Identification of genetic subtypes in follicular lymphoma.

Follicular lymphoma (FL) exhibits considerable variability in biological features and clinical trajectories across patients. To dissect the diversity of FL, we utilized a Bernoulli mixture model to identify genetic subtypes in 713 pre-treatment tumor tissue samples. Our analysis revealed the existence of five subtypes with unique genetic profiles that correlated with clinicopathological characteristics. The clusters were enriched in specific mutations as follows: CS (CREBBP and STAT6), TT (TNFAIP3 and TP53), GM (GNA13 and MEF2B), Q (quiescent, for low mutation burden), and AR (mutations of mTOR pathway-related genes). The subtype Q was enriched for patients with stage I disease and associated with a lower proliferative history than the other subtypes. The AR subtype was unique in its enrichment for IgM-expressing FL cases and was associated with advanced-stage and more than 4 nodal sites. The existence of subtypes was validated in an independent cohort of 418 samples from the GALLIUM trial. Notably, patients assigned to the TT subtype consistently experienced inferior progression-free survival when treated with immunochemotherapy. Our findings offer insight into core pathways distinctly linked with each FL cluster and are expected to be informative in the era of targeted therapies.

Travel Health-Related Preparation Practices of Institutions of Higher Education and Occurrence of Health-Related Events among Undergraduate Students Studying Abroad, 2018-2021.

Background: Knowledge of specific health-related events encountered by students studying abroad and the availability and use of pre-travel healthcare for these students is lacking. Methods: Anonymous web-based questionnaires were sent to study abroad offices, student health centers, and undergraduate students after studying abroad at eight institutions of higher education in the United States and Ireland from 2018-2021. Analyses were descriptive; relative risks and 95% confidence intervals were calculated for health-related events. Results: One study abroad office required a pre-travel consultation. All student health centers had pre-travel counseling available. Among 686 students, there were 307 infectious and 1,588 non-infectious health-related issues; 12 students (2%) were hospitalized. Duration of travel and timing of a pre-travel consultation impacted the risk of health-related events. Certain mental health conditions were associated with increased risk of alcohol and drug use. Conclusion: Future studies should address the optimal timing and best practices to optimize health for students studying abroad.

Finite mixtures of matrix variate Poisson-log normal distributions for three-way count data.

MOTIVATION: Three-way data structures, characterized by three entities, the units, the variables and the occasions, are frequent in biological studies. In RNA sequencing, three-way data structures are obtained when high-throughput transcriptome sequencing data are collected for n genes across p conditions at r occasions. Matrix variate distributions offer a natural way to model three-way data and mixtures of matrix variate distributions can be used to cluster three-way data. Clustering of gene expression data is carried out as means of discovering gene co-expression networks. RESULTS: In this work, a mixture of matrix variate Poisson-log normal distributions is proposed for clustering read counts from RNA sequencing. By considering the matrix variate structure, full information on the conditions and occasions of the RNA sequencing dataset is simultaneously considered, and the number of covariance parameters to be estimated is reduced. We propose three different frameworks for parameter estimation: a Markov chain Monte Carlo-based approach, a variational Gaussian approximation-based approach, and a hybrid approach. Various information criteria are used for model selection. The models are applied to both real and simulated data, and we demonstrate that the proposed approaches can recover the underlying cluster structure in both cases. In simulation studies where the true model parameters are known, our proposed approach shows good parameter recovery. AVAILABILITY AND IMPLEMENTATION: The GitHub R package for this work is available at https://github.com/anjalisilva/mixMVPLN and is released under the open source MIT license.

Laser Polishing of Additive Manufactured Aluminium Parts by Modulated Laser Power.

In this study a new approach to laser polishing with periodic modulated laser power in the kilohertz regime is introduced. By varying the modulation frequency and modulation time, different periodic laser power curves with varying minimum, peak and average laser power can be created. The feasibility of the method is shown by polishing of vertical built AlSi10Mg L-PBF parts with an initial roughness of Ra = 12.22 µm. One polishing pass revealed a decreasing surface roughness with increasing energy density on the surface up to Ra = 0.145 µm. An increasing energy density results in a rising remelting depth between 50 and 255 µm and a rising relative porosity of 0.3% to 4.6%. Furthermore, the thermal process stability, analysed by the melt pool length in scanning direction, reveals a steadily increasing melt pool dimension due to component heating. Multiple laser polishing passes offers a further reduced surface roughness, especially at higher modulation frequencies and provides an improved orientation independent roughness homogeneity. The process stability regarding varying initial surface roughness revealed an almost constant relative roughness reduction rate with an achievable roughness variation after two polishing passes between Ra = 0.13-0.26 µm from an initial state of Ra = 8.0-19.2 µm.

DNA Methylation-Based Classification of Small B-Cell Lymphomas: A Proof-of-Principle Study.

