(FGF-21), angiopoietin-like proteins 3 (ANGPTL3) and apolipoprotein A-II concentrations, were independent components of a model to detect FHTG compared with CHTG (AUC 0.948, 95%CI 0.901–0.970, 98.5% sensitivity, 92.2% specificity, P Conclusions The clinical and genetic differences observed between FHTG and CHTG supports the notion that FHTG is a unique entity, distinguishable from other causes of hypertriglyceridemia by the higher concentrations of insulin, FGF-21, ANGPTL3, apo A-II and lower levels of apo B. We propose the inclusion of these parameters as useful markers for differentiating FHTG from other causes of hypertriglyceridemia.

We report on the case of an 8-year-old Mexican male, with a 3-year-old clinical diagnosis of familial hypercholesterolemia, and the difficulties encountered in his treatment while in our care. His treatment started with a regimen consisting of ezetimibe/simvastatin, cholestyramine, and a dietary plan of 1600 calories, with a limited intake of 200 mg of cholesterol per day. Problems arose when the patient’s low-density lipoprotein cholesterol (LDL) levels did not meet ideal targets, which prompted the use of LDL cholesterol apheresis (not available in Mexico) for 6 months. As a last resort, PCSK9 inhibitors were administered but the LDL levels remained in the 600 mg/dL range. AmbryGenetics conducted a genetic test employing the Sanger method. The results suggested that there were 2 different mutations for each allele of the same LDL receptor gene (c.249delTinsGG and p.(Cys109Arg)), located in exons 3 and 4, respectively. We identified compound heterozygous mutations in our index case, with him having both the p.C109R mutation (from the maternal lineage), as well as a c.249delTinsGG mutation (from the paternal lineage). The p.C109R mutation has been previously reported, not only in Mexico, but in European regions (Germany, Czech Republic, Ireland, Italy) as well. Functional studies indicated a residual enzymatic activity of 15% to 30% for heterozygotes. To date, the variant c.249delTinsGG has not been reported. This case study illustrates the fact that in Mexico there are limited options available for treatment in such a scenario. As medical professionals, we are limited by the tools at our disposal.

OBJECTIVE To assess whether an ethnic-specific variant (p.E508K) in the maturity-onset diabetes of the young (MODY) gene hepatocyte nuclear factor-1α (HNF1A) found in Mexicans is associated with higher sensitivity to sulfonylureas, as documented in patients with MODY3. RESEARCH DESIGN AND METHODS We recruited 96 participants (46 variant carriers and 50 age- and sex-matched noncarriers). Response to glipizide (one 2.5–5.0-mg dose), metformin (four 500-mg doses), and an oral glucose challenge was evaluated using a previously validated protocol. Glucose and insulin levels and their areas under the curve (AUCs) were compared between groups. RESULTS Carriers of the p.E508K variant had a lower maximum insulin peak during the glipizide challenge as compared with noncarriers with diabetes (P < 0.05). Also, carriers had a lower insulin response after the oral glucose challenge. Following an oral glucose tolerance test in the presence of metformin, carriers of the p.E508K variant with diabetes had a lower maximum insulin peak and total and incremental insulin AUC value as compared with noncarriers with diabetes (P < 0.05). A similar but nonsignificant trend was seen in participants without type 2 diabetes. CONCLUSIONS Carriers of variant p.E508K in HNF1A have a reduced insulin response rather than the increased sensitivity to sulfonylureas seen in patients with MODY3.

Background Type 2 diabetes mellitus (T2D) is a leading cause of morbidity and mortality in Mexico. Here, we aimed to report incidence rates (IR) of type 2 diabetes in middle-aged apparently-healthy Mexican adults, identify risk factors associated to ID and develop a predictive model for ID in a high-risk population. Methods Prospective 3-year observational cohort, comprised of apparently-healthy adults from urban settings of central Mexico in whom demographic, anthropometric and biochemical data was collected. We evaluated risk factors for ID using Cox proportional hazard regression and developed predictive models for ID. Results We included 7636 participants of whom 6144 completed follow-up. We observed 331 ID cases (IR: 21.9 per 1000 person-years, 95%CI 21.37–22.47). Risk factors for ID included family history of diabetes, age, abdominal obesity, waist-height ratio, impaired fasting glucose (IFG), HOMA2-IR and metabolic syndrome. Early-onset ID was also high (IR 14.77 per 1000 person-years, 95%CI 14.21–15.35), and risk factors included HOMA-IR and IFG. Our ID predictive model included age, hypertriglyceridemia, IFG, hypertension and abdominal obesity as predictors (Dxy = 0.487, c-statistic = 0.741) and had higher predictive accuracy compared to FINDRISC and Cambridge risk scores. Conclusions ID in apparently healthy middle-aged Mexican adults is currently at an alarming rate. The constructed models can be implemented to predict diabetes risk and represent the largest prospective effort for the study metabolic diseases in Latin-American population. Electronic supplementary material The online version of this article (10.1186/s12902-019-0361-8) contains supplementary material, which is available to authorized users.

