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Syphilis is a chronic infectious disease, which dates back to the XV century and is caused by the spirochete treponema pallidum, capable of invading the central nervous system in any of its stages- Its incidence has increased in parallel to the HIV/AIDS pandemic, and the synergism between both pathologies is such. that it has become a public health problem in recent years. Here we present the case of a 31-year-old female patient, who consulted for headache associated with decreased visual acuity and provided an unenhanced head CT showing hypodense lesions in both thalamic regions, serological tests for syphilis were reactive and those for HIV were not reactive. The brain MRI with spectroscopy was reported in favor of cerebral toxoplasmosis, which was later ruled out with a study of cerebrospinal fluid. Management with penicillin G sodium IV for 6 weeks was indicated, achieving complete imaging resolution of her lesions., (© 2024 The Authors. Published by Elsevier Inc. on behalf of University of Washington.)

Introduction: Artificial intelligence (AI) can automate certain tasks to improve data collection. Models have been created to annotate the steps of Roux-en-Y Gastric Bypass (RYGB). However, model performance has not been compared with individual surgeon annotator performance. We developed a model that automatically labels RYGB steps and compares its performance to surgeons., Methods and Procedures: 545 videos (17 surgeons) of laparoscopic RYGB procedures were collected. An annotation guide (12 steps, 52 tasks) was developed. Steps were annotated by 11 surgeons. Each video was annotated by two surgeons and a third reconciled the differences. A convolutional AI model was trained to identify steps and compared with manual annotation. For modeling, we used 390 videos for training, 95 for validation, and 60 for testing. The performance comparison between AI model versus manual annotation was performed using ANOVA (Analysis of Variance) in a subset of 60 testing videos. We assessed the performance of the model at each step and poor performance was defined (F1-score < 80%)., Results: The convolutional model identified 12 steps in the RYGB architecture. Model performance varied at each step [F1 > 90% for 7, and > 80% for 2]. The reconciled manual annotation data (F1 > 80% for > 5 steps) performed better than trainee's (F1 > 80% for 2-5 steps for 4 annotators, and < 2 steps for 4 annotators). In testing subset, certain steps had low performance, indicating potential ambiguities in surgical landmarks. Additionally, some videos were easier to annotate than others, suggesting variability. After controlling for variability, the AI algorithm was comparable to the manual (p < 0.0001)., Conclusion: AI can be used to identify surgical landmarks in RYGB comparable to the manual process. AI was more accurate to recognize some landmarks more accurately than surgeons. This technology has the potential to improve surgical training by assessing the learning curves of surgeons at scale., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Head and neck cancer is the sixth most frequent cancer type. Drug resistance and toxicity are common challenges of the existing therapies, making the development of reliable preclinical models essential for the study of the involved molecular mechanisms as well as for eventual intervention approaches that improve the clinical outcome. Preclinical models of head and neck squamous cell carcinoma have been traditionally based on cell lines and murine models. In this review, we will go over the most frequently used preclinical models, from immortalised-cell and primary tumour cultures in monolayer or 3D, to the currently available animal models. We will scrutinise their efficiency in mimicking the molecular and cellular complexity of head and neck squamous cell carcinoma. Finally, the challenges and the opportunities of other envisaged putative approaches, as well as the potential of the preclinical models to further develop personalised therapies will be discussed., (© 2023. The Author(s).)

