Germline NPAT inactivating variants as cause of hereditary colorectal cancer.

Two independent exome sequencing initiatives aimed to identify new genes involved in the predisposition to nonpolyposis colorectal cancer led to the identification of heterozygous loss-of-function variants in NPAT, a gene that encodes a cyclin E/CDK2 effector required for S phase entry and a coactivator of histone transcription, in two families with multiple members affected with colorectal cancer. Enrichment of loss-of-function and predicted deleterious NPAT variants was identified in familial/early-onset colorectal cancer patients compared to non-cancer gnomAD individuals, further supporting the association with the disease. Previous studies in Drosophila models showed that NPAT abrogation results in chromosomal instability, increase of double strand breaks, and induction of tumour formation. In line with these results, colorectal cancers with NPAT somatic variants and no DNA repair defects have significantly higher aneuploidy levels than NPAT-wildtype colorectal cancers. In conclusion, our findings suggest that constitutional inactivating NPAT variants predispose to mismatch repair-proficient nonpolyposis colorectal cancer.

Mutational topography reflects clinical neuroblastoma heterogeneity.

Neuroblastoma is a pediatric solid tumor characterized by strong clinical heterogeneity. Although clinical risk-defining genomic alterations exist in neuroblastomas, the mutational processes involved in their generation remain largely unclear. By examining the topography and mutational signatures derived from all variant classes, we identified co-occurring mutational footprints, which we termed mutational scenarios. We demonstrate that clinical neuroblastoma heterogeneity is associated with differences in the mutational processes driving these scenarios, linking risk-defining pathognomonic variants to distinct molecular processes. Whereas high-risk MYCN-amplified neuroblastomas were characterized by signs of replication slippage and stress, homologous recombination-associated signatures defined high-risk non-MYCN-amplified patients. Non-high-risk neuroblastomas were marked by footprints of chromosome mis-segregation and TOP1 mutational activity. Furthermore, analysis of subclonal mutations uncovered differential activity of these processes through neuroblastoma evolution. Thus, clinical heterogeneity of neuroblastoma patients can be linked to differences in the mutational processes that are active in their tumors.

Exhaustive Variant Interaction Analysis using Multifactor Dimensionality Reduction.

One of the main goals of human genetics is to understand the connections between genomic variation and the predisposition to develop a complex disorder. These disease-variant associations are usually studied in a single independent manner, disregarding the possible effect derived from the interaction between genomic variants. In particular, in a background of complex diseases, these interactions can be directly linked to the disorder and may play an important role in disease development. Although their study has been suggested to help to complete the understanding of the genetic bases of complex diseases, this still represents a big challenge due to large computing demands. Here, we have taken advantage of High-Performance Computing technologies to tackle this problem using a combination of machine learning methods and statistical approaches. As a result, we have created a containerized framework that uses Multifactor Dimensionality Reduction to detect pairs of variants associated with Type 2 Diabetes (T2D). This methodology has been tested in the Northwestern University NUgene project cohort using a dataset of 1,883,192 variant pairs with a certain degree of association with T2D. Out of the pairs studied, we have identified 104 significant pairs, two of which exhibit a potential functional relationship with T2D.

A transposase-derived gene required for human brain development.

DNA transposable elements and transposase-derived genes are present in most living organisms, including vertebrates, but their function is largely unknown. PiggyBac Transposable Element Derived 5 (PGBD5) is an evolutionarily conserved vertebrate DNA transposase-derived gene with retained nuclease activity in cells. Vertebrate brain development is known to be associated with prominent neuronal cell death and DNA breaks, but their causes and functions are not well understood. Here, we show that PGBD5 contributes to normal brain development in mice and humans, where its deficiency causes disorder of intellectual disability, movement and seizures. In mice, Pgbd5 is required for the developmental induction of post-mitotic DNA breaks and recurrent somatic genome rearrangements in neurons. Together, these studies nominate PGBD5 as the long-hypothesized neuronal DNA nuclease required for brain function in mammals.

Sequence diversity of the uniparentally transmitted portions of the genome in the resident population of Catalonia.

