Human IgG Research Articles

FcRider: a recombinant Fc nanoparticle with endogenous adjuvant activities for hybrid immunization.

Active immunization (vaccination) induces long-lasting immunity with memory, which takes weeks to months to develop. Passive immunization (transfer of neutralizing antibodies) provides immediate protection, yet with high cost and effects being comparatively short-lived. No currently approved adjuvants are compatible with formulations to combine active and passive immunizations, not to mention their huge disparities in administration routes and dosage. To solve this, we engineered the Fc fragment of human IgG1 into a hexamer nanoparticle and expressed its afucosylated form in Fut8-/- CHO cells, naming it "FcRider." FcRider is highly soluble with long-term stability, easily produced at high levels equivalent to those of therapeutic antibodies, and is amenable to conventional antibody purification schemes. Most importantly, FcRider possesses endogenous adjuvant activities. Using SWHEL B cell receptor (BCR) transgenic mice, we found that HEL-FcRider induced GL7+ germinal center B cells and HEL-specific IgG. Similarly, immunizing mice with UFO-BG-FcRider, a fusion containing the stabilized human immunodeficiency virus-1 (HIV-1) Env protein as immunogen, promoted somatic hypermutation and generation of long CDR3 of the IgG heavy chains. Intramuscular injection of (Fba+Met6)3-FcRider, a fusion with two peptide epitopes from Candida albicans cell surface, stimulated strong antigen-specific IgG titers. In three different models, we showed that afucosylated FcRider functions as a multivalent immunogen displayer and stimulates antigen-specific B cells without any exogenous adjuvant. As an antibody derivative, afucosylated FcRider could be a novel platform combining vaccines and therapeutic antibodies, integrating active and passive immunizations into single-modality "hybrid immunization" to provide complete and long-lasting protection against infections, and may open new avenues in cancer immunotherapy as well.

A genetically engineered mouse/human chimeric antibody targeting CD99 enhances antibody-dependent cellular phagocytosis against human mantle cell lymphoma Z138 cells

Background: Mantle cell lymphoma (MCL) is an aggressive form of B-cell nonHodgkin lymphoma. The elimination of MCL cells via phagocytosis is essential for cancer eradication. Therefore, discovering novel targeted antibodies that can induce phagocytosis is needed. We have demonstrated that our in-houseproduced mouse anti-CD99 mAb clone MT99/3 could induce potent anticancer activities against MCL cell lines in both in vitro and in vivo mouse xenograft models. Nevertheless, for use in humans, the mouse mAb needs to be transformed into a mouse/human chimeric mAb that contains a human Fc region to activate human immune effector functions, especially macrophage-mediated phagocytosis. Antibody-dependent cellular phagocytosis (ADCP) mediated by mouse/human chimeric mAb MT99/3 against MCL has not been previously reported. Objective: This study aimed to genetically engineer a mouse/human chimeric antibody against human CD99 derived from mouse mAb MT99/3 and to evaluate its effect in mediating the ADCP mechanism for eradicating MCL cells in vitro using monocyte-derived macrophages. Materials and methods: The expression plasmid to produce chimeric anti-CD99 antibody, ChAbMT99/3, was constructed by fusing the variable domains of mouse mAb MT99/3 with the constant domains of human IgG1 and the constant domains of kappa light chain. ChAbMT99/3 was expressed in the stable human expression system based on HEK293T cells. ChAbMT99/3 was purified from the culture supernatant of ChAbMT99/3-expressing HEK293T cells using Protein G chromatography. The purity and structure of ChAbMT99/3 were verified by SDS-PAGE and western blotting. The binding specificity and activity were determined by staining with cells expressing recombinant and native human CD99. The anticancer activity of ChAbMT99/3 in mediating the ADCP mechanism against MCL cell line Z138 using human monocytederived macrophages was evaluated. Results: We successfully constructed the plasmid to produce ChAbMT99/3. Human HEK293T cells stably expressing ChAbMT99/3 were established. The ChAbMT99/3- expressing HEK293T cells could secrete ChAbMT99/3 into the culture supernatant. The high purity and complete IgG structure of ChAbMT99/3 were obtained from the purification process. Crucially, this chimeric antibody retained its binding reactivity to recombinant and native human CD99. In addition, the produced ChAbMT99/3, upon binding to MCL cells, significantly enhanced ADCP against MCL cell line Z138 in a dose-dependent manner. Conclusion: The production of a mouse/human chimeric antibody against human CD99 derived from mouse mAb MT99/3 was successful. The engineered antibody could mediate ADCP activity against MCL cells. The produced ChAbMT99/3 might be a promising therapeutic candidate for MCL treatment.

Advances in Understanding and Managing Alzheimer's Disease: From Pathophysiology to Innovative Therapeutic Strategies.

