Deficiency of nucleotide excision repair is associated with mutational signature observed in cancer.

Nucleotide excision repair (NER) is one of the main DNA repair pathways that protect cells against genomic damage. Disruption of this pathway can contribute to the development of cancer and accelerate aging. Mutational characteristics of NER-deficiency may reveal important diagnostic opportunities, as tumors deficient in NER are more sensitive to certain treatments. Here, we analyzed the genome-wide somatic mutational profiles of adult stem cells (ASCs) from NER-deficient Ercc1 -/Δ mice. Our results indicate that NER-deficiency increases the base substitution load twofold in liver but not in small intestinal ASCs, which coincides with the tissue-specific aging pathology observed in these mice. Moreover, NER-deficient ASCs of both tissues show an increased contribution of Signature 8 mutations, which is a mutational pattern with unknown etiology that is recurrently observed in various cancer types. The scattered genomic distribution of the base substitutions indicates that deficiency of global-genome NER (GG-NER) underlies the observed mutational consequences. In line with this, we observe increased Signature 8 mutations in a GG-NER-deficient human organoid culture, in which XPC was deleted using CRISPR-Cas9 gene-editing. Furthermore, genomes of NER-deficient breast tumors show an increased contribution of Signature 8 mutations compared with NER-proficient tumors. Elevated levels of Signature 8 mutations could therefore contribute to a predictor of NER-deficiency based on a patient's mutational profile.

Amplatzer Vascular Plug Embolization in Two Patients with Ruptured Infrarenal Aortic Aneurysm and Aortoiliac Occlusive Disease.

Mortality for ruptured abdominal aortic aneurysm (AAA) is known to be high. When left untreated, it is nearly always fatal. Standard treatment options include open surgery and endovascular aneurysm repair (EVAR), but both techniques have limitations. Owing to comorbidities and anatomical constraints, some patients are deemed unsuitable for both open surgery and EVAR. In these patients, alternative treatment strategies can be of special interest. To our knowledge, these are the first two cases reported using an Amplatzer Vascular Plug II for aortic embolization in patients with coexisting aneurysmatic and aorto-bi-iliac occlusive disease requiring urgent treatment for contained AAA rupture. Successful aneurysm exclusion was noted at follow-up ranging from 5 months to 3 years, and no procedure-related complications occurred. We therefore believe that in selected patients, this could be an elegant alternative in life-threatening situations with sustained occlusion in the mid-term.

How to choose between a pacemaker or defibrillator for resynchronization therapy?

OBJECTIVE: The choice between a resynchronization pacemaker (CRT-P) or defibrillator (CRT-D) is still a matter of debate. We hypothesised that when selecting patients based on co-morbidities and age as proposed by the ESC-guidelines, there would be no long-term survival benefit of CRT-D compared to CRT-P. METHODS: We performed a retrospective analysis of patients who received a CRT device at the University Hospitals Leuven between 2001 and 2007. For the analysis of the association between predictors and outcome, uni- and multivariate Cox regression analyses were performed. We present data from three multivariate models. RESULTS: A total of 144 CRT devices were implanted (CRT-D n=98, CRT-P n=46). Patients who received a CRT-P were older and had a higher prevalence of co-morbidities. Patients who received a CRT-D had a significant lower mortality. When applying incremental multivariate analysis using 1st variables with a P < 0.05 in univariate analysis, 2nd variables with a P < 0.10 and 3rd adding on top all the baseline variables that were significantly different between the two groups, the significance of a possible survival benefit for CRT-D over CRT-P disappeared: risk model 1, hazard ratio 2.21 (P = 0.008), risk model 2, HR 1.81 (P = 0.069), and risk model 3, HR 1.85 (P = 0.091). The use of amiodarone and the presence of COPD or renal insufficiency remained associated with a significant, higher mortality risk, while the use of beta blockers was protective in all three models. CONCLUSION: The choice of a CRT-D seemed a predictor of improved survival in simple but not in more complex multivariable analyses. The fact that the survival benefit strongly depended on the number of co-variables suggests that it is at most marginal.

Complex Dysautonomia in a Patient With Cerebral Cavernous Malformations Due to a KRIT1 Pleiotropic Gene Mutation.

