Relevant Antibodies Research Articles

Immunized Patients Face Reduced Access to Transplantation in the Eurotransplant Kidney Allocation System.

The presence of donor-specific HLA antibodies before transplantation is associated with poor transplantation outcomes. Unacceptable antigens can be assigned for Eurotransplant kidney transplant candidates to prevent kidney offers against which the candidate has developed clinically relevant HLA antibodies. This retrospective cohort study aimed to assess to what degree unacceptable antigens affect access to transplantation in the Eurotransplant Kidney Allocation System (ETKAS). Candidates who underwent kidney-only transplantation between 2016 and 2020 were included (n = 19 240). Cox regression was used to quantify the relationship between the relative transplantation rate and virtual panel-reactive antibodies (vPRAs), which is the percentage of the donor pool with unacceptable antigens. Models used accrued dialysis time as the timescale; were stratified by country and blood group of patient and were adjusted for nontransplantable status, patient age, sex, history of kidney transplantations, and prevalence of 0 HLA-DR-mismatched donors. Transplantation rates were 23% lower for vPRA 0.1% to 50%, 51% lower for vPRA 75% to 85%, and decreased rapidly for vPRA of >85%. Prior studies showed significantly lower ETKAS transplantation rates only for highly sensitized patients (vPRA of >85%). The inverse relationship between transplantation rate and vPRA is independent of Eurotransplant country, listing time, and 0 HLA-DR-mismatched donor availability. Results were similar when quantifying the relationship between vPRA and attainment of a sufficiently high rank for an ETKAS offer, suggesting lower transplantation rates for immunized patients are due to current ETKAS allocation. Immunized patients face lower transplantation rates across Eurotransplant. The current ETKAS allocation mechanism inadequately compensates immunized patients for reduced access to transplantation.

Response to COVID-19 Vaccination Post-CAR T Therapy in Patients With Non-Hodgkin Lymphoma and Multiple Myeloma.

COVID-19 adversely affects individuals with cancer. Several studies have found that seroconversion rates among patients with hematologic malignancies are suboptimal when compared to patients without cancer. Patients with non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) are immunocompromised due to impaired humoral and cellular immunity in addition to prescribed immunosuppressive therapy. Chimeric antigen receptor T-cell (CAR T) therapy is now widely used for NHL and MM, but little is known about seroconversion rates after COVID-19 vaccination among these populations. We evaluated SARS-CoV-2 spike-binding IgG antibody levels following COVID-19 vaccination among NHL and MM CAR T therapy recipients. Out of 104 CAR T infusions, 19 patients developed known COVID-19 infection post-CAR T. We tested 17 patients that received CAR T for antibody spike titers post COVID-19 vaccination, only 29 % (n=5) were able to mount a clinically relevant antibody response (>250 IU/mL).

RAFT Polymer-Antibody Conjugation: Squaramide Ester Chemistry Leads to Conjugates with a Therapeutic Anti-EGFR Antibody with Full Retention of Activity and Increased Tumor Uptake In Vivo.

Covalent conjugation of a biologically stable polymer to a therapeutic protein, e.g., an antibody, holds many benefits such as prolonged plasma exposure of the protein and improved tumor uptake. Generation of defined conjugates is advantageous in many applications, and a range of site-selective conjugation methods have been reported. Many current coupling methods lead to dispersity in coupling efficiencies with subsequent conjugates of less-well-defined structure, which impacts reproducibility of manufacture and ultimately may impact successful translation to treat or image diseases. We explored designing stable, reactive groups for polymer conjugation reactions that would lead to conjugates through the simplest and most abundant residue on most proteins, the lysine residue, yielding conjugates in high purity and demonstrating retention of mAb efficacy through surface plasmon resonance (SPR), cell targeting, and in vivo tumor targeting. We utilized squaric acid diesters as coupling agents for selective amidation of lysine residues and were able to selectively conjugate one, or two, high-molecular-weight polymers to a therapeutically relevant antibody, 528mAb, that subsequently retained full binding specificity. Water-soluble copolymers of N-(2-hydroxypropyl) methacrylamide (HPMA) and N-isopropylacrylamide (NIPAM) were prepared by Reversible Addition-Fragmentation chain-Transfer (RAFT) polymerization and we demonstrated that a dual-dye-labeled antibody-RAFT conjugate (528mAb-RAFT) exhibited effective tumor targeting in model breast cancer xenografts in mice. The combination of the precise and selective squaric acid ester conjugation method, with the use of RAFT polymers, leads to a promising strategic partnership for improved therapeutic protein-polymer conjugates having a very-well-defined structure.