Background: Homozygous familial hypercholesterolemia (HoFH), a very rare autosomal dominant disease, mainly caused by LDL receptor (LDL-R) alterations, with aortic valve compromise reaching nearly 100% and vascular calcifications that can progress independently of marked decrease in total and LDL cholesterol levels. Clinical case: A 21-year-old Hispanic woman diagnosed with HoFH at the age of 6 was referred to our hospital for follow up. Her father had died of acute myocardial infarction at the age of 40. Her mother had an LDLc of 309 mg/dl (8.06 mmol/l) and total cholesterol (TC) 400 mg/dl (10.44 mmol/l) with Achilles tendon xanthomas. She had one elder-brother and sister with elevated levels of TC without xanthomas. She had initially been treated with a statin and ezetimibe at age of 15, but had discontinued these drugs several years ago. Her lipid profile showed TC 700 mg/dl (18.2 mmol/l), triglycerides (Tg) 100 mg/dl (1.31 mmol/l), high density lipoprotein cholesterol (HDL-C) 32 mg/dl (0.83 mmol/l) and LDL-C 546 (14.2 mmol/l). Physical examination showed, xanthomas in Achilles tendons and elbows, and tuberous xanthomas on the knees. She had a systolic ejection murmur, which irradiated to the axillar region. Her EKG was normal. Carotid arterial intima-media thickness (cIMT) revealed non-obstructive plaque formation in both right and left carotid arteries. CT coronary angiography showed severe calcification in the right coronary, left anterior descending arteries, ascending and descending aorta with calcium score of 890 AU. Strain echocardiography showed thickened and flared valves in the aortic, tricuspid and mitral regions. PET/CT with 18F Sodium-Fluoride (18F-NaF) radiotracer showed significant aortic valve uptake with active tissue calcifications. The patient is currently receiving PCSK-9 inhibitor bi-monthly. Conclusion: Despite of the symptoms and cholesterol levels, the clinical course of atherosclerotic manifestations in HoHF individual is highly variable. Premature malignant atherosclerosis leading to aortic valve and root abnormalities is a recognized complication of HoFH and has a strong correlation with age and calcium scores. Greater access to imaging techniques are critical to define the clinical variability of HoFH that have important prognostic and therapeutic implications. Reference: M. Cuchel, E. Bruckert, H.N. Ginsberg, et al., Homozygous familial hypercholesterolaemia: new insights and guidance for clinicians to improve detection and clinical management. A position paper from the Consensus Panel on Familial Hypercholesterolaemia of the European Atherosclerosis Society, Eur. Heart J. 35 (32) (2014) 2146e2157 ehu274

Background Heterozygous germline TP53 gene mutations result in Li-Fraumeni Syndrome (LFS). Breast cancer (BC) is the most frequent tumor in young women with LFS. An important issue related to BC in the Mexican population is the average age at diagnosis, which is approximately 11 years younger than that of patients in the United States (U.S.) and Europe. The aim of this study was to determine the prevalence of germline mutations in TP53 among young Mexican BC patients. Methods We searched for germline mutations in the TP53 gene using targeted next-generation sequencing (NGS) in 78 BC patients younger than 45 years old (yo) who tested negative for BRCA1/2 mutations. A group of 509 Mexican women aged 45yo or older without personal or family BC history (parents/grandparents) was used as a control. Results We identified five patients with pathogenic variants in the TP53 gene, equivalent to 6.4% (5/78). Among patients diagnosed at age 36 or younger, 9.4% (5/55) had pathogenic TP53 mutations. Three of these variants were missense mutations (c.844C > T, c.517G > A, and c.604C > T), and the other two mutations were frameshifts (c.291delC and c.273dupC) and had not been reported previously. We also identified a variant of uncertain clinical significance (VUS), c.672G > A, which causes a putative splice donor site mutation. All patients with TP53 mutations had high-grade and HER2-positive tumors. None of the 509 patients in the healthy control group had mutations in TP53. Conclusions Among Mexican BC patients diagnosed at a young age, we identified a high proportion with germline mutations in the TP53 gene. All patients with the TP53 mutations had a family history suggestive of LFS. To establish the clinical significance of the VUS found, additional studies are needed. Pathogenic variants of TP53 may explain a substantial fraction of BC in young women in the Mexican population. Importantly, none of these mutations or other pathological variants in TP53 were found in the healthy control group.

Genome-wide association studies (GWAS) have made possible the identification of >175 loci that affect plasma lipid levels. Its results have been crucial to identify roles of new players in lipid metabolism (ie, apolipoprotein A5) or even to postulate potential treatment targets (ie, apolipoprotein C-III). However, a large proportion of the GWAS results has not been translated in clinically useful information because a large proportion of responsible single nucleotide polymorphisms (SNPs) are located either in noncoding regions or in genes without an obvious participation in any metabolic pathway.1 This is the case of the association between rs964184 and plasma triglycerides concentrations. This highly significant association has been a constant finding in the GWAS reports, regardless the sample size or the study sample composition.2 The frequency of the risk allele (G) varies between populations from 12% in whites to 27% in Mexicans. Also, it is common in Japanese (31%) and in Native American communities located in Central Mexico (≈50%). This SNP is nearby the 3′ untranslated region of the zinc finger gene ( ZPR1 ) and close to APOA5 . However, the identification of the functional variant that explains the association has not been possible. See Article by Hsueh et al The study of ethnic groups not included in the discovery cohorts, some methodological approaches (ie, admixture mapping or inception by descent [IBD] mapping), and deep genotyping are among the potential approaches to identify the variants responsible for a GWAS signal.3 The study of the Native American populations is an option because this group has not included in the majority of the lipids-related GWAS. The greater susceptibility of the populations derived from the Native American founding groups for having hypertriglyceridemia is well documented.4 These groups include the Amerindian communities living in Canada and the United States, mestizos living in the …