Although most small B-cell lymphomas (SBCLs) can be diagnosed using routine methods, challenges exist. For example, marginal zone lymphomas (MZLs) can be difficult to rule-in, in large part because no widely-used, sensitive, and specific biomarker is available for the marginal zone cell of origin. In this study, it was hypothesized that DNA methylation array profiling can assist with the classification of SBCLs, including MZLs. Extramedullary SBCLs, including challenging cases, were reviewed internally for pathology consensus and profiled. By combining the resulting array data set with data sets from other groups, a set of 26 informative probes was selected and used to train machine learning models to classify 4 common SBCLs: chronic lymphocytic leukemia/small lymphocytic lymphoma, follicular lymphoma, mantle cell lymphoma, and MZL. Prediction probability cutoff was used to separate classifiable from unclassifiable cases, and show that the trained model was able to classify 95% of independent test cases (n = 264/279). The concordance between model predictions and pathology diagnoses was 99.6% (n = 262/263) among classifiable test cases. One validation reference test case was reclassified based on model prediction. The model was also used to predict the diagnoses of two challenging SBCLs. Although the differential examined and data on difficult cases are limited, these results support accurate methylation-based classification of SBCLs. Furthermore, high specificities of predictions suggest that methylation signatures can be used to rule-in MZLs.

Combined EZH2 Inhibition and IKAROS Degradation Leads to Enhanced Antitumor Activity in Diffuse Large B-cell Lymphoma.

PURPOSE: The efficacy of EZH2 inhibition has been modest in the initial clinical exploration of diffuse large B-cell lymphoma (DLBCL), yet EZH2 inhibitors are well tolerated. Herein, we aimed to uncover genetic and pharmacologic opportunities to enhance the clinical efficacy of EZH2 inhibitors in DLBCL. EXPERIMENTAL DESIGN: We conducted a genome-wide sensitizing CRISPR/Cas9 screen with tazemetostat, a catalytic inhibitor of EZH2. The sensitizing effect of IKZF1 loss of function was then validated and leveraged for combination treatment with lenalidomide. RNA sequencing (RNA-seq) and chromatin immunoprecipitation sequencing analyses were performed to elucidate transcriptomic and epigenetic changes underlying synergy. RESULTS: We identified IKZF1 knockout as the top candidate for sensitizing DLBCL cells to tazemetostat. Treating cells with tazemetostat and lenalidomide, an immunomodulatory drug that selectively degrades IKAROS and AIOLOS, phenocopied the effects of the CRISPR/Cas9 screen. The combined drug treatment triggered either cell-cycle arrest or apoptosis in a broad range of DLBCL cell lines, regardless of EZH2 mutational status. Cell-line-based xenografts also showed slower tumor growth and prolonged survival in the combination treatment group. RNA-seq analysis revealed strong upregulation of interferon signaling and antiviral immune response signatures. Gene expression of key immune response factors such as IRF7 and DDX58 were induced in cells treated with lenalidomide and tazemetostat, with a concomitant increase of H3K27 acetylation at their promoters. Furthermore, transcriptome analysis demonstrated derepression of endogenous retroviruses after combination treatment. CONCLUSIONS: Our data underscore the synergistic interplay between IKAROS degradation and EZH2 inhibition on modulating epigenetic changes and ultimately enhancing antitumor effects in DLBCL.

A multivariate Poisson-log normal mixture model for clustering transcriptome sequencing data.

BACKGROUND: High-dimensional data of discrete and skewed nature is commonly encountered in high-throughput sequencing studies. Analyzing the network itself or the interplay between genes in this type of data continues to present many challenges. As data visualization techniques become cumbersome for higher dimensions and unconvincing when there is no clear separation between homogeneous subgroups within the data, cluster analysis provides an intuitive alternative. The aim of applying mixture model-based clustering in this context is to discover groups of co-expressed genes, which can shed light on biological functions and pathways of gene products. RESULTS: A mixture of multivariate Poisson-log normal (MPLN) model is developed for clustering of high-throughput transcriptome sequencing data. Parameter estimation is carried out using a Markov chain Monte Carlo expectation-maximization (MCMC-EM) algorithm, and information criteria are used for model selection. CONCLUSIONS: The mixture of MPLN model is able to fit a wide range of correlation and overdispersion situations, and is suited for modeling multivariate count data from RNA sequencing studies. All scripts used for implementing the method can be found at https://github.com/anjalisilva/MPLNClust .

Marine fish oil is more potent than plant-based n-3 polyunsaturated fatty acids in the prevention of mammary tumors.

