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Synovial sarcoma (SS) is an uncommon soft tissue tumor that occurs primarily in the extremities of young adults, especially in the periarticular region. In this report, we describe the rare occurrence of primary SS of the pleura in a 15-year-old boy. Histologically, the tumor demonstrated monophasic morphologic findings and showed positive staining with vimentin and Bcl-2 and focally for cytokeratin CK7. Fluorescent in situ hybridization identified t(X;18) translocation. The patient developed recurrences 20 months following resection of the tumor. The literature on this uncommon entity is reviewed, and its histogenesis, differential diagnoses, and cytologic features are also discussed. [ABSTRACT FROM AUTHOR]

In the last decade, there has been increased understanding of the pathologic features and biology of peripheral T cell lymphomas (PTCLs) through development of multi omics and molecular profiling techniques. In addition, international collaborations through multi center trials as well as prospective registry studies have improved our knowledge of host and tumor genomic factors and treatment factors affecting disease outcomes. In our review today, we aim to highlight the current epidemiology, latest advances in classification, disease biology and the evolving treatment landscape for nodal PTCLs., (© 2023 John Wiley & Sons Ltd.)

Natural products encompass a diverse range of compounds with high impact applications in consumer care, agriculture and most notably, therapeutics. However, despite the expansive chemical repertoire indicated in genomic information of microbes, only a small subset can be obtained under laboratory conditions. To increase accessible chemical space and realize Nature's full chemical potential, a multi-pronged genetic- and cultivation-based strategy has been employed to activate and upregulate natural product biosyntheses in native and heterologous strains. This data descriptor documents a characterized collection of 2,138 liquid chromatography-tandem mass spectrometry (LC/MS-MS) spectra of fermentation extracts from 54 native actinobacterial strains collected from soil and marine environments in Singapore, and their 459 activated mutants in 3 to 5 media. A total of 743 unique metabolites have been identified, with the activated mutants demonstrating an approximately 2-fold expansion in accessible chemical space over wild type strains. Interrogating this expanded chemical diversity with cheminformatic tools can provide direction for the discovery of novel natural products with desirable functional activity., (© 2024. The Author(s).)

Researchers are increasingly interested in discovering new pancreatic lipase inhibitors as anti-obesity ingredients. Medicine-and-food homology plants contain a diverse set of natural bioactive compounds with promising development potential. This study screened and identified potent pancreatic lipase inhibitors from 20 commonly consumed medicine-and-food homology plants using affinity ultrafiltration combined with spectroscopy and docking simulations. The results showed that turmeric exhibited the highest pancreatic lipase-inhibitory activity, and curcumin, demethoxycurcumin, and bisdemethoxycurcumin were discovered to be potent pancreatic lipase inhibitors within the turmeric extract, with IC 50 values of 0.52 ± 0.04, 1.12 ± 0.05, and 3.30 ± 0.08 mg/mL, respectively. In addition, the enzymatic kinetics analyses demonstrated that the inhibition type of the three curcuminoids was the reversible competitive model, and curcumin exhibited a higher binding affinity and greater impact on the secondary structure of pancreatic lipase than found with demethoxycurcumin or bisdemethoxycurcumin, as observed through fluorescence spectroscopy and circular dichroism. Furthermore, docking simulations supported the above experimental findings, and revealed that the three curcuminoids might interact with amino acid residues in the binding pocket of pancreatic lipase through non-covalent actions, such as hydrogen bonding and π-π stacking, thereby inhibiting the pancreatic lipase. Collectively, these findings suggest that the bioactive compounds of turmeric, in particular curcumin, can be promising dietary pancreatic lipase inhibitors for the prevention and management of obesity.

Techno-functional properties of protein isolates such as emulsification, foaming, and gelling serve as key indicators to determine their food applications. Conventional macro-volume techniques used to measure these techno-functional properties are usually time consuming, require large amounts of protein samples, and are impractical when diverse protein samples are handled at the early screening stage. To overcome these issues, we have developed scaled-down (miniaturized) assays to test techno-functional properties of protein samples. These assays are simple, efficient, and require <400 μl of protein solution. Specifically, the miniaturized emulsification and gelling assays require 25-fold less protein than conventional macro-volume techniques and the miniaturized foaming assay requires 100-fold less sample. The performance of these assays has been thoroughly validated using conventional techno-functional tests for each parameter. The protocols described herein offer high-throughput screening capabilities, accelerating the testing process for protein techno-functional properties and allowing for quick identification of samples of interest from diverse samples. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Miniaturized emulsification assay Alternate Protocol 1: Conventional macro-volume emulsification assay Basic Protocol 2: Miniaturized foaming assay Alternate Protocol 2: Conventional macro-volume foaming assay Basic Protocol 3: Miniaturized gelling assay Alternate Protocol 3: Conventional macro-volume gelling assay., (© 2024 Wiley Periodicals LLC.)

In recent years, CRISPR-Cas toolboxes for Streptomyces editing have rapidly accelerated natural product discovery and engineering. However, Cas efficiencies are oftentimes strain-dependent, and the commonly used Streptococcus pyogenes Cas9 (SpCas9) is notorious for having high levels of off-target toxicity effects. Thus, a variety of Cas proteins is required for greater flexibility of genetic manipulation within a wider range of Streptomyces strains. This study explored the first use of Acidaminococcus sp. Cas12j, a hypercompact Cas12 subfamily, for genome editing in Streptomyces and its potential in activating silent biosynthetic gene clusters (BGCs) to enhance natural product synthesis. While the editing efficiencies of Cas12j were not as high as previously reported efficiencies of Cas12a and Cas9, Cas12j exhibited higher transformation efficiencies compared to SpCas9. Furthermore, Cas12j demonstrated significantly improved editing efficiencies compared to Cas12a in activating BGCs in Streptomyces sp. A34053, a strain wherein both SpCas9 and Cas12a faced limitations in accessing the genome. Overall, this study expanded the repertoire of Cas proteins for genome editing in actinomycetes and highlighted not only the potential of recently characterized Cas12j in Streptomyces but also the importance of having an extensive genetic toolbox for improving the editing success of these beneficial microbes.