The burden caused by familial hypercholesterolemia (FH) varies among countries and ethnic groups. The prevalence and characteristics of FH in Latin American (LA) countries is largely unknown. We present a systematic review (following the PRISMA statement) of FH in LA countries. The epidemiology, genetics, screening, management, and unique challenges encountered in these countries are discussed. Published reports discussing FH in Hispanic or LA groups was considered for analysis. Thirty studies were included representing 10 countries. The bulk of the data was generated in Brazil and Mexico. Few countries have registries and there was little commonality in FH mutations between LA countries. LDL receptor mutations predominate; APOB and PCSK9 mutations are rare. No mutation was found in an FH gene in nearly 50% of cases. In addition, some country-specific mutations have been reported. Scant information exists regarding models of care, cascade screening, cost, treatment effectiveness, morbidity, and mortality. In conclusion, FH is largely underdiagnosed and undertreated in the LA region. The genetic admixture with indigenous populations, producing mestizo's groups, may influence the mutational findings in Latin America. Potential opportunities to close gaps in knowledge and health care are identified.

SummaryLarge-scale reference data sets of human genetic variation are critical for the medical and functional interpretation of DNA sequence changes. Here we describe the aggregation and analysis of high-quality exome (protein-coding region) sequence data for 60,706 individuals of diverse ethnicities generated as part of the Exome Aggregation Consortium (ExAC). The resulting catalogue of human genetic diversity contains an average of one variant every eight bases of the exome, and provides direct evidence for the presence of widespread mutational recurrence. We show that this catalogue can be used to calculate objective metrics of pathogenicity for sequence variants, and to identify genes subject to strong selection against various classes of mutation; we identify 3,230 genes with near-complete depletion of truncating variants, 72% of which have no currently established human disease phenotype. Finally, we demonstrate that these data can be used for the efficient filtering of candidate disease-causing variants, and for the discovery of human “knockout” variants in protein-coding genes.

Objective To test the hypothesis that persons with the R230C allele of ABCA1 show a decreased glycemic response to glyburide. This polymorphism is exclusively found in Ameri-indian populations and is associated with type 2 diabetes. Research design and methods This is a single blind controlled study including participants with type 2 diabetes (fasting glucose levels 126-250 mg/dl and HbA1c 7%–10%) managed with metformin and a lifestyle program. Each person with the risk allele (R230C) was matched by age, gender and BMI to three others with the wild type variant (R230R). Following a four week stabilization period, only participants with a greater than 70% adherence to metformin and a stable body weight were prescribed glyburide therapy for a further 16 weeks. The main outcome variable was the glyburide dose required to achieve treatment goals. Results No significant difference was observed in the glucose lowering effect of glyburide between subjects with the R230C and R230R alleles. However, the dose of sulfonylurea was significantly higher in the R230C participants compared with the R230R subjects (3.3 ± 2.1 vs 6.3 ± 5 mg/day, p

The etiology of Alzheimer's disease (AD) is complex. To date, molecular genetic studies in several families affected with AD have identified three genes associated with highly penetrant early-onset AD: Presenilin 1 (PSEN1), Presenilin 2 (PSEN2) and beta-amyloid precursor protein (APP); and one gene (apolipoprotein E) associated with late-onset AD. Molecular analysis of the PSEN1 gene was performed by direct sequencing of genomic DNA. The possible founder effect was investigated analyzing two highly polymorphic microsatellite markers flanking the PSEN1 gene. Twelve unrelated Mexican families with early-onset AD were analyzed. The Ala431Glu mutation in exon 12 of PSEN1 was found in nine (75%) of these families, which segregated showing autosomal dominant inheritance. Because all families bearing the mutation are from the State of Jalisco (located in Western Mexico), a founder effect was hypothesized. Microsatellite haplotype analysis suggested a common ancestor in these nine kindreds. In conclusion, the Ala431Glu mutation is a prevalent cause of early-onset familial Alzheimer's disease in families from the State of Jalisco, Mexico. Genetic evidence supports that it is a founder mutation descending from a single common ancestor. These findings have important implications for prompt diagnosis and genetic counseling for Mexican patients with familial AD from Jalisco.

