Predicting class switch recombination in B-cells from antibody repertoire data.

Statistical and machine learning methods have proved useful in many areas of immunology. In this paper, we address for the first time the problem of predicting the occurrence of class switch recombination (CSR) in B-cells, a problem of interest in understanding antibody response under immunological challenges. We propose a framework to analyze antibody repertoire data, based on clonal (CG) group representation in a way that allows us to predict CSR events using CG level features as input. We assess and compare the performance of several predicting models (logistic regression, LASSO logistic regression, random forest, and support vector machine) in carrying out this task. The proposed approach can obtain an unweighted average recall of 71 % $71\%$ with models based on variable region descriptors and measures of CG diversity during an immune challenge and, most notably, before an immune challenge.

Virtual Remote Pathology Education in Support of Virtual Remote Gynecologic-Oncology Training: The Open Pathology Education Network Pilot Proof of Concept Experience.

CONTEXT.­: The subspecialty workforce in pathology globally is inadequate for the demands of many modern therapies. The Open Pathology Education Network (OPEN) was formed to augment the global pathology workforce. The International Gynecologic Cancer Society (IGCS) virtual gynecologic-oncology (gyn-onc) fellowship training identified needs for higher-level pathology support. OBJECTIVE.­: To report on an OPEN-IGCS pilot project to support gyn-onc and pathology education efforts in a developing country. DESIGN.­: Curriculum with learning objectives and content from open sources was assembled. Mentoring sessions included bidirectional case sharing. Trainees received sequential curricula assignments and had options for communication outside mentoring sessions. Pretest and posttest digital slide assessments were included. Mentors attended the gynecology tumor board, allowing for the assessment of quality and accuracy of pathology diagnosis for cases discussed. RESULTS.­: Learners completing the pretest and posttest showed substantial improvement, with 2 practicing pathologists improving their diagnostic scores from 60% to an average of 95%. A third trainee-level participant also improved, but to a lesser degree. Qualitative assessments included increased confidence in presentation and an increased ability to anticipate questions, raise issues of expanded differential diagnoses, and articulate appropriate workup. Observations of clinicians who participated also noted increased confidence in participating pathologists. Secondary value included establishing an expanded network of support in other subspecialties for participants. Pathologic issues at the tumor board decreased, from more than 50% in the first 3 months of study to 0% in the last 3 months of study. The curriculum was embedded into a self-paced learning portal at courses.open-pathology.org. CONCLUSIONS.­: The OPEN-IGCS collaboration model shows the potential to provide subspecialty pathology training remotely.

Comprehensive characterization of genomic features and clinical outcomes following targeted therapy and secondary cytoreductive surgery in OCCC: a single center experience.

OBJECTIVE: Ovarian clear cell carcinoma (OCCC) is associated with chemoresistance. Limited data exists regarding the efficacy of targeted therapies such as immune checkpoint inhibitors (ICI) and bevacizumab, and the role of secondary cytoreductive surgery (SCS). METHODS: We retrospectively analyzed genomic features and treatment outcomes of 172 OCCC patients treated at our institution from January 2000 to May 2022. Next-generation sequencing (NGS) was performed where sufficient archival tissue was available. RESULTS: 64.0% of patients were diagnosed at an early stage, and 36.0% at an advanced stage. Patients with advanced/relapsed OCCC who received platinum-based chemotherapy plus bevacizumab followed by maintenance bevacizumab had a median first-line progression-free survival (PFS) of 12.2 months, compared with 9.3 months for chemotherapy alone (hazard ratio=0.69; 95% confidence interval [CI]=0.33, 1.45). In 27 patients who received an ICI, the overall response rate was 18.5% and median duration of response was 7.4 months (95% CI=6.5, 8.3). In 17 carefully selected patients with fewer than 3 sites of relapse, median PFS was 35 months (95% CI=0, 73.5) and median overall survival was 96.8 months (95% CI=44.6, 149.0) after SCS. NGS on 58 tumors revealed common mutations in ARID1A (48.3%), PIK3CA (46.6%), and KRAS (20.7%). Pathogenic alterations in PIK3CA, FGFR2, and NBN were associated with worse survival outcomes. Median tumor mutational burden was 3.78 (range, 0-16). All 26 patients with available loss of heterozygosity (LOH) scores had LOH <16%. CONCLUSION: Our study demonstrates encouraging outcomes with bevacizumab and ICI, and SCS in select relapsed OCCC patients. Prospective trials are warranted.

