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Evidence indicates that there are significant alterations in gut microbiota diversity and composition in patients with hematological malignancies. The present study investigated the oral and intestinal microbiome in patients with chronic lymphocytic leukemia (CLL) (n=81) and age-matched healthy volunteers (HVs; n=21) using 16S ribosomal RNA next-generation sequencing. Changes in both oral and gut microbiome structures were identified, with a high abundance of Proteobacteria and depletion of Bacteroidetes in CLL as compared to HVs. Oral and stool samples of patients with CLL revealed a significant change in the abundance of short-chain fatty acid-producing genera in comparison with HVs. Furthermore, the relative abundance of oral and intestine Bacteroidetes was significantly decreased in patients with CLL with negative prognostic features, including unmutated immunoglobulin heavy chain gene (IGHV). Notably, an increased abundance of gut Firmicutes was found to be associated with high expression of CD38. Finally, the present study suggested the log Firmicutes/Bacteroidota ratio as a novel intestinal microbiome signature associated with a shorter time to first treatment in individuals with CLL. The findings indicate that oral and gut microbial diversity in CLL might point to the inflammatory-related modulation of the clinical course of the disease. [ABSTRACT FROM AUTHOR]

The article discusses the use of next-generation sequencing and high DNA input to detect measurable residual disease (MRD) in the peripheral blood of patients with B-cell precursor acute lymphoblastic leukemia (ALL). Traditional MRD assessment in bone marrow samples may miss MRD in peripheral blood, but modern molecular methods can enhance detection. The study found that adjusting traditional MRD assessment strategies can improve MRD detection in peripheral blood, but it may not fully replace bone marrow monitoring. The findings suggest that high DNA input and next-generation sequencing can increase the sensitivity of MRD detection in peripheral blood, especially in cases where bone marrow samples are unavailable. [Extracted from the article]

The article discusses the impact of total lesion glycolysis (TLG) in predicting the response of follicular lymphoma (FL) to rituximab. FL is a common type of non-Hodgkin lymphoma characterized by the (14;18) translocation. The study evaluates whether early rituximab monotherapy could delay chemotherapy in FL patients with low tumor burden. Results suggest that patients with FL should be evaluated using 18F-FDG PET scan and stratified based on TMTV and TLG thresholds to guide treatment decisions. [Extracted from the article]

This letter discusses the characterization of MYC rearrangements in multiple myeloma (MM) using an optical genome mapping (OGM) approach. MYC rearrangements are genetic alterations that have been proposed as high-risk features in MM, but their prognostic implications are not fully established. The study used OGM in conjunction with other detection methods such as fluorescence in situ hybridization (FISH), karyotyping, and next-generation sequencing (NGS) to comprehensively analyze MYC rearrangements in MM. The results showed that MYC rearrangements were detected in a significant proportion of MM cases, particularly when using OGM. The study also found that MYC expression was uniformly high in MM samples with translocations involving immunoglobulin loci but variable in those with non-immunoglobulin partners. The authors suggest that combining OGM and MYC immunohistochemistry (IHC) could have practical implications for routine prognostic assessments in MM. Further studies are needed to clarify the prognostic impact of MYC rearrangements and to understand the dysregulation mechanisms associated with them. The study was supported by grants from the National Research Foundation of Korea, the Korea Medical Device Development Fund, and Korea University Guro Hospital. The authors declare no competing interests. Supplementary information and data availability are provided in the article. [Extracted from the article]

Summary The study of antibodies in jawed vertebrates (gnathostomes) provides every immunologist with a bird's eye view of how human immunoglobulins (Igs) came into existence and subsequently evolved into their present forms. It is a fascinating Darwinian history of conservation on the one hand and flexibility on the other, exemplified by the Ig heavy chain (H) isotypes IgM and IgD/W, respectively. The cartilaginous fish (e.g., sharks) Igs provide a glimpse of “how everything got off the ground,” while the amphibians (e.g., the model Xenopus) reveal how the adaptive immune system made an about face with the emergence of Ig isotype switching and IgG‐like structure/function. The evolution of mucosal Igs is a captivating account of malleability, convergence, and conservation, and a call to arms for future study! In between there are spellbinding chronicles of antibody evolution in each class of vertebrates and rather incredible stories of how antibodies can adapt to occupy niches, for example, single‐domain variable regions, cold‐adapted Igs, convergent mechanisms to dampen antibody function, provision of mucosal defense, and many more. The purpose here is not to provide an encyclopedic examination of antibody evolution, but rather to hit the high points and entice readers to appreciate how things “came to be.” [ABSTRACT FROM AUTHOR]

The purpose of this study was to develop a monoclonal antibody (mAb) that can identify porcine dendritic cells (DCs) that have differentiated from bone marrow progenitor cells. Hybridoma technology was used to obtain mAbs, and bone marrow-derived DCs (BMDCs) were employed as immunogens for producing antibodies. The generated PD9-9 mAbs exhibited considerable reactivity towards porcine BMDCs with applications in flow cytometry and immunostaining. The antibody was composed of heavy immunoglobulin gamma-1 chains and light kappa chains. The PD9-9 mAb recognized fully differentiated porcine BMDCs and cells undergoing DC differentiation. In contrast, bone marrow cells and macrophages were not recognized by PD9-9. In addition, the PD9-9 mAb promoted porcine DC proliferation. Consequently, the PD9-9 mAb may be a biomarker for porcine DCs and will be advantageous for investigating porcine DC biology. [ABSTRACT FROM AUTHOR]

The long-term value of efficient antigen discovery includes gaining insights into the variety of potential cancer neoantigens, effective vaccines lacking adverse effects, and adaptive immune receptor (IR) targets for blocking adaptive IR-antigen interactions in autoimmunity. While the preceding goals have been partially addressed via big data approaches to HLA (human leukocyte antigen)-epitope binding, there has been little such progress in the big data setting for adaptive IR-epitope binding. This delay in progress for the latter is likely due to, among other things, the much more complicated adaptive IR repertoire in an individual compared to individual HLA alleles. Thus, results described here represent the application of an algorithm for efficient assessment of immunoglobulin heavy chain complementarity determining region-3 (IGH CDR3)-gliadin epitope interactions, with a focus on epitopes known to be associated with an immune response in celiac disease. The hydrophobic, chemical complementarity between celiac case IGH CDR3s and known celiac epitopes was found to be greater in comparison to the hydrophobic, chemical complementarity between the same celiac case IGH CDR3s and a series of control epitopes. Thus, the approaches indicated here likely offer guidance for the development of conveniently applied algorithms for antigen verification and discovery. [ABSTRACT FROM AUTHOR]

The association between high-density lipoprotein (HDL) cholesterol and breast cancer (BC) remains controversial due to the high complexity of the HDL particle and its functionality. The HDL proteome was determined in newly diagnosed BC classified according to the molecular type [luminal A or B (LA or LB), HER2, and triple-negative (TN)] and clinical stage of the disease. Women (n = 141) aged between 18 and 80 years with BC, treatment-naïve, and healthy women [n = 103; control group (CT)], matched by age and body mass index, were included. Data-independent acquisition (DIA) proteomics was performed in isolated HDL (D = 1.063–1.21 g/mL). Results: Paraoxonase1, carnosine dipeptidase1, immunoglobulin mMu heavy chain constant region (IGHM), apoA-4, and transthyretin were reduced, and serum amyloid A2 and tetranectin were higher in BC compared to CT. In TNBC, apoA-1, apoA-2, apoC-2, and apoC-4 were reduced compared to LA, LB, and HER2, and apoA-4 compared to LA and HER2. ComplementC3, lambda immunoglobulin2/3, serpin3, IGHM, complement9, alpha2 lysine rich-glycoprotein1, and complement4B were higher in TNBC in comparison to all other types; complement factor B and vitamin D-binding protein were in contrast to LA and HER2, and plasminogen compared to LA and LB. In grouped stages III + IV, tetranectin and alpha2-macroglobulin were reduced, and haptoglobin-related protein; lecithin cholesterol acyltransferase, serum amyloid A1, and IGHM were increased compared to stages I + II. Conclusions: A differential proteomic profile of HDL in BC based on tumor molecular classification and the clinical stage of the disease may contribute to a better understanding of the association of HDL with BC pathophysiology, treatment, and outcomes. [ABSTRACT FROM AUTHOR]