A rare cause of congental adrenal hyperplasia is 17α-hydroxylase deficiency. It results in sexual infantilism, primary amenorrhea in females, pseudohermaphroditism in males, hypertension, and hypokalemia. We studied two female siblings from a rural community in Mexico. The cause of consultation was primary amenorrhea. The proband had low levels of estrogen, progesterone and cortisol. Deoxycorticosterone and corticosterone levels were elevated. The proband was homozygous for a transversion of cytosine to thymine at exon 4 (CGA→TGA), causing a premature stop codon at position 239 (R239X). Analysis of family members showed the presence of this heterozygous mutation in the mother, father and one healthy sibling. In summary, we describe a Mexican family with 17α-hydroxylase deficiency due to R239X mutation.

Thyroid hormone resistance is a syndrome characterized by a reduced response to thyroid hormone with different degrees of resistance at target tissues. We present the clinical features, physical findings, and study protocol in a woman with thyroid hormone resistance. An arginine to tryptophan mutation on the β isoform of the thyroid hormone receptor gene was demonstrated. Thyroid hormone resistance is an uncommon cause of thyroid dysfunction. It is necessary to perform an adequate study and confirmation to avoid an inadequate and ineffective treatment of this condition.

Background Alterations in postprandial metabolism have been described in familial combined hyperlipidemia (FCH); however, their underlying mechanisms are not well characterized. We aimed to identify factors related to the magnitude of postprandial lipemia and apolipoprotein (apo) A-V levels in subjects with FCH. Methods FCH cases (n = 99) were studied using a standardized meal test. Abdominal obesity was assessed using the waist to hip ratio (WHR). A linear regression model was performed to investigate the variables associated with the triglycerides incremental area under the curve (iAUC). Independent associations between metabolic variables and apo A-V iAUC were also investigated in a randomly selected subgroup (n = 44). The study sample was classified according to the presence of fasting hypertriglyceridemia (≥150 mg/dL) and abdominal obesity (WHR ≥0.92 in men and ≥0.85 in women) to explore differences in parameters. Results The fasting apo B-48 levels (r = 0.404), and the WHR (r = 0.359) were independent factors contributing to the triglycerides iAUC (r2 = 0.29, P

Steroid 21-hydroxylase deficiency is the underlying cause in over 90% of patients with congenital adrenal hyperplasia, an inherited metabolic disorder of adrenal steroidogenesis. We have characterized 94 mutant alleles from 47 unrelated Mexican patients and the corresponding mutant alleles in their parents by amplification of the functional CYP21 gene by PCR, followed by direct sequence analysis. The study included patients diagnosed with the three clinical forms of the disease. Our results revealed: (1) the presence of relatively few mutations or combinations of mutations associated with particular phenotypes; (2) the presence of putative new mutations; (3) the finding of identical genotypes in patients displaying discordant phenotypes; (4) the identification of patients lacking all previous reported mutations; and (5) an apparent high frequency of germ-line mutations. The absence of previously reported mutations in about 22% of the disease alleles, the finding of putative new mutations in some of the patients lacking previously known mutations, and the apparent high prevalence of germ-line mutations make evident the differences in the genetic background leading to this disorder between the Caucasian and the Mexican populations.

Most cases of congenital adrenal hyperplasia, the inherited inability to synthesize cortisol, are caused by mutations in the steroid 21-hydroxylase gene (CYP21). Steroid 21-hydroxylase deficiency is unusual among genetic diseases in that approximately 95% of the mutant alleles have apparently been generated by recombination between a normally active gene (CYP21) and a linked pseudogene (CYP21P). Approximately 20% of mutant alleles carry DNA deletions of 30 kb that have presumably been generated by unequal meiotic crossing-over, whereas 75% carry one or more mutations in CYP21 that are normally found in the CYP21P pseudogene. These latter mutations are termed "gene conversions," although the mechanism by which they are generated is not well understood. To assess the frequency at which these different recombination events occur, we have used PCR to detect de novo deletions and gene conversions in matched sperm and peripheral blood leukocyte DNA samples from normal individuals. Deletions with breakpoints in a 100-bp region in intron 2 and exon 3 were detected in sperm DNA samples with frequencies of approximately 1 in 10(5)-10(6) genomes but were never detected in the matching leukocyte DNA. Gene conversions in the same region occur in approximately 1 in 10(3)-10(5) genomes in both sperm and leukocyte DNA. These data suggest that whereas deletions occur exclusively in meiosis, gene conversions occur during both meiosis and mitosis, or perhaps only during mitosis. Thus, gene conversions must occur by a mechanism distinct from unequal crossing-over.

Several studies have identified nearly 40 different type 2 diabetes susceptibility loci, mainly in European populations, but few of them have been evaluated in the Mexican population. The aim of this study was to examine the extent to which 24 common genetic variants previously associated with type 2 diabetes are associated in Mexican Mestizos. Twenty-four single nucleotide polymorphisms (SNPs) in or near genes (KCNJ11, PPARG, TCF7L2, SLC30A8, HHEX, CDKN2A/2B, CDKAL1, IGF2BP2, ARHGEF11, JAZF1, CDC123/CAMK1D, FTO, TSPAN8/LGR5, KCNQ1, THADA, ADAMTS9, NOTCH2, NXPH1, RORA, UBQLNL, and RALGPS2) were genotyped in Mexican Mestizos. A case-control association study comprising 1,027 type 2 diabetic individuals and 990 control individuals was conducted. To account for population stratification, a panel of 104 ancestry-informative markers was analyzed. Association to type 2 diabetes was found for rs13266634 (SLC30A8), rs7923837 (HHEX), rs10811661 (CDKN2A/2B), rs4402960 (IGF2BP2), rs12779790 (CDC123/CAMK1D), and rs2237892 (KCNQ1). In addition, rs7754840 (CDKAL1) was associated in the nonobese type 2 diabetic subgroup, and for rs7903146 (TCF7L2), association was observed for early-onset type 2 diabetes. Lack of association for the rest of the variants may have resulted from insufficient power to detect smaller allele effects.