Marine-derived n-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been shown to inhibit mammary carcinogenesis. However, evidence regarding plant-based α-linolenic acid (ALA), the major n-3 PUFA in the Western diet, remains equivocal. The objective of this study was to examine the effect of lifelong exposure to plant- or marine-derived n-3 PUFAs on pubertal mammary gland and tumor development in MMTV-neu(ndl)-YD5 mice. It is hypothesized that lifelong exposure to n-3 PUFA reduces terminal end buds during puberty leading to delayed tumor onset, volume and multiplicity. It is further hypothesized that plant-derived n-3 PUFAs will exert dose-dependent effects. Harems of MMTV-FVB males were bred with wild-type females and fed either a (1) 10% safflower (10% SF, n-6 PUFA, control), (2) 10% flaxseed (10% FS), (3) 7% safflower plus 3% flaxseed (3% FS) or (4) 7% safflower plus 3% menhaden (3% FO) diet. Female offspring were maintained on parental diets. Compared to SF, 10% FS and 3% FO reduced (P<.05) terminal end buds at 6 weeks and tumor volume and multiplicity at 20 weeks. A dose-dependent reduction of tumor volume and multiplicity was observed in mice fed 3% and 10% FS. Antitumorigenic effects were associated with altered HER2, pHER-2, pAkt and Ki-67 protein expression. Compared to 10% SF, 3% FO significantly down-regulated expression of genes involved in eicosanoid synthesis and inflammation. From this, it can be estimated that ALA was 1/8 as potent as EPA+DHA. Thus, marine-derived n-3 PUFAs have greater potency versus plant-based n-3 PUFAs.

Proanthocyanidin accumulation and transcriptional responses in the seed coat of cranberry beans (Phaseolus vulgaris L.) with different susceptibility to postharvest darkening.

BACKGROUND: Edible dry beans (Phaseolus vulgaris L.) that darken during postharvest storage are graded lower and are less marketable than their non-darkened counterparts. Seed coat darkening in susceptible genotypes is dependent upon the availability of proanthocyanidins, and their subsequent oxidation to reactive quinones. Mature cranberry beans lacking this postharvest darkening trait tend to be proanthocyanidin-deficient, although the underlying molecular and biochemical determinants for this metabolic phenomenon are unknown. RESULTS: Seed coat proanthocyanidin levels increased with plant maturation in a darkening-susceptible cranberry bean recombinant inbred line (RIL), whereas these metabolites were absent in seeds of the non-darkening RIL plants. RNA sequencing (RNA-seq) analysis was used to monitor changes in the seed coat transcriptome as a function of bean development, where transcript levels were measured as fragments per kilobase of exon per million fragments mapped. A total of 1336 genes were differentially expressed between darkening and non-darkening cranberry bean RILs. Structural and regulatory genes of the proanthocyanidin biosynthesis pathway were upregulated in seed coats of the darkening RIL. A principal component analysis determined that changes in transcript levels for two genes of unknown function and three proanthocyanidin biosynthesis genes, FLAVANONE 3-HYDROXYLASE 1, DIHYDROFLAVONOL 4-REDUCTASE 1 and ANTHOCYANIDIN REDUCTASE 1 (PvANR1) were highly correlated with proanthocyanidin accumulation in seed coats of the darkening-susceptible cranberry bean RIL. HPLC-DAD analysis revealed that in vitro activity of a recombinant PvANR1 was NADPH-dependent and assays containing cyanidin yielded epicatechin and catechin; high cyanidin substrate levels inhibited the formation of both of these products. CONCLUSION: Proanthocyanidin oxidation is a pre-requisite for postharvest-related seed coat darkening in dicotyledonous seeds. In model plant species, the accumulation of proanthocyanidins is dependent upon upregulation of biosynthetic genes. In this study, proanthocyanidin production in cranberry bean seed coats was strongly associated with an increase in PvANR1 transcripts during seed maturation. In the presence of NADPH, PvANR1 converted the physiologically relevant substrate cyanidin to epicatechin and catechin.

Identification of multiple phosphorylation sites on maize endosperm starch branching enzyme IIb, a key enzyme in amylopectin biosynthesis.

Starch branching enzyme IIb (SBEIIb) plays a crucial role in amylopectin biosynthesis in maize endosperm by defining the structural and functional properties of storage starch and is regulated by protein phosphorylation. Native and recombinant maize SBEIIb were used as substrates for amyloplast protein kinases to identify phosphorylation sites on the protein. A multidisciplinary approach involving bioinformatics, site-directed mutagenesis, and mass spectrometry identified three phosphorylation sites at Ser residues: Ser(649), Ser(286), and Ser(297). Two Ca(2+)-dependent protein kinase activities were partially purified from amyloplasts, termed K1, responsible for Ser(649) and Ser(286) phosphorylation, and K2, responsible for Ser(649) and Ser(297) phosphorylation. The Ser(286) and Ser(297) phosphorylation sites are conserved in all plant branching enzymes and are located at opposite openings of the 8-stranded parallel ß-barrel of the active site, which is involved with substrate binding and catalysis. Molecular dynamics simulation analysis indicates that phospho-Ser(297) forms a stable salt bridge with Arg(665), part of a conserved Cys-containing domain in plant branching enzymes. Ser(649) conservation appears confined to the enzyme in cereals and is not universal, and is presumably associated with functions specific to seed storage. The implications of SBEIIb phosphorylation are considered in terms of the role of the enzyme and the importance of starch biosynthesis for yield and biotechnological application.