Tissue eosinophilia is seldom reported in B-cell lymphoma. It poses diagnostic challenges and frequently leads to the consideration of other diagnoses, particularly T-cell lymphomas. The scarce literature underscores the need for in-depth studies to enhance awareness and understanding of this phenomenon. We investigated 54 cases of B-cell lymphoma with notable tissue eosinophils, analyzing clinical information, hematoxylin and eosin staining, immunohistochemistry, and PCR-based clonality analysis. Nodal marginal zone lymphoma (NMZL) emerged as the most prevalent type (n=26), followed by B-cell lymphoma, not otherwise specified (n=13), diffuse large B-cell lymphoma (n=10), follicular lymphoma (n=2), chronic lymphocytic leukemia/small lymphocytic lymphoma (n=1), extranodal marginal zone lymphoma (n=1), and primary cutaneous marginal zone lymphoma (n=1). Shared features across different lymphoma types, best exemplified by NMZL, included plasmacytic differentiation (57.7%), increased vascularity (84.6%) with a tendency for perivascular distribution of neoplastic cells, and a tumor microenvironment abundant in T cells and histiocytes; some cases showed increased PD-1-positive cells. These features often raise consideration of angioimmunoblastic T-cell lymphoma. Along with clonality analysis, features supporting the diagnosis of B-cell lymphoma included cytological atypia in B cells rather than T cells, and the lack of follicular dendritic cell meshwork expansion. In addition, diffuse large B-cell lymphoma frequently exhibited interfollicular distribution and monocytoid appearance, indicating the possibility of transformed NMZL. Collectively, tissue eosinophilia can occur in diverse B-cell lymphomas but is most prevalent in tumors with a postgerminal stage of differentiation., Competing Interests: Conflicts of Interest and Source of Funding: This work was supported by funding from the Intramural Research Program of the Center for Cancer Research, National Cancer Institute, NIH. The authors have disclosed that they have no significant relationships with, or financial interest in, any commercial companies pertaining to this article., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)

Macrophages are abundant immune cells in the microenvironment of diffuse large B-cell lymphoma (DLBCL). Macrophage estimation by immunohistochemistry shows varying prognostic significance across studies in DLBCL, and does not provide a comprehensive analysis of macrophage subtypes. Here, using digital spatial profiling with whole transcriptome analysis of CD68+ cells, we characterize macrophages in distinct spatial niches of reactive lymphoid tissues (RLTs) and DLBCL. We reveal transcriptomic differences between macrophages within RLTs (light zone /dark zone, germinal center/ interfollicular), and between disease states (RLTs/ DLBCL), which we then use to generate six spatially-derived macrophage signatures (MacroSigs). We proceed to interrogate these MacroSigs in macrophage and DLBCL single-cell RNA-sequencing datasets, and in gene-expression data from multiple DLBCL cohorts. We show that specific MacroSigs are associated with cell-of-origin subtypes and overall survival in DLBCL. This study provides a spatially-resolved whole-transcriptome atlas of macrophages in reactive and malignant lymphoid tissues, showing biological and clinical significance., (© 2024. The Author(s).)

Background and Aims: ChatGPT is a powerful artificial intelligence (AI) chatbot developed by the OpenAI research laboratory which is capable of analysing human input and generating human-like responses. Early research into the potential application of ChatGPT in healthcare has focused mainly on clinical and administrative functions. The diagnostic ability and utility of ChatGPT in histopathology is not well defined. We benchmarked the performance of ChatGPT against pathologists in diagnostic histopathology, and evaluated the collaborative potential between pathologists and ChatGPT to deliver more accurate diagnoses., Methods and Results: In Part 1 of the study, pathologists and ChatGPT were subjected to a series of questions encompassing common diagnostic conundrums in histopathology. For Part 2, pathologists reviewed a series of challenging virtual slides and provided their diagnoses before and after consultation with ChatGPT. We found that ChatGPT performed worse than pathologists in reaching the correct diagnosis. Consultation with ChatGPT provided limited help and information generated from ChatGPT is dependent on the prompts provided by the pathologists and is not always correct. Finally, we surveyed pathologists who rated the diagnostic accuracy of ChatGPT poorly, but found it useful as an advanced search engine., Conclusions: The use of ChatGPT4 as a diagnostic tool in histopathology is limited by its inherent shortcomings. Judicious evaluation of the information and histopathology diagnosis generated from ChatGPT4 is essential and cannot replace the acuity and judgement of a pathologist. However, future advances in generative AI may expand its role in the field of histopathology., (© 2023 The Authors. Histopathology published by John Wiley & Sons Ltd.)

This study investigated the non-volatile metabolites and antioxidant activity of Douchi, an edible mushroom by-product. A total of 695 non-volatile metabolites were detected using UPLC-MS/MS-based metabolomics analysis, and the greatest impact on metabolite composition was observed during Koji-making and the first 5 days of post-fermentation. Throughout the fermentation process, 366 differential metabolites were identified, with flavonoids being the most prominent followed by amino acids and their derivatives, which were found to be important for the quality of edible mushroom by-product Douchi (EMD). The antioxidant capacity of EMD significantly increased with the longer fermentation time, which might be associated with the conversion of isoflavone glycosides to aglycones, amino acids and their derivatives, free fatty acids, group A saponins, and phenolic acids. These findings suggested that different fermentation phases of EMD significantly affected the non-volatile metabolite profile and antioxidant capacity., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Natural products possess significant therapeutic potential but remain underutilized despite advances in genomics and bioinformatics. While there are approaches to activate and upregulate natural product biosynthesis in both native and heterologous microbial strains, a comprehensive strategy to elicit production of natural products as well as a generalizable and efficient method to interrogate diverse native strains collection, remains lacking. Here, we explore a flexible and robust integrase-mediated multi-pronged activation approach to reliably perturb and globally trigger antibiotics production in actinobacteria. Across 54 actinobacterial strains, our approach yielded 124 distinct activator-strain combinations which consistently outperform wild type. Our approach expands accessible metabolite space by nearly two-fold and increases selected metabolite yields by up to >200-fold, enabling discovery of Gram-negative bioactivity in tetramic acid analogs. We envision these findings as a gateway towards a more streamlined, accelerated, and scalable strategy to unlock the full potential of Nature's chemical repertoire., (© 2024. The Author(s).)

Osmanthus fragrans ( O. fragrans ) has been cultivated in China for over 2,500 years as a traditional fragrant plant. Recently, O. fragrans has drawn increasing attention due to its unique aroma and potential health benefits. In this review, the aroma and functional components of O. fragrans are summarized, and their biosynthetic mechanism is discussed. The beneficial functions and related molecular mechanism of O. fragrans extract are then highlighted. Finally, potential applications of O. fragrans are summarized, and future perspectives are proposed and discussed. According to the current research, O. fragrans extracts and components have great potential to be developed into value-added functional ingredients with preventive effects on certain chronic diseases. However, it is crucial to develop efficient, large-scale, and commercially viable extraction methods to obtain the bioactive components from O. fragrans . Furthermore, more clinical studies are highly needed to explore the beneficial functions of O. fragrans and guide its development into functional food products.

Multiple beneficial effects have been attributed to green tea catechins (GTCs). However, the bioavailability of GTCs is generally low, with only a small portion directly absorbed in the small intestine. The majority of ingested GTCs reaches the large intestinal lumen, and are extensively degraded via biotransformation by gut microbiota, forming many low-molecular-weight metabolites such as phenyl-γ-valerolactones, phenolic acids, butyrate, and acetate. This process not only improves the overall bioavailability of GTC-derived metabolites but also enriches the biological activities of GTCs. Therefore, the intra- and inter-individual differences in human gut microbiota as well as the resulting biological contribution of microbial metabolites are crucial for the ultimate health benefits. In this review, the microbial degradation of major GTCs was characterized and an overview of the in vitro models used for GTC metabolism was summarized. The intra- and inter-individual differences of human gut microbiota composition and the resulting divergence in the metabolic patterns of GTCs were highlighted. Moreover, the potential beneficial effects of GTCs and their gut microbial metabolites were also discussed. Overall, the microbial metabolites of GTCs with higher bioavailability and bioactive potency are key factors for the observed beneficial effects of GTCs and green tea consumption.