Alzheimer's disease (AD) is a debilitating neurodegenerative disorder characterized by the presence of amyloid-β (Aβ) plaques and tau-containing neurofibrillary tangles, leading to cognitive and physical decline. Representing the majority of dementia cases, AD poses a significant burden on healthcare systems globally, with onset typically occurring after the age of 65. While most cases are sporadic, about 10% exhibit autosomal forms associated with specific gene mutations. Neurofibrillary tangles and Aβ plaques formed by misfolded tau proteins and Aβ peptides contribute to neuronal damage and cognitive impairment. Currently, approved drugs, such as acetylcholinesterase inhibitors and N-methyl D-aspartate receptor agonists, offer only partial symptomatic relief without altering disease progression. A promising development is using lecanemab, a humanized IgG1 monoclonal antibody, as an immune therapeutic approach. Lecanemab demonstrates selectivity for polymorphic Aβ variants and binds to large soluble Aβ aggregates, providing a potential avenue for targeted treatment. This shift in understanding the role of the adaptive immune response in AD pathogenesis opens new possibilities for therapeutic interventions aiming to address the disease's intricate mechanisms. This review aims to summarize recent advancements in understanding Alzheimer's disease pathophysiology and innovative therapeutic approaches, providing valuable insights for both researchers and clinicians.

A Randomized Placebo-Controlled Trial of Leronlimab in Mild-To-Moderate COVID-19

PurposeEarly in the course of the SARS-CoV-2 pandemic it was hypothesised that host genetics played a role in the pathophysiology of COVID-19 including a suggestion that the CCR5-Δ32 mutation may be protective in SARS-CoV-2 infection. Leronlimab is an investigational CCR5-specific humanized IgG4 monoclonal antibody currently in development for HIV-1 infection. We aimed to explore the impact of leronlimab on the severity of disease symptoms among participants with mild-to-moderate COVID-19. MethodsThe TEMPEST trial was a randomized, double-blind, placebo-controlled study in participants with mild-to-moderate COVID-19. Participants were randomly assigned in a 2:1 ratio to receive subcutaneous leronlimab (700 mg) or placebo on days 0 and 7. The primary efficacy endpoint was assessed by change in total symptom score based on fever, myalgia, dyspnea, and cough, at end of treatment (day 14). FindingsOverall, 84 participants were randomized and treated with leronlimab (n = 56) or placebo (n = 28). No difference was observed in change in total symptom score (P = 0.8184) or other pre-specified secondary endpoints between treatments. However, in a post hoc analysis, 50.0% of participants treated with leronlimab demonstrated improvements from baseline in National Early Warning Score 2 (NEWS2) at day 14, compared with 20·8% of participants in the placebo group (post hoc; p = 0.0223). Among participants in this trial with mild-to-moderate COVID-19 adverse events rates were numerically but not statistically significantly lower in leronlimab participants (33.9%) compared with placebo participants (50.0%). ImplicationsAt the time the TEMPEST trial was designed although CCR5 was known to be implicated in COVID-19 disease severity the exact pathophysiology of SARS-CoV-2 infection was poorly understood. Today it is well accepted that SARS-CoV-2 infection in asymptomatic-to-mild cases is primarily characterized by viral replication, with a heightened immune response, accompanied by diminished viral replication in moderate-to-severe disease and a peak in inflammatory responses with excessive production of pro-inflammatory cytokines in critical disease. It is therefore perhaps not surprising that no differences between treatments were observed in the primary endpoint or in pre-specified secondary endpoints among participants with mild-to-moderate COVID-19. However, the results of the exploratory post hoc analysis showing that participants in the leronlimab group had greater improvement in NEWS2 assessment compared to placebo provided a suggestion that leronlimab may be associated with a lower likelihood of people with mild-to-moderate COVID-19 progressing to more severe disease and needs to be confirmed in other appropriately designed clinical trials.ClinicalTrials.gov number, NCT04343651 https://classic.clinicaltrials.gov/ct2/show/NCT04343651

A study of multiple solid-state dewetting of sputtered Au ultra-thin films for chip-based LSPR sensor applications

This article proposes a lithography-free technique to fabricate Au NPs on glass substrates using multiple solid-state dewetting (SSD) of sputtered Au ultra-thin films for LSPR sensor chip applications. We studied the influence of initial film thickness and the number of repeated process cycles on the morphology and LSPR sensing performance. This fabrication process allowed control over particle size, gap spacing, and density of the Au NPs, which influenced the LSPR peak position as observed using field emission scanning electron microscopy (FE-SEM) and UV–Vis–NIR spectrophotometry. To demonstrate LSPR sensing performance, the refractive index (RI) sensitivity was evaluated by measuring the wavelength shift of the LSPR peak in a series of glycerol/phosphate-buffered saline (PBS) mixtures, varying the refractive index from 1.33909 to 1.37409. The results showed that RI sensitivity and the figure of merit (FOM) for all prepared samples ranged from 37.191 ± 12.26–73.592 ± 9.70 and 0.35 ± 0.12–0.75 ± 0.02, respectively. An increase in repeated process cycles tended to decrease RI sensitivity and FOM. The best LSPR performance was achieved with an 8 nm initial film thickness after the first cycle, with an RI sensitivity of 70.937 ± 2.60 and an FOM of 0.75 ± 0.02, attributed to optimal Au size and density. Additionally, the binding efficiency response to human IgG with high regeneration cycles was demonstrated, highlighting the potential for biosensor applications.