Background and aims: Aberrations in the immunoglobulin heavy chain (IgH) locus are associated with poor prognosis in pediatric precursor B-cell acute lymphoblastic leukemia (BCP-ALL) patients. The primary objective of this pilot study is to enhance our understanding of the IgH phenotype by exploring the intracellular chiral metabolome. Materials and methods: Leukemia cells were isolated from the bone marrow of BCP-ALL pediatric patients at diagnosis. The samples' metabolome and transcriptome were characterized using untargeted chiral metabolomic and next-generation sequencing transcriptomic analyses. Results: For the first time D-amino acids were identified in the leukemic cells' intracellular metabolome from the bone marrow niche. Chiral metabolic signatures at diagnosis was indicative of a resistant phenotype. Through integrated network analysis and Pearson correlation, confirmation was obtained regarding the association of the IgH phenotype with several genes linked to poor prognosis. Conclusion: The findings of this study have contributed to the understanding that the chiral metabolome plays a role in the poor prognosis observed in an exceptionally rare patient cohort. The findings include elevated D-amino acid incorporation in the IgH group, the emergence of several unknown, potentially enantiomeric, metabolites, and insights into metabolic pathways that all warrant further exploration. [ABSTRACT FROM AUTHOR]

The molecular landscape of chronic lymphocytic leukemia (CLL) has been extensively characterized, and various potent prognostic biomarkers were discovered. The genetic composition of the B-cell receptor (BCR) immunoglobulin (IG) was shown to be especially powerful for discerning indolent from aggressive disease at diagnosis. Classification based on the IG heavy chain variable gene (IGHV) somatic hypermutation status is routinely applied. Additionally, BCR IGH stereotypy has been implicated to improve risk stratification, through characterization of subsets with consistent clinical profiles. Despite these advances, it remains challenging to predict when CLL progresses to requiring first-line therapy, thus emphasizing the need for further refinement of prognostic indicators. Signaling pathways downstream of the BCR are essential in CLL pathogenesis, and dysregulated components within these pathways impact disease progression. Considering not only genomics but the entirety of factors shaping BCR signaling activity, this review offers insights in the disease for better prognostic assessment of CLL. [ABSTRACT FROM AUTHOR]

The severe acute respiratory syndrome coronavirus 2 variant JN.1 recently emerged as the dominant variant despite having only one amino acid change on the spike (S) protein receptor binding domain (RBD) compared with the ancestral BA.2.86, which never represented more than 5% of global variants. To define at the molecular level the JN.1 ability to spread globally, we interrogated a panel of 899 neutralizing human monoclonal antibodies. Our data show that the single leucine-455–to–serine mutation in the JN.1 spike protein RBD unleashed the global spread of JN.1, likely occurring by elimination of more than 70% of the neutralizing antibodies mediated by IGHV3-53/3-66 germlines. However, the resilience of class 3 antibodies with low neutralization potency but strong Fc functions may explain the absence of JN.1 severe disease. Editor's summary: New variants of SARS-CoV-2 continue to emerge and enable viral immune evasion. Using a panel of 899 human monoclonal neutralizing antibodies (nAbs), Paciello et al. examined antibody neutralization activity and effector functions against the recently dominant JN.1 variant. Compared with the ancestral BA.2.86 strain, around two-thirds of nAbs lost neutralization activity against JN.1, including most derived from the immunoglobulin heavy chain V gene (IGHV) 3-53 and 3-66 germlines. The remaining nAbs were only detected in individuals with a history of multiple vaccinations and Omicron breakthrough infections. Among nAbs that could bind the JN.1 spike protein, Fc functions were largely retained and primarily driven by class 3 antibodies that bind outside of the ACE2 binding region, possibly explaining the absence of JN.1 severe disease despite strong evasion of class 1/2 nAbs. —Claire Olingy [ABSTRACT FROM AUTHOR]

Background Methods Results Conclusion Antigen‐specific memory B cells play a key role in the induction of desensitization and remission to food allergens in oral immunotherapy and in the development of natural tolerance (NT). Here, we characterized milk allergen Bos d 9‐specific B cells in oral allergen‐specific immunotherapy (OIT) and in children spontaneously outgrowing cow's milk allergy (CMA) due to NT.Samples from children with CMA who received oral OIT (before, during, and after), children who naturally outgrew CMA (NT), and healthy individuals were received from Stanford biobank. Bos d 9‐specific B cells were isolated by flow cytometry and RNA‐sequencing was performed. Protein profile of Bos d 9‐specific B cells was analyzed by proximity extension assay.Increased frequencies of circulating milk allergen Bos d 9‐specific B cells were observed after OIT and NT. Milk‐desensitized subjects showed the partial acquisition of phenotypic features of remission, suggesting that desensitization is an earlier stage of remission. Within these most significantly expressed genes, IL10RA and TGFB3 were highly expressed in desensitized OIT patients. In both the remission and desensitized groups, B cell activation‐, Breg cells‐, BCR‐signaling‐, and differentiation‐related genes were upregulated. In NT, pathways associated with innate immunity characteristics, development of marginal zone B cells, and a more established suppressor function of B cells prevail that may play a role in long‐term tolerance. The analyses of immunoglobulin heavy chain genes in specific B cells demonstrated that IgG2 in desensitization, IgG1, IgA1, IgA2, IgG4, and IgD in remission, and IgD in NT were predominating. Secreted proteins from allergen‐specific B cells revealed higher levels of regulatory cytokines, IL‐10, and TGF‐β after OIT and NT.Allergen‐specific B cells are essential elements in regulating food allergy towards remission in OIT‐received and naturally resolved individuals. [ABSTRACT FROM AUTHOR]

Background: The effects of inhaled corticosteroids (ICS) on healthy airways are poorly defined. Objectives: To delineate the effects of ICS on gene expression in healthy airways, without confounding caused by changes in disease‐related genes and disease‐related alterations in ICS responsiveness. Methods: Randomized open‐label bronchoscopy study of high‐dose ICS therapy in 30 healthy adult volunteers randomized 2:1 to (i) fluticasone propionate 500 mcg bd daily or (ii) no treatment, for 4 weeks. Laboratory staff were blinded to allocation. Biopsies and brushings were analysed by immunohistochemistry, bulk RNA sequencing, DNA methylation array and metagenomics. Results: ICS induced small between‐group differences in blood and lamina propria eosinophil numbers, but not in other immunopathological features, blood neutrophils, FeNO, FEV1, microbiome or DNA methylation. ICS treatment upregulated 72 genes in brushings and 53 genes in biopsies, and downregulated 82 genes in brushings and 416 genes in biopsies. The most downregulated genes in both tissues were canonical markers of type‐2 inflammation (FCER1A, CPA3, IL33, CLEC10A, SERPINB10 and CCR5), T cell‐mediated adaptive immunity (TARP, TRBC1, TRBC2, PTPN22, TRAC, CD2, CD8A, HLA‐DQB2, CD96, PTPN7), B‐cell immunity (CD20, immunoglobulin heavy and light chains) and innate immunity, including CD48, Hobit, RANTES, Langerin and GFI1. An IL‐17‐dependent gene signature was not upregulated by ICS. Conclusions: In healthy airways, 4‐week ICS exposure reduces gene expression related to both innate and adaptive immunity, and reduces markers of type‐2 inflammation. This implies that homeostasis in health involves tonic type‐2 signalling in the airway mucosa, which is exquisitely sensitive to ICS. [ABSTRACT FROM AUTHOR]

