DNA from multiple viral species is associated with Alzheimer's disease risk.

INTRODUCTION: Multiple infectious agents, including viruses, bacteria, fungi, and protozoa, have been linked to Alzheimer's disease (AD) risk by independent lines of evidence. We explored this association by comparing the frequencies of viral species identified in a large sample of AD cases and controls. METHODS: DNA sequence reads that did not align to the human genome in sequences were mapped to viral reference sequences, quantified, and then were tested for association with AD in whole exome sequences (WES) and whole genome sequences (WGS) datasets. RESULTS: Several viruses were significant predictors of AD according to the machine learning classifiers. Subsequent regression analyses showed that herpes simplex type 1 (HSV-1) (odds ratio [OR] = 3.71, p = 8.03 × 10-4) and human papillomavirus 71 (HPV-71; OR = 3.56, p = 0.02), were significantly associated with AD after Bonferroni correction. The phylogenetic-related cluster of Herpesviridae was significantly associated with AD in several strata of the data (p < 0.01). DISCUSSION: Our results support the hypothesis that viral infection, especially HSV-1, is associated with AD risk.

Genome-wide association and multi-omics studies identify MGMT as a novel risk gene for Alzheimer's disease among women.

INTRODUCTION: Variants in the tau gene (MAPT) region are associated with breast cancer in women and Alzheimer's disease (AD) among persons lacking apolipoprotein E ε4 (ε4-). METHODS: To identify novel genes associated with tau-related pathology, we conducted two genome-wide association studies (GWAS) for AD, one among 10,340 ε4- women in the Alzheimer's Disease Genetics Consortium (ADGC) and another in 31 members (22 women) of a consanguineous Hutterite kindred. RESULTS: We identified novel associations of AD with MGMT variants in the ADGC (rs12775171, odds ratio [OR] = 1.4, P = 4.9 × 10-8 ) and Hutterite (rs12256016 and rs2803456, OR = 2.0, P = 1.9 × 10-14 ) datasets. Multi-omics analyses showed that the most significant and largest number of associations among the single nucleotide polymorphisms (SNPs), DNA-methylated CpGs, MGMT expression, and AD-related neuropathological traits were observed among women. Furthermore, promoter capture Hi-C analyses revealed long-range interactions of the MGMT promoter with MGMT SNPs and CpG sites. DISCUSSION: These findings suggest that epigenetically regulated MGMT expression is involved in AD pathogenesis, especially in women.

Targeting EIF4E signaling with ribavirin in infant acute lymphoblastic leukemia.

The poor outcomes in infant acute lymphoblastic leukemia (ALL) necessitate new treatments. Here we discover that EIF4E protein is elevated in most cases of infant ALL and test EIF4E targeting by the repurposed antiviral agent ribavirin, which has anticancer properties through EIF4E inhibition, as a potential treatment. We find that ribavirin treatment of actively dividing infant ALL cells on bone marrow stromal cells (BMSCs) at clinically achievable concentrations causes robust proliferation inhibition in proportion with EIF4E expression. Further, we find that ribavirin treatment of KMT2A-rearranged (KMT2A-R) infant ALL cells and the KMT2A-AFF1 cell line RS4:11 inhibits EIF4E, leading to decreases in oncogenic EIF4E-regulated cell growth and survival proteins. In ribavirin-sensitive KMT2A-R infant ALL cells and RS4:11 cells, EIF4E-regulated proteins with reduced levels of expression following ribavirin treatment include MYC, MCL1, NBN, BCL2 and BIRC5. Ribavirin-treated RS4:11 cells exhibit impaired EIF4E-dependent nuclear to cytoplasmic export and/or translation of the corresponding mRNAs, as well as reduced phosphorylation of the p-AKT1, p-EIF4EBP1, p-RPS6 and p-EIF4E signaling proteins. This leads to an S-phase cell cycle arrest in RS4:11 cells corresponding to the decreased proliferation. Ribavirin causes nuclear EIF4E to re-localize to the cytoplasm in KMT2A-AFF1 infant ALL and RS4:11 cells, providing further evidence for EIF4E inhibition. Ribavirin slows increases in peripheral blasts in KMT2A-R infant ALL xenograft-bearing mice. Ribavirin cooperates with chemotherapy, particularly L-asparaginase, in reducing live KMT2A-AFF1 infant ALL cells in BMSC co-cultures. This work establishes that EIF4E is broadly elevated across infant ALL and that clinically relevant ribavirin exposures have preclinical activity and effectively inhibit EIF4E in KMT2A-R cases, suggesting promise in EIF4E targeting using ribavirin as a means of treatment.