Purpose: The percentage of patients with metastatic non-small cell lung cancer (NSCLC) and melanoma who benefit from anti-programmed cell death protein 1 (anti-PD-1) is low owing to resistance mechanisms. SABR has a role in oligoprogressive disease and can improve responses to anti-PD-1. This multicenter prospective observational study aimed to determine whether concomitant anti-PD-1 and SABR to oligoprogressive sites enhance tumor response in metastatic NSCLC and melanoma., Methods and Materials: Patients with metastatic NSCLC or melanoma in progression to anti-PD-1 but continuing the same line owing to clinical benefit were referred for palliative SABR. All patients received concomitant pembrolizumab or nivolumab and SABR to 1 to 5 lesions, maintaining anti-PD-1 until further progression, unacceptable toxicity, or medical/patient decision. Objective response rate-complete responses and partial responses-was evaluated during all follow-up according to Response Evaluation Criteria in Solid Tumors 1.1. The abscopal response was evaluated 8 weeks after SABR as a ≥30% reduction in 1 to 2 predefined nonirradiated lesions., Results: Of the 61 patients enrolled, 50 could be analyzed. With a median follow-up of 32.8 months, objective response rate was 42% (30% complete responses and 12% partial responses). Median progression-free survival was 14.2 months (95% confidence interval, 6.9-29 months). Median overall survival since SABR was 37.4 months (95% confidence interval, 22.9 months-not reached). Abscopal response was 65%, evaluated in 40 patients who fulfilled the criteria., Conclusions: Combined anti-PD-1 and SABR in oligoprogressive metastatic NSCLC or melanoma can achieve high rates of response and extend the clinical benefit of immunotherapy by delaying further progression and a new systemic therapy. This approach should be assessed in larger randomized trials., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Cancer is the second leading cause of death worldwide being responsible for 9.6 million deaths in 2018. Epigenetic alterations are key in directing the aberrant expression of tumor-associated genes that drive cellular malignant transformation and cancer progression. Among epigenetic alterations, DNA methylation is the most deeply studied one in relation to environmental exposure. Tissue biopsies have traditionally been the main procedure by which a small sample of body tissue is excised to confirm cancer diagnosis or to indicate the primary site when cancer has spread. In contrast, the analysis of circulating tumor-derived material, or tumor circulome, by means of liquid biopsy of peripheral blood, urine, saliva or sputum is a noninvasive, fast and reproducible alternative to tissue biopsy. Recently, the assessment of epigenetic alterations such as DNA methylation and hydroxymethylation in circulating free DNA has been proved possible. These marks can be associated to prognosis and response to a variety of treatments including chemotherapy, hormonotherapy or immunotherapy. Epigenetic biomarkers may offer some advantages over RNA or genetic biomarkers given their stability in bodily fluids and their high tissue-specificity. While many challenges are still ahead, the unique advantages of these types of biomarkers is urging the scientific community to persevere in their clinical validation and integration into reliable prediction models. This review aims at recapitulating the emerging noninvasive DNA methylated biomarkers of importance for prediction of prognosis and drug response in cancer., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: To better comprehend transcriptional phenotypes of cancer cells, we globally characterised RNA-binding proteins (RBPs) to identify altered RNAs, including long non-coding RNAs (lncRNAs)., Design: To unravel RBP-lncRNA interactions in cancer, we curated a list of ~2300 highly expressed RBPs in human cells, tested effects of RBPs and lncRNAs on patient survival in multiple cohorts, altered expression levels, integrated various sequencing, molecular and cell-based data., Results: High expression of RBPs negatively affected patient survival in 21 cancer types, especially hepatocellular carcinoma (HCC). After knockdown of the top 10 upregulated RBPs and subsequent transcriptome analysis, we identified 88 differentially expressed lncRNAs, including 34 novel transcripts. CRISPRa-mediated overexpression of four lncRNAs had major effects on the HCC cell phenotype and transcriptome. Further investigation of four RBP-lncRNA pairs revealed involvement in distinct regulatory processes. The most noticeable RBP-lncRNA connection affected lipid metabolism, whereby the non-canonical RBP CCT3 regulated LINC00326 in a chaperonin-independent manner. Perturbation of the CCT3- LINC00326 regulatory network led to decreased lipid accumulation and increased lipid degradation in cellulo as well as diminished tumour growth in vivo ., Conclusions: We revealed that RBP gene expression is perturbed in HCC and identified that RBPs exerted additional functions beyond their tasks under normal physiological conditions, which can be stimulated or intensified via lncRNAs and affected tumour growth., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY. Published by BMJ.)