Automated Assignment of 15N And 13C Enrichment Levels in Doubly-Labeled Proteins.

Uniform enrichment of 15N and 13C in proteins is commonly employed for 2D heteronuclear NMR measurements of the three-dimensional protein structure. Achieving a high degree of enrichment of both elements is important for obtaining high quality data. Uniform labeling of proteins and glycoproteins expressed in higher organisms (yeast or mammalian cell lines) is more challenging than expression in Escherichia coli, a prokaryote that grows on simple, chemically defined media but does not provide appropriate eukaryotic modifications. It is difficult to achieve complete incorporation of both heavy isotopes, and quality control measures are important for quantitating the level of their enrichment. Mass spectrometry measurements of the isotopic distribution of the intact protein or its proteolytic fragments provide the means to assess the enrichment level. A mass accuracy of 1 ppm or better is shown to be required to distinguish the correct combination of 13C and 15N enrichment due to subtle shifts in peak centroids with differences in the underlying, but unresolved, isotopic fine structure. A simple computer program was developed to optimize the fitting of experimental isotope patterns to statistically derived distributions. This method can determine the isotopic abundance from isotope patterns and isotopologue masses in conventional MS data for peptides, intact proteins, and glycans. For this purpose, MATLAB's isotope simulator, isotopicdist, has been modified to permit the variation of 15N and 13C enrichment levels and to perform a two-dimensional grid search of enrichment levels of both isotopes. We also incorporated an alternate isotope simulator, js-emass, into MATLAB for use in the same fitting program. Herein we benchmark this technique on natural abundance ubiquitin and uniformly [15N,13C]-labeled ubiquitin at both the intact and peptide level, outline considerations for data quality and mass accuracy, and report several improvements we have made to the previously reported analysis of the [15N,13C]-enriched human IgG Fc domain, a glycoprotein that has been expressed in Saccharomyces cerevisiae.

The KEYVIBE program: vibostolimab and pembrolizumab for the treatment of advanced malignancies.

Vibostolimab, a humanized IgG1 monoclonal antibody, blocks the interaction between TIGITand its ligands, preventing the immunosuppressive effects of TIGIT. The addition of vibostolimab to the PD-1 inhibitor pembrolizumab has shown promising antitumor activity, warranting further exploration of vibostolimab as a potential therapeutic option. The KEYVIBE program consists of nine trials that will evaluate the safety and efficacy of vibostolimab monotherapy and vibostolimab-based combination therapy in advanced solid tumors and hematological malignancies. These studies will also evaluate coformulated immunotherapy addressing issues that occur with the sequential administration of immunotherapy. The KEYVIBE program will provide further insight into the clinical utility of vibostolimab-based therapy across multiple indications and various stages of disease.

Broad-Spectrum Legionaminic Acid-Specific Antibodies in Pooled Human IgGs Revealed by Glycan Microarrays with Chemoenzymatically Synthesized Nonulosonosides.

The presence and the level of antibodies in human sera against bacterial glycans are indications of prior encounters with similar antigens and/or the bacteria that express them by the immune system. An increasing number of pathogenic bacteria that cause human diseases have been shown to express polysaccharides containing a bacterial nonulosonic acid called 5,7-di-N-acetyllegionaminic acid (Leg5,7Ac2). To investigate the immune recognition of Leg5,7Ac2, which is critical for the fight against bacterial infections, a highly effective chemoenzymatic synthon strategy was applied to construct a library of α2-3/6-linked Leg5,7Ac2-glycans via their diazido-derivatives (Leg5,7diN3-glycans) formed by efficient one-pot three-enzyme (OP3E) synthetic systems from a diazido-derivative of a six-carbon monosaccharide precursor. Glycan microarray studies using this synthetic library of a Leg5,7Ac2-capped collection of diverse underlying glycan carriers and their matched sialoside counterparts revealed specific recognition of Leg5,7Ac2 by human IgG antibodies pooled from thousands of healthy donors (IVIG), suggesting prior human encounters with Leg5,7Ac2-expressing pathogenic bacteria at the population level. These biologically relevant Leg5,7Ac2-glycans and their immune recognition assays are important tools to begin elucidating their biological roles, particularly in the context of infection and host-pathogen interactions.

Digital Ion Trap Isolation and Mass Analysis of Macromolecular Analytes with Multiply Charged Ion Attachment.