Thioredoxin is a metabolic rheostat controlling regulatory B cells.

Metabolic programming is important for B cell fate, but the bioenergetic requirement for regulatory B (Breg) cell differentiation and function is unknown. Here we show that Breg cell differentiation, unlike non-Breg cells, relies on mitochondrial electron transport and homeostatic levels of reactive oxygen species (ROS). Single-cell RNA sequencing analysis revealed that TXN, encoding the metabolic redox protein thioredoxin (Trx), is highly expressed by Breg cells, unlike Trx inhibitor TXNIP which was downregulated. Pharmacological inhibition or gene silencing of TXN resulted in mitochondrial membrane depolarization and increased ROS levels, selectively suppressing Breg cell differentiation and function while favoring pro-inflammatory B cell differentiation. Patients with systemic lupus erythematosus (SLE), characterized by Breg cell deficiencies, present with B cell mitochondrial membrane depolarization, elevated ROS and fewer Trx+ B cells. Exogenous Trx stimulation restored Breg cells and mitochondrial membrane polarization in SLE B cells to healthy B cell levels, indicating Trx insufficiency underlies Breg cell impairment in patients with SLE.

The role of surgery in gestational trophoblastic disease: an overview.

Gestational trophoblastic disease comprises a group of rare, and potentially malignant, conditions that arise from abnormal trophoblastic proliferation. When there is invasion and evidence of metastatic disease, gestational trophoblastic neoplasia is used. While chemotherapy is the mainstay of treatment for gestational trophoblastic neoplasia, the role of surgery has come full circle in recent years. Before the introduction of highly effective systemic treatment options, surgery was the default treatment. Surgery for gestational trophoblastic neoplasia often yielded unsatisfactory results and mortality remained high. In recent years, the role of adjuvant surgery in the management of gestational trophoblastic neoplasia has been examined with great interest. We aim to provide an overview of the various surgical approaches employed in managing gestational trophoblastic neoplasia, including their indications, techniques, and outcomes. Additionally, we discuss whether there is a role to do less in surgery for gestational trophoblastic neoplasia and describe our experience with a modified surgical technique for its treatment. By summarizing the current evidence, this article highlights the significant contributions of surgery to the holistic management of patients with gestational trophoblastic neoplasia and provides a framework on which to base management and treatment programs.

Modelling the assembly and flexibility of antibody structures.

Antibodies are large protein assemblies capable of both specifically recognising antigens and engaging with other proteins and receptors to coordinate immune action. Traditionally, structural studies have been dedicated to antibody variable regions, but efforts to determine and model full-length antibody structures are emerging. Here we review the current knowledge on modelling the structures of antibody assemblies, focusing on their conformational flexibility and the challenge this poses to obtaining and evaluating structural models. Integrative modelling approaches, combining experiments (cryo-electron microscopy, mass spectrometry, etc.) and computational methods (molecular dynamics simulations, deep-learning based approaches, etc.), hold the promise to map the complex conformational landscape of full-length antibody structures.

sciCSR infers B cell state transition and predicts class-switch recombination dynamics using single-cell transcriptomic data.

Class-switch recombination (CSR) is an integral part of B cell maturation. Here we present sciCSR (pronounced 'scissor', single-cell inference of class-switch recombination), a computational pipeline that analyzes CSR events and dynamics of B cells from single-cell RNA sequencing (scRNA-seq) experiments. Validated on both simulated and real data, sciCSR re-analyzes scRNA-seq alignments to differentiate productive heavy-chain immunoglobulin transcripts from germline 'sterile' transcripts. From a snapshot of B cell scRNA-seq data, a Markov state model is built to infer the dynamics and direction of CSR. Applying sciCSR on severe acute respiratory syndrome coronavirus 2 vaccination time-course scRNA-seq data, we observe that sciCSR predicts, using data from an earlier time point in the collected time-course, the isotype distribution of B cell receptor repertoires of subsequent time points with high accuracy (cosine similarity ~0.9). Using processes specific to B cells, sciCSR identifies transitions that are often missed by conventional RNA velocity analyses and can reveal insights into the dynamics of B cell CSR during immune response.