Replication of progressive supranuclear palsy genome-wide association study identifies SLCO1A2 and DUSP10 as new susceptibility loci.

BACKGROUND: Progressive supranuclear palsy (PSP) is a parkinsonian neurodegenerative tauopathy affecting brain regions involved in motor function, including the basal ganglia, diencephalon and brainstem. While PSP is largely considered to be a sporadic disorder, cases with suspected familial inheritance have been identified and the common MAPT H1haplotype is a major genetic risk factor. Due to the relatively low prevalence of PSP, large sample sizes can be difficult to achieve, and this has limited the ability to detect true genetic risk factors at the genome-wide statistical threshold for significance in GWAS data. With this in mind, in this study we genotyped the genetic variants that displayed the strongest degree of association with PSP (P<1E-4) in the previous GWAS in a new cohort of 533 pathologically-confirmed PSP cases and 1172 controls, and performed a combined analysis with the previous GWAS data. RESULTS: Our findings validate the known association of loci at MAPT, MOBP, EIF2AK3 and STX6 with risk of PSP, and uncover novel associations with SLCO1A2 (rs11568563) and DUSP10 (rs6687758) variants, both of which were classified as non-significant in the original GWAS. CONCLUSIONS: Resolving the genetic architecture of PSP will provide mechanistic insights and nominate candidate genes and pathways for future therapeutic intervention strategies.

Unique Familial MLL(KMT2A)-Rearranged Precursor B-Cell Infant Acute Lymphoblastic Leukemia in Non-twin Siblings.

BACKGROUND: Infant acute lymphoblastic leukemia (ALL) has never occurred in families except for the ∼100% concordant cases in monozygous twins attributed to twin-to-twin metastases. We report the first kindred with infant ALL in non-twin siblings. The siblings were diagnosed with MLL-rearranged (MLL-R) ALL 26 months apart. The second affected sibling had an unaffected dichorionic monozygous co-twin. Both had fatal outcomes. PROCEDURES: Translocations were characterized by karyotype, FISH, multiplex FISH, and MLL breakpoint cluster region (bcr) Southern blot analysis. Breakpoint junctions and fusion transcripts were cloned by PCR. TP53 mutation and NADPH quinone oxidorecuctase 1 (NQO1) C609T analyses were performed, and pedigree history and parental occupations were ascertained. The likelihood of chance occurrence of infant ALL in non-twin siblings was computed based on a binomial distribution. Zygosity was determined by single nucleotide polymorphism (SNP) array. RESULTS: The translocations were not related or vertically transmitted. The complex karyotype of the proband's ALL had chromosome 2, 3, 4, and 11 abnormalities causing a 5'-MLL-AFF1-3' fusion and a non-productive rearrangement of 3'MLL with a chromosome 3q intergenic region. The affected twin's ALL exhibited a simple t(4;11). The complex karyotype of the proband's ALL suggested a genotoxic insult, but no exposure was identified. There was no germline TP53 mutation. The NQO1 C609T risk allele was absent. The likelihood of infant ALL occurring in non-twin siblings by chance alone is one in 1.198 × 10(9) families. CONCLUSIONS: Whether because of a deleterious transplacental exposure, novel predisposition syndrome, or exceedingly rare chance occurrence, MLL-R infant ALL can occur in non-twin siblings. The discordant occurrence of infant ALL in the monozygous twins was likely because they were dichorionic.

Identifying amyloid pathology-related cerebrospinal fluid biomarkers for Alzheimer's disease in a multicohort study.