Multiply charged ions produced by electrospray ionization (ESI) of heterogeneous mixtures of macromolecular analytes under native conditions are typically confined to relatively narrow ranges of mass-to-charge (m/z) ratio, often with extensive overlap. This scenario makes charge and mass assignments extremely challenging, particularly when individual charge states are unresolved. An ion/ion reaction strategy involving multiply charged ion attachment (MIA) to the mixture components in a narrow range of m/z can facilitate charge and mass assignment. In MIA operation, multiply charged reagent ions are attached to the analyte ions of opposite polarity to provide large m/z displacements resulting from both large changes in mass and charge. However, charge reduction of the high m/z ions initially generated under native ESI conditions requires the ability to isolate high m/z ions and to analyze even higher m/z product ions. Digital ion trap (DIT) operation offers means for both high m/z ion isolation and high m/z mass analysis, in addition to providing conditions for the reaction of oppositely charged ions. The feasibility of conducting MIA experiments in a DIT that takes advantage of high m/z ion operation is demonstrated here using a tandem 2D-3D DIT instrument. Proof-of-concept MIA experiments with cations derived from β-galactosidase using the 20- charge state of human serum immunoglobulin G (IgG, ∼149 kDa) as the reagent anion are described. MIA experiments involving mixtures of ions derived from the E. coli. ribosome are also described. For example, three components in a mixture of 70S particles (>2.2 MDa) were resolved and assigned with masses and charges following an MIA experiment involving the 20- charge state of human serum IgG.

IL1RAP-specific T cell engager depletes acute myeloid leukemia stem cells

BackgroundThe interleukin-1 receptor accessory protein (IL1RAP) is highly expressed on acute myeloid leukemia (AML) bulk blasts and leukemic stem cells (LSCs), but not on normal hematopoietic stem cells (HSCs), providing an opportunity to target and eliminate the disease, while sparing normal hematopoiesis. Herein, we report the activity of BIF002, a novel anti-IL1RAP/CD3 T cell engager (TCE) in AML.MethodsAntibodies to IL1RAP were isolated from CD138+ B cells collected from the immunized mice by optoelectric positioning and single cell sequencing. Individual mouse monoclonal antibodies (mAbs) were produced and characterized, from which we generated BIF002, an anti-human IL1RAP/CD3 TCE using Fab arm exchange. Mutations in human IgG1 Fc were introduced to reduce FcγR binding. The antileukemic activity of BIF002 was characterized in vitro and in vivo using multiple cell lines and patient derived AML samples.ResultsIL1RAP was found to be highly expressed on most human AML cell lines and primary blasts, including CD34+ LSC-enriched subpopulation from patients with both de novo and relapsed/refractory (R/R) leukemia, but not on normal HSCs. In co-culture of T cells from healthy donors and IL1RAPhigh AML cell lines and primary blasts, BIF002 induced dose- and effector-to-target (E:T) ratio-dependent T cell activation and leukemic cell lysis at subnanomolar concentrations. BIF002 administered intravenously along with human T cells led to depletion of leukemic cells, and significantly prolonged survival of IL1RAPhigh MOLM13 or AML patient-derived xenografts with no off-target side effects, compared to controls. Of note, BiF002 effectively redirects T cells to eliminate LSCs, as evidenced by the absence of disease initiation in secondary recipients of bone marrow (BM) from BIF002+T cells-treated donors (median survival not reached; all survived > 200 days) compared with recipients of BM from vehicle- (median survival: 26 days; p = 0.0004) or isotype control antibody+T cells-treated donors (26 days; p = 0.0002).ConclusionsThe novel anti-IL1RAP/CD3 TCE, BIF002, eradicates LSCs and significantly prolongs survival of AML xenografts, representing a promising, novel treatment for AML.

Triple Combinations of AAV9-Vectors Encoding Anti-HIV bNAbs Provide Long-Term In Vivo Expression of Human IgG Effectively Neutralizing Pseudoviruses from HIV-1 Global Panel.

Anti-human immunodeficiency virus (HIV) broadly neutralizing antibodies (bNAbs) offer a promising approach for the treatment of HIV-1. The current paradigm for antibody therapy involves passive antibody transfer, requiring regular delivery of bNAbs in treating chronic diseases such as HIV-1. An alternative strategy is to use AAV-mediated gene transfer to enable in vivo production of desirable anti-HIV-1 antibodies. In this study, we investigated two sets of triple combinations of AAV9-vectors encoding different bNAbs: N6, 10E8, 10-1074 (CombiMab1), and VRC07-523, PGDM1400, 10-1074 (CombiMab2). We used CBAxC57Bl and C57BL/6 mouse models to characterize rAAV-induced antibody expression and to evaluate the neutralization capacity of mouse sera against a global panel of HIV-1 viral strains. rAAV9-mediated IgG expression varied between bNAb clones and mouse strains, with C57BL/6 mice exhibiting higher bNAb titers following rAAV delivery. Although CombiMab2 treatment elicited a higher IgG titer than CombiMab1, both combinations resulted in neutralization of all the viral strains from the global HIV-1 panel. Our data highlight the potential of AAV vectors as a long-term option for HIV-1 therapy.