Key Clinical Message: Heavy‐chain deposition disease (HCDD), a rare monoclonal immunoglobulin deposition disease, involves truncated heavy‐chain deposition in kidneys. Limited long‐term data exist. We report a case of renal and cardiac failure with favorable outcomes post bortezomib‐based therapy. Stable renal function observed over 4 years suggests efficacy in HCDD with multisystem involvement. Heavy‐chain deposition disease (HCDD) is an extremely rare form of monoclonal immunoglobulin deposition disease (MIDD) that involves the deposition of truncated immunoglobulin heavy chains in the kidneys. Only a few cases of HCDD with a favorable long‐term renal prognosis have been reported, resulting in limited long‐term follow‐up data for this patient population. In this report, we present the case of a 52‐year‐old patient with nephrotic syndrome who experienced renal failure and cardiac failure. Renal biopsy confirmed the presence of γ3‐HCDD and monoclonal Immunoglobulin G (IgG)κ in the serum. The patient exhibited low voltage on electrocardiogram (ECG) and unexplained left ventricular hypertrophy on cardiac ultrasound. The patient underwent eight cycles of bortezomib‐based chemotherapy, which led to hematological remission. After 4 years of follow‐up, the patient's renal function remained stable, with serum creatinine levels ranging from 0.7 to 0.9 mg/dL and proteinuria of 0.3–0.5 g/24 h. Our findings suggest that bortezomib‐based chemotherapy is equally effective in HCDD patients with combined multisystem damage. [ABSTRACT FROM AUTHOR]

The Mycoplasma Immunoglobulin Binding/Protease (MIB-MIP) system is a candidate 'virulence factor present in multiple pathogenic species of the Mollicutes, including the fast-growing species Mycoplasma feriruminatoris. The MIB-MIP system cleaves the heavy chain of host immunoglobulins, hence affecting antigen-antibody interactions and potentially facilitating immune evasion. In this work, using -omics technologies and 5'RACE, we show that the four copies of the M. feriruminatoris MIB-MIP system have different expression levels and are transcribed as operons controlled by four different promoters. Individual MIB-MIP gene pairs of M. feriruminatoris and other Mollicutes were introduced in an engineered M. feriruminatoris strain devoid of MIB-MIP genes and were tested for their functionality using newly developed oriC-based plasmids. The two proteins are functionally expressed at the surface of M. feriruminatoris, which confirms the possibility to display large membrane-associated proteins in this bacterium. However, functional expression of heterologous MIB-MIP systems introduced in this engineered strain from phylogenetically distant porcine Mollicutes like Mesomycoplasma hyorhinis or Mesomycoplasma hyopneumoniae could not be achieved. Finally, since M. feriruminatoris is a candidate for biomedical applications such as drug delivery, we confirmed its safety in vivo in domestic goats, which are the closest livestock relatives to its native host the Alpine ibex. A microbiology study on the creation of plasmids for the candidate vaccine chassis Mycoplasma feriruminatoris and its use to study anti-Ig systems. The study also confirms safety of M. feriruminatoris when used in domestic goats. [ABSTRACT FROM AUTHOR]

Mutated or unmutated immunoglobulin heavy chain (IGHV) gene is an important prognostic factor in chronic lymphocytic leukemia (CLL). However, a small fraction of patients with CLL are classified as borderline (BL)-IGHV. Few data are available on this subgroup of CLL. In this paper, we retrospectively report and analyze data from 21 patients with BL-IGHV CLL, showing the heterogeneity of this subgroup of CLL and paving the way for more research focusing on this entity to optimize the management and treatment of patients with Borderline- IGHV CLL. [ABSTRACT FROM AUTHOR]

Introduction: Somatic hypermutation (SHM) of immunoglobulin variable (V) regions by activation induced deaminase (AID) is essential for robust, longterm humoral immunity against pathogen and vaccine antigens. AID mutates cytosines preferentially within WRCH motifs (where W=A or T, R=A or G and H=A, C or T). However, it has been consistently observed that the mutability of WRCH motifs varies substantially, with large variations in mutation frequency even between multiple occurrences of the same motif within a single V region. This has led to the notion that the immediate sequence context of WRCH motifs contributes to mutability. Recent studies have highlighted the potential role of local DNA sequence features in promoting mutagenesis of AGCT, a commonly mutated WRCH motif. Intriguingly, AGCT motifs closer to 5' ends of V regions, within the framework 1 (FW1) sub-region1, mutate less frequently, suggesting an SHM-suppressing sequence context. Methods: Here, we systematically examined the basis of AGCT positional biases in human SHM datasets with DeepSHM, a machine-learning model designed to predict SHM patterns. This was combined with integrated gradients, an interpretability method, to interrogate the basis of DeepSHM predictions. Results: DeepSHM predicted the observed positional differences in mutation frequencies at AGCT motifs with high accuracy. For the conserved, lowly mutating AGCT motifs in FW1, integrated gradients predicted a large negative contribution of 5'C and 3'G flanking residues, suggesting that a CAGCTG context in this location was suppressive for SHM. CAGCTG is the recognition motif for Ebox transcription factors, including E2A, which has been implicated in SHM. Indeed, we found a strong, inverse relationship between E-box motif fidelity and mutation frequency. Moreover, E2A was found to associate with the V region locale in two human B cell lines. Finally, analysis of human SHM datasets revealed that naturally occurring mutations in the 3'G flanking residues, which effectively ablate the E-box motif, were associated with a significantly increased rate of AGCT mutation. Discussion: Our results suggest an antagonistic relationship between mutation frequency and the binding of E-box factors like E2A at specific AGCT motif contexts and, therefore, highlight a new, suppressive mechanism regulating local SHM patterns in human V regions. [ABSTRACT FROM AUTHOR]

Chronic lymphocytic leukemia (CLL) is the most common leukemia in adults. One of the best established CLL prognostic markers is the somatic hypermutational status of the IGHV gene which is a part of the immunoglobulin heavy chain variable region. Technology for IGHV genotyping has been optimized and has been applied in routine diagnostics for the first time in Bulgaria. A total of 105 patients with CLL from different Bulgarian regions were tested. IGHV mutational status was determined by Sanger sequencing on total genomic DNA (gDNA) or RNA extracted from mononuclear cells. All sequencing profiles were analyzed with the IMGT/V-QUEST tool. Within the course of the analysis a high percentage of IGHV unmutated status was established in the Varna district on the Black Sea (Northeast Bulgaria). In addition, the IGHV genotyping performed on gDNA revealed a rare case with multiple rearrangements. The present data from IGHV genotyping will help in choosing the proper treatment for the benefit of Bulgarian CLL patients. [ABSTRACT FROM AUTHOR]

2,3,7,8‐Tetrachlorodibenzo‐p‐dioxin (TCDD) is a persistent environmental contaminant and high affinity ligand for the aryl hydrocarbon receptor (AhR). In animal models, AhR activation by TCDD generally inhibits antibody secretion. However, it is less clear if this translates to human antibody production. Using a human Burkitt lymphoma B-cell line (CL-01) that can be stimulated to secrete Ig and undergo class switch recombination to other Ig isotypes, the current study evaluated the effects of AhR activation or antagonism on the human Ig isotypic expression profile with CD40L+IL-4 stimulation. Our results suggest that AhR agonists (TCDD and indirubin) have little to no effect on IgM or IgA secretion, which were also not induced with stimulation. However, AhR activation significantly inhibited stimulation-induced IgG secretion, an effect reversed by the AhR antagonist CH223191. Evaluation of Ig heavy chain (IgH) constant region gene expression (ie Cμ, Cγ1-4, Cα1-2, and Cε that encode for IgM, IgG1-4, IgA1-2, and IgE, respectively) demonstrated differential effects. While Cμ and Cα2 transcripts were unaffected by stimulation or AhR agonists, AhR activation significantly inhibited stimulation-induced Cγ2-4 and Cε mRNA transcripts, which was reversed by AhR antagonism. Notably, AhR antagonism in the absence of exogenous AhR ligands significantly increased IgG and IgA secretion as well as the expression of Cγ2-4 and Cε. These results suggest that modulation of AhR activity differentially alters the IgH isotypic expression profile and antibody secretion that may be partly dependent on cellular stimulation. Since a variety of chemicals from anthropogenic, industrial, pharmaceutical, dietary, and bacterial sources bind the AhR, the ability of environmental exposures to alter AhR activity (i.e. activate or inhibit) may have a direct influence on immune function and antibody-relevant disease conditions. [ABSTRACT FROM AUTHOR]