Dysautonomia is a disruption of the body's autonomic processes. Symptoms vary among patients, depending on the underlying disease pathways. Given that symptoms can affect all organ functions, dysautonomia often significantly impacts quality of life. However, due to its complex and varied presentation, early recognition of dysautonomia remains a challenge, yet it is crucial for improving patient outcomes. We report a case of a patient with a KRIT1 mutation presenting with dysautonomia causing urological, sexual, and bowel dysfunction. We hypothesize that the patient's symptoms are due to a pontine cavernous malformation (CM) caused by the KRIT1 mutation. A literature review was conducted to establish a link between pontine CM and dysautonomia.

Cystic Adventitial Disease of the (ilio) Femoral Artery with a Connection to the Hip Joint: Case Report and a Review of the Literature.

Objective: Cystic adventitial disease (CAD) is an uncommon non-atherosclerotic peripheral vessel disease, most often seen in the popliteal artery. Only a small number of cases involving the (ilio) femoral artery have been reported. The case of a 48 year old female with CAD of the left femoral artery with a connection of the disease to the hip joint on pre-operative imaging confirmed during surgery is described. A literature review of CAD of the (ilio) femoral artery with patient demographic data, symptoms, management, presence of a joint connection, and long term outcomes was performed. Methods: Multiple databases (Medline, CINAHL, EMBASE) were searched and each article was cross referenced to collect the literature on CAD of the (ilio) femoral artery. Case studies or series of CAD of the (ilio) femoral artery in English between 1995 and 2021 were included. Results: Sixteen case reports with 17 patients were included; 71% were male. CAD was unilateral in all case reports, with 53% on the right side. Patients presented with vascular symptoms including claudication (88%), a palpable pulsating mass (18%), acute limb ischaemia (6%) or limb swelling (8%). Computed tomography angiography (CTA) (76%) and duplex ultrasonography (47%) were the most commonly used imaging modalities. The common femoral artery was the most affected site (88%). Reported treatments were cyst resection and autologous vein reconstruction (six, one recurrence), cyst resection and patch repair (five, one recurrence), cyst resection with synthetic graft reconstruction (three, no recurrence), cyst resection (two, one recurrence), and cyst incision and decompression (one, one recurrence). In 18% of the cases, a connection between the CAD and hip joint was seen. Conclusion: Cyst resection and ligation with interposition of an autologous vein graft, synthetic graft or patch repair (in only locally affected arteries) seems to be the preferred treatment, with a low reported recurrence rate. CTA and magnetic resonance imaging are the imaging modalities of choice when suspecting CAD to determine an appropriate pre-operative plan and identify joint connections.

GRIDSS2: comprehensive characterisation of somatic structural variation using single breakend variants and structural variant phasing.

GRIDSS2 is the first structural variant caller to explicitly report single breakends-breakpoints in which only one side can be unambiguously determined. By treating single breakends as a fundamental genomic rearrangement signal on par with breakpoints, GRIDSS2 can explain 47% of somatic centromere copy number changes using single breakends to non-centromere sequence. On a cohort of 3782 deeply sequenced metastatic cancers, GRIDSS2 achieves an unprecedented 3.1% false negative rate and 3.3% false discovery rate and identifies a novel 32-100 bp duplication signature. GRIDSS2 simplifies complex rearrangement interpretation through phasing of structural variants with 16% of somatic calls phasable using paired-end sequencing.

Precancerous liver diseases do not cause increased mutagenesis in liver stem cells.

Inflammatory liver disease increases the risk of developing primary liver cancer. The mechanism through which liver disease induces tumorigenesis remains unclear, but is thought to occur via increased mutagenesis. Here, we performed whole-genome sequencing on clonally expanded single liver stem cells cultured as intrahepatic cholangiocyte organoids (ICOs) from patients with alcoholic cirrhosis, non-alcoholic steatohepatitis (NASH), and primary sclerosing cholangitis (PSC). Surprisingly, we find that these precancerous liver disease conditions do not result in a detectable increased accumulation of mutations, nor altered mutation types in individual liver stem cells. This finding contrasts with the mutational load and typical mutational signatures reported for liver tumors, and argues against the hypothesis that liver disease drives tumorigenesis via a direct mechanism of induced mutagenesis. Disease conditions in the liver may thus act through indirect mechanisms to drive the transition from healthy to cancerous cells, such as changes to the microenvironment that favor the outgrowth of precancerous cells.