Efficacy and safety of bintrafusp alfa in 2 phase I expansion cohorts with advanced HCC.

Simultaneous inhibition of the TGF-β and programmed cell death 1 ligand 1 pathways provides a potential novel treatment approach. Bintrafusp alfa, a first-in-class bifunctional fusion protein composed of the extracellular domain of TGF-βRII (a TGF-β "trap") fused to a human IgG1 monoclonal antibody blocking programmed cell death 1 ligand 1, was evaluated in patients with advanced HCC. In this global, open-label, phase I study (NCT02517398), patients with programmed cell death 1 ligand 1-unselected HCC who failed or were intolerant to ≥1 line of sorafenib received bintrafusp alfa 1200mg every 2 weeks in a dose-escalation (n = 38) or dose-expansion (n = 68) cohort until confirmed progression, unacceptable toxicity, or trial withdrawal. The primary endpoint was the best overall response per Response Evaluation Criteria in Solid Tumors version 1.1 by an independent review committee. Secondary endpoints included investigator-assessed best overall response, safety, and pharmacokinetics. Median follow-up times (range) were 41.4 (39.8-44.2) and 38.6 (33.5-39.7) months in the dose-escalation and dose-expansion cohorts, respectively. The objective response rate was below the prespecified 20% objective response rate threshold set to evaluate the efficacy of bintrafusp alfa in both cohorts (10.5% and 8.8%, respectively). Median overall survival and progression-free survival, respectively, were 13.8 and 1.5 months in the dose-escalation cohort and 13.5 and 1.4 months in the dose-expansion cohort. Treatment-related adverse events occurred in 78.9% and 64.7% of patients in the respective cohorts (grade ≥3 in 18.4% and 25.0% of patients). Bintrafusp alfa showed moderate clinical activity and a safety profile consistent with previous studies of bintrafusp alfa in patients with advanced HCC.

Updated efficacy of anti-TROP2 ADC ESG401 for first-line metastatic TNBC in phase 1b study.

2 Background: ESG401, a novel ADC comprising a humanized anti-TROP2 IgG1 monoclonal antibody conjugated to SN-38, with a Drug-Antibody Ratio of 8 and a stable cleavable linker, demonstrated preliminary efficacy and tolerability in the Phase Ia trial (Ma, ASCO 2023). This report presents updated results from the first-line mTNBC cohort of the Phase Ib trial and results from patients with brain metastases. Methods: Patients (pts) aged ≥18 years with confirmed local advanced/unresectable or metastatic TNBC, without prior metastatic treatment, received ESG401 (16 mg/kg IV on Day 1,8, and 15, with a 28-day cycle) until unacceptable toxicity, progressive disease, or consent withdrawal. Results: As of the cutoff date (Apr 7th, 2024), 24 pts in the cohort received ≥1 dose of ESG401. Median age was 53 years (range: 33-73), 33.3% were ECOG 0, 25.0% were de novo stage IV, 70.8% had visceral disease (8.3% brain, 33.3% liver, 50.0% lung). Updated efficacy results are shown in the Table. Thirteen pts (68.4% of 19 efficacy evaluable pts) remained on treatment, with the longest on-treatment duration being 10.3 mons. Median PFS has not been reached. Two pts in this cohort had brain metastases, with one achieving a complete intracranial response and an overall response of PR, and the other demonstrating overall PR with a -13.8% reduction in brain lesion size. Totally 16 pts with brain metastases were enrolled in the entire Phase Ia/1b trials, for these pts, the best overall intracranial response rate was 31%, with an intracranial disease control rate of 75%, while the corresponding overall response rate and disease control rate were 50% and 69%, respectively. The response of intracranial lesions to the treatment is consistent with that of extracranial lesions. The safety profile of ESG401 remained consistent with previous reports, showing no new or unexpected safety signals. Conclusions: ESG401 monotherapy demonstrates promising antitumor activity in 1L mTNBC pts, characterized by a high response rate and durable response. The ORR numerically exceeded that reported in the BEGONIA Study arm 7 in which pts with the same indication were treated with TROP-2 ADC and PD-L1 combination therapy. Additionally, ESG401 shows clear potential for treating HER2-negative breast cancer with brain metastases, including mTNBC and HR+/HER2- BC. These results support further clinical investigation of ESG401. Clinical trial information: NCT04892342 . [Table: see text]

Efficacy and safety of surufatinib plus toripalimab in previously treated advanced tumors progressing after immunotherapy.

