Immune imprinting, breadth of variant recognition, and germinal center response in human SARS-CoV-2 infection and vaccination.

During the SARS-CoV-2 pandemic, novel and traditional vaccine strategies have been deployed globally. We investigated whether antibodies stimulated by mRNA vaccination (BNT162b2), including third-dose boosting, differ from those generated by infection or adenoviral (ChAdOx1-S and Gam-COVID-Vac) or inactivated viral (BBIBP-CorV) vaccines. We analyzed human lymph nodes after infection or mRNA vaccination for correlates of serological differences. Antibody breadth against viral variants is lower after infection compared with all vaccines evaluated but improves over several months. Viral variant infection elicits variant-specific antibodies, but prior mRNA vaccination imprints serological responses toward Wuhan-Hu-1 rather than variant antigens. In contrast to disrupted germinal centers (GCs) in lymph nodes during infection, mRNA vaccination stimulates robust GCs containing vaccine mRNA and spike antigen up to 8 weeks postvaccination in some cases. SARS-CoV-2 antibody specificity, breadth, and maturation are affected by imprinting from exposure history and distinct histological and antigenic contexts in infection compared with vaccination.

Epstein-Barr virus latency patterns in polymorphic lymphoproliferative disorders and lymphomas in immunodeficiency settings: Diagnostic implications.

Epstein-Barr virus (EBV) is responsible for many B cell lymphoproliferative disorders (LPD) spanning subclinical infection to immunodeficiency-related neoplasms. EBV establishes a latent infection in the host B cell as defined histologically by the expression of EBV latent membrane proteins and nuclear antigens. Herein, we characterize the latency patterns of immunodeficiency-related neoplasms including post-transplant lymphoproliferative disorders (PTLD) and therapy-related LPD (formerly iatrogenic) with latent membrane protein-1 (LMP-1) and EBV nuclear antigen-2 (EBNA-2) immunohistochemistry. The latency pattern was correlated with immunodeficiency and dysregulation (IDD) status and time from transplant procedure. 38 cases of EBV+ PTLD in comparison to 27 cases of classic Hodgkin lymphoma (CHL) and diffuse large B cell lymphoma (DLBCL) arising in either the therapy-related immunodeficiency setting (n = 12) or without an identified immunodeficiency (n = 15) were evaluated for EBV-encoded small RNAs by in situ hybridization (EBER-ISH) and for LMP-1 and EBNA-2 by immunohistochemistry. A full spectrum of EBV latency patterns was observed across PTLD in contrast to CHL and DLBCL arising in the therapy-related immunodeficiency setting. Polymorphic-PTLD (12 of 16 cases, 75 %) and DLBCL-PTLD (9 of 11 cases, 82 %) showed the greatest proportion of cases with latency III pattern. Whereas, EBV+ CHL in an immunocompetent patient showed exclusively latency II pattern (13 of 13 cases, 100 %). The majority of EBV+ PTLD occurred by three years of transplant procedure date and were enriched for latency III pattern (21 of 22 cases, 95 %). Immunohistochemical identification of EBV latency by LMP-1 and EBNA-2 can help classify PTLD in comparison to other EBV+ B cell LPD and lymphomas arising in therapy-related immunodeficiency and non-immunodeficiency settings.

Detection of Aberrant CD58 Expression in a Wide Spectrum of Lymphoma Subtypes: Implications for Treatment Resistance.