Neoadjuvant Chemotherapy (NAC) in Breast Cancer (BC) has proved useful for the reduction in tumor burden prior to surgery, allowing for a more extensive breast preservation and the eradication of subjacent micrometastases. However, the impact on prognosis is highly dependent on the establishment of Pathological Complete Response (pCR), in particular for Triple Negative (TN) and Hormonal Receptor negative/Human Epidermal growth factor Receptor 2 positive (HR-/HER2+) subtypes. Several pCR predictors, such as PAM50, Integrative Cluster (IntClust), mutations in PI3KCA, or the Trastuzumab Risk model (TRAR), are useful molecular tools for estimating response to treatment and are prognostic. Major evolution events during BC NAC that feature the Residual Disease (RD) are the loss of HR and HER2, which are prognostic of bad outcome, and stemness and immune depletion-related gene expression aberrations. This dynamic nature of the determinants of response to BC NAC, together with the extensive heterogeneity of BC, raises the need to discern the individual and subtype-specific determinants of resistance. Moreover, refining the current approaches for a comprehensive monitoring of tumor evolution during treatment, RD, and eventual recurrences is essential for identifying new actionable alterations and the integral best management of the disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Recently, it was published that CYP3A5 contributes to chemotherapeutic drug resistance in a wide range of solid tumors, including hepatocellular carcinoma. However, CYP3A5 is highly polymorphic and 90% of Caucasians are homozygous for the loss-of-function allele CYP3A5*3 . Here, we evaluate the relationship between CYP3A5 genotype and expression level of both CYP3A5 transcripts and protein in biopsies from 19 pairs of liver tumors and corresponding peritumoral tissue. We find that CYP3A5 transcript levels are reduced compared with peritumoral controls. Moreover, we do not detect CYP3A5 protein in homozygous CYP3A5*3 carriers and no relative increase of CYP3A5 in tumoral tissue of CYP3A5*1 carriers. We conclude that anticancer drug resistance is unlikely to be caused by increased CYP3A5 expression.