The anaplastic lymphoma kinase (ALK, CD247) is a potential target for antibody-based therapy. However, no antibody-based therapeutics targeting ALK have entered clinical trials, necessitating the development of novel antibodies with unique therapeutic merits. Single-domain antibodies (sdAb) bear therapeutic advantages compared to the full-length antibody including deeper tumor penetration, cost-effective production and fast washout from normal tissues. In this study, we identified a human immunoglobulin heavy chain variable domain (VH domain) (VH20) from an in-house phage library. VH20 exhibits good developability and high specificity with no off-target binding to ~6000 human membrane proteins. VH20 efficiently bound to the glycine-rich region of ALK with an EC50 of 0.4 nM and a KD of 6.54 nM. Both VH20-based bispecific T cell engager (TCE) and chimeric antigen receptor T cells (CAR Ts) exhibited potent cytolytic activity to ALK-expressing tumor cells in an ALK-dependent manner. VH20 CAR Ts specifically secreted proinflammatory cytokines including IL-2, TNFα and IFNγ after incubation with ALK-positive cells. To our knowledge, this is the first reported human single-domain antibody against ALK. Our in vitro characterization data indicate that VH20 could be a promising ALK-targeting sdAb with potential applications in ALK-expressing tumors, including neuroblastoma (NBL) and non-small cell lung cancer. [ABSTRACT FROM AUTHOR]

Venetoclax is active in both frontline and relapsed/refractory settings for the treatment of chronic lymphocytic leukemia (CLL). Although the prevalence and severity of tumor lysis syndrome (TLS) are well characterized in clinical trials, laboratory and clinical TLS remain relatively unexplored in real-world clinical practice. In this prospective, real-world observational study, we aimed to determine the incidence and outcomes of TLS in patients with CLL receiving venetoclax outside a clinical trial. The study (VeRVe) was conducted in centers in Austria, Germany, and Switzerland. Two hundred and thirty-nine patients were treated according to local label with at least one dose of venetoclax. Patient demographics, baseline characteristics, and blood chemistry at baseline were documented, and descriptive statistical analyses were conducted. Seventy eight patients (33%) were treated with venetoclax monotherapy, 101 (42%) with venetoclax in combination with rituximab and 60 (25%) with venetoclax in combination with obinutuzumab. In all cases, the TLS risk mitigation strategy adhered to the ramp-up protocol. Median age was 73 years and 66% of patients were male. The majority of patients (75%) had relapsed/refractory CLL, 63/192 (32.8%) patients tested had a del(17p) and 93/134 (69.4%) patients tested had unmutated immunoglobulin heavy chain variable region gene (IGHV). Clinical TLS occurred in 5 patients (2.1%) and laboratory TLS occurred in 15 patients (6.3%). Ten patients received specific treatment, of which 6 were hospitalized. There were no deaths due to a TLS event and venetoclax was well-tolerated. Of the 5 clinical TLS events reported, none were fatal or resulted in renal failure (NCT03342144, registered on Nov 10, 2017). [ABSTRACT FROM AUTHOR]

Due to their exceptional solubility and stability, nanobodies have emerged as powerful building blocks for research tools and therapeutics. However, their generation in llamas is cumbersome and costly. Here, by inserting an engineered llama immunoglobulin heavy chain (IgH) locus into IgH-deficient mice, we generate a transgenic mouse line, which we refer to as 'LamaMouse'. We demonstrate that LamaMice solely express llama IgH molecules without association to Igκ or λ light chains. Immunization of LamaMice with AAV8, the receptor-binding domain of the SARS-CoV-2 spike protein, IgE, IgG2c, and CLEC9A enabled us to readily select respective target-specific nanobodies using classical hybridoma and phage display technologies, single B cell screening, and direct cloning of the nanobody-repertoire into a mammalian expression vector. Our work shows that the LamaMouse represents a flexible and broadly applicable platform for a facilitated selection of target-specific nanobodies. Nanobodies are normally made from immunized camelids, Ig transgenic mice or synthetic libraries. In this study, the authors introduce the llama Ig heavy chain locus into mice lacking this locus, thereby generating a line in which nanobodies can be made by direct immunization in the absence of an endogenous antibody repertoire. [ABSTRACT FROM AUTHOR]

This article discusses new insights into the mechanisms of V(D)J recombination in different immunoglobulin loci. The study reveals that recombination signal sequences (RSS) within the IGKV locus have evolved to be more efficient mediators of recombination compared to the same elements in the IGH locus. The difference in RSS strength is proposed to be driven by different molecular mechanisms for recombination between the two loci. The findings contribute to our understanding of the generation of adaptive receptor genes and the diversity of the immune system. [Extracted from the article]

Introduction: Lymphomas are among the most important and common malignant tumors in cats. Differentiating lymphomas from reactive lymphoid proliferations can be challenging, so additional tools such as clonality assessment by PCR are important in diagnosis finding. Several PCR assays have been developed to assess clonality in feline lymphomas. For T-cell lymphomas TRG (T-cell receptor gamma) genes are the preferred target whereas for B-cell lymphomas most primer sets target immunoglobulin heavy chain (IGH) genes. Here we compare commonly used diagnostic primer sets for the assessment of clonality in feline lymphomas under controlled conditions (i.e., identical sample set, PCR setup, amplicon detection system). Methods: Formalin-fixed and paraffin-embedded samples from 31 feline T-cell lymphomas, 29 B-cell lymphomas, and 11 non-neoplastic controls were analyzed by PCR combined with capillary electrophoresis. Results and discussion: We show that the combination of the primer sets published by Weiss et al. and Mochizuki et al. provided the best results for T-cell clonality, i.e., correctly assigns most populations as clonal or polyclonal. For B-cell clonality, the combination of the primer sets by Mochizuki et al. and Rout et al. gave the best results when omitting the Kde gene rearrangement due to its low specificity. This study rigorously evaluated various primer sets under uniform experimental conditions to improve accuracy of lymphoma diagnostic and provides a recommendation for achieving the highest diagnostic precision in lymphoma clonality analysis. [ABSTRACT FROM AUTHOR]

Background: Richter transformation refers to the progression of an initially slowgrowing small lymphocytic lymphoma/chronic lymphocytic leukemia (SLL/CLL) into an aggressive lymphoma, typically diffuse large B-cell lymphoma (DLBCL) or Hodgkin lymphoma. Case presentation: The patient presented with a rapid onset of localized cervical swelling, accompanied by monoclonal B-cell lymphocytosis displaying a CLL immunophenotype. The histopathological analysis identified a Burkitt lymphoma (BL) located in the submandibular gland and adjacent lymph node. The patient's bone marrow displayed a minor infiltration of monoclonal B-cells with a CLL immunophenotype (< 10%). Molecular analysis demonstrated the presence of the same monoclonal rearrangement in the framework region (FR3 region) of the immunoglobulin heavy chain (IGH) locus. High-throughput sequencing of the immunoglobulin heavy and light chains also confirmed the presence of the same rearrangement in SLL/CLL and in the Burkitt lymphoma sample, but also highlighted the presence of a second rearrangement in the Burkitt lymphoma cells, not shared with the SLL/CLL cells in the bone marrow. The patient was treated with DA-EPOCH-R, which lead to a complete metabolic response. Conclusion: This report provides an exceptionally rare description of a CLL-type monoclonal B-cell lymphocytosis transforming into a very aggressive Burkitt lymphoma in a treatment naïve patient. [ABSTRACT FROM AUTHOR]