INTRODUCTION: The dynamic range of cerebrospinal fluid (CSF) amyloid ß (Aß1-42) measurement does not parallel to cognitive changes in Alzheimer's disease (AD) and cognitively normal (CN) subjects across different studies. Therefore, identifying novel proteins to characterize symptomatic AD samples is important. METHODS: Proteins were profiled using a multianalyte platform by Rules Based Medicine (MAP-RBM). Due to underlying heterogeneity and unbalanced sample size, we combined subjects (344 AD and 325 CN) from three cohorts: Alzheimer's Disease Neuroimaging Initiative, Penn Center for Neurodegenerative Disease Research of the University of Pennsylvania, and Knight Alzheimer's Disease Research Center at Washington University in St. Louis. We focused on samples whose cognitive and amyloid status was consistent. We performed linear regression (accounted for age, gender, number of APOE e4 alleles, and cohort variable) to identify amyloid-related proteins for symptomatic AD subjects in this largest ever CSF-based MAP-RBM study. ANOVA and Tukey's test were used to evaluate if these proteins were related to cognitive impairment changes as measured by mini-mental state examination (MMSE). RESULTS: Seven proteins were significantly associated with Aß1-42 levels in the combined cohort (false discovery rate adjusted P < .05), of which lipoprotein a (Lp(a)), prolactin (PRL), resistin, and vascular endothelial growth factor (VEGF) have consistent direction of associations across every individual cohort. VEGF was strongly associated with MMSE scores, followed by pancreatic polypeptide and immunoglobulin A (IgA), suggesting they may be related to staging of AD. DISCUSSION: Lp(a), PRL, IgA, and tissue factor/thromboplastin have never been reported for AD diagnosis in previous individual CSF-based MAP-RBM studies. Although some of our reported analytes are related to AD pathophysiology, others' roles in symptomatic AD samples worth further explorations.

Solutions to Peto's paradox revealed by mathematical modelling and cross-species cancer gene analysis.

Whales have 1000-fold more cells than humans and mice have 1000-fold fewer; however, cancer risk across species does not increase with the number of somatic cells and the lifespan of the organism. This observation is known as Peto's paradox. How much would evolution have to change the parameters of somatic evolution in order to equalize the cancer risk between species that differ by orders of magnitude in size? Analysis of previously published models of colorectal cancer suggests that a two- to three-fold decrease in the mutation rate or stem cell division rate is enough to reduce a whale's cancer risk to that of a human. Similarly, the addition of one to two required tumour-suppressor gene mutations would also be sufficient. We surveyed mammalian genomes and did not find a positive correlation of tumour-suppressor genes with increasing body mass and longevity. However, we found evidence of the amplification of TP53 in elephants, MAL in horses and FBXO31 in microbats, which might explain Peto's paradox in those species. Exploring parameters that evolution may have fine-tuned in large, long-lived organisms will help guide future experiments to reveal the underlying biology responsible for Peto's paradox and guide cancer prevention in humans.

HAMR: high-throughput annotation of modified ribonucleotides.

RNA is often altered post-transcriptionally by the covalent modification of particular nucleotides; these modifications are known to modulate the structure and activity of their host RNAs. The recent discovery that an RNA methyl-6 adenosine demethylase (FTO) is a risk gene in obesity has brought to light the significance of RNA modifications to human biology. These noncanonical nucleotides, when converted to cDNA in the course of RNA sequencing, can produce sequence patterns that are distinguishable from simple base-calling errors. To determine whether these modifications can be detected in RNA sequencing data, we developed a method that can not only locate these modifications transcriptome-wide with single nucleotide resolution, but can also differentiate between different classes of modifications. Using small RNA-seq data we were able to detect 92% of all known human tRNA modification sites that are predicted to affect RT activity. We also found that different modifications produce distinct patterns of cDNA sequence, allowing us to differentiate between two classes of adenosine and two classes of guanine modifications with 98% and 79% accuracy, respectively. To show the robustness of this method to sample preparation and sequencing methods, as well as to organismal diversity, we applied it to a publicly available yeast data set and achieved similar levels of accuracy. We also experimentally validated two novel and one known 3-methylcytosine (3mC) sites predicted by HAMR in human tRNAs. Researchers can now use our method to identify and characterize RNA modifications using only RNA-seq data, both retrospectively and when asking questions specifically about modified RNA.

Potent obatoclax cytotoxicity and activation of triple death mode killing across infant acute lymphoblastic leukemia.