Background: Social behaviors such as altruism, where one self-sacrifices for collective benefits, critically influence an organism's survival and responses to the environment. Such behaviors are widely exemplified in nature but have been underexplored in cancer cells which are conventionally seen as selfish competitive players. This multidisciplinary study explores altruism and its mechanism in breast cancer cells and its contribution to chemoresistance., Methods: MicroRNA profiling was performed on circulating tumor cells collected from the blood of treated breast cancer patients. Cancer cell lines ectopically expressing candidate miRNA were used in co-culture experiments and treated with docetaxel. Ecological parameters like relative survival and relative fitness were assessed using flow cytometry. Functional studies and characterization performed in vitro and in vivo include proliferation, iTRAQ-mass spectrometry, RNA sequencing, inhibition by small molecules and antibodies, siRNA knockdown, CRISPR/dCas9 inhibition and fluorescence imaging of promoter reporter-expressing cells. Mathematical modeling based on evolutionary game theory was performed to simulate spatial organization of cancer cells., Results: Opposing cancer processes underlie altruism: an oncogenic process involving secretion of IGFBP2 and CCL28 by the altruists to induce survival benefits in neighboring cells under taxane exposure, and a self-sacrificial tumor suppressive process impeding proliferation of altruists via cell cycle arrest. Both processes are regulated concurrently in the altruists by miR-125b, via differential NF-κB signaling specifically through IKKβ. Altruistic cells persist in the tumor despite their self-sacrifice, as they can regenerate epigenetically from non-altruists via a KLF2/PCAF-mediated mechanism. The altruists maintain a sparse spatial organization by inhibiting surrounding cells from adopting the altruistic fate via a lateral inhibition mechanism involving a GAB1-PI3K-AKT-miR-125b signaling circuit., Conclusions: Our data reveal molecular mechanisms underlying manifestation, persistence and spatial spread of cancer cell altruism. A minor population behave altruistically at a cost to itself producing a collective benefit for the tumor, suggesting tumors to be dynamic social systems governed by the same rules of cooperation in social organisms. Understanding cancer cell altruism may lead to more holistic models of tumor evolution and drug response, as well as therapeutic paradigms that account for social interactions. Cancer cells constitute tractable experimental models for fields beyond oncology, like evolutionary ecology and game theory., (© 2023. The Author(s).)

Background: Overexpressed EZH2 is oncogenically involved in the pathogenesis of different cancerous contexts including extranodal natural killer/T cell lymphoma (ENKTL). However, the underlying mechanisms of EZH2 upregulation have not been fully clarified and it is still difficult to target EZH2 in ENKTL., Results: Current study identifies an E3 ligase TRIP12 that triggers K63-linked polyubiquitination of EZH2 in ENKTL and unexpectedly, stabilizes EZH2. As determined by gene expression profiling (GEP), TRIP12 and EZH2 levels correlate with each other in ENKTL patient samples. Aided by quantitative mass spectrometry (MS) and follow-up analysis, we identify K634 as the ubiquitination site of EZH2. Further study confirms that TRIP12-mediated EZH2 K634 ubiquitination enhances the interaction between EZH2 and SUZ12 or CDK1 and increases the level of EZH2 T487 phosphorylation. This study further demonstrates the TRIP12-EZH2 signaling might be regulated by cytoplasmic HSP60. Importantly, the TRIP12-EZH2 axis mediates ENKTL cell migration via accelerating epithelial-mesenchymal transition (EMT). Moreover, our study finds out dexamethasone treatment manipulates TRIP12-EZH2 signaling and may represent a novel therapeutic strategy against ENKTL metastasis., Conclusions: Altogether, TRIP12 induces K63-linked site-specific polyubiquitination of EZH2 for stabilization, which promotes ENKTL cell migration and could be targeted by dexamethasone treatment., (© 2023. The Author(s).)

The 2022 European Association for Haematopathology/Society for Hematopathology lymphoma workshop session on cavity-based lymphomas included sixty-eight cases in seven sections. The disease entities discussed include primary effusion lymphomas (PEL), extracavitary primary effusion lymphomas and confounding entities (ECPEL), HHV8-negative B-lineage lymphomas-effusion based (EBV-negative, EBV-positive, and plasmablastic types), diffuse large B-cell lymphoma associated with chronic inflammation, fibrin-associated diffuse large B-cell lymphoma (FA-DLBCL), breast implant-associated anaplastic large cell lymphoma (BIA-ALCL), and other lymphomas presenting as an effusion. All entities above are discussed; however, three are delved into greater detail given the challenges with classification: ECPEL, HHV8-negative effusion-based lymphomas, and FA-DLBCL. Cases exemplifying the diagnostic difficulty in differentiating ECPEL from HHV8-positive diffuse large B-cell lymphoma and germinotropic lymphoproliferative disorder were discussed. The more recently recognized effusion-based HHV8-negative large B-cell lymphoma is explored, with several cases submitted raising the question if this subset should be carved out as a specific entity, and if so, what should be the refining diagnostic criteria. Case submissions to the FA-DLBCL section yielded one of the largest case series to date, including classic cases, cases furthering the discussion on disease sites and prognosis, as well as novel concepts to be considered in this entity. The 2022 EA4HP/SH workshop cases allowed for further confirmation of the characteristics of some of the more historically accepted cavity-based lymphomas, as well as further inquiry and debate on relatively new or evolving entities., (© 2023. The Author(s).)

Cytotoxic peripheral T-cell lymphomas and EBV-positive T/NK-cell lymphoproliferative diseases were discussed at the 2022 European Association for Haematopathology/Society for Hematopathology lymphoma workshop held in Florence, Italy. This session focused on (i) primary nodal EBV-positive T and NK-cell lymphomas (primary nodal-EBV-TNKL), (ii) extranodal EBV-positive T/NK lymphoproliferative diseases (LPD) in children and adults, (iii) cytotoxic peripheral T-cell lymphomas, NOS (cPTCL-NOS), EBV-negative, and (iv) miscellaneous cases. Primary nodal-EBV-TNKL is a newly recognized entity which is rare, aggressive, and associated with underlying immune deficiency/immune dysregulation. All cases presented with lymphadenopathy but some demonstrated involvement of tonsil/Waldeyer's ring and extranodal sites. The majority of tumors are of T-cell lineage, and the most frequent mutations involve the epigenetic modifier genes, such as TET2 and DNMT3A, and JAK-STAT genes. A spectrum of EBV-positive T/NK LPD involving extranodal sites were discussed and highlight the diagnostic challenge with primary nodal-EBV-TNKL when these extranodal EBV-positive T/NK LPD cases demonstrate predominant nodal disease either at presentation or during disease progression from chronic active EBV disease. The majority of cPTCL-NOS demonstrated the TBX21 phenotype. Some cases had a background of immunosuppression or immune dysregulation. Interestingly, an unexpected association of cPTCL-NOS, EBV-positive and negative, with TFH lymphomas/LPDs was observed in the workshop cases. Similar to a published literature, the genetic landscape of cPTCL-NOS from the workshop showed frequent mutations in epigenetic modifiers, including TET2 and DNMT3A, suggesting a role of clonal hematopoiesis in the disease pathogenesis., (© 2023. The Author(s).)