Background: Anti-programmed death-1 (PD-1) treatment for advanced non-small-cell lung cancer (NSCLC) has improved the survival of patients. However, a substantial percentage of patients do not respond to this treatment. We examined the use of DNA methylation profiles to determine the efficacy of anti-PD-1 treatment in patients recruited with current stage IV NSCLC., Methods: In this multicentre study, we recruited adult patients from 15 hospitals in France, Spain, and Italy who had histologically proven stage IV NSCLC and had been exposed to PD-1 blockade during the course of the disease. The study structure comprised a discovery cohort to assess the correlation between epigenetic features and clinical benefit with PD-1 blockade and two validation cohorts to assess the validity of our assumptions. We first established an epigenomic profile based on a microarray DNA methylation signature (EPIMMUNE) in a discovery set of tumour samples from patients treated with nivolumab or pembrolizumab. The EPIMMUNE signature was validated in an independent set of patients. A derived DNA methylation marker was validated by a single-methylation assay in a validation cohort of patients. The main study outcomes were progression-free survival and overall survival. We used the Kaplan-Meier method to estimate progression-free and overall survival, and calculated the differences between the groups with the log-rank test. We constructed a multivariate Cox model to identify the variables independently associated with progression-free and overall survival., Findings: Between June 23, 2014, and May 18, 2017, we obtained samples from 142 patients: 34 in the discovery cohort, 47 in the EPIMMUNE validation cohort, and 61 in the derived methylation marker cohort (the T-cell differentiation factor forkhead box P1 [FOXP1]). The EPIMMUNE signature in patients with stage IV NSCLC treated with anti-PD-1 agents was associated with improved progression-free survival (hazard ratio [HR] 0·010, 95% CI 3·29 × 10 -4 -0·0282; p=0·0067) and overall survival (0·080, 0·017-0·373; p=0·0012). The EPIMMUNE-positive signature was not associated with PD-L1 expression, the presence of CD8+ cells, or mutational load. EPIMMUNE-negative tumours were enriched in tumour-associated macrophages and neutrophils, cancer-associated fibroblasts, and senescent endothelial cells. The EPIMMUNE-positive signature was associated with improved progression-free survival in the EPIMMUNE validation cohort (0·330, 0·149-0·727; p=0·0064). The unmethylated status of FOXP1 was associated with improved progression-free survival (0·415, 0·209-0·802; p=0·0063) and overall survival (0·409, 0·220-0·780; p=0·0094) in the FOXP1 validation cohort. The EPIMMUNE signature and unmethylated FOXP1 were not associated with clinical benefit in lung tumours that did not receive immunotherapy., Interpretation: Our study shows that the epigenetic milieu of NSCLC tumours indicates which patients are most likely to benefit from nivolumab or pembrolizumab treatments. The methylation status of FOXP1 could be associated with validated predictive biomarkers such as PD-L1 staining and mutational load to better select patients who will experience clinical benefit with PD-1 blockade, and its predictive value should be evaluated in prospective studies., Funding: "Obra Social" La Caixa, Cellex Foundation, and the Health and Science Departments of the Generalitat de Catalunya., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Pharmacological treatment and exposure to xenobiotics can cause substantial changes in epigenetic signatures. The majority of these epigenetic changes, caused by the compounds in question, occur downstream of transcriptional activation mechanisms, whereby the epigenetic alterations can create a transcriptional memory and stably modulate cell function. The increasing understanding of epigenetic mechanisms and their importance in disease has prompted the development of therapeutic interventions that target epigenetic modulatory mechanisms, particularly in oncology where inhibitors of epigenetic-modifying proteins (epidrugs) have been successfully used in treatment, mostly in combination with standard-of-care chemotherapy, either provoking direct cytotoxicity or inhibiting resistance to anticancer drugs. In addition, emerging methods for detecting epigenetically modified DNA in bodily fluids may provide information about tumor phenotype or drug treatment success. However, it is important to note that many technical pitfalls, such as the nondeconvolution of methylcytosine and hydroxymethylcytosine, compromise epigenetic analyses and the interpretation of results. In this review, we provide an update on the field, with an emphasis on the novel therapeutic opportunities made possible by epidrugs.

To improve the epigenomic analysis of tissues rich in 5-hydroxymethylcytosine (hmC), we developed a novel protocol called TAB-Methyl-SEQ, which allows for single base resolution profiling of both hmC and 5-methylcytosine by targeted next-generation sequencing. TAB-Methyl-SEQ data were extensively validated by a set of five methodologically different protocols. Importantly, these extensive cross-comparisons revealed that protocols based on Tet1-assisted bisulfite conversion provided more precise hmC values than TrueMethyl-based methods. A total of 109 454 CpG sites were analyzed by TAB-Methyl-SEQ for mC and hmC in 188 genes from 20 different adult human livers. We describe three types of variability of hepatic hmC profiles: (i) sample-specific variability at 40.8% of CpG sites analyzed, where the local hmC values correlate to the global hmC content of livers (measured by LC-MS), (ii) gene-specific variability, where hmC levels in the coding regions positively correlate to expression of the respective gene and (iii) site-specific variability, where prominent hmC peaks span only 1 to 3 neighboring CpG sites. Our data suggest that both the gene- and site-specific components of hmC variability might contribute to the epigenetic control of hepatic genes. The protocol described here should be useful for targeted DNA analysis in a variety of applications., (© The Author 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Statin treatment of hypercholesterolemia can lead to chronic myotoxicity which is, in most cases, alleviated by drug withdrawal. Cellular and molecular mechanisms of this adverse effect have been elusive, in particular because of the lack of in vitro models suitable for long-term exposures. We have taken advantage of the properties of human pluripotent stem cell-derived mesodermal precursors, that can be maintained unaltered in vitro for a long period of time, to develop a model of repeated exposures to simvastatin during more than 2 weeks. This approach unveiled major differences, both in functional and molecular terms, in response to single versus repeated-dose exposures to simvastatin. The main functional effect of the in vitro simvastatin-induced long-term toxicity was a loss of proliferative capacity in the absence of concomitant cell death, revealing that cytostatic effect could be a major contributor to statin-induced myotoxicity. Comparative analysis of molecular modifications induced by simvastatin short-term versus prolonged exposures demonstrated powerful adaptive cell responses, as illustrated by the dramatic decrease in the number of differentially expressed genes, distinct biological pathway enrichments, and distinct patterns of nutrient transporters expressed at the cell surface. This study underlines the potential of derivatives of human pluripotent stem cells for developing new approaches in toxicology, in particular for chronic toxicity testing., (© 2015 AlphaMed Press.)