Introduction. Familial hypobetalipoproteinemia (FHBL) is an autosomal dominant disease characterized by abnormally low levels of apolipoprotein-B (apoB) containing lipoproteins. FHBL is caused by APOB, PCSK9 or ANGPTL3 mutations or is associated with loci located in chromosomes 10 and 3p21. However, other genes should be involved. This study describes the kinetic parameters of the apoB containing lipoproteins and sequence abnormalities of the APOB and PCSK9 genes of FHBL patients identified in a large hospital based survey. Material and methods. Cases with primary or secondary causes of hypobetalipoproteinemia were identified. ApoB kinetics were measured in cases with primary forms in whom truncated forms of apoB were not present in VLDL (n = 4). A primed constant infusion of [ 13 C] leucine was administered, VLDL and LDL apoB production and catabolic rates measured by a multicompartmental model and compared to normolipemic controls. In addition, these subjects had an abdominal ultrasound and direct sequencing was carried out for the PCSK9 and apoB genes. Results. Three individuals had normal apoB production with increased catabolic rate; the remaining had reduced synthetic and catabolic rates. Various polymorphisms, some of them previously unreported (*), in the PCSK9 gene (R46L, A53V, I474V, D480N*, E498K*) and in the apoB gene (N441D*, Y1395C, P2712L, D2285E*, I2286V, T3540S*, T3799M*) were found in the FHBL patients. We found hepatic ultrasound changes of hepatic steatosis in only one of the four probands. Conclusion. FHBL without truncated apoB is a heterogeneous disease from a metabolic and a genetic perspective. Hypobetalipoproteinemia is a risk factor but not an obligate cause of steatosis.

Familial combined hyperlipidemia (FCHL) is the most frequent primary dyslipidemia. Its manifestations include hypercholesterolemia, hypertriglyceridemia or the combination of both abnormalities. In spite of its high frequency, the proper diagnosis is rarely done. For this purpose, the measurement of a lipid profile is required in at least three first-degree relatives. A critical review of the current literature in this field is presented in this paper. Prospective studies have confirmed the atherogenicity of the disease. It is possible to identify the FCHL causal genes with the current methodology because it is an oligogenic disease. Based on the use of new technologies, several loci that regulate apolipoprotein B concentrations have been identified. In addition it was demostrated that variations of the activity or the expression of various nuclear factors (USF1, TCF7L2, HNF4alfa) have a major role in the pathophysiology of FCHL. These nuclear factors regulate the expression of multiple genes involved in the metabolism of lipids or carbohydrates.

To search for an association between the non-synonymous Arg230Cys variant (R230C) of the ATP-binding cassette transporter A1 and low HDL cholesterol levels in a Mexican, population-based nation wide survey.The 2000 National Health Survey is a cross sectional study that included individuals from 400 cities. All individuals who had a 9-12-h fasted blood sample and a DNA sample were selected (n = 1729). These cases were randomly distributed; no bias was detected for sex, education, region or socioeconomic status. The R230C variant was genotyped using TaqMan assays.In individuals with the R230C/C230C genotypes (39.03 mg/dl (36.63-41.43)) lower HDL-C levels (p0.001) were observed compared to those with the R230R genotype (44.7 mg/dl (43.31-46.24)). The difference remained significant after adjusting for gender, body mass index and waist circumference; the mean difference in HDL cholesterol levels between alleles was 5.73 ± 1.4 mg/dl. The magnitude of the effect was significantly greater in males. The C230 allele of ABCA1 was associated with an increased risk for hypoalphalipoproteinemia (OR 1.66 (95%CI 1.08-2.54), p0.05). The population attributable risk (PAR) for having hypoalphalipoproteinemia of the C230 allele of the ABCA1, after considering the confounding effect of waist circumference and gender, was 12.2% (95%CI 1.4-24.2%).The non-synonymous Arg230Cys variant of ABCA1 is associated with low levels of HDL cholesterol levels in Mexican adults. The HDL cholesterol lowering effect of the variant is greater in males. The size of the effect is greater compared to that reported for other ABCA1 variants.