Follicular helper T-cell lymphomas (TFH lymphomas) were discussed in session V of the lymphoma workshop of the European Association for Haematopathology (EA4HP)/Society for Hematopathology (SH) 2022 meeting in Florence, Italy. The session focused on the morphologic spectrum of TFH lymphoma, including its three subtypes: angioimmunoblastic-type (AITL), follicular-type, and not otherwise specified (NOS). The submitted cases encompassed classic examples of TFH lymphoma and unusual cases such as those with early or indolent presentations, associated B-cell proliferations, or Hodgkin/Reed-Sternberg-like cells. The relationship between TFH lymphoma and clonal hematopoiesis was highlighted by several cases documenting divergent evolution of myeloid neoplasm and AITL from shared clonal mutations. The distinction between TFH lymphoma and peripheral T-cell lymphoma, not otherwise specified (PTCL, NOS), was stressed, and many challenging examples were presented. Various cases highlighted the difficulties of differentiating TFH lymphoma from other established types of lymphoma and reactive conditions. Cutaneous T-cell lymphoma expressing TFH markers, particularly when resulting in lymph node involvement, should be distinguished from TFH lymphomas. Additional immunophenotyping and next-generation sequencing studies were performed on various cases in this session, highlighting the importance of these technologies to our current understanding and classification of TFH lymphomas., (© 2023. The Author(s).)

Session 3 of the lymphoma workshop of the XXI joint meeting of the European Association for Haematopathology and the Society for Hematopathology took place in Florence, Italy, on September 22, 2022. The topics of this session were splenic and nodal marginal zone lymphomas, transformation in marginal zone lymphomas, and pediatric nodal marginal zone lymphomas and their differential diagnosis as well as related entities. Forty-two cases in these categories were submitted to the workshop, including splenic lymphomas (marginal zone and diffuse red pulp lymphomas), transformed marginal zone lymphomas (splenic and nodal), nodal marginal zone lymphomas with increased TFH-cells, and pediatric nodal marginal zone lymphomas. The case review highlighted some of the principal problems in the diagnosis of marginal zone lymphomas, including the difficulties in the distinction between splenic marginal zone lymphoma, splenic diffuse red pulp lymphoma, and hairy cell leukemia variant/splenic B-cell lymphoma with prominent nucleoli which requires integration of clinical features, immunophenotype, and morphology in blood, bone marrow, and spleen; cases of marginal zone lymphoma with markedly increased TFH-cells, simulating a T-cell lymphoma, where molecular studies (clonality and mutation detection) can help to establish the final diagnosis; the criteria for transformation of marginal zone lymphomas, which are still unclear and might require the integration of morphological and molecular data; the concept of an overlapping spectrum between pediatric nodal marginal zone lymphoma and pediatric-type follicular lymphoma; and the distinction between pediatric nodal marginal zone lymphoma and "atypical" marginal zone hyperplasia, where molecular studies are mandatory to correctly classify cases., (© 2023. The Author(s).)

Emerging entities and molecular subgroups in large B-cell lymphomas (LBCLs) were discussed during the 2022 European Association for Haematopathology/Society for Hematopathology workshop in Florence, Italy. This session focused on newly recognized diseases and their diagnostic challenges. High-grade/large B-cell lymphoma with 11q aberration (HG/LBCL-11q) is defined by chromosome 11q-gains and telomeric loss. FISH analysis is recommended for the diagnosis. HG/LBCL-11q can occur in the setting of immunodeficiency, including ataxia-telangiectasia, and predominates in children. The morphological spectrum of these cases is broader than previously thought with often Burkitt-like morphology and coarse apoptotic bodies. It has a Burkitt-like immunophenotype (CD10+, BCL6+, BCL2-) but MYC expression is weak or negative, lacks MYC rearrangement, and is in contrast to Burkitt lymphoma 50% of the cases express LMO2. LBCL with IRF4 rearrangement (LBCL-IRF4) occurs mainly in the pediatric population but also in adults. LBCL-IRF4 has an excellent prognosis, with distinguishing molecular findings. IRF4 rearrangements, although characteristic of this entity, are not specific and can be found in association with other chromosomal translocations in other large B-cell lymphomas. Other molecular subgroups discussed included primary bone diffuse large B-cell lymphoma (PB-DLBCL), which has distinctive clinical presentation and molecular findings, and B-acute lymphoblastic leukemia (B-ALL) with IGH::MYC translocation recently segregated from Burkitt lymphoma with TdT expression. This latter disorder has molecular features of precursor B-cells, often tetrasomy 1q and recurrent NRAS and KRAS mutations. In this report, novel findings, recommendations for diagnosis, open questions, and diagnostic challenges raised by the cases submitted to the workshop will be discussed., (© 2023. The Author(s).)

Natural products have long been used as a source of antimicrobial agents against various microorganisms. Actinobacteria are a group of bacteria best known to produce a wide variety of bioactive secondary metabolites, including many antimicrobial agents. In this study, four actinobacterial strains found in Singapore terrestrial soil were investigated as potential sources of new antimicrobial compounds. Large-scale cultivation, chemical, and biological investigation led to the isolation of a previously undescribed tetronomycin A ( 1 ) that demonstrated inhibitory activities against both Gram-positive bacteria Staphylococcus aureus (SA) and methicillin-resistant Staphylococcus aureus (MRSA) (i.e., MIC 90 of 2-4 μM and MBC 90 of 9-12 μM), and several known antimicrobial compounds, namely nonactin, monactin, dinactin, 4E-deacetylchromomycin A3, chromomycin A2, soyasaponin II, lysolipin I, tetronomycin, and naphthomevalin. Tetronomycin showed a two- to six-fold increase in antibacterial activity (i.e., MIC 90 and MBC 90 of 1-2 μM) as compared to tetronomycin A ( 1 ), indicating the presence of an oxy-methyl group at the C-27 position is important for antibacterial activity.