Genomic reference databases of residing populations are available in different countries and regions. Since they represent the whole genetic diversity of a geographical region, they have wide applications, from biomedical studies to forensic identifications. Uniparentally transmitted portions of the genome specifically are highly suitable for kinship analyses, mixed DNA cases and geographical ancestry inferences. We have sampled 808 individuals currently residing in Catalonia within the GCAT cohort, from which we have generated 808 high-quality whole mitochondrial DNA (mtDNA) genomes and 399 sequences of the male-specific part of the Y chromosome (MSY). We observe higher genetic diversity than in classical population genetics datasets. We test the robustness of whole sequences for unequivocal identifications, and we found that they have higher resolution than mitochondrial control region and Y chromosome short tandem repeats (Y-STRs), and that most of the variants they present are at low frequencies, increasing the discrimination capacity between individuals. These results confirm the forensic applicability of whole uniparental sequences and provide one of the largest high-quality reference datasets ever published.

Detection of early seeding of Richter transformation in chronic lymphocytic leukemia.

Richter transformation (RT) is a paradigmatic evolution of chronic lymphocytic leukemia (CLL) into a very aggressive large B cell lymphoma conferring a dismal prognosis. The mechanisms driving RT remain largely unknown. We characterized the whole genome, epigenome and transcriptome, combined with single-cell DNA/RNA-sequencing analyses and functional experiments, of 19 cases of CLL developing RT. Studying 54 longitudinal samples covering up to 19 years of disease course, we uncovered minute subclones carrying genomic, immunogenetic and transcriptomic features of RT cells already at CLL diagnosis, which were dormant for up to 19 years before transformation. We also identified new driver alterations, discovered a new mutational signature (SBS-RT), recognized an oxidative phosphorylation (OXPHOS)high-B cell receptor (BCR)low-signaling transcriptional axis in RT and showed that OXPHOS inhibition reduces the proliferation of RT cells. These findings demonstrate the early seeding of subclones driving advanced stages of cancer evolution and uncover potential therapeutic targets for RT.

Clustering and graph mining techniques for classification of complex structural variations in cancer genomes.

For many years, a major question in cancer genomics has been the identification of those variations that can have a functional role in cancer, and distinguish from the majority of genomic changes that have no functional consequences. This is particularly challenging when considering complex chromosomal rearrangements, often composed of multiple DNA breaks, resulting in difficulties in classifying and interpreting them functionally. Despite recent efforts towards classifying structural variants (SVs), more robust statistical frames are needed to better classify these variants and isolate those that derive from specific molecular mechanisms. We present a new statistical approach to analyze SVs patterns from 2392 tumor samples from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium and identify significant recurrence, which can inform relevant mechanisms involved in the biology of tumors. The method is based on recursive KDE clustering of 152,926 SVs, randomization methods, graph mining techniques and statistical measures. The proposed methodology was able not only to identify complex patterns across different cancer types but also to prove them as not random occurrences. Furthermore, a new class of pattern that was not previously described has been identified.

GCAT|Panel, a comprehensive structural variant haplotype map of the Iberian population from high-coverage whole-genome sequencing.

The combined analysis of haplotype panels with phenotype clinical cohorts is a common approach to explore the genetic architecture of human diseases. However, genetic studies are mainly based on single nucleotide variants (SNVs) and small insertions and deletions (indels). Here, we contribute to fill this gap by generating a dense haplotype map focused on the identification, characterization, and phasing of structural variants (SVs). By integrating multiple variant identification methods and Logistic Regression Models (LRMs), we present a catalogue of 35 431 441 variants, including 89 178 SVs (≥50 bp), 30 325 064 SNVs and 5 017 199 indels, across 785 Illumina high coverage (30x) whole-genomes from the Iberian GCAT Cohort, containing a median of 3.52M SNVs, 606 336 indels and 6393 SVs per individual. The haplotype panel is able to impute up to 14 360 728 SNVs/indels and 23 179 SVs, showing a 2.7-fold increase for SVs compared with available genetic variation panels. The value of this panel for SVs analysis is shown through an imputed rare Alu element located in a new locus associated with Mononeuritis of lower limb, a rare neuromuscular disease. This study represents the first deep characterization of genetic variation within the Iberian population and the first operational haplotype panel to systematically include the SVs into genome-wide genetic studies.

Recessive Genome-Wide Meta-analysis Illuminates Genetic Architecture of Type 2 Diabetes.