Vibriosis, caused by Vibrio, seriously affects the health of fish, shellfish, and shrimps, causing large economic losses. Teleosts are represent the first bony vertebrates with both innate and adaptive immune responses against pathogens. Aquatic animals encounter hydraulic pressure and more pathogens, compared to terrestrial animals. The skin is the first line of defense in fish, constituting the skin-associated lymphoid tissue (SALT), which belongs to the main mucosa-associated lymphoid tissues (MALT). However, little is known about the function of immunity related proteins in fish. Therefore, this study used iTRAQ (isobaric tags for relative and absolute quantitation) to compare the skin proteome between the resistant and susceptible families of Cynoglossus semilaevis. The protein integrin beta-2, the alpha-enolase isoform X1, subunit B of V-type proton ATPase, eukaryotic translation initiation factor 6, and ubiquitin-like protein ISG15, were highly expressed in the resistant family. The 16S sequencing of the skin tissues of the resistant and susceptible families showed significant differences in the microbial communities of the two families. The protein-microbial interaction identified ten proteins associated with skin microbes, including immunoglobulin heavy chain gene (IGH), B-cell lymphoma/leukemia 10 (BCL10) and pre-B-cell leukemia transcription factor 1 isoform X2 (PBX2). This study highlights the interaction between skin proteins and the microbial compositions of C. semilaevis and provides new insights into understanding aquaculture breeding research. [ABSTRACT FROM AUTHOR]

Primary vitreoretinal lymphoma is a potentially aggressive intraocular malignancy with poor systemic prognosis and sometimes significant diagnostic delays as it may masquerade as chronic uveitis. Despite the variety of diagnostic techniques, it is unclear which modality is most accurate in the diagnosis of PVRL. A systematic literature search was conducted on Ovid MEDLINE, EMBASE and the Cochrane Controlled Register of Trials for studies published between January, 2000, and June, 2023. Randomized controlled trials (RCTs) reporting on the following diagnostic tools used to diagnose patients with PVRL were included: cytology, flow cytometry, MYD88 L265P mutation, CD79B mutation, interleukin 10/interleukin-6 (IL-10/IL-6) ratio, polymerase chain reaction (PCR) for monoclonal immunoglobulin heavy chain (IgH) and immunoglobulin kappa light chain (IgK) rearrangements, and imaging findings. The aggregated sensitivity of each diagnostic modality was reported and compared using the chi-squared (χ2) test. A total of 662 eyes from 29 retrospective studies reporting on patients diagnosed with PVRL were included. An IL-10/IL-6 ratio greater than 1 had the highest sensitivity (89.39%, n = 278/311 eyes, n = 16 studies) for PVRL, where the sensitivity was not significantly different when only vitreous samples were drawn (88.89%, n = 232/261 eyes, n = 13 studies) compared to aqueous samples (83.33%, n = 20/24, n = 2) (p = 0.42). Flow cytometry of vitreous samples gave a positive result in 66/75 eyes (88.00%, n = 6 studies) with PVRL, and monoclonal IgH rearrangements on PCR gave a positive result in 354/416 eyes (85.10%, n = 20 studies) with PVRL. MYD88 L265P and CD79B mutation analysis performed poorly, yielding a positive result in 63/90 eyes (70.00%, n = 8 studies) with PVRL, and 20/57 eyes (35.09%, n = 4 studies) with PVRL, respectively. Overall, our systematic review found that an IL-10/IL-6 ratio greater or equal to one may provide the highest sensitivity in identifying patients with PVRL. Future studies are needed to employ multiple diagnostic tools to aid in the detection of PVRL and to further establish nuanced guidelines when determining the optimal diagnostic tool to use in diverse patient populations. [ABSTRACT FROM AUTHOR]

Whey, a component of milk and a useful by-product of the dairy industry's casein and cheese-making, has been used for generations to augment animal feed. It contains a range of proteins, including α-lactalbumin, β-lactoglobulin, bovine serum albumin, heavy and light chain immunoglobulins, lactoferrin, glycomacropeptide, and lactoperoxidase. Whey proteins exhibit great potential as biopolymers for creating bioactive delivery systems owing to their distinct health-enhancing characteristics and the presence of numerous amino acid groups within their structures. Whey has considerable factors such as antitumor, anti-inflammatory, antihypertensive, hypolipidemic, antiviral, and antibacterial properties in addition to chelating. The global market of whey protein stood at USD 5.33 billion in 2021, with a projected compound annual growth rate of 10.48% spanning the interval from 2022 to 2030. The escalating demand for whey protein is intrinsically linked to the amplifying consciousness surrounding healthy lifestyles. Notably, protein supplements are recurrently endorsed by fitness and sports establishments, thereby accentuating the focal point of customers toward whey protein. This review focuses on nutritional composition, whey bioactives, and their bioavailability with potential health benefits. [ABSTRACT FROM AUTHOR]

Recent studies have demonstrated a role for Ten-Eleven Translocation-2 (TET2), an epigenetic modulator, in regulating germinal center formation and plasma cell differentiation in B-2 cells, yet the role of TET2 in regulating B-1 cells is largely unknown. Here, B-1 cell subset numbers, IgM production, and gene expression were analyzed in mice with global knockout of TET2 compared to wildtype (WT) controls. Results revealed that TET2-KO mice had elevated numbers of B-1a and B-1b cells in their primary niche, the peritoneal cavity, as well as in the bone marrow (B-1a) and spleen (B-1b). Consistent with this finding, circulating IgM, but not IgG, was elevated in TET2-KO mice compared to WT. Analysis of bulk RNASeq of sort purified peritoneal B-1a and B-1b cells revealed reduced expression of heavy and light chain immunoglobulin genes, predominantly in B-1a cells from TET2-KO mice compared to WT controls. As expected, the expression of IgM transcripts was the most abundant isotype in B-1 cells. Yet, only in B-1a cells there was a significant increase in the proportion of IgM transcripts in TET2-KO mice compared to WT. Analysis of the CDR3 of the BCR revealed an increased abundance of replicated CDR3 sequences in B-1 cells from TET2-KO mice, which was more clearly pronounced in B-1a compared to B-1b cells. V-D-J usage and circos plot analysis of V-J combinations showed enhanced usage of VH11 and VH12 pairings. Taken together, our study is the first to demonstrate that global loss of TET2 increases B-1 cell number and IgM production and reduces CDR3 diversity, which could impact many biological processes and disease states that are regulated by IgM. [ABSTRACT FROM AUTHOR]

Human herpesvirus 8 (HHV8)-associated diseases include Kaposi sarcoma (KS), multicentric Castleman disease (MCD), germinotropic lymphoproliferative disorder (GLPD), Kaposi sarcoma inflammatory cytokine syndrome (KICS), HHV8-positive diffuse large B-cell lymphoma (HHV8+ DLBCL), primary effusion lymphoma (PEL), and extra-cavitary PEL (ECPEL). We report the case of a human immunodeficiency virus (HIV)-negative male treated for cutaneous KS, who developed generalized lymphadenopathy, hepatosplenomegaly, pleural and abdominal effusions, renal insufficiency, and pancytopenia. The excised lymph node showed features of concomitant involvement by micro-KS and MCD, with aggregates of HHV8+, Epstein Barr virus (EBV)-negative, IgM+, and lambda+ plasmablasts reminiscent of microlymphoma. Molecular investigations revealed a somatically hypermutated (SHM) monoclonal rearrangement of the immunoglobulin heavy chain (IGH), accounting for 4% of the B-cell population of the lymph node. Mutational analyses identified a pathogenic variant of KMT2D and variants of unknown significance in KMT2D, FOXO1, ARID1A, and KMT2A. The patient died shortly after surgery. The histological features (HHV8+, EBV−, IgM+, Lambda+, MCD+), integrated with the molecular findings (monoclonal IGH, SHM+, KMT2D mutated), supported the diagnosis of a monoclonal HHV8+ microlymphoma, with features intermediate between an incipient HHV8+ DLBCL and an EBV-negative ECPEL highlighting the challenges in the accurate classification of HHV8-driven lymphoid proliferations. [ABSTRACT FROM AUTHOR]