Cancers often overexpress multiple clinically relevant oncogenes, but it is not known if combinations of oncogenes in cellular subpopulations within a cancer influence clinical outcomes. Using quantitative multispectral imaging of the prognostically relevant oncogenes MYC, BCL2, and BCL6 in diffuse large B-cell lymphoma (DLBCL), we show that the percentage of cells with a unique combination MYC+BCL2+BCL6- (M+2+6-) consistently predicts survival across four independent cohorts (n = 449), an effect not observed with other combinations including M+2+6+. We show that the M+2+6- percentage can be mathematically derived from quantitative measurements of the individual oncogenes and correlates with survival in IHC (n = 316) and gene expression (n = 2,521) datasets. Comparative bulk/single-cell transcriptomic analyses of DLBCL samples and MYC/BCL2/BCL6-transformed primary B cells identify molecular features, including cyclin D2 and PI3K/AKT as candidate regulators of M+2+6- unfavorable biology. Similar analyses evaluating oncogenic combinations at single-cell resolution in other cancers may facilitate an understanding of cancer evolution and therapy resistance., Significance: Using single-cell-resolved multiplexed imaging, we show that selected subpopulations of cells expressing specific combinations of oncogenes influence clinical outcomes in lymphoma. We describe a probabilistic metric for the estimation of cellular oncogenic coexpression from IHC or bulk transcriptomes, with possible implications for prognostication and therapeutic target discovery in cancer. This article is highlighted in the In This Issue feature, p. 1027., (©2023 The Authors; Published by the American Association for Cancer Research.)

Background: Nature has provided unique molecular scaffolds for applications including therapeutics, agriculture, and food. Due to differences in ecological environments and laboratory conditions, engineering is often necessary to uncover and utilize the chemical diversity. Although we can efficiently activate and mine these often complex 3D molecules, sufficient production of target molecules for further engineering and application remain a considerable bottleneck. An example of these bioactive scaffolds is armeniaspirols, which are potent polyketide antibiotics against gram-positive pathogens and multi-resistance gram-negative Helicobacter pylori. Here, we examine the upregulation of armeniaspirols in an alternative Streptomyces producer, Streptomyces sp. A793., Results: Through an incidental observation of enhanced yields with the removal of a competing polyketide cluster, we observed seven-fold improvement in armeniaspirol production. To further investigate the improvement of armeniaspirol production, we examine upregulation of armeniaspirols through engineering of biosynthetic pathways and primary metabolism; including perturbation of genes in biosynthetic gene clusters and regulation of triacylglycerols pool., Conclusion: With either overexpression of extender unit pathway or late-stage N-methylation, or the deletion of a competing polyketide cluster, we can achieve seven-fold to forty nine-fold upregulation of armeniaspirol production. The most significant upregulation was achieved by expression of heterologous fatty acyl-CoA synthase, where we observed not only a ninety seven-fold increase in production yields compared to wild type, but also an increase in the diversity of observed armeniaspirol intermediates and analogs., (© 2023. The Author(s).)

Background: Extranodal natural killer/T-cell lymphoma (NKTL) is an aggressive type of non-Hodgkin lymphoma with dismal outcome. A better understanding of disease biology and key oncogenic process is necessary for the development of targeted therapy. Super-enhancers (SEs) have been shown to drive pivotal oncogenes in various malignancies. However, the landscape of SEs and SE-associated oncogenes remain elusive in NKTL., Methods: We used Nano-ChIP-seq of the active enhancer marker histone H3 lysine 27 acetylation (H3K27ac) to profile unique SEs NKTL primary tumor samples. Integrative analysis of RNA-seq and survival data further pinned down high value, novel SE oncogenes. We utilized shRNA knockdown, CRISPR-dCas9, luciferase reporter assay, ChIP-PCR to investigate the regulation of transcription factor (TF) on SE oncogenes. Multi-color immunofluorescence (mIF) staining was performed on an independent cohort of clinical samples. Various function experiments were performed to evaluate the effects of TOX2 on the malignancy of NKTL in vitro and in vivo., Results: SE landscape was substantially different in NKTL samples in comparison with normal tonsils. Several SEs at key transcriptional factor (TF) genes, including TOX2, TBX21(T-bet), EOMES, RUNX2, and ID2, were identified. We confirmed that TOX2 was aberrantly overexpressed in NKTL relative to normal NK cells and high expression of TOX2 was associated with worse survival. Modulation of TOX2 expression by shRNA, CRISPR-dCas9 interference of SE function impacted on cell proliferation, survival and colony formation ability of NKTL cells. Mechanistically, we found that RUNX3 regulates TOX2 transcription by binding to the active elements of its SE. Silencing TOX2 also impaired tumor formation of NKTL cells in vivo. Metastasis-associated phosphatase PRL-3 has been identified and validated as a key downstream effector of TOX2-mediated oncogenesis., Conclusions: Our integrative SE profiling strategy revealed the landscape of SEs, novel targets and insights into molecular pathogenesis of NKTL. The RUNX3-TOX2-SE-TOX2-PRL-3 regulatory pathway may represent a hallmark of NKTL biology. Targeting TOX2 could be a valuable therapeutic intervene for NKTL patients and warrants further study in clinic., (© 2023. The Author(s).)

With the advent of rapid automated in silico identification of biosynthetic gene clusters (BGCs), genomics presents vast opportunities to accelerate natural product (NP) discovery. However, prolific NP producers, Streptomyces , are exceptionally GC-rich (>80%) and highly repetitive within BGCs. These pose challenges in sequencing and high-quality genome assembly which are currently circumvented via intensive sequencing. Here, we outline a more cost-effective workflow using multiplex Illumina and Oxford Nanopore sequencing with hybrid long-short read assembly algorithms to generate high quality genomes. Our protocol involves subjecting long read-derived assemblies to up to 4 rounds of polishing with short reads to yield accurate BGC predictions. We successfully sequenced and assembled 8 GC-rich Streptomyces genomes whose lengths range from 7.1 to 12.1 Mb with a median N50 of 8.2 Mb. Taxonomic analysis revealed previous misrepresentation among these strains and allowed us to propose a potentially new species, Streptomyces sydneybrenneri . Further comprehensive characterization of their biosynthetic, pan-genomic and antibiotic resistance features especially for molecules derived from type I polyketide synthase (PKS) BGCs reflected their potential as alternative NP hosts. Thus, the genome assemblies and insights presented here are envisioned to serve as gateway for the scientific community to expand their avenues in NP discovery., Competing Interests: The authors declare no competing interests., (© 2023 The Authors.)

Adaptation to a wide variety of habitats allows fungi to develop unique abilities to produce diverse secondary metabolites with diverse bioactivities. In this study, 30 Ascomycetes fungi isolated from St. John's Island, Singapore were investigated for their general biosynthetic potential and their ability to produce antimicrobial secondary metabolites (SMs). All the 30 fungal isolates belong to the Phylum Ascomycota and are distributed into 6 orders and 18 genera with Order Hypocreales having the highest number of representative (37%). Screening for polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) genes using degenerate PCR led to the identification of 23 polyketide synthases (PKSs) and 5 nonribosomal peptide synthetases (NRPSs) grouped into nine distinct clades based on their reduction capabilities. Some of the identified PKSs genes share high similarities between species and known reference genes, suggesting the possibility of conserved biosynthesis of closely related compounds from different fungi. Fungal extracts were tested for their antimicrobial activity against S. aureus , Methicillin-resistant S. aureus (MRSA), and Candida albicans . Bioassay-guided fractionation of the active constituents from two promising isolates resulted in the isolation of seven compounds: Penilumamides A, D, and E from strain F4335 and xanthomegnin, viomellein, pretrichodermamide C and vioxanthin from strain F7180. Vioxanthin exhibited the best antibacterial activity with IC 50 values of 3.0 μM and 1.6 μM against S. aureus and MRSA respectively. Viomellein revealed weak antiproliferative activity against A549 cells with an IC 50 of 42 μM. The results from this study give valuable insights into the diversity and biosynthetic potential of fungi from this unique habitat and forms a background for an in-depth analysis of the biosynthetic capability of selected strains of interest with the aim of discovering novel fungal natural products.