Post-translational covalent assembly of CAR and synNotch receptors for programmable antigen targeting

Chimeric antigen receptors (CARs) and synthetic Notch (synNotch) receptors are engineered cell-surface receptors that sense a target antigen and respond by activating T cell receptor signaling or a customized gene program, respectively. Here, to expand the targeting capabilities of these receptors, we develop “universal” receptor systems for which receptor specificity can be directed post-translationally via covalent attachment of a co-administered antibody bearing a benzylguanine (BG) motif. A SNAPtag self-labeling enzyme is genetically fused to the receptor and reacts with BG-conjugated antibodies for covalent assembly, programming antigen recognition. We demonstrate that activation of SNAP-CAR and SNAP-synNotch receptors can be successfully targeted by clinically relevant BG-conjugated antibodies, including anti-tumor activity of SNAP-CAR T cells in vivo in a human tumor xenograft mouse model. Finally, we develop a mathematical model to better define the parameters affecting universal receptor signaling. SNAP receptors provide a powerful strategy to post-translationally reprogram the targeting specificity of engineered cells.

Mechanistically Weighted Metric to Predict In Vivo Antibody-Receptor Occupancy: An Analytical Approach.

In situ clinical measurement of receptor occupancy (RO) is challenging, particularly for solid tumors, necessitating the use of mathematical models that predict tumor receptor occupancy to guide dose decisions. A potency metric, average free tissue target to initial target ratio (AFTIR), was previously described based on a mechanistic compartmental model and is informative for near-saturating dose regimens. However, the metric fails at clinically relevant subsaturating antibody doses, as compartmental models cannot capture the spatial heterogeneity of distribution faced by some antibodies in solid tumors. Here we employ a partial differential equation (PDE) Krogh cylinder model to simulate spatiotemporal receptor occupancy and derive an analytical solution, a mechanistically weighted global AFTIR, that can better predict receptor occupancy regardless of dosing regimen. In addition to the four key parameters previously identified, a fifth key parameter, the absolute receptor density (targets/cell), is incorporated into the mechanistic AFTIR metric. Receptor density can influence equilibrium intratumoral drug concentration relative to whether the dose is saturating or not, thereby influencing the tumor penetration depth of the antibody. We derive mechanistic RO predictions based on distinct patterns of antibody tumor penetration, presented as a global AFTIR metric guided by a Thiele Modulus and a local saturation potential (drug equivalent of binding potential for positron emissions tomography imaging) and validate the results using rigorous global and local sensitivity analysis. This generalized AFTIR serves as a more accurate analytical metric to aid clinical dose decisions and rational design of antibody-based therapeutics without the need for extensive PDE simulations. SIGNIFICANCE STATEMENT: Determining antibody-receptor occupancy (RO) is critical for dosing decisions in pharmaceutical development, but direct clinical measurement of RO is often challenging and invasive, particularly for solid tumors. Significant efforts have been made to develop mathematical models and simplified analytical metrics of RO, but these often require complex computer simulations. Here we present a mathematically rigorous but simplified analytical model to accurately predict RO across a range of affinities, doses, drug, and tumor properties.