Optimizing Nanopore sequencing-based detection of structural variants enables individualized circulating tumor DNA-based disease monitoring in cancer patients.

Here, we describe a novel approach for rapid discovery of a set of tumor-specific genomic structural variants (SVs), based on a combination of low coverage cancer genome sequencing using Oxford Nanopore with an SV calling and filtering pipeline. We applied the method to tumor samples of high-grade ovarian and prostate cancer patients and validated on average ten somatic SVs per patient with breakpoint-spanning PCR mini-amplicons. These SVs could be quantified in ctDNA samples of patients with metastatic prostate cancer using a digital PCR assay. The results suggest that SV dynamics correlate with and may improve existing treatment-response biomarkers such as PSA. https://github.com/UMCUGenetics/SHARC .

Publisher Correction: The mutational impact of culturing human pluripotent and adult stem cells.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The mutational impact of culturing human pluripotent and adult stem cells.

Genetic changes acquired during in vitro culture pose a risk for the successful application of stem cells in regenerative medicine. To assess the genetic risks induced by culturing, we determined all mutations in individual human stem cells by whole genome sequencing. Individual pluripotent, intestinal, and liver stem cells accumulate 3.5 ± 0.5, 7.2 ± 1.1 and 8.3 ± 3.6 base substitutions per population doubling, respectively. The annual in vitro mutation accumulation rate of adult stem cells is nearly 40-fold higher than the in vivo mutation accumulation rate. Mutational signature analysis reveals that in vitro induced mutations are caused by oxidative stress. Reducing oxygen tension in culture lowers the mutational load. We use the mutation rates, spectra, and genomic distribution to model the accumulation of oncogenic mutations during typical in vitro expansion, manipulation or screening experiments using human stem cells. Our study provides empirically defined parameters to assess the mutational risk of stem cell based therapies.

Twenty Years of Ferroportin Disease: A Review or An Update of Published Clinical, Biochemical, Molecular, and Functional Features.

Iron overloading disorders linked to mutations in ferroportin have diverse phenotypes in vivo, and the effects of mutations on ferroportin in vitro range from loss of function (LOF) to gain of function (GOF) with hepcidin resistance. We reviewed 359 patients with 60 ferroportin variants. Overall, macrophage iron overload and low/normal transferrin saturation (TSAT) segregated with mutations that caused LOF, while GOF mutations were linked to high TSAT and parenchymal iron accumulation. However, the pathogenicity of individual variants is difficult to establish due to the lack of sufficiently reported data, large inter-assay variability of functional studies, and the uncertainty associated with the performance of available in silico prediction models. Since the phenotypes of hepcidin-resistant GOF variants are indistinguishable from the other types of hereditary hemochromatosis (HH), these variants may be categorized as ferroportin-associated HH, while the entity ferroportin disease may be confined to patients with LOF variants. To further improve the management of ferroportin disease, we advocate for a global registry, with standardized clinical analysis and validation of the functional tests preferably performed in human-derived enterocytic and macrophagic cell lines. Moreover, studies are warranted to unravel the definite structure of ferroportin and the indispensable residues that are essential for functionality.

5-Fluorouracil treatment induces characteristic T>G mutations in human cancer.

5-Fluorouracil (5-FU) is a chemotherapeutic drug commonly used for the treatment of solid cancers. It is proposed that 5-FU interferes with nucleotide synthesis and incorporates into DNA, which may have a mutational impact on both surviving tumor and healthy cells. Here, we treat intestinal organoids with 5-FU and find a highly characteristic mutational pattern that is dominated by T>G substitutions in a CTT context. Tumor whole genome sequencing data confirms that this signature is also identified in vivo in colorectal and breast cancer patients who have received 5-FU treatment. Taken together, our results demonstrate that 5-FU is mutagenic and may drive tumor evolution and increase the risk of secondary malignancies. Furthermore, the identified signature shows a strong resemblance to COSMIC signature 17, the hallmark signature of treatment-naive esophageal and gastric tumors, which indicates that distinct endogenous and exogenous triggers can converge onto highly similar mutational signatures.