Background: The liver plays a central role in the maintenance of homeostasis and health in general. However, there is substantial inter-individual variation in hepatic gene expression, and although numerous genetic factors have been identified, less is known about the epigenetic factors., Results: By analyzing the methylomes and transcriptomes of 14 fetal and 181 adult livers, we identified 657 differentially methylated genes with adult-specific expression, these genes were enriched for transcription factor binding sites of HNF1A and HNF4A. We also identified 1,000 genes specific to fetal liver, which were enriched for GATA1, STAT5A, STAT5B and YY1 binding sites. We saw strong liver-specific effects of single nucleotide polymorphisms on both methylation levels (28,447 unique CpG sites (meQTL)) and gene expression levels (526 unique genes (eQTL)), at a false discovery rate (FDR) < 0.05. Of the 526 unique eQTL associated genes, 293 correlated significantly not only with genetic variation but also with methylation levels. The tissue-specificities of these associations were analyzed in muscle, subcutaneous adipose tissue and visceral adipose tissue. We observed that meQTL were more stable between tissues than eQTL and a very strong tissue-specificity for the identified associations between CpG methylation and gene expression., Conclusions: Our analyses generated a comprehensive resource of factors involved in the regulation of hepatic gene expression, and allowed us to estimate the proportion of variation in gene expression that could be attributed to genetic and epigenetic variation, both crucial to understanding differences in drug response and the etiology of liver diseases.

Human pluripotent stem cells (hPSC) have the potential to become important tools for the establishment of new models for in vitro drug testing of, for example, toxicity and pharmacological effects. Late-stage attrition in the pharmaceutical industry is to a large extent caused by selection of drug candidates using nonpredictive preclinical models that are not clinically relevant. The current hepatic in vivo and in vitro models show clear limitations, especially for studies of chronic hepatotoxicity. For these reasons, we evaluated the potential of using hPSC-derived hepatocytes for long-term exposure to toxic drugs. The differentiated hepatocytes were incubated with hepatotoxic compounds for up to 14 days, using a repeated-dose approach. The hPSC-derived hepatocytes became more sensitive to the toxic compounds after extended exposures and, in addition to conventional cytotoxicity, evidence of phospholipidosis and steatosis was also observed in the cells. This is, to the best of our knowledge, the first report of a long-term toxicity study using hPSC-derived hepatocytes, and the observations support further development and validation of hPSC-based toxicity models for evaluating novel drugs, chemicals, and cosmetics., (Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.)