Cerebellar Degeneration Causing Saccadic Dysmetria and Ocular Flutter: A Seronegative Paraneoplastic Syndrome (P4-5.015)

<h3>Objective:</h3> To report a challenging case of a woman who developed eye movement abnormalities, progressive truncal, and gait ataxia due to a paraneoplastic syndrome. <h3>Background:</h3> Progressive cerebellar degeneration is a well-known paraneoplastic syndrome. However, it can be challenging to define phenotypically and diagnose in the absence of relevant antibodies. Response to treatment is often poor. <h3>Design/Methods:</h3> Case Report. <h3>Results:</h3> A 68-year-old woman presented with several months of moderately progressive truncal, gait ataxia, followed by development of saccadic hypermetria and ocular flutter. Her metastatic small cell lung cancer was in remission, and she no longer showed bilateral cerebellar metastases on brain MRI, following treatment with chemotherapy, durvalumab, and whole brain radiation. CSF analysis revealed lymphocytic pleocytosis (8/μL), normal protein, normal glucose, and 4 oligoclonal bands. Paraneoplastic antibodies from serum and CSF were negative. She was diagnosed with a probable paraneoplastic syndrome, based on a PNS-Care score of 7. She was treated with high dose intravenous methylprednisolone and plasmapheresis, with resolution of eye movement abnormalities, and improvement of ataxia. <h3>Conclusions:</h3> Subacute development of saccadic dysmetria and ocular flutter in this case helped define a new onset cerebellar syndrome in a patient with possible residual cerebellar dysfunction from prior cerebellar metastatic disease. Saccadic hypermetria is typically caused by dysfunction of the fastigial oculomotor region (FOR). In our case, ocular flutter might have been caused by indirect instability of the brainstem network due to impairment of the FOR projections to the omnipause neurons (OPN) in the pontine saccadic burst generator area. Even in the absence of relevant antibodies and in the setting of confounders such as prior cerebellar metastases, use of checkpoint inhibitors, clinical presentation and CSF findings should raise concern for a paraneoplastic disorder. Cerebellar paraneoplastic syndromes may respond to Immunotherapy with methylprednisolone and plasmapheresis. <b>Disclosure:</b> Dr. Zampino has nothing to disclose. Dr. Figgie has nothing to disclose. Dr. Shaikh has received personal compensation in the range of $10,000-$49,999 for serving on a Speakers Bureau for Acorda. Dr. Serra has received personal compensation in the range of $10,000-$49,999 for serving on a Speakers Bureau for Biogen Idec. Dr. Serra has received personal compensation in the range of $5,000-$9,999 for serving on a Speakers Bureau for Bristol Myers Squibb.

Immunomodulatory Aspects of Therapeutic Plasma Exchange in Neurological Disorders—A Pilot Study

Therapeutic plasma exchange (TPE) is used for drug-resistant neuroimmunological disorders, but its mechanism of action remains poorly understood. We therefore prospectively explored changes in soluble, humoral, and cellular immune components associated with TPE. We included ten patients with neurological autoimmune disorders that underwent TPE and assessed a panel of clinically relevant pathogen-specific antibodies, total serum immunoglobulin (Ig) levels, interleukin-6 (IL-6, pg/mL), C-reactive protein (CRP, mg/dL), procalcitonin (PCT, µg/L) and major lymphocyte subpopulations (cells/µL). Blood was collected prior to TPE (pre-TPE, baseline), immediately after TPE (post-TPE), as well as five weeks (follow-up1) and 130 days (follow-up2) following TPE. Pathogen-specific antibody levels were reduced by -86% (p < 0.05) post-TPE and recovered to 55% (follow-up1) and 101% (follow-up2). Ig subclasses were reduced by -70-89% (p < 0.0001) post-TPE with subsequent complete (IgM/IgA) and incomplete (IgG) recovery throughout the follow-ups. Mean IL-6 and CRP concentrations increased by a factor of 3-4 at post-TPE (p > 0.05) while PCT remained unaffected. We found no alterations in B- and T-cell populations. No adverse events related to TPE occurred. TPE induced a profound but transient reduction in circulating antibodies, while the investigated soluble immune components were not washed out. Future studies should explore the effects of TPE on particular cytokines and assess inflammatory lymphocyte lineages to illuminate the mode of action of TPE beyond autoantibody removal.

Decreased immunoreactivity of hepatitis E virus antigen following treatment with Sakhalin spruce (Picea glehnii) essential oil.