Survival in infants younger than 1 year who have acute lymphoblastic leukemia (ALL) is inferior whether MLL is rearranged (R) or germline (G). MLL translocations confer chemotherapy resistance, and infants experience excess complications. We characterized in vitro sensitivity to the pan-antiapoptotic BCL-2 family inhibitor obatoclax mesylate in diagnostic leukemia cells from 54 infants with ALL/bilineal acute leukemia because of the role of prosurvival BCL-2 proteins in resistance, their imbalanced expression in infant ALL, and evidence of obatoclax activity with a favorable toxicity profile in early adult leukemia trials. Overall, half maximal effective concentrations (EC50s) were lower than 176 nM (the maximal plasma concentration [Cmax] with recommended adult dose) in 76% of samples, whether in MLL-AF4, MLL-ENL, or other MLL-R or MLL-G subsets, and regardless of patients' poor prognostic features. However, MLL status and partner genes correlated with EC50. Combined approaches including flow cytometry, Western blot, obatoclax treatment with death pathway inhibition, microarray analyses, and/or electron microscopy indicated a unique killing mechanism involving apoptosis, necroptosis, and autophagy in MLL-AF4 ALL cell lines and primary MLL-R and MLL-G infant ALL cells. This in vitro obatoclax activity and its multiple killing mechanisms across molecular cytogenetic subsets provide a rationale to incorporate a similarly acting compound into combination strategies to combat infant ALL.

Selection of individual VH genes occurs at the pro-B to pre-B cell transition.

B cells are subjected to selection at multiple checkpoints during their development. The selection of Ab H chains is difficult to study because of the large diversity of the CDR3. To study the selection of individual Ab H chain V region genes (V(H)), we performed CDR3 spectratyping of ∼ 75-300 rearrangements per individual V(H) in C57BL6/J mice. We measured the fraction of rearrangements that were in-frame in B cell DNA. We demonstrate that individual V(H)s have different fractions of in-frame rearrangements (IF fractions) ranging from 10 to 90% and that these IF fractions are reproducible in different mice. For most V(H)s, the IF fraction in pro-B cells approximated 33% and then shifted to the nearly final (mature) B cell value by the cycling pre-B cell stage. The frequency of high in-frame (IF) V(H) usage increased in cycling pre-B cells compared with that in pro-B cells, whereas this did not occur for low IF V(H)s. The IF fraction did not shift as much in BCR-expressing B cells and was minimally affected by L chain usage for most V(H). High IF clan II/III V(H)s share more positively charged CDR2 sequences, whereas high IF clan I J558 CDR2 sequences are diverse. These data indicate that individual V(H)s are subjected to differential selection, that V(H) IF fraction is mainly established through pre-BCR-mediated selection, that it may operate differently in clan I versus II/III V(H)s, and that it has a lasting influence on the Ab repertoire.

Polyploidy, aneuploidy and the evolution of cancer.

Aneuploidy is a ubiquitous feature of cancer and pre-cancerous lesions, yet its significance is poorly characterized. In this chapter, we review the role oftetraploidy and aneuploidy in progression. We examine how aneuploidy may contribute to the evolutionary dynamics prevalent in neoplastic progression, considering whether aneuploidy itself is selectively neutral or advantageous or if it simply acts as a mechanism for the more rapid accumulation of mutations increasing survival and reproduction of cancer cells. We also review evidence from Barrett's esophagus, a pre-malignant condition, demonstrating that tetraploidy and aneuploidy are correlated with an increased risk of progression to cancer. Ultimately, we aim provide testable hypotheses and methods for understanding the role of aneuploidy in cancer.

Design of comprehensive Alzheimer's disease centers to address unmet national needs.