Class switch recombination (CSR) plays an important role in adaptive immune response by enabling mature B cells to replace the initial IgM by another antibody class (IgG, IgE or IgA). CSR is preceded by transcription of the IgH constant genes and is controlled by the super-enhancer 3′ regulatory region (3′RR) in an activation-specific manner. The 3'RR is composed of four enhancers (hs3a, hs1-2, hs3b and hs4). In mature B cells, 3'RR activity correlates with transcription of its enhancers. CSR can also occur in primary developing B cells though at low frequency, but in contrast to mature B cells, the transcriptional elements that regulate the process in developing B cells are ill-known. In particular, the role of the 3'RR in the control of constant genes' transcription and CSR has not been addressed. Here, by using a mouse line devoid of the 3'RR and a culture system that highly enriches in pro-B cells, we show that the 3'RR activity is indeed required for switch transcription and CSR, though its effect varies in an isotype-specific manner and correlates with transcription of hs4 enhancer only. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the efficacy of next-generation sequencing (NGS) in minimal-residual-disease (MRD) monitoring in Chinese patients with multiple myeloma (MM). Methods: This study analyzed 60 Chinese MM patients. During MRD monitoring in these patients' post-therapy, clonal immunoglobulin heavy chain (IGH) rearrangements were detected via NGS using LymphoTrack assays. MRD monitoring was performed using NGS or next-generation flow cytometry (NGF), and the results were compared. Additionally, the sensitivity and reproducibility of the NGS method were assessed. Results: The MRD detection range of the NGS method was 10–6–10–1, which suggested good linearity, with a Pearson correlation coefficient of 0.985 and a limit of detection of 10–6. Intra- and inter-assay reproducibility analyses showed that NGS exhibited 100% reproducibility with low variability in clonal cells. At diagnosis, unique clones were found in 42 patients (70.0%) with clonal IGH rearrangements, which were used as clonality markers for MRD monitoring post-therapy. Comparison of NGS and NGF for MRD monitoring showed 79.1% concordance. No samples that tested MRD-positive via NGF were found negative via NGS, indicating the higher sensitivity of NGS. MRD could be detected using NGS in 6 of 7 samples before autologous hematopoietic stem-cell transplantation, and 5 of them tested negative post-transplantation. In contrast, the NGF method could detect MRD in only 1 sample pre-transplantation. Conclusion: Compared with NGF, NGS exhibits higher sensitivity and reproducibility in MRD detection and can be an effective strategy for MRD monitoring in Chinese MM patients. [ABSTRACT FROM AUTHOR]

Vaccine priming immunogens that activate germline precursors for broadly neutralizing antibodies (bnAbs) have promise for development of precision vaccines against major human pathogens. In a clinical trial of the eOD-GT8 60mer germline-targeting immunogen, higher frequencies of vaccine-induced VRC01-class bnAb-precursor B cells were observed in the high dose compared to the low dose group. Through immunoglobulin heavy chain variable (IGHV) genotyping, statistical modeling, quantification of IGHV1-2 allele usage and B cell frequencies in the naive repertoire for each trial participant, and antibody affinity analyses, we found that the difference between dose groups in VRC01-class response frequency was best explained by IGHV1-2 genotype rather than dose and was most likely due to differences in IGHV1-2 B cell frequencies for different genotypes. The results demonstrate the need to define population-level immunoglobulin allelic variations when designing germline-targeting immunogens and evaluating them in clinical trials. [ABSTRACT FROM AUTHOR]

B cells and their secreted antibodies are fundamental for host-defense against pathogens. The generation of high-affinity class switched antibodies results from both somatic hypermutation (SHM) of the immunoglobulin (Ig) variable region genes of the B-cell receptor and class switch recombination (CSR) which alters the Ig heavy chain constant region. Both of these processes are initiated by the enzyme activation-induced cytidine deaminase (AID), encoded by AICDA. Deleterious variants in AICDA are causal of hyper-IgM syndrome type 2 (HIGM2), a B-cell intrinsic primary immunodeficiency characterised by recurrent infections and low serum IgG and IgA levels. Biallelic variants affecting exons 2, 3 or 4 of AICDA have been identified that impair both CSR and SHM in patients with autosomal recessive HIGM2. Interestingly, B cells from patients with autosomal dominant HIGM2, caused by heterozygous variants (V186X, R190X) located in AICDA exon 5 encoding the nuclear export signal (NES) domain, show abolished CSR but variable SHM. We herein report the immunological and functional phenotype of two related patients presenting with common variable immunodeficiency who were found to have a novel heterozygous variant in AICDA (L189X). This variant led to a truncated AID protein lacking the last 10 amino acids of the NES at the C-terminal domain. Interestingly, patients' B cells carrying the L189X variant exhibited not only greatly impaired CSR but also SHM in vivo, as well as CSR and production of IgG and IgA in vitro. Our findings demonstrate that the NES domain of AID can be essential for SHM, as well as for CSR, thereby refining the correlation between AICDA genotype and SHM phenotype as well as broadening our understanding of the pathophysiology of HIGM disorders. [ABSTRACT FROM AUTHOR]

Despite extensive research on severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) vaccination responses in healthy individuals, there is comparatively little known beyond antibody titers and T‐cell responses in the vulnerable cohort of patients after allogeneic hematopoietic stem cell transplantation (ASCT). In this study, we assessed the serological response and performed longitudinal multimodal analyses including T‐cell functionality and single‐cell RNA sequencing combined with T cell receptor (TCR)/B cell receptor (BCR) profiling in the context of BNT162b2 vaccination in ASCT patients. In addition, these data were compared to publicly available data sets of healthy vaccinees. Protective antibody titers were achieved in 40% of patients. We identified a distorted B‐ and T‐cell distribution, a reduced TCR diversity, and increased levels of exhaustion marker expression as possible causes for the poorer vaccine response rates in ASCT patients. Immunoglobulin heavy chain gene rearrangement after vaccination proved to be highly variable in ASCT patients. Changes in TCRα and TCRβ gene rearrangement after vaccination differed from patterns observed in healthy vaccinees. Crucially, ASCT patients elicited comparable proportions of SARS‐CoV‐2 vaccine‐induced (VI) CD8+ T‐cells, characterized by a distinct gene expression pattern that is associated with SARS‐CoV‐2 specificity in healthy individuals. Our study underlines the impaired immune system and thus the lower vaccine response rates in ASCT patients. However, since protective vaccine responses and VI CD8+ T‐cells can be induced in part of ASCT patients, our data advocate early posttransplant vaccination due to the high risk of infection in this vulnerable group. [ABSTRACT FROM AUTHOR]

Introduction: Analysis of an individual's immunoglobulin (IG) gene repertoire requires the use of high-quality germline gene reference sets. When sets only contain alleles supported by strong evidence, AIRR sequencing (AIRR-seq) data analysis is more accurate and studies of the evolution of IG genes, their allelic variants and the expressed immune repertoire is therefore facilitated. Methods: The Adaptive Immune Receptor Repertoire Community (AIRR-C) IG Reference Sets have been developed by including only human IG heavy and light chain alleles that have been confirmed by evidence from multiple high-quality sources. To further improve AIRR-seq analysis, some alleles have been extended to deal with short 3' or 5' truncations that can lead them to be overlooked by alignment utilities. To avoid other challenges for analysis programs, exact paralogs (e.g. IGHV1-69*01 and IGHV1-69D*01) are only represented once in each set, though alternative sequence names are noted in accompanyingmetadata. Results and discussion: The Reference Sets include less than half the previously recognised IG alleles (e.g. just 198 IGHV sequences), and also include a number of novel alleles: 8 IGHV alleles, 2 IGKV alleles and 5 IGLV alleles. Despite their smaller sizes, erroneous calls were eliminated, and excellent coverage was achieved when a set of repertoires comprising over 4 million V(D)J rearrangements from 99 individuals were analyzed using the Sets. The versiontracked AIRR-C IG Reference Sets are freely available at the OGRDB website (https://ogrdb.airr-community.org/germline_sets/Human) and will be regularly updated to include newly observed and previously reported sequences that can be confirmed by new high-quality data. [ABSTRACT FROM AUTHOR]