The hepatitis E virus (HEV) causes a common infectious disease that infects pigs, wild boars, deer, and humans. In most cases, humans are infected by eating raw meat. Some essential oils have been reported to exhibit antiviral activities. In this study, in order to investigate the anti-HEV properties of essential oils, the immunoreactivities of HEV antigen proteins against the relevant antibodies were analyzed after the HEV antigens underwent treatment with various essential oils. The essential oils extracted from the tea tree, which was previously reported to exhibit antiviral activity, lavender, and lemon had strongly reduced activity. We found that treatment with the essential oil prepared from Sakhalin spruce was associated with the strongest reduction in immunoreactivity of HEV antigen protein(s) among the tested substances. The main volatile constituents of Sakhalin spruce essential oil were found to be bornyl acetate (32.30%), α-pinene (16.66%), camphene (11.14%), camphor (5.52%), β-phellandrene (9.09%), borneol (4.77%), and limonene (4.57%). The anti-HEV properties of the various components of the essential oils were examined: treatment with bornyl acetate, the main component of Sakhalin spruce oil, a-pinene, the main component of tea tree oil, and limonene, the main component of lemon oil, resulted in a strong reduction in HEV antigen immunoreactivity. These results indicate that each main component of the essential oils plays an important role in the reduction of the immunoreactivity of HEV antigen protein(s); they also suggest that Sakhalin spruce essential oil exhibits anti-HEV activity. In a formulation with the potential to eliminate the infectivity of HEV in foodborne infections, this essential oil can be applied as an inactivating agent for meat processing and cooking utensils, such as knives and chopping boards.

Nano-LYTACs for Degradation of Membrane Proteins and Inhibition of CD24/Siglec-10 Signaling Pathway.

Lysosome-targeting chimeras (LYTACs) are an emerging therapeutic modality that effectively degrade cancer cell membranes and extracellular target proteins. In this study, a nanosphere-based LYTAC degradation system is developed. The amphiphilic peptide-modified N-acetylgalactosamine (GalNAc) can self-assemble into nanospheres with a strong affinity for asialoglycoprotein receptor targets. They can degrade different membranes and extracellular proteins by linking with the relevant antibodies. CD24, a heavily glycosylated glycosylphosphatidylinositol-anchored surface protein, interacts with Siglec-10 to modulate the tumor immune response. The novel Nanosphere-AntiCD24, synthesized by linking nanospheres with CD24 antibody, accurately regulates the degradation of CD24 protein and partially restores the phagocytic function of macrophages toward tumor cells by blocking the CD24/Siglec-10 signaling pathway. When Nanosphere-AntiCD24 is combined with glucose oxidase, an enzyme promoting the oxidative decomposition of glucose, the combination not only effectively restores the function of macrophages in vitro but also suppresses tumor growth in xenograft mouse models without detectable toxicity to normal tissues. The results indicate that GalNAc-modified nanospheres, as a part of LYTACs, can be successfully internalized and are an effective drug-loading platform and a modular degradation strategy for the lysosomal degradation of cell membrane and extracellular proteins, which can be broadly applied in the fields of biochemistry and tumor therapeutics.

Structure of the Inmazeb cocktail and resistance to Ebola virus escape

Monoclonal antibodies can provide important pre- or post-exposure protection against infectious disease for those not yet vaccinated or in individuals that fail to mount a protective immune response after vaccination. Inmazeb (REGN-EB3), a three-antibody cocktail against Ebola virus, lessened disease and improved survival in a controlled trial. Here, we present the cryo-EM structure at 3.1Å of the Ebola virus glycoprotein, determined without symmetry averaging, in a simultaneous complex with the antibodies in the Inmazeb cocktail. This structure allows the modeling of previously disordered portions of the glycoprotein glycan cap, maps the non-overlapping epitopes of Inmazeb, and illuminates the basis for complementary activities and residues critical for resistance to escape by these and other clinically relevant antibodies. We further provide direct evidence that Inmazeb protects against the rapid emergence of escape mutants, whereas monotherapies even against conserved epitopes do not, supporting the benefit of a cocktail versus a monotherapy approach.