La hiperlipidemia familiar combinada (HLFC) es la dislipidemia primarias más frecuente en México. Se manifiesta por hipercolesterolemia, hipertrigliceridemia o la combinación de estos defectos. Pese a su frecuencia, pocas veces es diagnosticada con certidumbre. Se requiere del estudio de al menos tres familiares directos para corroborar el diagnóstico. En esta revisión se describen los avances ocurridos en los últimos años sobre la fisiopatología y diagnóstico de la hiperlipidemia familiar combinada. Estudios prospectivos han confirmado la aterogenicidad de la enfermedad. Por ser una enfermedad oligogénica es factible identificar los genes involucrados con los recursos ahora existentes. Gracias al uso de nuevas tecnologías, se han identificado regiones cromosómicas que determinan la concentración de la apolipoproteína B y se demostró que diferencias en la actividad o concentración de varios factores nucleares (USF1, TCF7L2, HNF4alfa) juegan un papel importante en la fisiopatología de la HLFC. Los factores nucleares antes mencionados regulan la expresión de múltiples genes que participan en el metabolismo de lípidos y carbohidratos. Familial combined hyperlipidemia (FCHL) is the most frequent primary dyslipidemia. Its manifestations include hypercholesterolemia, hypertriglyceridemia or the combination of both abnormalities. In spite of its high frequency, the proper diagnosis is rarely done. For this purpose, the measurement of a lipid profile is required in at least three first-degree relatives. A critical review of the current literature in this field is presented in this paper. Prospective studies have confirmed the atherogenicity of the disease. It is possible to identify the FCHL causal genes with the current methodology because it is an oligogenic disease. Based on the use of new technologies, several loci that regulate apolipoprotein B concentrations have been identified. In addition it was demostrated that variations of the activity or the expression of various nuclear factors (USF1, TCF7L2, HNF4alfa) have a major role in the pathophysiology of FCHL. These nuclear factors regulate the expression of multiple genes involved in the metabolism of lipids or carbohydrates.

Genotyping for 10 mutations in the CYP21 gene was performed in 88 families with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Southern blot analysis was used to detect CYP21 deletions or large gene conversions, and allele-specific hybridizations were performed with DNA amplified by the polymerase chain reaction to detect smaller mutations. Mutations were detected on 95% of chromosomes examined. The most common mutations were an A----G change in the second intron affecting pre-mRNA splicing (26%), large deletions (21%), Ile-172----Asn (16%), and Val-281----Leu (11%). Patients were classified into three mutation groups based on degree of predicted enzymatic compromise. Mutation groups were correlated with clinical diagnosis and specific measures of in vivo 21-hydroxylase activity, such as 17-hydroxyprogesterone, aldosterone, and sodium balance. Mutation group A (no enzymatic activity) consisted principally of salt-wasting (severely affected) patients, group B (2% activity) of simple virilizing patients, and group C (10-20% activity) of nonclassic (mildly affected) patients, but each group contained patients with phenotypes either more or less severe than predicted. These data suggest that most but not all of the phenotypic variability in 21-hydroxylase deficiency results from allelic variation in CYP21. Accurate prenatal diagnosis should be possible in most cases using the described strategy.

Steroid 21-hydroxylase deficiency is the most common enzymatic defect causing congenital adrenal hyperplasia, an inherited disorder of cortisol biosynthesis. All mutations thus far characterized that cause this disorder appear to result from recombinations between the gene encoding the enzyme, CYP21B (CYP21), and the adjacent pseudogene, CYP21A (CYP21P). These are either deletions caused by unequal crossing-over during meiosis or apparent transfers of deleterious sequences from CYP21A to CYP21B, a phenomenon termed gene conversion. However, a small percentage of alleles do not carry such a mutation. We analyzed DNA from a patient with the mild, nonclassic form of 21-hydroxylase deficiency, who carried one allele that had no gene conversions detectable by hybridization with oligonucleotide probes. Sequence analysis revealed that this allele carried two missense mutations, R339H and P453S, neither of which has been previously observed in CYP21A or CYP21B. Each of these mutations was introduced into CYP21 cDNA which was then expressed in COS1 cells using a vaccinia virus system. Each mutation reduced the ability of the enzyme to 21-hydroxylate 17-hydroxyprogesterone to 50% of normal and the ability to metabolize progesterone to 20% of normal. Thus, each of these mutations represents a potential nonclassic 21-hydroxylase deficiency allele that is not the result of an apparent gene conversion.

Numerous efforts for the development of basic and clinical research in obesity are being made by the National Institutes of Health and Federal Reference Hospitals in Mexico. However, greater interaction among researchers and stronger efforts towards the dissemination of the results are needed. The document outlines the general ideas and proposals of the Academic Group for the Study, Prevention and Treatment of Obesity and Metabolic Syndrome of the Coordinating Committee of the National Institutes of Health and High Specialty Hospitals (CCINSHAE). This is the first step in developing common objectives, with the aim of understanding the effect of these entities in public health and to establish guidelines to limit and eventually overcome them. We discuss the appropriateness of analyzing obesity and the metabolic syndrome together, and the current management of these entities at the National Institutes of Health in Mexico. The problems that arise in clinical practice lead to the need to generate a new model of medical care, including a new health worker and a new patient. It is imperative to establish permanent lines of communication and education with health personnel and with patients. The group proposes an integrated approach for research in these areas. Finally, a master plan that links the National Institutes of Health, particularly in the areas of research and programs within the institutions, is required as a first step in seeking answers useful in solving the problem. The second step would be linking the first and second levels of care through concrete actions needed to limit and reduce obesity and metabolic syndrome in the population.