36 Background: Immunotherapy has resulted in significant and lasting clinical responses in non-small lung cancer (NSCLC), esophagus cancer, gastric cancer, colorectal cancer etc. However, most patients (pts) inevitably lose efficacy due to acquired resistance. This study (NCT04169672) aimed to investigate the efficacy and safety of surufatinib (a multi-kinase inhibitor of VEGFR1-3, FGFR1 and CSF1R) plus toripalimab (monoclonal humanized IgG4 programmed death 1 [PD-1] antibody) in pts with advanced solid tumors. Here, we reported the results from pts who bore different tumors and failed in previous treatments containing the checkpoint inhibitors. Methods: Eligible pts were 18~75 years old who had advanced solid tumors and received prior PD-1/programmed death-ligand 1 (PD-L1) antibody therapy for at least 3 months (m) before disease progression. Enrolled pts received surufatinib 250 mg orally once daily and toripalimab 240 mg intravenously every three weeks in a 21-day cycle. The primary endpoint was the objective response rate (ORR) per RECIST v1.1. Secondary endpoints include disease control rate (DCR), progression free survival (PFS), overall survival (OS) and safety. Results: As of data cutoff (Feb 28, 2023), a total of 28 pts were enrolled and received at least one dose of study treatment. Pts with PD-L1 combined positive score (CPS) ≥1 accounted for 67.9%. 9 pts were diagnosed with gastric cancer /gastroesophageal junction adenocarcinoma (GC/GEJ), 4 pts each with esophageal cancer, NSCLC or intestinal cancer, 2 pts with biliary tract cancer, 1 pt each with other 5 tumors. 39.3% of pts received ≥2 previous anti-tumor therapies. A variety of PD-1 antibodies were used before enrollment, which included tislelizumab (25.0%), sintilimab (17.9%), camrelizumab (14.3%), toripalimab (14.3%), pembrolizumab (7.1%), nivolumab (3.6%), etc. The median exposure duration was 4.1m for study treatment. In 27 tumor evaluable pts, the confirmed ORR and DCR were 7.4 % and 66.7%, respectively. In 28 intent-to-treat pts, the median PFS and OS were 3.94m (95% CI 1.38, 4.21) and 13.04m (95% CI 8.64, 21.59), respectively. The median survival follow-up duration was 20.90m (95% CI 16.10, 25.03). In 9 pts with GC/GEJ, the median PFS and median OS were 2.72m and 13.69m, respectively. Treatment related adverse events (TRAE) occurred in 89.3% of pts. Grade≥3 TRAEs occurred in 39.3% of pts with blood pressure increased (10.7%) and liver injury (7.1%) reported most commonly. 28.6% of pts had immune-related adverse events, among which immune-related pneumonitis (7.1%) and immune-related diabetes (7.1%) reached G3. Two TRAEs led to surufatinib discontinuation. No deaths related to either surufatinib or toripalimab. Conclusions: Surufatinib plus toripalimab showed antitumor activity and a manageable safety profile in pts with advanced solid tumors progressing after immunotherapy, especially in GC/GEJ.

Lack of alpha CGRP exacerbates the development of atherosclerosis in ApoE-knockout mice

The effects of calcitonin gene-related peptide (CGRP) on atherosclerosis remain unclear. We used apolipoprotein E-deficient (ApoE−/−) mice to generate double-knockout ApoE−/−:CGRP−/− mice lacking alpha CGRP. ApoE−/−:CGRP−/− mice exhibited larger atherosclerotic plaque areas, peritoneal macrophages with enhanced migration functions, and elevated levels of the inflammatory cytokine tumor necrosis factor (TNF)-⍺. Thus, we also explored whether inhibiting TNF-⍺ could improve atherosclerosis in ApoE−/−:CGRP−/− mice by administering etanercept intraperitoneally once a week (5 mg/kg) alongside a high-fat diet for 2 weeks. This treatment led to significant reductions in aortic root lesion size, atherosclerotic plaque area and macrophage migration in ApoE−/−:CGRP−/− mice compared with mice treated with human IgG (5 mg/kg). We further examined whether results observed in ApoE−/−:CGRP−/− mice could similarly be obtained by administering a humanized monoclonal CGRP antibody, galcanezumab, to ApoE−/− mice. ApoE−/− mice were subcutaneously administered galcanezumab at an initial dose of 50 mg/kg, followed by a dose of 30 mg/kg in the second week. Galcanezumab administration did not affect systolic blood pressure, serum lipid levels, or macrophage migration but led to a significant increase in lipid deposition at the aortic root. These findings suggest that alpha CGRP plays a critical role in inhibiting the progression of atherosclerosis.