Summary: Chronic lymphocytic leukaemia (CLL) is a clonal B‐cell malignancy and remains a chronic disease despite improvements in clinical outcomes since the use of targeted therapies. Both clinical and biological parameters are important for determining prognosis. Unlike other mature B‐cell lymphomas, translocations involving the immunoglobulin heavy chain (IGH) locus are uncommon in CLL. There have been few case reports of CLL harbouring t(14;18)/IGH::BCL2 and t(14;19)/IGH::BCL3. Here we describe the first two cases of patients with CLL with documented t(14;18)(q32;q21)/IGH::MALT1. Both cases in this report were associated with lower‐risk biological parameters. Thus, FISH testing for MALT1 in cases with unknown IGH translocation partners in the setting of CLL should be implemented in clinical practice to better define such cases. [ABSTRACT FROM AUTHOR]

Lymphoplasmacytic lymphoma (LPL) is a rare variant of non‐Hodgkin lymphoma, accounting for <1% of cases. Skin involvement in LPL is quite rare—accounting for approximately 5% of extramedullary disease—and includes a variety of clinical morphologies, such as erythematous‐to‐violaceous plaques, violaceous nodules or tumors, and ulceration at various anatomical sites. Herein, we report the case of a 45‐year‐old Korean woman who presented with generalized erythematous indurated plaques and pendulous skin growths, which were asymptomatic, with marked diffuse infiltration of lymphocytes and plasma cells in the dermis. Immunohistochemical studies revealed that the lymphoid cells expressed CD3, CD79a, and cytoplasmic IgG, but lacked CD10 and IgM. Moreover, kappa light chain restriction and monoclonal immunoglobulin heavy chain gene rearrangement were observed. Upon further workup, lymphoma involvement was reported in multiple lymph nodes, including those in the cervical and axillary regions. This case shows a unique form of cutaneous LPL clinically presenting as acquired cutis laxa, emphasizing the dermatologists' need to be vigilant for variant forms of this disease. [ABSTRACT FROM AUTHOR]

During B cell development, pre-B cell receptor (pre-BCR), comprising the immunoglobulin heavy chain (HC) and surrogate light chain (SLC), plays a crucial role. The expression of pre-BCR serves as a certification of HC quality, confirming its ability to associate with the SLC and light chain (LC). In mice lacking SLC, the absence of this quality control mechanism leads to a distorted repertoire of HCs in the spleen and bone marrow. In this study, we conducted a comparative analysis of the immunoglobulin gene repertoire in peripheral blood cells of both wild-type mice and pre-BCR–deficient mice. Our findings reveal differences not only in the μ HC repertoire but also in the α HC and κ LC repertoires of the pre-BCR–deficient mice. These results suggest that the pre-BCR–mediated quality check of HC influences the selection of class-switched HC and LC repertoires. To further explore the impact of pre-BCR deficiency, we immunized these mice with thymus-dependent antigens and compared the antigen-responding repertoires. Our observations indicate that the affinity maturation pathways remain consistent between wild-type mice and pre-BCR–deficient mice, albeit with variations in the degree of maturation. • Antibody repertoires of peripheral blood of pre-BCR–deficient mice were analyzed. • Lack of pre-BCR distorts heavy and light chain repertoires. • Affinity maturation proceeds along the same pathways but to a different degree. [ABSTRACT FROM AUTHOR]

Introduction: Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease. Anti-B-cell-activating factor (BAFF) therapy effectively depletes B cells and reduces SLE disease activity. This research aimed to evaluate the effect of BAFF blockade on B cell receptor (BCR) repertoire and gene expression. Methods: Through next-generation sequencing, we analyzed gene expression and BCR repertoire in MRL/lpr mice that received long-term anti-BAFF therapy. Based on gene expression profiles, we predicted the relative proportion of immune cells using ImmuCellAI-mouse, validating our predictions via flow cytometry and FluoroSpot. Results: The loss of BCR repertoire diversity and richness, along with increased clonality and differential frequency distribution of the immunoglobulin heavy chain variable (IGHV) segment gene usage, were observed in BAFF-blockade mice. Meanwhile, the distribution of complementarity-determining region 3 (CDR3) length and CDR3 amino acid usage remained unaffected. BAFF blockade resulted in extensive changes in gene expression, particularly that of genes related to B cells and immunoglobulins. Besides, the tumor necrosis factor (TNF)-a responses and interferon (IFN)-α/γ were downregulated, consistent with the decrease in IFN-γ and TNF-α serum levels following anti-BAFF therapy. In addition, BAFF blockade significantly reduced B cell subpopulations and plasmacytoid dendritic cells, and caused the depletion of antibodysecreting cells. Discussion: Our comparative BCR repertoire and transcriptome analyses of MRL/lpr mice subjected to BAFF blockade provide innovative insights into the molecular pathophysiology of SLE. [ABSTRACT FROM AUTHOR]

One gene, the immunoglobulin heavy chain (IgH) gene, is responsible for the expression of all the different antibody isotypes. Transcriptional regulation of the IgH gene is complex and involves several regulatory elements including a large element at the 3' end of the IgH gene locus (3'RR). Animal models have demonstrated an essential role of the 3'RR in the ability of B cells to express high affinity antibodies and to express different antibody classes. Additionally, environmental chemicals such as aryl hydrocarbon receptor (AhR) ligands modulate mouse 3'RR activity that mirrors the effects of these chemicals on antibody production and immunocompetence in mouse models. Although first discovered as a mediator of the toxicity induced by the high affinity ligand 2,3,7,8-tetracholordibenzo-p-dioxin (dioxin), understanding of the AhR has expanded to a physiological role in preserving homeostasis and maintaining immunocompetence. We posit that the AhR also plays a role in human antibody production and that the 3'RR is not only an IgH regulatory node but also an environmental sensor receiving signals through intrinsic and extrinsic pathways, including the AhR. This review will 1) highlight the emerging role of the AhR as a key transducer between environmental signals and altered immune function; 2) examine the current state of knowledge regarding IgH gene regulation and the role of the AhR in modulation of Ig production; 3) describe the evolution of the IgH gene that resulted in species and population differences; and 4) explore the evidence supporting the environmental sensing capacity of the 3'RR and the AhR as a transducer of these cues. This review will also underscore the need for studies focused on human models due to the premise that understanding genetic differences in the human population and the signaling pathways that converge at the 3'RR will provide valuable insight into individual sensitivities to environmental factors and antibody-mediated disease conditions, including emerging infections such as SARS-CoV-2. [ABSTRACT FROM AUTHOR]

Chronic lymphocytic leukemia is a lymphoproliferative disorder marked by the expansion of monoclonal, mature CD5+CD23+ B cells in peripheral blood, secondary lymphoid tissues, and bone marrow. The disease exhibits significant heterogeneity, with numerous somatic genetic alterations identified in the neoplastic clone, notably mutated TP53 and immunoglobulin heavy chain mutational statuses. Recent studies emphasize the pivotal roles of genetics and patient fragility in treatment decisions. This complexity underscores the need for a personalized approach, tailoring interventions to individual genetic profiles for heightened efficacy. The era of personalized treatment in CLL signifies a transformative shift, holding the potential for improved outcomes in the conquest of this intricate hematologic disorder. This review plays a role in elucidating the evolving CLL treatment landscape, encompassing all reported genetic factors. Through a comprehensive historical analysis, it provides insights into the evolution of CLL management. Beyond its retrospective nature, this review could be a valuable resource for clinicians, researchers, and stakeholders, offering a window into the latest advancements. In essence, it serves as a dynamic exploration of our current position and the promising prospects on the horizon. [ABSTRACT FROM AUTHOR]