Most genome-wide association studies (GWAS) of complex traits are performed using models with additive allelic effects. Hundreds of loci associated with type 2 diabetes have been identified using this approach. Additive models, however, can miss loci with recessive effects, thereby leaving potentially important genes undiscovered. We conducted the largest GWAS meta-analysis using a recessive model for type 2 diabetes. Our discovery sample included 33,139 case subjects and 279,507 control subjects from 7 European-ancestry cohorts, including the UK Biobank. We identified 51 loci associated with type 2 diabetes, including five variants undetected by prior additive analyses. Two of the five variants had minor allele frequency of <5% and were each associated with more than a doubled risk in homozygous carriers. Using two additional cohorts, FinnGen and a Danish cohort, we replicated three of the variants, including one of the low-frequency variants, rs115018790, which had an odds ratio in homozygous carriers of 2.56 (95% CI 2.05-3.19; P = 1 × 10-16) and a stronger effect in men than in women (for interaction, P = 7 × 10-7). The signal was associated with multiple diabetes-related traits, with homozygous carriers showing a 10% decrease in LDL cholesterol and a 20% increase in triglycerides; colocalization analysis linked this signal to reduced expression of the nearby PELO gene. These results demonstrate that recessive models, when compared with GWAS using the additive approach, can identify novel loci, including large-effect variants with pathophysiological consequences relevant to type 2 diabetes.

TIGER: The gene expression regulatory variation landscape of human pancreatic islets.

Genome-wide association studies (GWASs) identified hundreds of signals associated with type 2 diabetes (T2D). To gain insight into their underlying molecular mechanisms, we have created the translational human pancreatic islet genotype tissue-expression resource (TIGER), aggregating >500 human islet genomic datasets from five cohorts in the Horizon 2020 consortium T2DSystems. We impute genotypes using four reference panels and meta-analyze cohorts to improve the coverage of expression quantitative trait loci (eQTL) and develop a method to combine allele-specific expression across samples (cASE). We identify >1 million islet eQTLs, 53 of which colocalize with T2D signals. Among them, a low-frequency allele that reduces T2D risk by half increases CCND2 expression. We identify eight cASE colocalizations, among which we found a T2D-associated SLC30A8 variant. We make all data available through the TIGER portal (http://tiger.bsc.es), which represents a comprehensive human islet genomic data resource to elucidate how genetic variation affects islet function and translates into therapeutic insight and precision medicine for T2D.

Impact of Transcriptome and Gut Microbiome on the Response of HIV-1 Infected Individuals to a Dendritic Cell-Based HIV Therapeutic Vaccine.

Therapeutic vaccines based on dendritic cells offer a good approach to HIV-specific T-cell responses and partial control of the viral load after antiretroviral therapy interruption. The aim of the present study was to identify mRNA expression profiles and to assess the impact of the gut microbiome composition for predicting the viral load control after antiretroviral therapy interruption. We enrolled 29 patients to receive either placebo or a monocyte-derived dendritic cell vaccine. Patients with a decrease in their viral load of >0.5 log10 copies/mL by 12 weeks after antiretroviral therapy interruption were considered responders. In total, 66 genes were considered differentially expressed between responders and non-responders. Enrichment analysis revealed several upregulated pathways involved in the host defense response to a virus via the type I interferon signaling pathway. Regarding the gut microbiota, responders showed enriched levels of Bacteroidetes (p < 0.005) and Verrucomicrobia (p = 0.017), while non-responders were enriched with Tenericutes (p = 0.049) and Actinobacteria (p < 0.005). We also found important differences at the genus level. However, we did not discover any effect of the dendritic cell vaccine on the transcriptome or the gut microbiota. An alternative analysis did characterize that the microbiota from responders were associated with the metabolic production of short-chain fatty acids, which are key metabolites in the regulation of intestinal homeostasis. The evidence now consistently shows that short-chain fatty acid depletion occurs in HIV-infected individuals receiving antiretroviral treatment.

Adherence to a Supplemented Mediterranean Diet Drives Changes in the Gut Microbiota of HIV-1-Infected Individuals.

OBJECTIVE: The health effects of a supplemented Mediterranean diet (SMD) with extra-virgin olive oil (EVOO) and nuts are well documented in non-HIV-infected individuals. We hypothesised that the benefits of an SMD could be mediated by changes in the gut microbiota, even in those with an altered intestinal microbiota such as people living with HIV. DESIGN: Individuals living with HIV (n = 102) were randomised to receive an SMD with 50 g/day of EVOO and 30 g/day of walnuts (SMD group) or continue with their regular diet (control group) for 12 weeks. METHODS: Adherence to the Mediterranean diet was assessed using the validated 14-item MD-Adherence-Screener (MEDAS) from the PREDIMED study. A sub-study classifying the participants according to their MEDAS scores was performed. RESULTS: The lipid profile was improved in the SMD group vs. that in the control group (delta-total cholesterol and delta-B-lipoprotein). The immune activation (CD4+HLADR+CD38+ and CD8+HLADR+CD38+ cells) and IFN-γ-producing T-cells significantly decreased at week 12 compared to the baseline in the SMD group but not in the control group. The gut microbiota in those from the high-adherence group presented significantly high diversity and richness at the end of the intervention. Succinivibrio and Bifidobacterium abundances were influenced by the adherence to the MD and significantly correlated with Treg cells. CONCLUSION: The Mediterranean diet improved metabolic parameters, immune activation, Treg function, and the gut microbiota composition in HIV-1-infected individuals. Further, Mediterranean diet increased the Bifidobacterium abundances after the intervention, and it was associated to a beneficial profile.