CD58 or lymphocyte function-associated antigen-3, is a ligand for CD2 receptors on T and NK cells and is required for their activation and target cell killing. We recently showed a trend toward higher frequency of CD58 aberrations in patients with diffuse large B-cell lymphoma (DLBCL) who progressed on chimeric antigen receptor-T-cell treatment compared with those who responded. Given that CD58 status may be an important measure of T-cell-mediated therapy failure, we developed a CD58 immunohistochemical assay and evaluated CD58 status in 748 lymphomas. Our results show that CD58 protein expression is downregulated in a significant proportion of all subtypes of B-, T-, and NK-cell lymphomas. CD58 loss is significantly related to poor prognostic indicators in DLBCL and to ALK and DUSP22 rearrangements in anaplastic large-cell lymphoma. However, it is not associated with overall or progression-free survival in any of the lymphoma subtypes. As eligibility for chimeric antigen receptor-T-cell therapy is being extended to a broader spectrum of lymphomas, mechanisms of resistance, such as target downregulation and CD58 loss, may limit therapeutic success. CD58 status is therefore an important biomarker in lymphoma patients who may benefit from next-generation T-cell-mediated therapies or other novel approaches that mitigate immune escape.

Conditional expression of HGAL leads to the development of diffuse large B-cell lymphoma in mice.

Diffuse large B-cell lymphomas (DLBCLs) are clinically and genetically heterogeneous tumors. Deregulation of diverse biological processes specific to B cells, such as B-cell receptor (BCR) signaling and motility regulation, contribute to lymphomagenesis. Human germinal center associated lymphoma (HGAL) is a B-cell-specific adaptor protein controlling BCR signaling and B lymphocyte motility. In normal B cells, it is expressed in germinal center (GC) B lymphocytes and promptly downregulated upon further differentiation. The majority of DLBCL tumors, primarily GC B-cell types, but also activated types, express HGAL. To investigate the consequences of constitutive expression of HGAL in vivo, we generated mice that conditionally express human HGAL at different stages of hematopoietic development using 3 restricted Cre-mediated approaches to initiate expression of HGAL in hematopoietic stem cells, pro-B cells, or GC B cells. Following immune stimulation, we observed larger GCs in mice in which HGAL expression was initiated in GC B cells. All 3 mouse strains developed DLBCL at a frequency of 12% to 30% starting at age 13 months, leading to shorter survival. Immunohistochemical studies showed that all analyzed tumors were of the GC B-cell type. Exon sequencing revealed mutations reported in human DLBCL. Our data demonstrate that constitutive enforced expression of HGAL leads to DLBCL development.

Point-of-Care and Dual-Response Detection of Hydrazine/Hypochlorite-Based on a Smart Hydrogel Sensor and Applications in Information Security and Bioimaging.

A novel dual-response fluorescence probe (XBT-CN) was developed by using a fluorescence priming strategy for quantitative monitoring and visualization of hydrazine (N2H4) and hypochlorite (ClO-). With the addition of N2H4/ClO-, the cleavage reaction of C=C bond initiated by N2H4/ClO- was transformed into corresponding hydrazone and aldehyde derivatives, inducing the probe XBT-CN appeared a fluorescence "off-on" response, which was verified by DFT calculation. HRMS spectra were also conducted to confirm the sensitive mechanism of XBT-CN to N2H4 and ClO-. The probe XBT-CN had an obvious fluorescence response to N2H4 and ClO-, which caused a significant color change in unprotected eyes. In addition, the detection limits of XBT-CN for N2H4 and ClO- were 27 nM and 34 nM, respectively. Interference tests showed that other competitive analytes could hardly interfere with the detection of N2H4 and ClO- in a complex environment. In order to realize the point-of-care detection of N2H4 and ClO-, an XBT-CN@hydrogel test kit combined with a portable smartphone was developed. Furthermore, the portable test kit has been applied to the detection of N2H4 and ClO- in a real-world environment and food samples, and a series of good results have been achieved. Attractively, we demonstrated that XBT-CN@hydrogel was successfully applied as an encryption ink in the field of information security. Finally, the probe can also be used to monitor and distinguish N2H4 and ClO- in living cells, exhibiting excellent biocompatibility and low cytotoxicity.

Penta-Twin Destruction by Coordinated Twin Boundary Deformation.