Assessment and management of allosensitization following heart transplant in adults

Immunological injury to the allograft, specifically by antibodies to de novo donor specific human leukocyte antigen (dnDSA) and antibody mediated injury and rejection are the major limitations to graft survival after heart transplantation (HT). As such, our approach to allosensitization remains limited by the inability of contemporaneous immunoassays to unravel pathogenic potential of dnDSA. Additionally, the role of dnDSA is continuously evaluated with emerging methods to detect rejection. Moreover, the timing and frequency of dnDSA monitoring for early detection and risk mitigation as well as management of dnDSA remain challenging. A strategic approach to dnDSA employs diagnostic assays to determine relevant antibodies in conjunction with clinical presentation and injury/rejection of allograft to tailor therapeutics. In this review, we aim to outline contemporary knowledge involving detection, monitoring and management of dnDSA after HT. Subsequently, we propose a diagnostic and therapeutic approach that may mitigate morbidity and mortality while balancing adverse reactions from pharmacotherapy.

Modelling acute antibody-mediated rejection of human kidney transplants using ex-vivo warm machine perfusion.

Transplant rejection is a major cause of graft loss and morbidity. Currently, no human models of antibody-mediated rejection (AMR) exist, limiting mechanistic investigation and organ-specific targeted therapy. Here, using 12 human kidneys and ex-vivo normothermic machine perfusion, we demonstrate phenotypes of AMR after addition of antibodies against either human HLA class I or blood group antigens (A, B), thus modelling clinical AMR that can follow HLA incompatible (HLAi) or blood group incompatible (ABOi) transplantation. Discarded human kidneys with wide ranging demographics and cold ischaemia times (11-54h) were perfused with red blood cells and fresh frozen plasma (FFP) as a source of complement/coagulation factors. For the HLAi model, 600μg of W6/32 anti-class 1 HLA antibody was added to the circuit (time '0'). For the ABOi model, high titre FFP of the relevant blood group antibody was added. Renal blood flow index (RBFi, mL/min/100 g), C3 desArg, prothrombin fragments 1+2 and histology were determined. Our endpoints included haemodynamic changes, thrombosis, and biopsy proven complement deposition. Compared to control kidneys perfused without anti-donor antibodies, both models demonstrated haemodynamic collapse after antibody perfusion with only the HLAi model showing glomerular C4d deposition. We show that a clinically relevant human kidney model of AMR is feasible, and anticipate that these models, with refinements, could provide a basis to test different strategies to prevent AMR. The Rosetrees and Stonygate Trust, The Royal College of Surgeons of England Fellowship Grant, NIHR Biomedical Research Centre/KCL Early Career Grant, Kidney Research U.K.

Bioluminescence Goes Dark: Boosting the Performance of Bioluminescent Sensor Proteins Using Complementation Inhibitors.

Bioluminescent sensor proteins have recently gained popularity in both basic research and point-of-care diagnostics. Sensor proteins based on intramolecular complementation of split NanoLuc are particularly attractive because their intrinsic modular design enables for systematic tuning of sensor properties. Here we show how the sensitivity of these sensors can be enhanced by the introduction of catalytically inactive variants of the small SmBiT subunit (DarkBiTs) as intramolecular inhibitors. Starting from previously developed bioluminescent antibody sensor proteins (LUMABS), we developed single component, biomolecular switches with a strongly reduced background signal for the detection of three clinically relevant antibodies, anti-HIV1-p17, cetuximab (CTX), and an RSV neutralizing antibody (101F). These new dark-LUMABS sensors showed 5-13-fold increases in sensitivity which translated into lower limits of detection. The use of DarkBiTs as competitive intramolecular inhibitor domains is not limited to the LUMABS sensor family and might be used to boost the performance of other bioluminescent sensor proteins based on split luciferase complementation.

Zika Virus Infection and Antibody Neutralization in FcRn Expressing Placenta and Engineered Cell Lines.