A case-control study of the association of toxoplasmosis with antirheumatic therapy in rheumatoid arthritis

Background & Objective: Toxoplasmosis is a globally prevalent opportunistic zoonotic infection. Rheumatoid arthritis (RA) is an autoimmune disease which is manifested as joint inflammation, pain, stiffness, and swelling. The patients may contract toxoplasmosis, especially when receiving antirheumatic immunosuppressive therapy. However conflicting results have been reported on toxoplasmosis in RA patients. We aimed to evaluate Toxoplasma gondii positivity and associated risk factors in patients with RA in Duhok province, Iraq, and to assess the role of antirheumatic therapy in toxoplasmosis and the possibility of its reactivation. Methodology: This case-control study was carried out in Duhok, Kurdistan Region, Iraq, from February 2022 to June 2023. A total of 88 RA patients, with and without concurrent antirheumatic therapy, were recruited. The diagnosis of RA was according to the 2010 American College of Rheumatology/European League. RA-patients were categorized into three groups: Group 1 - no-therapy (n = 14); Group 2 - conventional therapy (n = 49) and Group 3 - biologics in combination with conventional medications (n = 25). Healthy subjects (n = 61) were also selected as controls. Plasma samples obtained from the participants were screened for T. gondii infection using Human anti-Toxoplasma gondii antibody IgG. IgG positive samples were tested utilizing human anti-Toxoplasma gondii antibody IgM ELISA Kit and the avidity T. gondii IgG ELISA kit was used to determine the state of infection. Results: Of the 88 RA patients, with or without antirheumatic therapy, 76 (86.36%) were females and 12 (13.64%) were males. Patients on therapy (n = 74) received several conventional antirheumatic drugs of which methotrexate was the most frequently used one (86.36%). Among biological therapies, etanercept, a TNF-alpha inhibitor, was mostly used (13.51%). The toxoplasmosis was significantly less in the older age group (> 50 y) compared to individuals under 30 y. Gender had no significant effect. Toxoplasma IgG positivity was found to be significantly higher in the three groups of RA patients in comparison with healthy controls. Further assessment of anti-Toxoplasma IgM positivity and Toxoplasma IgG avidity results to identify the state of T. gondii infection in anti-Toxoplasma IgG positive subjects revealed that the frequency of reactivation of toxoplasmosis in RA patient groups was not significantly different from that of the control counterparts. Conclusion: The study showed higher seroprevalence of anti-Toxoplasma antibodies in patients with RA irrespective of therapy; therefore, we could consider toxoplasmosis interaction with RA in the pathogenesis of both diseases. This seroprevalence was higher in younger age groups, but with no significant role of gender, duration of disease, duration of therapy and its type. No significant difference was noted in rates of reactivation of latent T. gondii infection among RA patients and healthy controls. Abbreviations: CI - Confidence interval; O - Odds ratio; RA - Rheumatoid arthritis; SD - Standard deviation; T. gondii - Toxoplasma gondii; TNF - Tumor necrosis factor. Keywords: Autoimmune disease; Rheumatoid arthritis; Antirheumatic drugs; Toxoplasma gondii; Toxoplasma seroprevalence; Toxoplasma IgG avidity. Citation: Abdullah RG, Eassa SH, Mohammad FK. A case-control study of the association of toxoplasmosis with antirheumatic therapy in rheumatoid arthritis. Anaesth, pain intensive care 2024;28(4):674−680; DOI: 10.35975/apic.v28i4.2510 Received: May 14, 2024; Reviewed: June 28, 2024; Accepted: June 28, 2024

Dynamics of scFv-targeted VAP2 correlating with IL-16, MIF and IL-1Ra in ANCA-associated vasculitis

Background and hypothesisUsing a mouse model of MPA with microvascular lesion with a clone (VasSF) of recombinant single chain fragments of the variable region of human IgG, we previously showed that vasculitis-associated apolipoprotein A2 (VAP2) may be a therapeutic target for vasculitis. The present study estimated the target molecules for VasSF and the association between VAP2 and cytokine levels in patient sera in terms of microvascular lesion severity. MethodsSera and clinical information were collected from patients with microscopic polyangiitis and granulomatosis with polyangiitis (MPA/GPA) and infectious disease. Neutrophil counts, levels of C-reactive protein (CRP), creatinine, total cholesterol associated with microvascular lesion, HDL cholesterol, low-density lipoprotein cholesterol, triglycerides, glomerular filtration rate (eGFR), and cytokines were estimated. Serum VAP2 signals were determined with Western blotting. ResultsVasSF bound to a 24 kDa molecule in the serum of active MPA/GPA patients. Anti-AP2 antibody also bound with the same 24 kDa molecule, named VAP2, because of size difference from normal APOA2. The VAP2 signal was significantly stronger in the active-disease group but significantly weakened in remission. The signal correlated positively with eGFR but not with the Birmingham Vasculitis Activity Score, CRP, MPO-ANCA, or PR3-ANCA levels. It correlated negatively with MPO activity, IL-16, MIF, and IL-1Ra. Moreover, VasSF bound to a 17 kDa molecule in the remission phase. ConclusionThe 24 kDa VAP2 molecule may be associated with neutrophil functions because of its inverse correlation with MPO activity, IL-16, MIF, and IL-1Ra, suggesting that VAP2-APOA1 formation in HDL triggers microvascular injury. VasSF may reverse the injury by removing APOA1-VAP2 heterodimers from peripheral blood vessels.