Thiopeptides are macrocyclic natural products with potent bioactivity. Nine new natural thiopeptides (1−9) were obtained from a Nonomuraea jiangxiensis isolated from a terrestrial soil sample collected in Singapore. Even though some of these compounds were previously synthesized or isolated from engineered strains, herein we report the unprecedented isolation of these thiopeptides from a native Nonomuraea jiangxiensis. A comparison with the literature and a detailed analysis of the NMR and HRMS of compounds 1−9 was conducted to assign their chemical structures. The structures of all new compounds were highly related to the thiopeptide antibiotics GE2270, with variations in the substituents on the thiazole and amino acid moieties. Thiopeptides 1−9 exhibited a potent antimicrobial activity against the Gram-positive bacteria, Staphylococcus aureus with MIC90 values ranging from 2 µM to 11 µM. In addition, all compounds were investigated for their cytotoxicity against the human cancer cell line A549, none of the compounds were cytotoxic.

Angucyclines are a family of structurally diverse, aromatic polyketides with some members that exhibit potent bioactivity. Angucyclines have also attracted considerable attention due to the intriguing biosynthetic origins that underlie their structural complexity and diversity. Balmoralmycin (compound 1) represents a unique group of angucyclines that contain an angular benz[ α ]anthracene tetracyclic system, a characteristic C-glycosidic bond-linked deoxy-sugar (d-olivose), and an unsaturated fatty acid chain. In this study, we identified a Streptomyces strain that produces balmoralmycin and seven biosynthetically related coproducts (compounds 2-8). Four of the coproducts (compounds 5-8) are novel compounds that feature a highly oxygenated or fragmented lactone ring, and three of them (compounds 3-5) exhibited cytotoxicity against the human pancreatic cancer cell line MIA PaCa-2 with IC 50 values ranging from 0.9 to 1.2 μg/mL. Genome sequencing and CRISPR/dCas9-assisted gene knockdown led to the identification of the ~43 kb balmoralmycin biosynthetic gene cluster ( bal BGC). The bal BGC encodes a type II polyketide synthase (PKS) system for assembling the angucycline aglycone, six enzymes for generating the deoxysugar d-olivose, and a hybrid type II/III PKS system for synthesizing the 2,4-decadienoic acid chain. Based on the genetic and chemical information, we propose a mechanism for the biosynthesis of balmoralmycin and the shunt products. The chemical and genetic studies yielded insights into the biosynthetic origin of the structural diversity of angucyclines. IMPORTANCE Angucyclines are structurally diverse aromatic polyketides that have attracted considerable attention due to their potent bioactivity and intriguing biosynthetic origin. Balmoralmycin is a representative of a small family of angucyclines with unique structural features and an unknown biosynthetic origin. We report a newly isolated Streptomyces strain that produces balmoralmycin in a high fermentation titer as well as several structurally related shunt products. Based on the chemical and genetic information, a biosynthetic pathway that involves a type II polyketide synthase (PKS) system, cyclases/aromatases, oxidoreductases, and other ancillary enzymes was established. The elucidation of the balmoralmycin pathway enriches our understanding of how structural diversity is generated in angucyclines and opens the door for the production of balmoralmycin derivatives via pathway engineering.

BCL11B is an essential transcription factor for T-cell lineage commitment and differentiation, and its dysregulation has been shown to be associated with T-cell tumourigenesis. In this study, we investigated BCL11B expression by immunohistochemical analysis in 120 cases of mature T-cell lymphoma, 34 B-cell lymphomas, 11 NK-cell neoplasms and 17 reactive cutaneous conditions. All cases of mycosis fungoides (n=23), primary cutaneous CD4+ small/medium T-cell lymphoproliferative disorder (n=8) and T-prolymphocytic leukaemia (n=6) were positive for BCL11B and the staining intensity was higher than that of reactive T-cells. Fourteen of 15 (93%) cases of angioimmunoblastic T-cell lymphoma, 10 of 12 (83%) T-large granular lymphocytic leukaemia and 14 of 20 (70%) peripheral T-cell lymphoma, not otherwise specified, were also positive for BCL11B with an intensity comparable to reactive T-cells. Other T-cell neoplasms were uncommonly positive including one of three (33%) cases of primary cutaneous gamma delta T-cell lymphoma, one of four (25%) cases of subcutaneous panniculitis-like T-cell lymphoma, one of four (25%) cases of hepatosplenic T-cell lymphoma, and one of 20 (5%) cases of anaplastic large cell lymphoma (8 ALK-positive, 12 ALK-negative). T-cells in reactive cutaneous infiltrates were also positive for BCL11B, but staining intensity was much weaker than in mycosis fungoides. All NK-cell (n=11) and B-cell neoplasms (n=34) were negative for BCL11B. In conclusion, BCL11B shows a distinct expression pattern in various T-cell neoplasms. BCL11B appears to have utility as another T-cell marker and may be useful in the differential diagnosis of lymphoid neoplasms., (Copyright © 2022 Royal College of Pathologists of Australasia. Published by Elsevier B.V. All rights reserved.)

Large scale cultivation and chemical investigation of an extract obtained from Actimonadura sp. resulted in the identification of six previously undescribed spirotetronates (pyrrolosporin B and decatromicins C-G; 7 - 12 ), along with six known congeners, namely decatromicins A-B ( 1 - 2 ), BE-45722B-D ( 3 - 5 ), and pyrrolosporin A ( 6 ). The chemical structures of compounds 1 - 12 were characterized via comparison with previously reported data and analysis of 1D/2D NMR and MS data. The structures of all new compounds were highly related to the spirotetronate type compounds, decatromicin and pyrrolosporin, with variations in the substituents on the pyrrole and aglycone moieties. All compounds were evaluated for antibacterial activity against the Gram-negative bacteria, Acinetobacter baumannii and Gram-positive bacteria, Staphylococcus aureus and were investigated for their cytotoxicity against the human cancer cell line A549. Of these, decatromicin B ( 2 ), BE-45722B ( 3 ), and pyrrolosporin B ( 7 ) exhibited potent antibacterial activities against both Gram-positive (MIC 90 between 1-3 μM) and Gram-negative bacteria (MIC 90 values ranging from 12-36 μM) with weak or no cytotoxic activity against A549 cells.