Scalable Workflows and Reproducible Data Analysis for Genomics.

Biological, clinical, and pharmacological research now often involves analyses of genomes, transcriptomes, proteomes, and interactomes, within and between individuals and across species. Due to large volumes, the analysis and integration of data generated by such high-throughput technologies have become computationally intensive, and analysis can no longer happen on a typical desktop computer.In this chapter we show how to describe and execute the same analysis using a number of workflow systems and how these follow different approaches to tackle execution and reproducibility issues. We show how any researcher can create a reusable and reproducible bioinformatics pipeline that can be deployed and run anywhere. We show how to create a scalable, reusable, and shareable workflow using four different workflow engines: the Common Workflow Language (CWL), Guix Workflow Language (GWL), Snakemake, and Nextflow. Each of which can be run in parallel.We show how to bundle a number of tools used in evolutionary biology by using Debian, GNU Guix, and Bioconda software distributions, along with the use of container systems, such as Docker, GNU Guix, and Singularity. Together these distributions represent the overall majority of software packages relevant for biology, including PAML, Muscle, MAFFT, MrBayes, and BLAST. By bundling software in lightweight containers, they can be deployed on a desktop, in the cloud, and, increasingly, on compute clusters.By bundling software through these public software distributions, and by creating reproducible and shareable pipelines using these workflow engines, not only do bioinformaticians have to spend less time reinventing the wheel but also do we get closer to the ideal of making science reproducible. The examples in this chapter allow a quick comparison of different solutions.

Prioritization of genes driving congenital phenotypes of patients with de novo genomic structural variants.

BACKGROUND: Genomic structural variants (SVs) can affect many genes and regulatory elements. Therefore, the molecular mechanisms driving the phenotypes of patients carrying de novo SVs are frequently unknown. METHODS: We applied a combination of systematic experimental and bioinformatic methods to improve the molecular diagnosis of 39 patients with multiple congenital abnormalities and/or intellectual disability harboring apparent de novo SVs, most with an inconclusive diagnosis after regular genetic testing. RESULTS: In 7 of these cases (18%), whole-genome sequencing analysis revealed disease-relevant complexities of the SVs missed in routine microarray-based analyses. We developed a computational tool to predict the effects on genes directly affected by SVs and on genes indirectly affected likely due to the changes in chromatin organization and impact on regulatory mechanisms. By combining these functional predictions with extensive phenotype information, candidate driver genes were identified in 16/39 (41%) patients. In 8 cases, evidence was found for the involvement of multiple candidate drivers contributing to different parts of the phenotypes. Subsequently, we applied this computational method to two cohorts containing a total of 379 patients with previously detected and classified de novo SVs and identified candidate driver genes in 189 cases (50%), including 40 cases whose SVs were previously not classified as pathogenic. Pathogenic position effects were predicted in 28% of all studied cases with balanced SVs and in 11% of the cases with copy number variants. CONCLUSIONS: These results demonstrate an integrated computational and experimental approach to predict driver genes based on analyses of WGS data with phenotype association and chromatin organization datasets. These analyses nominate new pathogenic loci and have strong potential to improve the molecular diagnosis of patients with de novo SVs.

MutationalPatterns: comprehensive genome-wide analysis of mutational processes.

BACKGROUND: Base substitution catalogues represent historical records of mutational processes that have been active in a cell. Such processes can be distinguished by various characteristics, like mutation type, sequence context, transcriptional and replicative strand bias, genomic distribution and association with (epi)-genomic features. RESULTS: We have created MutationalPatterns, an R/Bioconductor package that allows researchers to characterize a broad range of patterns in base substitution catalogues to dissect the underlying molecular mechanisms. Furthermore, it offers an efficient method to quantify the contribution of known mutational signatures within single samples. This analysis can be used to determine whether certain DNA repair mechanisms are perturbed and to further characterize the processes underlying known mutational signatures. CONCLUSIONS: MutationalPatterns allows for easy characterization and visualization of mutational patterns. These analyses willsupport fundamental research into mutational mechanisms and may ultimately improve cancer diagnosis and treatment strategies. MutationalPatterns is freely available at http://bioconductor.org/packages/MutationalPatterns .