An antibody molecule that can bind to multiple distinct antigens is defined as polyreactive. In the present study, we performed statistical analyses to assess sequence correlates of polyreactivity of >600 antibodies cloned from different Bcell types of healthy humans. The data revealed several sequence patterns of variable regions of heavy and light immunoglobulin chains that determine polyreactivity. The most prominent identified patterns were increased number of basic amino acid residues, reduced frequency of acidic residues, increased number of aromatic and hydrophobic residues, and longer length of CDR L1. Importantly, our study revealed that antibodies isolated from different B-cell populations used distinct sequence patterns (or combinations of them) for polyreactive antigen binding. Furthermore, we combined the data from sequence analyses with molecular modeling of selected polyreactive antibodies and demonstrated that human antibodies can use multiple pathways for achieving antigen-binding promiscuity. These data reconcile some contradictions in the literature regarding the determinants of antibody polyreactivity. Moreover, our study demonstrates that the mechanism of polyreactivity of antibodies evolves during immune response and might be tailored to specific functional properties of different B-cell compartments. Finally, these data can be of use for efforts in the development and engineering of therapeutic antibodies. [ABSTRACT FROM AUTHOR]

Objective: At present, little is known about the immune mechanism of liver transplantation caused by decompensated cirrhosis. Lymphocytes play an essential important role in the immune rejection of liver transplantation. In this study, we aimed to comprehensively analyze changes in complementary determinant 3 (CDR3) repertoire of T cell receptor β chain (TRβ) and immunoglobulin heavy chain (IGH) in liver transplantation patients and healthy controls (HC). Methods: High-throughput sequencing technology was used to study the characteristics of TRβ/IGH CDR3 repertoire, and identify the amino acid sequences of TRβ and IGH associated with liver transplantation patients and HC. Results: We found that some TRβ and IGH CDR3 repertoire characteristics differed between liver transplant patients and HC. The diversity of TRβ CDR3 increased in the liver transplantation group. First and seven days after live transplantation patients showed a lower degree of T cell clone amplification compared to the HC group. The CDR3 repertoire of the TRβ/IGH chain was certainly biased in the use of some V, D, and J gene segments, TRβ/IGH V-J combined frequency was also skewed and TRβ CDR3 clonotypes were shared at a higher degree in the liver transplantation patients. Importantly, one amino acid sequence in the decompensated cirrhosis group was significantly higher than that in the healthy group. It should be noted that the frequency of some CDR3 sequences is closely correlated with the different stages of liver transplantation, and these sequences may play a key role in liver transplantation. Conclusion: Based on the above results, we can better understand the dynamic changes of TCβ/IGH CDR3 repertoire in patients during liver transplantation. [ABSTRACT FROM AUTHOR]

Extranodal NK/T‐cell lymphoma (ENKTL) is a subtype of non‐Hodgkin lymphoma mainly derived from NK cells and, uncommonly, T‐cells. A diagnostic challenge is presented when an atypical phenotype and gene rearrangement are encountered. Herein, we report a case of ENKTL with CD20 expression and IGH gene rearrangement, which is extremely rare. A 57‐year‐old female patient was seen in 2021 due to a nodule on her left leg and simultaneously impaired eyesight for 6 months. Skin biopsy and immunohistochemistry were performed. The lymphoid cells were positive for CD3, CD56, granzyme B, and TIA‐1, partially positive for CD2, and mildly positive for CD20. In situ hybridization for Epstein–Barr virus was positive. Molecular studies revealed immunoglobulin heavy chain (IGH) gene rearrangement, while no T‐cell receptor gene rearrangement was detected. The positron emission tomography scan showed that the lymphoma affected bilateral adrenal glands, pelvic cavity, peritoneal cavity, small intestine, skin, and subcutis of the bilateral lower extremities of the patient. Her disease progressed despite eight cycles of chemotherapy and radiation therapy. The importance of this case lies in the atypical phenotype and IGH gene rearrangements, necessitating comprehensive interpretation of clinicopathological data. [ABSTRACT FROM AUTHOR]

This article discusses a case of pseudolymphoma, a benign reactive lymphoproliferation, on the face involving the conjunctiva. Pseudolymphoma typically presents as nodules or papules on the body, with the face being the most commonly affected area. While pseudolymphoma in the ocular region is rare, this case is unique because it involves both the skin and the conjunctiva. The patient was treated with lesional injections of corticosteroid, which resulted in regression of both the facial and ocular lesions. This is the first reported case of pseudolymphoma with facial skin and conjunctival involvement in the same patient. [Extracted from the article]

Preterm labor (PTL) is the leading cause of neonatal morbidity and mortality worldwide. Whereas many studies have investigated the maternal immune responses that cause PTL, fetal immune cell activation has recently been raised as an important contributor to the pathogenesis of PTL. In this study, we analyzed lymphocyte receptor repertoires in maternal and cord blood from 14 term and 10 preterm deliveries, hypothesizing that the high prevalence of infection in patients with PTL may result in specific changes in the T cell and B cell repertoires. We analyzed TCR β-chain (TCR-β) and IgH diversity, CDR3 lengths, clonal sharing, and preferential usage of variable and joining gene segments. Both TCR-β and IgH repertoires had shorter CDR3s compared with those in maternal blood. In cord blood samples, we found that CDR3 lengths correlated with gestational age, with shorter CDR3s in preterm neonates suggesting a less developed repertoire. Preterm cord blood displayed preferential usage of a number of genes. In preterm pregnancies, we observed significantly higher prevalence of convergent clones between mother/baby pairs than in term pregnancies. Together, our results suggest the repertoire of preterm infants displays a combination of immature features and convergence with maternal TCR-β clones compared with that of term infants. The higher clonal convergence in PTL could represent mother and fetus both responding to a shared stimulus like an infection. These data provide a detailed analysis of the maternal-fetal immune repertoire in term and preterm patients and contribute to a better understanding of neonate immune repertoire development and potential changes associated with PTL.

PurposeAdenosine deaminase (ADA) deficiency causes severe combined immunodeficiency (SCID) through an accumulation of toxic metabolites within lymphocytes. Recently, ADA deficiency has been successfully treated using lentiviral-transduced autologous CD34+ cells carrying the ADA gene. T and B cell function appears to be fully restored, but in many patients' B cell numbers remain low, and assessments of the immunoglobulin heavy (IgHV) repertoire following gene therapy are lacking.MethodsWe performed deep sequencing of IgHV repertoire in peripheral blood lymphocytes from a child following lentivirus-based gene therapy for ADA deficiency and compared to the IgHV repertoire in healthy infants and adults.ResultsAfter gene therapy, Ig diversity increased over time as evidenced by V, D, and J gene usage, N-additions, CDR3 length, extent of somatic hypermutation, and Ig class switching. There was the emergence of predominant IgHM, IgHG, and IgHA CDR3 lengths after gene therapy indicating successful oligoclonal expansion in response to antigens. This provides proof of concept for the feasibility and utility of molecular monitoring in following B cell reconstitution following gene therapy for ADA deficiency.ConclusionBased on deep sequencing, gene therapy resulted in an IgHV repertoire with molecular diversity similar to healthy infants.

The Neuroscience Monoclonal Antibody Sequencing Initiative (NeuroMabSeq) is a concerted effort to determine and make publicly available hybridoma-derived sequences of monoclonal antibodies (mAbs) valuable to neuroscience research. Over 30 years of research and development efforts including those at the UC Davis/NIH NeuroMab Facility have resulted in the generation of a large collection of mouse mAbs validated for neuroscience research. To enhance dissemination and increase the utility of this valuable resource, we applied a high-throughput DNA sequencing approach to determine immunoglobulin heavy and light chain variable domain sequences from source hybridoma cells. The resultant set of sequences was made publicly available as a searchable DNA sequence database (neuromabseq.ucdavis.edu) for sharing, analysis and use in downstream applications. We enhanced the utility, transparency, and reproducibility of the existing mAb collection by using these sequences to develop recombinant mAbs. This enabled their subsequent engineering into alternate forms with distinct utility, including alternate modes of detection in multiplexed labeling, and as miniaturized single chain variable fragments or scFvs. The NeuroMabSeq website and database and the corresponding recombinant antibody collection together serve as a public DNA sequence repository of mouse mAb heavy and light chain variable domain sequences and as an open resource for enhancing dissemination and utility of this valuable collection of validated mAbs. [ABSTRACT FROM AUTHOR]