As a developmental toxicant, Zika virus (ZIKV) attacks both the growing nervous system, causing congenital Zika syndrome, and the placenta, resulting in pathological changes and associated adverse fetal outcomes. There are no vaccines, antibodies, or other treatments for ZIKV, despite the potential for its re-emergence. Multiple studies have highlighted the risk of antibodies for enhancing ZIKV infection, including during pregnancy, but the mechanisms for such effects are not fully understood. We have focused on the ability of the neonatal Fc receptor (FcRn) to interact with ZIKV in the presence and absence of relevant antibodies. We found that ZIKV replication was higher in Marvin Darby Canine Kidney (MDCK) cells that overexpress FcRn compared to those that do not, and knocking down FcRn decreased ZIKV RNA production. In the placenta trophoblast BeWo cell line, ZIKV infection itself downregulated FcRn at the mRNA and protein levels. Addition of anti-ZIKV antibodies to MDCK/FcRn cells resulted in non-monotonous neutralization curves with neutralization attenuation and even enhancement of infection at higher concentrations. Non-monotonous neutralization was also seen in BeWo cells at intermediate antibody concentrations. Our studies highlight the underappreciated role FcRn plays in ZIKV infection and may have implications for anti-ZIKV prophylaxis and therapy in pregnant women.

Immunoglobulin Class Profiles of ABO Antibodies in Saliva and Serum of Healthy Individuals

Introduction: The coronavirus disease (COVID-19) pandemic gave rise to studies investigating the association of ABO blood group with COVID-19 susceptibility. It is hypothesized that ABO antibodies might play a role in neutralizing SARS-CoV-2. However, ABO antibodies were exclusively analyzed in blood samples. Investigation of ABO antibodies in saliva, an easy-to-obtain surrogate for respiratory secretions, may provide novel insights into mucosal immunity crucial in early defense against respiratory pathogens. Methods: In this study, saliva and serum samples from healthy individuals with known blood groups were investigated using a flow cytometric method for separate anti-A/anti-B IgA, IgM, and IgG class antibody detection. Saliva samples were additionally tested using hemagglutination-based neutral and indirect anti-human globulin test gel cards. This method comparison was complemented by dilution experiments with a high-titer anti-A/anti-B WHO standard. Results: In saliva, IgA was the most abundant ABO antibody class, followed by IgM; IgG was detected only in low levels in all non-AB blood types. In serum, IgM was the predominant ABO antibody class in all non-AB blood types, followed by IgA and IgG, the latter mainly detected in group O individuals. Saliva and serum samples of group O individuals yielded the highest variability of ABO-specific antibody levels. Regardless of sample material and blood type, major interindividual differences in ABO antibody reactivities were recorded. Antibody levels correlated moderately between these two body fluids. There were no significant sex and age-group differences in ABO antibody levels in both serum and saliva. WHO standard dilution experiments yielded technique-specific limits of detection, illustrating the inherent differences of immunofluorescence versus agglutination. Conclusion: For the first time, salivary ABO antibodies were investigated by separate detection of the three most relevant antibody classes IgA, IgM, and IgG in a healthy cohort. This study opens new perspectives regarding mucosal ABO antibody class profiles and their potential influence on respiratory infections.

Single-Cell Multiomics Reveal Hypomethylating Agent Partially Restores Hematopoiesis in a LR-MDS Patient at the Cost of Leukemic Transformation