The steroid 11 beta-hydroxylase (P450c11) enzyme is responsible for the conversion of 11-deoxycortisol to cortisol in the zona fasciculata of the adrenal cortex. Animal studies have suggested that this enzyme or a closely related isozyme is also responsible for the successive 11 beta- and 18-hydroxylation and 18-oxidation of deoxycorticosterone required for aldosterone synthesis in the zona glomerulosa. There are two distinct 11 beta-hydroxylase genes in man, CYP11B1 and CYP11B2, which are predicted to encode proteins with 93% amino acid identity. We used a sensitive assay based on the polymerase chain reaction to analyze the expression of the CYP11B1 and B2 genes. Transcripts of CYP11B1 were detected at high levels in surgical specimens of normal adrenals and also in an aldosterone-secreting adrenal tumor. Transcripts of CYP11B2 were found at low levels in normal adrenals, but at a much higher level in the aldosterone-secreting tumor. CYP11B2 mRNA levels were increased in cultured zona glomerulosa cells by physiological levels of angiotensin-II. The entire coding regions of both CYP11B1 and B2 cDNAs were cloned from the tumor mRNA. Expression of these cDNAs in cultured COS-1 cells demonstrated that the CYP11B1 product could only 11 beta-hydroxylate 11-deoxycortisol or deoxycorticosterone, whereas the CYP11B2 product could also 18-hydroxylate cortisol or corticosterone. A small amount of aldosterone was synthesized from deoxycorticosterone only in cells expressing CYP11B2 cDNA. These data demonstrate that the product of CYP11B2 is required for the final steps in the synthesis of aldosterone.

To describe the contributions ofthree Mexican institutions (Instituto Nacional de Ciencias Médicas y Nutrición "Salvador Zubirán", Universidad Nacional Autónoma de México and the Centro de Encuestas Nacionales de Salud of the Instituto Nacional de Salud Pública) in the study, of chronic degenerative disorders.The most relevant group contributions include: the identification of a risk allele for metabolic syndrome and diabetes, specific for the Mexican population (the R230C variant of the ABC-A1 transporter); the design and validation of a population-based definition of metabolic syndrome which is useful to predict the risk of incident diabetes; the description of the molecular epidemiology of familial hypercholesterolemia in Mexico and the identification of several loci associated with familial combined hyperlipidemia. In addition, members of these institutions have participated in the description of the epidemiology of diabetes, metabolic syndrome and lipid abnormalities.The complementary approach of these research groups has facilitated successful collaborations. Our results will be useful for the future development of diagnostic tests and preventive programs.

Steroid 21-hydroxylase (P450c21) is absent or defective in more than 90% of patients with congenital adrenal hyperplasia. This disorder of cortisol biosynthesis occurs in a wide spectrum of clinical severity; specific mutations in the 21-hydroxylase gene (CYP21) have been found in association with particular clinical phenotypes. To determine the functional effects of mutations causing amino acid substitutions, normal P450c21 and three mutagenized P450c21 enzymes were expressed at high levels in cultured COS-1 cells using recombinant vaccinia virus. A single amino acid substitution (Val281----Leu) present in patients with mild "nonclassical" 21-hydroxylase deficiency resulted in an enzyme with 20-50% of normal activity. A mutation (Ile172----Asn) identified in patients with the "simple virilizing" form (poor cortisol synthesis but adequate aldosterone synthesis) resulted in an enzyme with less than 2% of normal activity. Finally, a cluster mutation (Ile-Val-Glu-Met234-238----Asn-Glu-Glu-Lys) found in a patient with severe "salt wasting" 21-hydroxylase deficiency (inadequate aldosterone synthesis) results in an enzyme with no detectable activity. These data indicate that the severity of 21-hydroxylase deficiency correlates with the degree of enzymatic compromise.

Background— Congenital long-QT syndrome (LQTS) is potentially lethal secondary to malignant ventricular arrhythmias and is caused predominantly by mutations in genes that encode cardiac ion channels. Nearly 25% of patients remain without a genetic diagnosis, and genes that encode cardiac channel regulatory proteins represent attractive candidates. Voltage-gated sodium channels have a pore-forming α-subunit associated with 1 or more auxiliary β-subunits. Four different β-subunits have been described. All are detectable in cardiac tissue, but none have yet been linked to any heritable arrhythmia syndrome. Methods and Results— We present a case of a 21-month-old Mexican-mestizo female with intermittent 2:1 atrioventricular block and a corrected QT interval of 712 ms. Comprehensive open reading frame/splice mutational analysis of the 9 established LQTS-susceptibility genes proved negative, and complete mutational analysis of the 4 Na vβ -subunits revealed a L179F (C535T) missense mutation in SCN4B that cosegregated properly throughout a 3-generation pedigree and was absent in 800 reference alleles. After this discovery, SCN4B was analyzed in 262 genotype-negative LQTS patients (96% white), but no further mutations were found. L179F was engineered by site-directed mutagenesis and heterologously expressed in HEK293 cells that contained the stably expressed SCN5A -encoded sodium channel α-subunit (hNa V 1.5). Compared with the wild-type, L179F-β4 caused an 8-fold (compared with SCN5A alone) and 3-fold (compared with SCN5A + WT-β4) increase in late sodium current consistent with the molecular/electrophysiological phenotype previously shown for LQTS-associated mutations. Conclusions— We provide the seminal report of SCN4B -encoded Na vβ 4 as a novel LQT3-susceptibility gene.