Penta-twinned nanomaterials often exhibit unique mechanical properties. However, the intrinsic deformation behavior of penta-twins remains largely unclear, especially under the condition of high shear stress. In this study, we show that the deformation of penta-twins often subject to a structural destruction via dislocation-mediated coordinated twin boundary (TB) deformation, resulting in a reconstructed pentagon-shaped core. This reconstructed core region is mainly induced by the coordinated TB migration along different directions (for the nucleation and growth) and accelerated by the TB sliding (for the growth). The destructed penta-twin core can effectively accommodate the intrinsic disclination of the penta-twin, which further collapses beyond a critical size, as predicted by an energy-based criterion. These intrinsic deformation behaviors of penta-twins would enable the possibility of controlling the morphology of penta-twinned nanomaterials with unique properties.

KLF1/EKLF expression in acute leukemia is correlated with chromosomal abnormalities.

KLF1 (EKLF) is a master regulator of erythropoiesis and controls expression of a wide array of target genes. We interrogated human tissue microarray samples via immunohistological analysis to address whether levels of KLF1 protein are associated with leukemia. We have made the unexpected findings that higher KLF1 levels are correlated with cells containing abnormal chromosomes, and that high KLF1 expression is not limited to acute myeloid leukemia (AML) associated with erythroid/megakaryoblastic differentiation. Expression of KLF1 is associated with poor survival. Further analyses reveal that KLF1 directly regulates a number of genes that play a role in chromosomal integrity. Together these results suggest that monitoring KLF1 levels may provide a new marker for risk stratification and prognosis in patients with AML.

Integrated systems analysis reveals a molecular network underlying autism spectrum disorders.

Autism is a complex disease whose etiology remains elusive. We integrated previously and newly generated data and developed a systems framework involving the interactome, gene expression and genome sequencing to identify a protein interaction module with members strongly enriched for autism candidate genes. Sequencing of 25 patients confirmed the involvement of this module in autism, which was subsequently validated using an independent cohort of over 500 patients. Expression of this module was dichotomized with a ubiquitously expressed subcomponent and another subcomponent preferentially expressed in the corpus callosum, which was significantly affected by our identified mutations in the network center. RNA-sequencing of the corpus callosum from patients with autism exhibited extensive gene mis-expression in this module, and our immunochemical analysis showed that the human corpus callosum is predominantly populated by oligodendrocyte cells. Analysis of functional genomic data further revealed a significant involvement of this module in the development of oligodendrocyte cells in mouse brain. Our analysis delineates a natural network involved in autism, helps uncover novel candidate genes for this disease and improves our understanding of its molecular pathology.

Identification of LMO2 transcriptome and interactome in diffuse large B-cell lymphoma.

LMO2 regulates gene expression by facilitating the formation of multipartite DNA-binding complexes. In B cells, LMO2 is specifically up-regulated in the germinal center (GC) and is expressed in GC-derived non-Hodgkin lymphomas. LMO2 is one of the most powerful prognostic indicators in diffuse large B-cell (DLBCL) patients. However, its function in GC B cells and DLBCL is currently unknown. In this study, we characterized the LMO2 transcriptome and transcriptional complex in DLBCL cells. LMO2 regulates genes implicated in kinetochore function, chromosome assembly, and mitosis. Overexpression of LMO2 in DLBCL cell lines results in centrosome amplification. In DLBCL, the LMO2 complex contains some of the traditional partners, such as LDB1, E2A, HEB, Lyl1, ETO2, and SP1, but not TAL1 or GATA proteins. Furthermore, we identified novel LMO2 interacting partners: ELK1, nuclear factor of activated T-cells (NFATc1), and lymphoid enhancer-binding factor1 (LEF1) proteins. Reporter assays revealed that LMO2 increases transcriptional activity of NFATc1 and decreases transcriptional activity of LEF1 proteins. Overall, our studies identified a novel LMO2 transcriptome and interactome in DLBCL and provides a platform for future elucidation of LMO2 function in GC B cells and DLBCL pathogenesis.

Spatial phenotyping of nodular lymphocyte predominant Hodgkin lymphoma and T-cell/histiocyte-rich large B-cell lymphoma.

Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) is a rare lymphoma with sparse tumor B-cells and a favorable prognosis. Variant growth patterns of NLPHL, however, often show advanced stage, progression to T-cell/histiocyte-rich large B-cell lymphoma (THRLBCL) and a worse prognosis. We studied the tumor microenvironment (TME) of NLPHL and THRLBCL using highplex imaging and spatial profiling at the single cell level. Our findings show distinct differences in TME composition and spatial configuration that differ among typical and variant NLPHL and THRLBCL. Typical NLPHL show abundant helper T-cell subsets, while THRLBCL show abundant cytotoxic T-cells and macrophages. Tumor B-cell size and content is lowest in typical NLPHL, followed by variant NLPHL, and highest in THRLBCL, whereas an opposite trend characterized TME B-cells. CD4/CD8 double-positive T-cells are seen in all NLPHL but not in the majority of THRLBCL and are spatially distant from LP-cells and TFH-rosettes. The differences in macrophage/monocyte content in distinguishing NLPHL pattern E from THRLBCL is further corroborated in independent cohorts of cases. Our results validate the current approach to classification and in addition provide novel insights that could be leveraged to refine clinical management for patients with this spectrum of lymphomas.

TIGIT is Frequently Expressed in the Tumor Microenvironment of Select Lymphomas: Implications for Targeted Therapy.

Immune checkpoint inhibitors against Programmed Cell Death Protein 1/Programmed Cell (PD-1/PD-L1) and CTLA-4/B7 axes have had limited success in hematologic malignancies, requiring the need to explore alternative targets such as T-cell immunoreceptor with Ig and ITIM domains (TIGIT)/CD155 to improve durable clinical responses. We undertook this study to investigate the expression profile of TIGIT such that the potential efficacy of TIGIT blockade could be mapped among lymphoma subtypes. We validated an immunohistochemical assay for TIGIT and evaluated its expression in lymphoma and tumor microenvironment (TME) cells in 661 lymphoma/leukemia biopsies. Multiplex immunofluorescence was used for correlation with normal TME cell subsets. Tumor or TME TIGIT-positivity was defined as moderate to strong membrane staining in at least 10% of tumor or TME cells, respectively. TME TIGIT expression was correlated with overall survival and progression-free survival and comparison with PD-L1 expression. In most cases, lymphoma cells were TIGIT-negative except for angioimmunoblastic and peripheral T-cell lymphomas, which showed 91% and 47% positivity, respectively. A high proportion of small B-cell lymphoma and anaplastic large cell lymphoma cases had TIGIT-positive TME cells. Chronic lymphocytic leukemia/small lymphocytic lymphoma patients with TIGIT-negative TME cells showed significantly shorter overall survival ( P =0.04). No other statistically significant differences were found. When TIGIT was expressed in TME cells, there were a comparable number of TIGIT-positive only and dual TIGIT/PD-L1 positive cases except for more TIGIT-positive only cases in CLL/SLL. TIGIT expression shows distinctive profiles among lymphoma subtypes. Chronic lymphocytic leukemia/small lymphocytic lymphoma and anaplastic large cell lymphoma demonstrated high TME TIGIT expression compared with PD-L1, with a high proportion of dual TIGIT and PD-L1-positivity. Our results are likely to contribute to the design and correlative study of therapeutic response in clinical trials targeting TIGIT alone or in combination with PD1/PDL1.

A phenothiazine-based ratiometric fluorescence probe for the detection of hydroxylamine in real water and living cells.