Correction: Chua et al. Cost-Effectiveness Analysis of HPV Extended versus Partial Genotyping for Cervical Cancer Screening in Singapore. Cancers 2023, 15, 1812.

The authors wish to revise two words in Table 1 row 3, and the first paragraph of Section 2 [...].

In Silico and In Vitro Analysis of IL36RN Alterations Reveals Critical Residues for the Function of the Interleukin-36 Receptor Complex.

Generalized pustular psoriasis is a potentially life-threatening skin disease, associated with IL36RN disease alleles. IL36RN encodes the IL-36 receptor antagonist (IL-36Ra), a protein that downregulates the activity of IL-36 cytokines by blocking their receptor (IL-36R). Although generalized pustular psoriasis can be treated with IL-36R inhibitors, the structural underpinnings of the IL-36Ra/IL-36R interaction remain poorly understood. In this study, we sought to address this question by systematically investigating the effects of IL36RN sequence changes. We experimentally characterized the effects of 30 IL36RN variants on protein stability. In parallel, we used a machinelearning tool (Rhapsody) to analyze the IL-36Ra three-dimensional structure and predict the impact of all possible amino acid substitutions. This integrated approach identified 21 amino acids that are essential for IL-36Ra stability. We next investigated the effects of IL36RN changes on IL-36Ra/IL-36R binding and IL-36R signaling. Combining invitro assays and machine learning with a second program (mCSM), we identified 13 amino acids that are critical for IL-36Ra/IL36R engagement. Finally, we experimentally validated three representative predictions, further confirming the reliability of Rhapsody and mCSM. These findings shed light on the structural determinants of IL-36Ra activity, with potential to facilitate the design of new IL-36 inhibitors and aid the interpretation of IL36RN variants in diagnostic settings.

B cell profiles, antibody repertoire and reactivity reveal dysregulated responses with autoimmune features in melanoma.

B cells are known to contribute to the anti-tumor immune response, especially in immunogenic tumors such as melanoma, yet humoral immunity has not been characterized in these cancers to detail. Here we show comprehensive phenotyping in samples of circulating and tumor-resident B cells as well as serum antibodies in melanoma patients. Memory B cells are enriched in tumors compared to blood in paired samples and feature distinct antibody repertoires, linked to specific isotypes. Tumor-associated B cells undergo clonal expansion, class switch recombination, somatic hypermutation and receptor revision. Compared with blood, tumor-associated B cells produce antibodies with proportionally higher levels of unproductive sequences and distinct complementarity determining region 3 properties. The observed features are signs of affinity maturation and polyreactivity and suggest an active and aberrant autoimmune-like reaction in the tumor microenvironment. Consistent with this, tumor-derived antibodies are polyreactive and characterized by autoantigen recognition. Serum antibodies show reactivity to antigens attributed to autoimmune diseases and cancer, and their levels are higher in patients with active disease compared to post-resection state. Our findings thus reveal B cell lineage dysregulation with distinct antibody repertoire and specificity, alongside clonally-expanded tumor-infiltrating B cells with autoimmune-like features, shaping the humoral immune response in melanoma.

Cost-Effectiveness Analysis of HPV Extended versus Partial Genotyping for Cervical Cancer Screening in Singapore.