The problem of Alzheimer's disease (AD) exemplifies the challenges of dealing with a broad range of aging-related chronic disorders that require long-term, labor-intensive, and expensive care. As the baby boom generation ages and brain diseases become more prevalent, the need to confront the pending health care crisis is more urgent than ever before. Indeed, there is now a critical need to expand significantly the national effort to solve the problem of AD, with special focus on prevention. The Campaign to Prevent Alzheimer's Disease by 2020 (PAD2020) aims to create a new paradigm for planning and supporting the organization of worldwide cooperative research networks to develop new technologies for early detection and treatments of aging-related memory and motor impairments. PAD 2020 is developing an implementation plan to justify (1) increasing the federal budget for research, (2) developing novel national resources to discover new interventions for memory and motor disorders, and (3) creating innovative and streamlined decision-making processes for selecting and supporting new ideas. Since 1978 the National Institute on Aging or National Institute of Health (NIH) established an extensive national network of AD research facilities at academic institutions including AD Centers (ADCs), Consortium to Establish a Registry for AD, AD Cooperative Study (ADCS), AD Drug Discovery Program, National Alzheimer's Coordinating Center, National Cell Repository for AD, and AD Neuroimaging Initiative. However, despite the success of these programs and their critical contributions, they are no longer adequate to meet the challenges presented by AD. PAD 2020 is designed to address these changes by improving the efficiency and effectiveness of these programs. For example, the ADCs (P30s and P50s) can be enhanced by converting some into Comprehensive Alzheimer's Disease Centers (CADCs) to support not only research, but also by being demonstration projects on care/treatment, clinical trials, and education as well as by seamlessly integrating multisite collaborative studies (ADCS, AD Neuroimaging Initiative, Patient Registries, Clinical Data Banks, etc) into a cohesive structure that further enhances the original mission of the National Institute on Aging ADCs. Regional CADCs offer greater efficiency and cost savings while serving as coordinating hubs of existing ADCs, thereby offering greater economies of scale and programmatic integration. The CADCs also broaden the scope of ADC activities to include research on interventions, diagnosis, imaging, prevention trials, and other longitudinal studies that require long-term support. Thus, CADCs can address the urgent need to identify subjects at high risk of AD for prevention trials and very early in the course of AD for clinical trials of disease modification. The enhanced CADCs will allow more flexibility among ADCs by supporting collaborative linkages with other institutions and drawing on a wider expertise from different locations. This perspective article describes the University of Pennsylvania (Penn) CADC Model as an illustrative example of how an existing ADC can be converted into a CADC by better utilization of Penn academic resources to address the wide range of problems concerning AD. The intent of this position paper is to stimulate thinking and foster the development of other or alternative models for a systematic approach to the study of dementia and movement disorders.

Enhancing CD8 T-cell memory by modulating fatty acid metabolism.

CD8 T cells, which have a crucial role in immunity to infection and cancer, are maintained in constant numbers, but on antigen stimulation undergo a developmental program characterized by distinct phases encompassing the expansion and then contraction of antigen-specific effector (T(E)) populations, followed by the persistence of long-lived memory (T(M)) cells. Although this predictable pattern of CD8 T-cell responses is well established, the underlying cellular mechanisms regulating the transition to T(M) cells remain undefined. Here we show that tumour necrosis factor (TNF) receptor-associated factor 6 (TRAF6), an adaptor protein in the TNF-receptor and interleukin-1R/Toll-like receptor superfamily, regulates CD8 T(M)-cell development after infection by modulating fatty acid metabolism. We show that mice with a T-cell-specific deletion of TRAF6 mount robust CD8 T(E)-cell responses, but have a profound defect in their ability to generate T(M) cells that is characterized by the disappearance of antigen-specific cells in the weeks after primary immunization. Microarray analyses revealed that TRAF6-deficient CD8 T cells exhibit altered expression of genes that regulate fatty acid metabolism. Consistent with this, activated CD8 T cells lacking TRAF6 display defective AMP-activated kinase activation and mitochondrial fatty acid oxidation (FAO) in response to growth factor withdrawal. Administration of the anti-diabetic drug metformin restored FAO and CD8 T(M)-cell generation in the absence of TRAF6. This treatment also increased CD8 T(M) cells in wild-type mice, and consequently was able to considerably improve the efficacy of an experimental anti-cancer vaccine.

A Bayesian approach to efficient differential allocation for resampling-based significance testing.

BACKGROUND: Large-scale statistical analyses have become hallmarks of post-genomic era biological research due to advances in high-throughput assays and the integration of large biological databases. One accompanying issue is the simultaneous estimation of p-values for a large number of hypothesis tests. In many applications, a parametric assumption in the null distribution such as normality may be unreasonable, and resampling-based p-values are the preferred procedure for establishing statistical significance. Using resampling-based procedures for multiple testing is computationally intensive and typically requires large numbers of resamples. RESULTS: We present a new approach to more efficiently assign resamples (such as bootstrap samples or permutations) within a nonparametric multiple testing framework. We formulated a Bayesian-inspired approach to this problem, and devised an algorithm that adapts the assignment of resamples iteratively with negligible space and running time overhead. In two experimental studies, a breast cancer microarray dataset and a genome wide association study dataset for Parkinson's disease, we demonstrated that our differential allocation procedure is substantially more accurate compared to the traditional uniform resample allocation. CONCLUSION: Our experiments demonstrate that using a more sophisticated allocation strategy can improve our inference for hypothesis testing without a drastic increase in the amount of computation on randomized data. Moreover, we gain more improvement in efficiency when the number of tests is large. R code for our algorithm and the shortcut method are available at http://people.pcbi.upenn.edu/~lswang/pub/bmc2009/.