In this study, a phenothiazine-based ratiometric fluorescent probe PCHO was developed for highly sensitive and specific detection of hydroxylamine (HA). In the presence of HA, the aldehyde group on the PCHO molecule underwent a specific nucleophilic addition with HA to form an oxime group, accompanied by significant changes in fluorescence from green to blue. This detection mechanism was well supported by 1H NMR titration, HRMS and DFT calculations. The probe PCHO exhibited high sensitivity for HA detection (LOD was 0.19 µM) with a rapid response time (1 min), high selectivity and strong anti-interference performance. Surprisingly, the probe PCHO could selectively distinguish HA from its similar competing agents such as hydrazine and amines. Moreover, paper strips loaded with PCHO were prepared and combined with a smartphone to achieve point-of-care and visual detection of HA. The probe PCHO was further applied for the detection of HA in real water samples, achieving a recovery rate of 98.90% to 104.86% and an RSD of 0.86% to 2.44%, confirming the accuracy and reliability of the method. Additionally, the probe PCHO was used for imaging analysis of HA in living cells, providing a powerful visualization tool for exploring the physiological functions of HA in vivo.

CD137 is expressed in follicular dendritic cell tumors and in classical Hodgkin and T-cell lymphomas: diagnostic and therapeutic implications.

CD137 (also known as 4-1BB and TNFRSF9) is a member of the tumor necrosis factor receptor superfamily. Originally identified as a costimulatory molecule expressed by activated T cells and NK cells, CD137 is also expressed by follicular dendritic cells, monocytes, mast cells, granulocytes, and endothelial cells. Anti-CD137 immunotherapy has recently shown promise as a treatment for solid tumors and lymphoid malignancies in preclinical models. We defined the expression of CD137 protein in both normal and neoplastic hematolymphoid tissue. CD137 protein is expressed by follicular dendritic cells in the germinal center and scattered paracortical T cells, but not by normal germinal-center B cells, bone marrow progenitor cells, or maturing thymocytes. CD137 protein is expressed by a select group of hematolymphoid tumors, including classical Hodgkin lymphoma, T-cell and NK/T-cell lymphomas, and follicular dendritic cells neoplasms. CD137 is a novel diagnostic marker of these tumors and suggests a possible target for tumor-directed antibody therapy.

Prediction of survival in diffuse large B-cell lymphoma based on the expression of 2 genes reflecting tumor and microenvironment.

Several gene-expression signatures predict survival in diffuse large B-cell lymphoma (DLBCL), but the lack of practical methods for genome-scale analysis has limited translation to clinical practice. We built and validated a simple model using one gene expressed by tumor cells and another expressed by host immune cells, assessing added prognostic value to the clinical International Prognostic Index (IPI). LIM domain only 2 (LMO2) was validated as an independent predictor of survival and the "germinal center B cell-like" subtype. Expression of tumor necrosis factor receptor superfamily member 9 (TNFRSF9) from the DLBCL microenvironment was the best gene in bivariate combination with LMO2. Study of TNFRSF9 tissue expression in 95 patients with DLBCL showed expression limited to infiltrating T cells. A model integrating these 2 genes was independent of "cell-of-origin" classification, "stromal signatures," IPI, and added to the predictive power of the IPI. A composite score integrating these genes with IPI performed well in 3 independent cohorts of 545 DLBCL patients, as well as in a simple assay of routine formalin-fixed specimens from a new validation cohort of 147 patients with DLBCL. We conclude that the measurement of a single gene expressed by tumor cells (LMO2) and a single gene expressed by the immune microenvironment (TNFRSF9) powerfully predicts overall survival in patients with DLBCL.

Palladium-catalyzed divergent reactions of 1,6-enyne carbonates: synthesis of vinylidenepyridines and vinylidenepyrrolidines.

A method for preparing five- or six-membered heterocyclic compounds from enyne carbonates via palladium catalysis was developed. Enyne carbonates were transformed into 3-vinylidene-1-tosylpyridines 2 in the presence of PdI(2) as the catalyst. Using Pd(dba)(2) as the catalyst, 3-vinylidene-1-tosylpyrrolidines 3 were obtained. Further functionalizations of compounds 3 were carried out in a one-pot manner.

Human Germinal Center-associated Lymphoma (HGAL) Is a Reliable Marker of Normal and Neoplastic Follicular Helper T Cells Including Angioimmunoblastic T-Cell Lymphoma.