The present study investigated the molecular phylogeny, antimicrobial and cytotoxic activities of fungal endophytes obtained from the A*STAR Natural Organism Library (NOL) and previously isolated from Sungei Buloh Wetland Reserve, Singapore. Phylogenetic analysis based on ITS2 gene suggests that these isolates belong to 46 morphotypes and are affiliated to 23 different taxa in 17 genera of the Ascomycota phylum. Colletotrichum was the most dominant fungal genus accounting for 37% of all the isolates, followed by Diaporthe (13%), Phyllosticta (10.9%) and Diplodia (8.7%) . Chemical elicitation using 5-azacytidine, a DNA methyltransferase inhibitor and suberoylanilide hydroxamic acid, a histone deacetylase inhibitor resulted in an increase in the number of active strains. Bioassay-guided isolation and structural elucidation yielded pestahivin and two new analogues from Bartalinia sp. F9447. Pestahivin and its related analogues did not exhibit antibacterial activity against Staphylococcus aureus but displayed strong antifungal activities against Candida albicans and Aspergillus brasiliensis, with IC 50 values ranging from 0.46 ± 0.06 to 144 ± 18 µM. Pestahivin and its two analogues furthermore exhibited cytotoxic activity against A549 and MIA PACA-2 cancer cell lines with IC 50 values in the range of 0.65 ± 0.12 to 42 ± 5.2 µM. The finding from this study reinforces that chemical epigenetic induction is a promising approach for the discovery of bioactive fungal secondary metabolites encoded by cryptic gene clusters., 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., (Copyright © 2022 Munusamy, Ching, Yang, Crasta, Gakuubi, Chee, Wibowo, Leong, Kanagasundaram and Ng.)

Although combination therapy is the standard of care for relapsed/refractory non-Hodgkin's lymphoma (RR-NHL), combination treatment chosen for an individual patient is empirical, and response rates remain poor in individuals with chemotherapy-resistant disease. Here, we evaluate an experimental-analytic method, quadratic phenotypic optimization platform (QPOP), for prediction of patient-specific drug combination efficacy from a limited quantity of biopsied tumor samples. In this prospective study, we enrolled 71 patients with RR-NHL (39 B cell NHL and 32 NK/T cell NHL) with a median of two prior lines of treatment, at two academic hospitals in Singapore from November 2017 to August 2021. Fresh biopsies underwent ex vivo testing using a panel of 12 drugs with known efficacy against NHL to identify effective single and combination treatments. Individualized QPOP reports were generated for 67 of 75 patient samples, with a median turnaround time of 6 days from sample collection to report generation. Doublet drug combinations containing copanlisib or romidepsin were most effective against B cell NHL and NK/T cell NHL samples, respectively. Off-label QPOP-guided therapy offered at physician discretion in the absence of standard options ( n  = 17) resulted in five complete responses. Among patients with more than two prior lines of therapy, the rates of progressive disease were lower with QPOP-guided treatments than with conventional chemotherapy. Overall, this study shows that the identification of patient-specific drug combinations through ex vivo analysis was achievable for RR-NHL in a clinically applicable time frame. These data provide the basis for a prospective clinical trial evaluating ex vivo-guided combination therapy in RR-NHL.

Lophiotrema is a genus of ascomycetous fungi within the family Lophiotremataceae . Members of this genus have been isolated as endophytes from a wide range of host plants and also from plant debris within terrestrial and marine habitats, where they are thought to function as saprobes. Lophiotrema sp. F6932 was isolated from white mangrove ( Avicennia officinalis ) in Pulau Ubin Island, Singapore. Crude extracts from the fungus exhibited strong antibacterial activity, and bioassay-guided isolation and structure elucidation of bioactive constituents led to the isolation of palmarumycin C 8 and a new analog palmarumycin CP 30 . Whole-genome sequencing analysis resulted in the identification of a putative type 1 iterative PKS (iPKS) predicated to be involved in the biosynthesis of palmarumycins. To verify the involvement of palmarumycin (PAL) gene cluster in the biosynthesis of these compounds, we employed ribonucleoprotein (RNP) - mediated CRISPR-Cas9 to induce targeted deletion of the ketosynthase (KS) domain in PAL. Double-strand breaks (DSBs) upstream and downstream of the KS domain was followed by homology-directed repair (HDR) with a hygromycin resistance cassette flanked by a 50 bp of homology on both sides of the DSBs. The resultant deletion mutants displayed completely different phenotypes compared to the wild-type strain, as they had different colony morphology and were no longer able to produce palmarumycins or melanin. This study, therefore, confirms the involvement of PAL in the biosynthesis of palmarumycins, and paves the way for implementing a similar approach in the characterization of other gene clusters of interest in this largely understudied fungal strain., 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., (Copyright © 2022 Gakuubi, Ching, Munusamy, Wibowo, Lim, Ma, Liang, Kanagasundaram and Ng.)

Genome-wide analysis of cell-free DNA methylation profile is a promising approach for sensitive and specific detection of many cancers. However, scaling such assays for clinical translation is impractical because of the high cost of whole-genome bisulfite sequencing. We show that the small fraction of GC-rich genome is highly enriched in CpG sites and disproportionately harbors most of the cancer-specific methylation signature. Here, we report on the simple and effective heat enrichment of CpG-rich regions for bisulfite sequencing (Heatrich-BS) platform that allows for focused methylation profiling in these highly informative regions. Our novel method and bioinformatics algorithm enable accurate tumor burden estimation and quantitative tracking of colorectal cancer patient's response to treatment at much reduced sequencing cost suitable for frequent monitoring. We also show tumor epigenetic subtyping using Heatrich-BS, which could enable patient stratification. Heatrich-BS holds great potential for highly scalable screening and monitoring of cancer using liquid biopsy.

With lowering costs of sequencing and genetic profiling techniques, genetic drivers can now be detected readily in tumors but current prognostic models for Natural-killer/T cell lymphoma (NKTCL) have yet to fully leverage on them for prognosticating patients. Here, we used next-generation sequencing to sequence 260 NKTCL tumors, and trained a genomic prognostic model (GPM) with the genomic mutations and survival data from this retrospective cohort of patients using LASSO Cox regression. The GPM is defined by the mutational status of 13 prognostic genes and is weakly correlated with the risk-features in International Prognostic Index (IPI), Prognostic Index for Natural-Killer cell lymphoma (PINK), and PINK-Epstein-Barr virus (PINK-E). Cox-proportional hazard multivariate regression also showed that the new GPM is independent and significant for both progression-free survival (PFS, HR: 3.73, 95% CI 2.07-6.73; p < .001) and overall survival (OS, HR: 5.23, 95% CI 2.57-10.65; p = .001) with known risk-features of these indices. When we assign an additional risk-score to samples, which are mutant for the GPM, the Harrell's C-indices of GPM-augmented IPI, PINK, and PINK-E improved significantly (p < .001, χ 2 test) for both PFS and OS. Thus, we report on how genomic mutational information could steer toward better prognostication of NKTCL patients., (© 2022 The Authors. American Journal of Hematology published by Wiley Periodicals LLC.)