Electrocardiographic-guided tip positioning technique for peripherally inserted central catheters in a Dutch teaching hospital: Feasibility and cost-effectiveness analysis in a prospective cohort study.

INTRODUCTION:: Peripherally inserted central catheters are venous devices intended for short to medium periods of intravenous treatment. Positioning of the catheter tip at the cavoatrial junction is necessary for optimum performance of a peripherally inserted central catheter. In this study, safety, effectiveness and cost-effectiveness of electrocardiographic-guided peripherally inserted central catheter positioning in a Dutch teaching hospital were evaluated. METHODS:: All patients who received a peripherally inserted central catheter in 2016 using electrocardiographic guidance were compared to those where fluoroscopy guidance was used in a prospective non-randomized cohort study. Relevant data were extracted from electronic health records. Cost-effectiveness analysis was performed. RESULTS:: A total of 162 patients received a peripherally inserted central catheter using fluoroscopy guidance and 103 patients using electrocardiographic guidance in 2016. No significant difference was found in malposition, infection or other complications between these groups. Due to personnel reduction and omission of fluoroscopy costs, cost reduction for each catheter insertion was €120 in the first year and, as a result of discounted acquisition costs, €190 in subsequent years. DISCUSSION:: The positioning results and complication rate are comparable to the previously reported literature. The cost reduction may vary in different hospitals. Other benefits of the electrocardiographic-guided technique are omission of X-ray exposure and improved patient service. CONCLUSION:: Implementation of electrocardiographic-guided tip positioning for peripherally inserted central catheter was safe and effective in this study and led to an improved high value and cost-conscious care.

Measuring mutation accumulation in single human adult stem cells by whole-genome sequencing of organoid cultures.

Characterization of mutational processes in adult stem cells (ASCs) will improve our understanding of aging-related diseases, such as cancer and organ failure, and may ultimately help prevent the development of these diseases. Here, we present a method for cataloging mutations in individual human ASCs without the necessity of using error-prone whole-genome amplification. Single ASCs are expanded in vitro into clonal organoid cultures to generate sufficient DNA for accurate whole-genome sequencing (WGS) analysis. We developed a data-analysis pipeline that identifies with high confidence somatic variants that accumulated in vivo in the original ASC. These genome-wide mutation catalogs are valuable resources for the characterization of the underlying mutational mechanisms. In addition, this protocol can be used to determine the effects of culture conditions or mutagen exposure on mutation accumulation in ASCs in vitro. Here, we describe a protocol for human liver ASCs that can be completed over a period of 3-4 months with hands-on time of ∼5 d.

Motor impairment and its relationship to fitness in children.

OBJECTIVES: The aim of this work was to explore the physiological and perceptual limits to exercise in children with varying degrees of motor impairment, and the relationships to measures of health. DESIGN AND METHODS: In a group comparison design, 35 boys aged 12-15 years completed the Movement ABC test for the assessment of motor impairment, followed by an incremental cycle ergometer test to exhaustion for the assessment of maximal oxygen uptake (VO2peak), respiratory exchange ratio (RER), heart rate (HR) and rating of perceived exertion (RPE). Ten participants classified as having either high or no motor impairment also performed a maximal voluntary isometric contraction (MVIC) for the assessment of lower limb extensor strength. RESULTS: 18 boys were classified as having high motor impairment. There was a significant difference in peak (34.9 vs 48.5 mL kg/min), workload (12.5 vs 10.0 mL W), maximal HR (176 vs 188 bpm), maximal oxygen pulse (12.1 vs 15.9 mL beat) and MVIC (5.7 vs 9.1 Nm kg) between the high and non-motor impaired participants, respectively, (p<0.05). There was no significant difference in the RER or RPE between groups. CONCLUSIONS: When performing cycling ergometry, perceived exertion was not a limiting factor in children with high motor impairment. The lower maximal HR, coupled with reduced movement efficiency and muscle strength reported in this group, suggests that exercise is limited by impairment at the muscular level. This finding was supported by high RER values despite low maximal HR values attained at exercise cessation and reduced maximal strength. Perception of effort is not heightened in children with high motor impairment and future-exercise interventions should be focused on improving muscular condition in these participants to enable them to be better prepared to engage in physical activity for health.