There are pronounced interindividual variations in drug metabolism, drug responses, and the incidence of adverse drug reactions. To a certain extent such variability can be explained by genetic factors, but epigenetic modifications, which are relatively scarcely described so far, also contribute. It is known that a novel class of drugs termed epidrugs intervene in the epigenetic control of gene expression, and many of these are now in clinical trials for disease treatment. In addition, disease prognosis and drug treatment success can be monitored using epigenetic biomarkers. Here we review these novel aspects in pharmacology and address intriguing future opportunities for gene-specific epigenetic editing., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Aim: This study aims to investigate the possible association between the risk of colorectal cancer (CRC) and allelic variants of CYP2W1 and their functional properties., Materials & Methods: The distribution of three different CYP2W1 alleles (CYP2W1*1, CYP2W1*2 and CYP2W1*6) in 1785 CRC patients and 1761 healthy blood donors was determined using the TaqMan(®) (Applied Biosystems, CA, USA) allelic discrimination assay or allele-specific amplification. Corresponding gene products (CYP2W1.1, CYP2W1.2 and CYP2W1.6) were expressed in human colon cancer SW480 cells and their activities towards two different substrates, the duocarmycin analogs ICT2706 and ICT2726, were monitored., Results: No significant differences in the distribution of CYP2W1*1, CYP2W1*2 and CYP2W1*6 alleles were found between CRC patients and controls. The CYP2W1.1, CYP2W1.2 and CYP2W1.6 variant enzymes were expressed at the similar levels in the transfected SW480 cells and had comparable kinetics in terms of the metabolism of the duocarmycin ICT2726, as well as in the bioactivation of ICT2706 into a cytotoxic product., Conclusion: These epidemiological data obtained from a large population of CRC patients and controls cannot confirm the previously suggested decreased risk for CRC among carriers of CYP2W1*2. On the molecular level, this conclusion is further supported by the similar catalytic characteristics of the CYP2W1.1, CYP2W1.2 and CYP2W1.6 variants of CYP2W1.

Background: Interindividual differences in liver functions such as protein synthesis, lipid and carbohydrate metabolism and drug metabolism are influenced by epigenetic factors. The role of the epigenetic machinery in such processes has, however, been barely investigated. 5-hydroxymethylcytosine (5hmC) is a recently re-discovered epigenetic DNA modification that plays an important role in the control of gene expression., Results: In this study, we investigate 5hmC occurrence and genomic distribution in 8 fetal and 7 adult human liver samples in relation to ontogeny and function. LC-MS analysis shows that in the adult liver samples 5hmC comprises up to 1% of the total cytosine content, whereas in all fetal livers it is below 0.125%. Immunohistostaining of liver sections with a polyclonal anti-5hmC antibody shows that 5hmC is detected in most of the hepatocytes. Genome-wide mapping of the distribution of 5hmC in human liver samples by next-generation sequencing shows significant differences between fetal and adult livers. In adult livers, 5hmC occupancy is overrepresented in genes involved in active catabolic and metabolic processes, whereas 5hmC elements which are found in genes exclusively in fetal livers and disappear in the adult state, are more specific to pathways for differentiation and development., Conclusions: Our findings suggest that 5-hydroxymethylcytosine plays an important role in the development and function of the human liver and might be an important determinant for development of liver diseases as well as of the interindividual differences in drug metabolism and toxicity.