Primary Epstein-Barr virus (EBV)-positive nodal T/NK-cell lymphoma (PTCL-EBV) is a poorly understood disease which shows features resembling extranodal NK/T-cell lymphoma (ENKTL) and is currently not recognized as a distinct entity but categorized as a variant of primary T-cell lymphoma not otherwise specified (PTCL-NOS). Herein, we analyzed copynumber aberrations (n=77) with a focus on global measures of genomic instability and homologous recombination deficiency and performed gene expression (n=84) and EBV miRNA expression (n=24) profiling as well as targeted mutational analysis (n=16) to further characterize PTCL-EBV in relation to ENKTL and PTCL-NOS. Multivariate analysis revealed that patients with PTCL-EBV had a significantly worse outcome compared to patients with PTCL-NOS (P=0.002) but not to those with ENKTL. Remarkably, PTCL-EBV exhibited significantly lower genomic instability and homologous recombination deficiency scores compared to ENKTL and PTCL-NOS. Gene set enrichment analysis revealed that many immune-related pathways, interferon α/γ response, and IL6&#95;JAK&#95;STAT3 signaling were significantly upregulated in PTCLEBV and correlated with lower genomic instability scores. We also identified that NFκB-associated genes, BIRC3, NFKB1 (P50) and CD27, and their proteins are upregulated in PTCL-EBV. Most PTCL-EBV demonstrated a type 2 EBV latency pattern and, strikingly, exhibited downregulated expression of most EBV miRNA compared to ENKTL and their target genes were also enriched in immune-related pathways. PTCL-EBV also showed frequent mutations of TET2, PIK3CD and STAT3, and are characterized by microsatellite stability. Overall, poor outcome, low genomic instability, upregulation of immune pathways and downregulation of EBV miRNA are distinctive features of PTCL-EBV. Our data support the concept that PTCL-EBV could be considered as a distinct entity, provide novel insights into the pathogenesis of the disease and offer potential new therapeutic targets for this tumor.

We herein present an overview of the upcoming 5 th edition of the World Health Organization Classification of Haematolymphoid Tumours focussing on lymphoid neoplasms. Myeloid and histiocytic neoplasms will be presented in a separate accompanying article. Besides listing the entities of the classification, we highlight and explain changes from the revised 4 th edition. These include reorganization of entities by a hierarchical system as is adopted throughout the 5 th edition of the WHO classification of tumours of all organ systems, modification of nomenclature for some entities, revision of diagnostic criteria or subtypes, deletion of certain entities, and introduction of new entities, as well as inclusion of tumour-like lesions, mesenchymal lesions specific to lymph node and spleen, and germline predisposition syndromes associated with the lymphoid neoplasms., (© 2022. The Author(s).)

Endophytic microorganisms are an important source of bioactive secondary metabolites. In this study, fungal endophytes obtained from A*STAR's Natural Product Library (NPL) and previously isolated from different habitats of Singapore were investigated for their diversity, antimicrobial, and cytotoxic activities. A total of 222 fungal strains were identified on the basis of sequence analysis of ITS region of the rDNA gene. The identified fungal strains belong to 59 genera distributed in 20 orders. Majority of the identified strains (99%; 219 strains) belong to the phylum Ascomycota , while two strains belonged to the phylum Basidiomycota , and only one strain was from Mucoromycota phylum. The most dominant genus was Colletotrichum accounting for 27% of all the identified strains. Chemical elicitation using 5-azacytidine and suberoylanilide hydroxamic acid (SAHA) and variation of fermentation media resulted in the discovery of more bioactive strains. Bioassay-guided isolation and structure elucidation of active constituents from three prioritized fungal strains: Lophiotrema sp. F6932, Muyocopron laterale F5912, and Colletotrichum tropicicola F10154, led to the isolation of a known compound; palmarumycin C 8 and five novel compounds; palmarumycin CP 30 , muyocopronol A-C and tropicicolide. Tropicicolide displayed the strongest antifungal activity against Aspergillus fumigatus with an IC 50 value of 1.8 μg/ml but with a weaker activity against the Candida albicans presenting an IC 50 of 7.1 μg/ml. Palmarumycin C 8 revealed the best antiproliferative activity with IC 50 values of 1.1 and 2.1 μg/ml against MIA PaCa-2 and PANC-1 cells, respectively., 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., (Copyright © 2022 Gakuubi, Ching, Munusamy, Wibowo, Liang, Kanagasundaram and Ng.)

Intelligent health diagnosis for young children aims at maintaining and promoting the healthy development of young children, aiming to make young children have a healthy state and provide a better future for their physical and mental health development. The biological basis of intelligence is the structure and function of human brain and the key to improve the intelligence level of infants is to improve the quality of brain development, especially the early development of brain. Based on machine learning and health information statistics, this paper studies the development of infant health diagnosis and intelligence, physical and mental health. Pre-process the sample data, and use the filtering method based on machine learning and health information statistics for feature screening. Compared with traditional statistical methods, machine learning and health information statistical methods can better obtain the hidden information in the big data of children's physical and mental health development, and have better learning ability and generalization ability. The machine learning theory is used to analyze and mine the infant's health diagnosis and intelligence development, establish a health state model, and intuitively show people the health status of their infant's physical and mental health development by means of data. Moreover, the accumulation of these big data is very important in the field of medical and health research driven by big data., 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., (Copyright © 2022 Wang, Li and Ng.)

The intestinal immune system, which must ensure appropriate immune responses to both pathogens and commensal microflora, comprises innate lymphoid cells and various T-cell subsets, including intra-epithelial lymphocytes (IELs). An example of innate lymphoid cells is natural killer cells, which may be classified into tissue-resident, CD56 bright NK-cells that serve a regulatory function and more mature, circulating CD56 dim NK-cells with effector cytolytic properties. CD56 bright NK-cells in the gastrointestinal tract give rise to indolent NK-cell enteropathy and lymphomatoid gastropathy, as well as the aggressive extranodal NK/T cell lymphoma, the latter following activation by EBV infection and neoplastic transformation. Conventional CD4+ TCRαβ+ and CD8αβ+ TCRαβ+ T-cells are located in the lamina propria and the intraepithelial compartment of intestinal mucosa as type 'a' IELs. They are the putative cells of origin for CD4+ and CD8+ indolent T-cell lymphoproliferative disorders of the gastrointestinal tract and intestinal T-cell lymphoma, NOS. In addition to such conventional T-cells, there are non-conventional T-cells in the intra-epithelial compartment that express CD8αα and innate lymphoid cells that lack TCRs. The central feature of type 'b' IELs is the expression of CD8αα homodimers, seen in monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL), which primarily arises from both CD8αα+ TCRαβ+ and CD8αα+ TCRγδ+ IELs. EATL is the other epitheliotropic T-cell lymphoma in the GI tract, a subset of which arises from the expansion and reprograming of intracytoplasmic CD3+ innate lymphoid cells, driven by IL15 and mutations of the JAK-STAT pathway.