Human papillomavirus (HPV) partial genotyping (PGT) identifies HPV16 and HPV18 individually, alongside 12 other high-risk HPV genotypes (hrHPV) collectively. HPV extended genotyping (XGT) identifies four additional hrHPV individually (HPV31, 45, 51, and 52), and reports the remaining eight in three groups (HPV33|58; 56|59|66; 35|39|68). Quality-adjusted life years (QALY), health care resource use, and costs of XGT were compared to PGT for cervical cancer screening in Singapore using DICE simulation. Women with one of the three hrHPV identified by XGT (HPV35|39|68; 56|59|66; 51), and atypical squamous cells of undetermined significance (ASCUS) on cytology, are recalled for a repeat screening in one year, instead of undergoing an immediate colposcopy with PGT. At the repeat screening, the colposcopy is performed only for persistent same-genotype infections in XGT, while with PGT, all the women with persistent HPV have a colposcopy. Screening 500,122 women, aged 30-69, with XGT, provided an incremental cost-effectiveness ratio (ICER) versus PGT of SGD 16,370/QALY, with 7130 (19.4%) fewer colposcopies, 6027 (7.0%) fewer cytology tests, 9787 (1.6%) fewer clinic consultations, yet 2446 (0.5%) more HPV tests. The XGT ICER remains well below SGD 100,000 in sensitivity analyses, (-SGD 17,736/QALY to SGD 50,474/QALY). XGT is cost-effective compared to PGT, utilizes fewer resources, and provides a risk-based approach as the primary cervical cancer screening method.

COVID-19 Mask Mandates in NY and Their Effect on the Incidence of Flu.

Objective: The goal of the study is to quantify the reduction in the cases of influenza and how this decrease in incidence correlates with the execution of masking requirements in public as well as social distancing. Methods: Influenza statistics were collected from Northwell Health, a 23-hospital system located throughout New York State. Positive influenza results were collected representing the 2018-2019 Flu season, 2019-2020 Flu season, and compared to the 2020-2021 Flu season, which corresponded to the mask mandates and social distancing measures implemented in NYS. Results: Our data showed a dramatic decrease in influenza rates during the 2020-2021 Flu season, which corresponded to NYS's strict social distancing and mask requirements during the pandemic. This shows a steep decline correlating with the implementation of public health mandates directed at decreasing the spread of aerosolized particles between members of the population. Conclusion: Our data show a significant decrease in the number of positive influenza tests during the same period of time when COVID-19 social distancing and mask-wearing requirements were in effect.

Investigations on the Role of Iron (III) and Silica-Iron (III) for DNA Protection Against Highly Intense UV Radiation: Tracking the Connection of Prebiotic Chemistry to Biology.

The mineral reaction pathways that yield organic compounds of increasing complexity would have required a means of protective screening against strong ultraviolet radiation for macromolecular assembly on early Earth. In this study, a bacterial chromosomal plasmid DNA was used as a model biomolecule that represents a complex polymeric nucleic acid containing genetic information. The plasmid DNA was exposed to UV radiation through a medium containing air, water, iron (Fe3+), or silica-iron rich aqueous solutions. Our results demonstrate that the plasmid DNA underwent covalent breakage in an aqueous solution when exposed to UV radiation but was shielded against damage due to the presence of iron and silica. It is demonstrated that a suspension of ca. 40 nm colloidal particles of silica gel embedded with Fe3+ ions adsorbed on silanol groups that formed nanoclusters of noncrystalline iron hydroxide is an extremely efficient shelter against intense UV radiation. The implications for our understanding of primitive Earth and Earth-like planets, moons, and asteroids are discussed. The stability of a chromosomal DNA molecule against UV radiation in the presence of iron and silica may provide support on how macromolecules endured early Earth environments and brought forth important implications on early molecular survival against UV radiation.

Diversification of immunoglobulin genes by gene conversion in the domestic chicken (Gallus gallus domesticus).

Sustainable modern poultry production depends on effective protection against infectious diseases and a diverse range of antibodies is key for an effective immune response. In the domestic chicken, somatic gene conversion is the dominant process in which the antibody immunoglobulin genes are diversified. Affinity maturation by somatic hypermutation (SHM) also occurs, but the relative contribution of gene conversion versus somatic hypermutation to immunoglobulin (Ig) gene diversity is poorly understood. In this study, we use high throughput long-read sequencing to study immunoglobulin diversity in multiple immune-associated tissues in Rhode Island Red chickens. To better understand the impact of genetic diversification in the chicken, a novel gene conversion identification software was developed (BrepConvert). In this study, BrepConvert enabled the identification of over 1 million gene conversion events. Mapping the occurrence of putative somatic gene conversion (SGC) events throughout the variable gene region revealed repetitive and highly restricted patterns of genetic insertions in both the antibody heavy and light chains. These patterns coincided with the locations of genetic variability in available pseudogenes and align with antigen binding sites, predominately the complementary determining regions (CDRs). We found biased usage of pseudogenes during gene conversion, as well as immunoglobulin heavy chain diversity gene (IGHD) preferences during V(D)J gene rearrangement, suggesting that antibody diversification in chickens is more focused than the genetic potential for diversity would suggest.