Background: There is ongoing interest in the possible interaction of the serotonin-transporter-linked polymorphic region (5-HTTLPR) with environmental factors in determining Major Depressive Disorder (MDD). The current study contributes to this research area by comprehensively examining the interaction-effects and direct-effects of 5-HTTLPR and five environmental factors on MDD prevalence and course in a well-characterized longitudinal sample., Methods: The sample consisted of 1625 patients with a CIDI-confirmed diagnosis of MDD and 1698 screened controls from the Netherlands. Four MDD outcomes were studied as dependent variables: one main MDD prevalence-outcome (all MDD), two more severe MDD prevalence-outcomes (suicidal and chronic MDD), and one MDD course outcome (chronic versus non-chronic MDD). Because SNP rs25531 modifies the effect of 5-HTTLPR, haplotypes of 5-HTTLPR and rs25531 were measured. For the four MDD outcome measures, we examined the direct effects of 5-HTTLPR/rs25531-haplotypes, five environmental factors (lifetime and recent stressful life-events, sexual abuse, low educational attainment, and childhood trauma) and their interaction in logistic regression models., Results: The environmental factors had large and consistent effects on all four MDD outcomes, including course of MDD. The 5-HTTLPR/rs25531-haplotype had a suggestive effect on course of MDD, but not on presence of MDD. Gene-by-environment interaction was significant (<0.05) for one of the 20 tests performed, which is not more than expected by chance., Limitations: Environmental factors were not assessed before the onset of MDD., Conclusions: Environmental factors had a strong impact on the presence and course of MDD, but no evidence for gene-by-environment interaction was found., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Purpose: Recently, a novel gene was cloned for autosomal recessive retinitis pigmentosa (arRP), EYS, on 6q12. This study was conducted to determine the spectrum and frequency of EYS mutations in 195 unrelated patients with autosomal recessive and autosomal dominant RP (adRP)., Methods: All cases had a complete ophthalmic examination, and the clinical diagnosis of RP was based on visual acuity, fundus photography, and electroretinography findings. The DNA extracted from all participants was subjected to molecular genetic analysis entailing amplification of the coding regions and exon-intron boundaries of EYS by polymerase chain reaction, followed by direct sequencing. Bioinformatics analysis was undertaken to study the effect of the identified mutations on protein structure and function., Results: Eleven novel missense, nonsense, and splice site mutations were identified within EYS in 10 unrelated arRP patients, with probable allele frequency of 11%. However, no mutations were observed in the adRP panel. In addition, 53 single-nucleotide polymorphisms (SNPs) were found, of which 12 were previously unreported. Bioinformatics analyses revealed that all mutations were highly conserved across other species and/or involved important domains on protein structure. Intrafamilial phenotypic variability was also observed in a family with double heterozygous mutations., Conclusions: This is the first report of molecular genetic analysis of EYS in a cohort of unrelated British and Chinese patients with RP. The results further the initial hypothesis that EYS is a major causative gene for recessive RP and emphasize the role of different types of mutations in disrupting the function of EYS.

Autosomal-recessive retinitis pigmentosa (arRP) was recently associated with mutations in a novel gene EYS, spanning over 2 Mb, making it the largest known gene expressed in the human eye. The purpose of this study was to establish the prevalence and nature of EYS mutations in a clinically well-characterized cohort of 239 sporadic and arRP French cases. Direct sequencing of EYS was performed in 186 subjects for whom known mutations had previously been excluded by applying microarray technology. We mostly identified novel mutations in EYS in a total of 29 patients: Fifteen of the mutations were predicted to create premature stop codons and two represent exonic deletions. In addition, twenty missense, silent or splice-site mutations were detected. Patients revealed homozygous or compound heterozygous mutations and in some cases, only a single mutation. Most patients showed classical signs of RP with relatively preserved central vision and visual field until late in the course of the disorder. One patient showed predominance of the disease in the inferior part of the retina suggesting potential phenotypic variability. With a prevalence of 12% or more we provide evidence that EYS is a major gene for RP in France and probably elsewhere., ((c) 2010 Wiley-Liss, Inc.)

Using a positional cloning approach supported by comparative genomics, we have identified a previously unreported gene, EYS, at the RP25 locus on chromosome 6q12 commonly mutated in autosomal recessive retinitis pigmentosa. Spanning over 2 Mb, this is the largest eye-specific gene identified so far. EYS is independently disrupted in four other mammalian lineages, including that of rodents, but is well conserved from Drosophila to man and is likely to have a role in the modeling of retinal architecture.