Background Myelodysplastic syndromes (MDS) are a heterogeneous group of diseases characterized by ineffective hematopoiesis. Hypomethylating agents (HMA) such as decitabine and azacytidine (AZA) induce clinical responses in both lower-risk (LR) and higher-risk groups; however, the treatment eventually fails and can be followed by leukemic transformation. The mechanism underlying transient clinical benefit from low-dose HMA (restoration of hematopoiesis) is not fully understood. Here we used cutting-edge single-cell multi-omic approaches to describe the abnormal clonal architecture and transcriptome in LR-MDS and how they are altered following HMA therapy. Results First, we defined genomic differences between aged and young healthy hematopoietic stem and progenitor cells (HSCPs) by building a single-cell atlas of human HSCP with molecularly-defined concentrations of relevant CITE seq antibodies. 350k high-quality bone marrow nucleated cells were obtained from four young donors; varying race and gender. A 6-fold molecular titration of 278 CITE-seq antibodies determined those which are functional and which contribute to clustering. Next, we repeated the captures on samples from aged healthy people (n=3), MDS patients (n=4) and conducted differential analysis on cell states and transcriptomes. Compared to young marrow, healthy marrow from elderly donors revealed global upregulation of inflammatory signals and innate immunity. In erythroid progenitors (ERP) we identified abnormally high expression of the transcription factors KLF1 and GATA1, but lower than normal expression of their hemoglobin target genes. At the same time altered erythropoiesis in the elderly appeared to be compensated by increased frequency of multilineage progenitors and more mature erythroblasts mitigating clinical anemia. Comparing MDS to aged marrow, while most cell populations from MDS showed significant gene expression changes, the amplitude was dwarfed by abnormal mRNA processing gene expression in ERP and erythroblasts. Moreover, in de novo LR-MDS samples and an anemia sample (without known mutations) further downregulation of hemoglobin genes in the erythroid lineage was accompanied by decreased erythroblast numbers leading to anemia. To study the impact of AZA on LR-MDS, we longitudinally profiled a single MDS patient (pre-HMA, post-HMA, sAML transformation, cycle 2 VEN+AZA) (Panel 1). We found that 6 cycles of AZA treatment depleted HSC; whereas lymphoid-primed multipotential progenitors (LMPP), megakaryocyte-erythroid progenitors (MEP), and less mature erythroblasts were enriched after AZA therapy (Panel 2 left). Using genotyping of the transcriptome (GoT) (Panel 2 right), we defined cells with mutations to find that not only SRSF2-relevant pathways (eg. mRNA processing, and mRNA splicing) are molecularly corrected (downregulated) but also there are fewer mutant cells among primitive stem cells and lineage-committed progenitors. However, mutant ERP and erythroblasts accumulated and pathways were more active suggesting the blockade of differentiation at terminal erythropoiesis is not rescued, consistent with the clinical observation that the patient was still RBC transfusion-dependent despite platelet recovery. Notably, these mutant erythroid cells, while defective, never transformed to leukemia. The expansion of mutant multilineage progenitors and LMPP preceded AML transformation and were eliminated (as evidenced by flow and CITE-seq) following two cycles of VEN+AZA. Conclusions: Age-matched references are critical in differential analysis of hematological diseases like AML or MDS. With matched elderly donor data sets, we were able to molecularly isolate functional disruption of erythropoiesis and megakaryopoiesis underlying anemia and thrombocytopenia, respectively, observed in an LR-MDS patient. AZA rescued the dysregulated signals in SRSF2-mutated LR-MDS patient HSPCs, but not in ERP and erythroblasts, and therefore only partially restored hematopoiesis. However, as AZA drove differentiation of both wildtype and mutant HSCP, accumulation of mutant multilineage progenitors and LMPPs contributed to progression to AML. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Formation of anti-infectious and post-vaccination anti-SARS-CoV-2 humoral immunity in medical workers of the perinatal center