Chickens, more than humans, focus the diversity of their immunoglobulin genes on the complementarity-determining region but utilise amino acids, indicative of a more cross-reactive antibody repertoire.

The mechanisms of B-cell diversification differ greatly between aves and mammals, but both produce B cells and antibodies capable of supporting an effective immune response. To see how differences in the generation of diversity might affect overall repertoire diversity, we have compared the diversity characteristics of immunoglobulin genes from domestic chickens to those from humans. Both use V(D)J gene rearrangement and somatic hypermutation, but only chickens use somatic gene conversion. A range of diversity analysis tools were used to investigate multiple aspects of amino acid diversity at both the germline and repertoire levels. The effect of differing amino acid usages on antibody characteristics was assessed. At both the germline and repertoire levels, chickens exhibited lower amino acid diversity in comparison to the human immunoglobulin genes, especially outside of the complementarity-determining region (CDR). Chickens were also found to possess much larger and more hydrophilic CDR3s with a higher predicted protein binding potential, suggesting that the antigen-binding site in chicken antibodies is more flexible and more polyreactive than that seen in human antibodies.

A Case Report of Anti-phospholipid Syndrome With Lower Extremity Arterial Thrombosis That Didn't Respond to Heparin and Direct Oral Anticoagulation: Ultimately, the Patient Agreed to Oral Warfarin.

In the absence of known thrombophilia or factors associated with thrombotic tendency, clinicians are more likely to think of antiphospholipid syndrome in patients presenting with venous thrombosis than in those with arterial thrombosis. We present a case of acute lower extremity arterial ischemia in a female smoker. Despite multiple surgical interventions and treatment with several different anticoagulants, our patient developed bilateral lower extremity thrombi. Ultimately, after developing a pulmonary embolism, she accepted to be on warfarin. She switched to warfarin without recurrence of her arterial thrombosis. We describe the challenging management of her critical limb ischemia and review the pertinent literature on the controversy surrounding optimal anticoagulation in such patients.

Health care provider's experience and perspective of cervical cancer screening in Singapore: A qualitative study.

Background: In Singapore, the current cervical cancer screening (CCS) coverage rate of 48% falls below the national target of 70%. Health care providers (HCPs) play a critical role in promoting CCS uptake. However, there is limited understanding of the perspectives of HCPs regarding CCS. Hence, we aimed to understand the challenges encountered by HCPs delivering CCS in different care settings in the Singapore health system. We also aimed to explore perspectives on newer features of CCS such as self-sampling and HPV genotyping. Methods: Physicians, nurses, program administrators and laboratory technicians involved with CCS were invited for a one-on-one semi-structured interview conducted over Zoom between May to August 2021. The interviews were transcribed and analyzed using thematic analysis. Results: Eighteen HCPs from 12 institutions were interviewed. Most participants were women (61.1%) and worked in public health institutions (72.2%). For factors influencing CCS, nine key themes were identified and organized into four categories: (1) patient factors, (2) HCP factors, (3) health system factors and (4) health promotion factors. Key themes commonly highlighted by study participants were related to patients' preferences and acceptance for screening, the processes of delivering CCS, the national priority for cervical cancer and the effectiveness of existing health promotion efforts. Five key themes were identified for CCS innovations. Self-sampling was viewed favorably to increase CCS uptake, while primary HPV screening with HPV partial genotyping had higher sensitivities to detect pre-cancers and cancers compared to cytology. Extended HPV genotyping beyond HPV16/18 could play an important role in CCS with increasing HPV vaccination coverage, as well as in the management of persistent HPV infection. Conclusion: In Singapore, HCPs face multiple challenges for CCS in practice. Insights from this study are directly relevant to, and useful for developing policies around national CCS programs and treatment guidelines.