Earlier studies have indicated that the degradation rate of poly(lactic acid) (PDLLA) can be modified by using 2,2'-bis(2-oxazoline) as a chain extender in polymer synthesis to form a lactic acid-based poly(ester-amide) (PEA). In the present study, the effect of an incorporated drug on the degradation rate of the PEA was evaluated. The model drugs, neutral guaifenesin, acidic sodium salicylate (pK(a) 3.0) and basic timolol (pK(a) 9.2), were incorporated into solvent cast PDLLA and PEA films. The drug content in the films was 2% (w/w). The degradation studies were carried out in PBS (pH 7.4, 37 degrees C); the resulting decrease in molecular weight of polymers was determined by size exclusion chromatography and the weight loss of films was measured. In addition, the drug release from the films in PBS (pH 7.4, 37 degrees C) was studied. The model drugs were released from the PDLLA and PEA films in a biphasic or triphasic manner. The final fast release phase of the drugs from both PDLLA and PEA films started when the molecular weight (M(n)) of the polymer had decreased close to 15,000 g/mol. The degradation rate of the PDLLA films was clearly enhanced by incorporated sodium salicylate or timolol. Whereas, the degradation rate of the PEA film was not enhanced by the incorporated drugs. The present results indicate that when compared to the PDLLA film, degradation rate of the PEA film in the presence of the drug is more predictable.

To identify the disease gene in 6 Spanish families with autosomal recessive retinitis pigmentosa linked to the RP25 locus, mutation screening of 4 candidate genes, KHDRBS2, PTP4A1, KIAA1411 and OGFRL1, was undertaken based on their expression or functional relevance to the retina. Twenty-six single nucleotide polymorphisms were identified, of which 14 were novel. Even though no pathological mutations were detected, these genes however remain as good candidates for other retinal degenerations mapping to the same chromosomal region.

Purpose: To identify the disease gene in five Spanish families with autosomal recessive retinitis pigmentosa (arRP) linked to the RP25 locus. Two candidate genes, EEF1A1 and IMPG1, were selected from the region between D6S280 and D6S1644 markers where the families are linked. The genes were selected as good candidates on the basis of their function, tissue expression pattern, and/or genetic data., Methods: A molecular genetic study was performed on DNA extracted from one parent and one affected member of each studied family. The coding exons, splice sites, and the 5' UTR of the genes were amplified by polymerase chain reaction (PCR). For mutation detection, direct sequence analysis was performed using the ABI 3100 automated sequencer. Segregation of an IMPG1 single nucleotide polymorphism (SNP) in all the families studied was analyzed by restriction enzyme digest of the amplified gene fragments., Results: In total, 15 SNPs were identified of which 7 were novel. Of the identified SNPs, one was insertion, two were deletions, five were intronic, six were missense, and one was located in the 5' UTR. These changes, however, were also identified in unaffected members of the families and/or 50 control Caucasians. The examined known IMPG1 SNP was not segregating with the disease phenotype but was correlating with the genetic data in all families studied., Conclusions: Our results indicate that neither EEF1A1 nor IMPG1 could be responsible for RP25 in the studied families due to absence of any pathogenic variants. However, it is important to notice that the methodology used in this study cannot detect larger deletions that lie outside the screened regions or primer site mutations that exist in the heterozygous state. A role of both genes in other inherited forms of RP and/or retinal degenerations needs to be elucidated.

Retinitis pigmentosa (RP) is the most common form of retinal dystrophy. It is featured by a great clinical and genetic heterogeneity. Different patterns of inheritance exist, such as autosomal dominant and recessive, X-linked and digenic. RP25, a locus for autosomal recessive retinitis pigmentosa (arRP), the most frequently inherited form of RP, was mapped to chromosome 6q between D6S257 and D6S1644 microsatellite markers. ELOVL5, SMAP1 and GLULD1 were selected on the basis of their location, tissue expression and/or function. ELOVL5 is implicated in the elongation of long chain fatty acids, including docosahexanoic acid (DHA), which constitutes 50% of the fatty acids of the outer segment of the photoreceptor. SMAP1 (stromal membrane associated protein 1) was found to be located within RP25 locus and is expressed in retina. GLULD1, glutamate-ammonia ligase (glutamine synthase) domain containing 1, plays a key role in the uptake and metabolism of glutamate in the retina. The absence of pathogenic mutations after molecular analysis argues against the implication of ELOVL5, SMAP1 and GLULD1 in the development of RP25 phenotype. Nevertheless, we could not rule them out as good candidates for other retinal degeneration mapping to the same chromosomal region.