The impact of non-additive genetic associations on age-related complex diseases.

Genome-wide association studies (GWAS) are not fully comprehensive, as current strategies typically test only the additive model, exclude the X chromosome, and use only one reference panel for genotype imputation. We implement an extensive GWAS strategy, GUIDANCE, which improves genotype imputation by using multiple reference panels and includes the analysis of the X chromosome and non-additive models to test for association. We apply this methodology to 62,281 subjects across 22 age-related diseases and identify 94 genome-wide associated loci, including 26 previously unreported. Moreover, we observe that 27.7% of the 94 loci are missed if we use standard imputation strategies with a single reference panel, such as HRC, and only test the additive model. Among the new findings, we identify three novel low-frequency recessive variants with odds ratios larger than 4, which need at least a three-fold larger sample size to be detected under the additive model. This study highlights the benefits of applying innovative strategies to better uncover the genetic architecture of complex diseases.

Enhancer hijacking determines extrachromosomal circular MYCN amplicon architecture in neuroblastoma.

MYCN amplification drives one in six cases of neuroblastoma. The supernumerary gene copies are commonly found on highly rearranged, extrachromosomal circular DNA (ecDNA). The exact amplicon structure has not been described thus far and the functional relevance of its rearrangements is unknown. Here, we analyze the MYCN amplicon structure using short-read and Nanopore sequencing and its chromatin landscape using ChIP-seq, ATAC-seq and Hi-C. This reveals two distinct classes of amplicons which explain the regulatory requirements for MYCN overexpression. The first class always co-amplifies a proximal enhancer driven by the noradrenergic core regulatory circuit (CRC). The second class of MYCN amplicons is characterized by high structural complexity, lacks key local enhancers, and instead contains distal chromosomal fragments harboring CRC-driven enhancers. Thus, ectopic enhancer hijacking can compensate for the loss of local gene regulatory elements and explains a large component of the structural diversity observed in MYCN amplification.

Author Correction: PGBD5 promotes site-specific oncogenic mutations in human tumors.

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

Genomic and epigenomic insights into the origin, pathogenesis, and clinical behavior of mantle cell lymphoma subtypes.

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm initially driven by CCND1 rearrangement with 2 molecular subtypes, conventional MCL (cMCL) and leukemic non-nodal MCL (nnMCL), that differ in their clinicobiological behavior. To identify the genetic and epigenetic alterations determining this diversity, we used whole-genome (n = 61) and exome (n = 21) sequencing (74% cMCL, 26% nnMCL) combined with transcriptome and DNA methylation profiles in the context of 5 MCL reference epigenomes. We identified that open and active chromatin at the major translocation cluster locus might facilitate the t(11;14)(q13;32), which modifies the 3-dimensional structure of the involved regions. This translocation is mainly acquired in precursor B cells mediated by recombination-activating genes in both MCL subtypes, whereas in 8% of cases the translocation occurs in mature B cells mediated by activation-induced cytidine deaminase. We identified novel recurrent MCL drivers, including CDKN1B, SAMHD1, BCOR, SYNE1, HNRNPH1, SMARCB1, and DAZAP1. Complex structural alterations emerge as a relevant early oncogenic mechanism in MCL, targeting key driver genes. Breakage-fusion-bridge cycles and translocations activated oncogenes (BMI1, MIR17HG, TERT, MYC, and MYCN), generating gene amplifications and remodeling regulatory regions. cMCL carried significant higher numbers of structural variants, copy number alterations, and driver changes than nnMCL, with exclusive alterations of ATM in cMCL, whereas TP53 and TERT alterations were slightly enriched in nnMCL. Several drivers had prognostic impact, but only TP53 and MYC aberrations added value independently of genomic complexity. An increasing genomic complexity, together with the presence of breakage-fusion-bridge cycles and high DNA methylation changes related to the proliferative cell history, defines patients with different clinical evolution.