In the context of the global spread of the new coronavirus infection, studies aimed at investigating formation of anti-infectious and post-vaccination immunity are of special importance, which is necessary to prevent and reduce morbidity and mortality due to SARS-CoV-2 infection. Purpose: to assess anti-infectious immunity against SARS-CoV-2 in various forms of the disease and development of post-vaccination humoral reactions in medical workers of the perinatal center.&#x0D; Materials and methods. A study of blood serum was carried out to assess SARS-CoV-2-specific IgM and IgG antibodies in 119 medical workers recovered after COVID-19, divided into groups based on the disease severity (mild, moderate and asymptomatic), as well as in 62 vaccinated employees, divided into groups according to age. Semi-quantitative measurement of virus-specific antibodies was carried out by ELISA with test systems SARS-CoV-2-IgG-ELISA-BEST and SARS-CoV-2-IgM-ELISA-BEST. Statistical processing of the research results was carried out using Microsoft Excel 2010 and Statistica 6. Quantitative characteristics were presented as median (ME), lower and upper quartiles (LQ1-UQ3); qualitative parameters as absolute value and relative number (%). Difference between groups was analyzed by using the 2 test (qualitative) and the MannWhitney U-test (quantitative).&#x0D; Results. The results of the study showed that the majority of employees with a moderate-severe form of SARS-CoV-2 had a high level of IgG (PR a positivity rate of more than 9.0 arbitrary units) 9 months after the disease compared to those who suffered from mild or asymptomatic (83.3% versus 25.8% and 13.3%, p 0.017) infection. The duration of IgG circulation after former illness had no relation to its severity and patient age. The effectiveness of the primary vaccination Sputnik V and revaccination with Sputnik Light and KoviVak was 100% after inoculating the vaccine second component. The lowest level of antibodies after the first vaccination is recorded in persons over 60 years old (1.48 (1.123.25 versus PR = 8.48(5.7810.11) and 9.27 (5.8410.31) arbitrary units, p 0.017)), in comparison with young and middle-age subjects. The speed SARS-CoV-2 elimination of IgG at 6, 9 or more months after vaccination depends on relevant initial peak antibody concentration. Subjects who were initially vaccinated with the KoviVac vaccine, IgG was not detected 2 months after vaccination. The protective effect of Sputnik V, Sputnik Light, KoviVac after re-infection with SARS-CoV-2 averages 71.2%.&#x0D; Conclusion. Thus, the results obtained on assessing anti-infectious and post-vaccination immunity against SARS-CoV-2 emphasize the need for further studies on a larger patient cohort, especially in those with asymptomatic infection as well as the elderly subjects.

Fourth dose of the SARS-CoV-2 vaccine in kidney transplant recipients with previously impaired humoral antibody response

Fourth dose of the SARS-CoV-2 vaccine in kidney transplant recipients with previously impaired humoral antibody response

Stable High-Concentration Monoclonal Antibody Formulations Enabled by an Amphiphilic Copolymer Excipient.

Monoclonal antibodies are a staple in modern pharmacotherapy. Unfortunately, these biopharmaceuticals are limited by their tendency to aggregate in formulation, resulting in poor stability and often requiring low concentration drug formulations. Moreover, existing excipients designed to stabilize these formulations are often limited by their toxicity and tendency to form particles such as micelles. Here, we demonstrate the ability of a simple "drop-in", amphiphilic copolymer excipient to enhance the stability of high concentration formulations of clinically-relevant monoclonal antibodies without altering their pharmacokinetics or injectability. Through interfacial rheology and surface tension measurements, we demonstrate that the copolymer excipient competitively adsorbs to formulation interfaces. Further, through determination of monomeric composition and retained bioactivity through stressed aging, we show that this excipient confers a significant stability benefit to high concentration antibody formulations. Finally, we demonstrate that the excipient behaves as an inactive ingredient, having no significant impact on the pharmacokinetic profile of a clinically relevant antibody in mice. This amphiphilic copolymer excipient demonstrates promise as a simple formulation additive to create stable, high concentration antibody formulations, thereby enabling improved treatment options such as a route-of-administration switch from low concentration intravenous (IV) to high concentration subcutaneous (SC) delivery while reducing dependence on the cold chain.

Therapeutic plasma exchange in adolescent and adult patients with autoimmune neuropsychiatric disorders associated with streptococcal infections.

Pediatric autoimmune neuropsychiatric disorders associated with streptococcal (PANDAS) infections and pediatric acute-onset neuropsychiatric syndrome (PANS) are typically diagnosed in childhood. Therapeutic plasma exchange (TPE) has been recommended to remove relevant antibodies and treat symptomatic presentations in children and adolescents, but there are no studies that evaluate the use of TPE in patients who are diagnosed later in life. It is therefore unclear if using an accepted treatment for pediatric PANS/PANDAS patients would be beneficial in adults with prolonged PANDAS/PANS symptomatic histories. This study investigated 16 late adolescent and adult PANDAS/PANS patients' responses to TPE. Improvement was noted in over half of the patients with available follow-up information.