BACKGROUND: The cytogenetic aspects of acute leukemia have been studied for over 30 years. Genomic sequencing has permitted the identification of recurrent mutations that have been associated with the heterogeneous behavior of acute leukemia. Currently, the genomic profile of patients with acute leukemia is important regarding risk classification and therapeutic approach. At this moment there are scarce studies of the genomic profile in adult patients with acute leukemia in Mexico. OBJECTIVES: To describe genomic alterations found in adult patients with acute leukemia in a cancer reference hospital in Mexico. Evaluate the efficacy of a commercial multiple PCR and nested qualitative PCR kit (HemaVision DNA Technology A/S, HV01-28N), for the detection of 28 translocations on acute leukemia. To assess by a quantitative method with RT-PCR the detection of the most frequently reported mutations in acute myeloid leukemia(38%) [t(15;17) PML-RARa, inv(16) CBF-MYH11, t(8;21) RUNX1-RUNX1T1 and acute lymphoid leukemia (32%) [t(9;22) BCR/ABL, t(4;11) MLL-AFF1, t(1;19) TCF3-PBX1]. Compare the results between the commercial kit, RT-PCR and conventional cytogenetic studies. METHODS: Patients with recent diagnosis of acute leukemia (myeloid and lymphoid) according to World Health Administration (WHO) 2008 criteria were included between the period of 03/25/2013 and 12/15/13. All patients were given informed consent form and approval of the local Ethics Committee We performed bone marrow smear and bone marrow biopsy of all patients. 5 mL of bone marrow were collected on heparin for cytogenetic study including karyotipe and FISH analysis. 5 mL of bone marrow were collected for molecular biology studies including PCR-kit and genome sequencing technique. 5mL of peripheral blood with heparin were obtained for karyotype in case of not finding metaphases for the analysis on bone marrow specimen. RESULTS: 24 patients were enrolled on the study. The mean age was 42 years (17-69). 8/24 were male (33.3%) and 16/24 were female (66.6%). Acute myeloid leukemia 11/24 (45.8%), acute lymphoblastic leukemia 8/24 (33.3%), acute promyeolocytic leukemia 3/24 (12.5%), acute biphenotypic leukemia 1/24 (4.2%) and one case of blastic phase of chronic myeloid leukemia (4.2%). The commercial PCR platform detected only two of the 28 translocations in 4/24 patients (16.6%). 2/24 (8.3%) BCR-ABL and 2/24 (8.3%) PML-RARa. In contrast to RT-PCR that detected 7/24 patients (29%). 4/24 (16%) BCR-ABL, 2/24 (8.3%) PML-RARa and 1/24 (4.1%) CBFB-MYH11. FISH analysis detected the 4/24 (16%) translocations of the commercial kit. Karyotype detected structural chromosomal anomalies in 9/24 patients (37.5%). After the analysis of the 24 samples by quantitative RT-PCR for the six most frequent rearrangements, we also identified 2 additional cases of BCR-ABL. CONCLUSIONS: The commercial qualitative PCR kit was inferior on detecting genomic alterations (16.6%) than quantitative PCR (29%) or conventional cytogenetic study (37.5%). The RT-PCR quantitative method with specific probe for the most frequent genomic alterations in acute leukemia in addition to cytogenetic analysis seems to be a better cost-effective strategy than the commercial multiple PCR platforms. Disclosures No relevant conflicts of interest to declare.

GENEHUNTER and SimWalk2 are among the most commonly used software for parametric multipoint linkage analysis. In the context of extended kindred analysis, GENEHUNTER has a limitation in terms of the number of individuals it can handle. One solution is to manually split the kindred into smaller pedigrees. SimWalk2 can handle a much larger number of individuals. However, its major drawback is the time it takes to process the data when compared to GENEHUNTER. Aside from the limitations of each program, when studying extended kindreds researchers are typically confronted with missing data. In this work we used simulated genotype data based on the structure of a real extended pedigree in order to compare the results obtained through GENEHUNTER and SimWalk2, evaluate the effect of discarding individuals and splitting the kindred on the logarithm of odds (lod) score, and to assess how missing data affect the performance of each program. Our results show that (1) for pedigrees of a moderate size, GENEHUNTER and SimWalk2 produce nearly the same results; (2) when using GENEHUNTER, either splitting the kindred into smaller sub-pedigrees or discarding individuals has an adverse effect when compared to the results obtained when using SimWalk2 with the whole pedigree; and (3) the performance of both programs is qualitatively similar in the missing data scenario. These conclusions are based on the sample distributions of the lod score values and of the estimates of the recombination fraction.