Preclinical Efficacy of VTX-0811: A Humanized First-in-Class PSGL-1 mAb Targeting TAMs to Suppress Tumor Growth.

Omnipresent suppressive myeloid populations in the tumor microenvironment limit the efficacy of T-cell-directed immunotherapies, become more inhibitory after administration of T-cell checkpoint inhibitors, and are overall associated with worse survival of cancer patients. In early clinical trials, positive outcomes have been demonstrated for therapies aimed at repolarizing suppressive myeloid populations in the tumor microenvironment. We have previously described the key role of P-selectin glycoprotein ligand-1 (PSGL-1) in maintaining an inhibitory state of tumor-associated macrophages (TAMs), most of which express high levels of PSGL-1. Here we describe a novel, first-in-class humanized high-affinity monoclonal antibody VTX-0811 that repolarizes human macrophages from an M2-suppressive phenotype towards an M1 inflammatory phenotype, similar to siRNA-mediated knockdown of PSGL-1. VTX-0811 binds to PSGL-1 of human and cynomolgus macaque origins without inhibiting PSGL-1 interaction with P- and L-Selectins or VISTA. In multi-cellular assays and in patient-derived human tumor cultures, VTX-0811 leads to the induction of pro-inflammatory mediators. RNAseq data from VTX-0811 treated ex vivo tumor cultures and M2c macrophages show similar pathways being modulated, indicating that the mechanism of action translates from isolated macrophages to tumors. A chimeric version of VTX-0811, consisting of the parental murine antibody in a human IgG4 backbone, inhibits tumor growth in a humanized mouse model of cancer. VTX-0811 is exceptionally well tolerated in NHP toxicology assessment and is heading into clinical evaluation after successful IND clearance.

Multivalent CXCR4-targeting nanobody formats differently affect affinity, receptor clustering, and antagonism

The chemokine receptor CXCR4 is involved in the development and migration of stem and immune cells but is also implicated in tumor progression and metastasis for a variety of cancers. Antagonizing ligand (CXCL12)-induced CXCR4 signaling is, therefore, of therapeutic interest. Currently, there are two small-molecule CXCR4 antagonists on the market for the mobilization of hematopoietic stem cells. Other molecules with improved potencies and safety profiles are being developed for different indications, including cancer. Moreover, multiple antagonistic nanobodies targeting CXCR4 displayed similar or better potencies as compared to the CXCR4-targeting molecule AMD3100 (Plerixafor), which was further enhanced through avid binding of bivalent derivatives. In this study, we aimed to compare the affinities of various multivalent nanobody formats which might be differently impacted by avidity. By fusion to a flexible GS-linker, Fc-region of human IgG1, different C4bp/CLR multimerization domains, or via site-directed conjugation to a trivalent linker scaffold, we generated different types of multivalent nanobodies with varying valencies ranging from bivalent to decavalent. Of these, C-terminal fusion, especially to human Fc, was most advantageous with a 2-log-fold and 3-log-fold increased potency in inhibiting CXCL12-mediated Gαi- or β-arrestin recruitment, respectively. Overall, we describe strategies for generating multivalent and high-potency CXCR4 antagonistic nanobodies able to induce receptor clustering and conclude that fusion to an Fc-tail results in the highest avidity effect irrespective of the hinge linker.

Pembrolizumab related perforated appendicitis.

Pembrolizumab is a humanized monoclonal antibody IgG4 programmed cell death protein 1 antagonist, and its use in oncology has been increasing in recent years, providing durable and favorable responses and tolerable toxicity profiles in various types of cancer. We describe a case of pembrolizumab related perforated appendicitis in a patient with stage 3C malignant melanoma (MM). A 70-year-old male patient who had no known disease was diagnosed with MM as a result of the excision of the mass on his right shoulder. The disease stage was stage 3C (pT4aN1bM0). Subsequently, adjuvant pembrolizumab treatment was started. A few days after the fourth maintenance course, he presented to the emergency department complaining of abdominal pain, nausea and vomiting. Emergency abdominal tomography showed a significant increase in the diameter of the appendix vermiformis, peritoneal thickening and appendiceal wall defects that could be significant in terms of perforation. The mentioned finding and given the clinical presentation, was attributed to a perporating of the appendix, so the patient was hospitalized in the Department of Surgery and the patient underwent emergency appendectomy. Histological findings were consistent with appendicitis. After a day in the hospital, the abdominal pain subsided, C-reactive protein tended to decrease and the patient was discharged. In patients who develop acute abdominal pain with or without diarrhea during immunotherapy, urgent imaging, endoscopic and clinical evaluation should be performed, and bowel perforation, although rare, should be considered as a potential complication of any immunotherapy.