The diagnosis of angioimmunoblastic T-cell lymphoma (AITL) is complex and requires the demonstration of a T-follicular helper (TFH) phenotype. Immunophenotypic markers that detect the TFH phenotype are highly variable, thereby necessitating the use of 3 to 5 TFH markers to substantiate a TFH phenotype. We tested the utility of germinal center markers human germinal center-associated lymphoma (HGAL) and LIM-domain only 2 (LMO2) in detecting a TFH phenotype. We compared their staining to that of 6 TFH markers in current use, PD-1, ICOS, CXCL13, SAP, CD10, and BCL6, in a cohort of 23 AITL. Our results show that although both markers can detect a TFH phenotype, HGAL was superior to LMO2 in the percent of cells stained and the intensity of staining, 2 variables used to generate H-scores. Using H-scores as the metric, HGAL was most comparable to BCL6 among the currently used TFH markers and was more sensitive than CXCL13, SAP, CD10, and LMO2. PD-1 and ICOS emerged as the most robust of the 8 markers tested in this study in detecting a TFH phenotype. We conclude that HGAL is a reliable marker of TFH cells and can aid in the diagnosis of lymphomas of TFH derivation, particularly in the recognition of early patterns of AITL.

Hierarchical twinning governed by defective twin boundary in metallic materials.

Dense networks of deformation twins endow metals and alloys with unprecedented mechanical properties. However, the formation mechanism of these hierarchical twin structures remains under debate, especially their relations with the imperfect nature of twin boundaries (TBs). Here, we investigate the intrinsic deformability of defective TBs in face-centered cubic metallic materials, where the inherent kinks on a set of primary TBs are demonstrated to facilitate the formation of secondary and hierarchical nanotwins. This defect-driven hierarchical twinning propensity is critically dependent on the kink height, which proves to be generally applicable in a variety of metals and alloys with low stacking fault energies. As a geometric extreme, a fivefold twin can be constructed via this self-activated hierarchical twinning mechanism. These findings differ from the conventional twinning mechanisms, enriching our understanding of twinning-mediated plasticity in metallic materials.

Pd(0)-catalyzed [1,5]-sigmatropic hydrogen shift of propargylic esters toward substituent naphthylamines.

A novel and convenient carboannulation method for the synthesis of highly substituted naphthylamine derivatives has been developed though a Pd(0)-catalyzed [1,5]-sigmatropic hydrogen shift and cyclization reaction of propargyl esters.

Copper-catalyzed direct thiolation of azoles with aliphatic thiols.

Cu(II)-catalyzed direct thiolation of azoles with thiols is described via intermolecular C-S bond formation/C-H functionalization under oxidative conditions. Both aryl thiols and aliphatic thiols are used as coupling partners, and furnished the thiolation products in moderate to good yields. The reaction is compatible with a wide range of heterocycles including oxazole, thiazole, imidazole and oxadiazole.

Classic Hodgkin lymphoma in Guatemalan children of age less than six years: analysis of immune regulatory pathways and the tumor microenvironment.

Classic Hodgkin lymphoma (cHL) in young children (ages 0-6) is rare in high income countries (HICs) but is more prevalent in low- and middle-income countries (LMICs) like Guatemala. Given that the majority of cHL studies have evaluated adolescent/adults, and the immune system changes with age, we sought to characterize Epstein-Barr virus (EBV) expression, immune regulatory pathway markers and the tumor microenvironment in 42 children ages 0-6 with cHL from Guatemala. We found a very high frequency of EBV expression (97.5%). Hodgkin cells showed increased expression of PD1 ligands and CD137, indicative of shared immune regulatory mechanisms with adult cHL. Pediatric cHL also showed an increase in CD8+ tumor infiltrating lymphocytes and tumor associated macrophages within the tumor microenvironment. Despite 25 having high risk disease, only 4 patients died from progressive disease, relapse or infection.