Felipe Andrade

Malignant ascites provides a window into cancer-immune interactions in the peritoneal cavity. Using Cytometry by Time of Flight (CyTOF), we analyzed ascitic immune cells collected from patients with advanced gastrointestinal cancers, nonmalignant forms of hepatitis (NMH), and healthy control peripheral blood mononuclear cells (HC PBMCs). Compositionally, ascitic immune cells shared a myeloid-rich, B and naïve T cell-low signature relative to HC PBMCs. Underneath the shared ascitic compositional signature, three major functional immunophenotypes emerged in response to exogenous stimulation along a suppressive to cytotoxic axis, aligning with Luminex measurements of ascitic cytokine levels: IL-10 enriched, Nonpolarized, and Granzyme B (GZMB) enriched. In our cohort, the IL-10 enriched functional immunophenotype was exclusive to samples from patients with cancer while the GZMB enriched response was dominated by NMH samples. Ascites from patients with pancreatic ductal adenocarcinoma (PDAC) contained a persistent anti-cancer humoral response characterized by IgG that bound to autologous ascitic PDAC cells and multiple allogeneic PDAC cell lines.

Chimeric antigen receptor (CAR)-T cell therapies can induce drug-free remission in systemic lupus erythematosus (SLE), but indiscriminate B cell targeting causes immunosuppression, unnecessary infections, and cytokine toxicities that preclude widespread use. Here, we overcome this by targeting the 9G4 idiotope, a shared structural feature of pathogenic B cell receptors encoded by the IGHV4-34 gene. We engineered anti-9G4 CAR-T cells and chimeric TCR-T cells to selectively eliminate autoreactive B cells while preserving protective immunity. Both platforms eradicated autoreactive B cells and autoantibodies in vitro and in vivo, spared normal B cells, and markedly reduced cytokine release compared to conventional CAR-T cells. This precision extended to cold agglutinin disease and lymphoma. These findings establish a framework for IGHV idiotope-directed cellular therapies for treating autoimmune and neoplastic diseases while preserving immune competence.

Objective: Autoantibodies targeting peptidylarginine deiminase 4 (PAD4), an enzyme involved in protein citrullination, are found in a subset of patients with rheumatoid arthritis (RA) with severe joint disease. However, the mechanisms by which anti‐PAD4 antibodies participate in disease pathogenesis are incompletely defined.Methods: We investigated the role of anti‐PAD4 monoclonal antibodies derived from patients with RA using a collagen‐induced arthritis (CIA) mouse model and human monocyte in vitro cultures. The cellular targets of anti‐PAD4 antibodies were identified using mouse knee joint cells and human peripheral blood mononuclear cells. In addition, PAD4 gene and protein expression was assessed using human fibroblast‐like synoviocyte in vitro cultures and a single‐cell RNA sequencing data set obtained from patients with RA.Results: We show that anti‐PAD4 antibody treatment augmented disease severity in the CIA mouse model, with increased joint damage, myeloid cell infiltration, and synovial fibroblast activation. Arthritic mice administered with anti‐PAD4 antibodies had an increased proportion of interleukin‐17A (IL‐17A), tumor necrosis factor α (TNFα), and interferon‐γ (IFNγ)–producing T cells. Anti‐PAD4 antibodies preferentially bound monocytes in both humans and mice, eliciting proinflammatory chemokine production by human monocytes in vitro. T cell cytokines enhanced by anti‐PAD4 antibodies in the CIA model (ie, IL‐17A, TNFα, and IFNγ) synergized to induce a proinflammatory phenotype in human fibroblast‐like synoviocytes.Conclusion: Our findings suggest a model in which anti‐PAD4 antibody binding to monocytes triggers an inflammatory cascade that promotes immune cell recruitment to the joint and T cell activation, culminating in synovial fibroblast activation and the development of more severe arthritis. image

Why autoantibodies in rheumatoid arthritis (RA) primarily target physiologically modified proteins, called citrullinated proteins, is unknown. Recognizing the inciting event in the production of anti-citrullinated protein antibodies (ACPAs) may shed light on the origin of RA. Here, we demonstrate that ACPAs originate from germline-encoded antibodies targeting a distinct but structurally similar modification, called carbamylation, which is pathogenic and environmentally driven. The transition from anti-carbamylated protein (anti-CarP) antibodies to ACPAs results from somatic hypermutations, indicating that the change in reactivity is acquired via antigen-driven affinity maturation. During this process, a single germline anti-CarP antibody transitions from anti-CarP to double positive (anti-CarP/ACPA) to ACPA according to the pattern and number of somatic hypermutations, explaining their coexistence and diverse specificity in RA. Artificial intelligence-based structural modeling revealed that an ACPA and its germline precursor exhibit distinct structural and biophysical properties, and pointed to heavy-chain tryptophan 48 (H-W48) as a critical residue in the differential recognition of citrullinated vs. carbamylated proteins. Indeed, a single methionine substitution in H-W48 changes the antibody specificity from ACPA to anti-CarP. These data indicate that the existence of germline-encoded anti-CarP antibodies is most likely the first event in the production of ACPAs during the early stages of RA development.

Background: Critically ill hospitalized patients with COVID-19 have greater antibody titers than those with mild to moderate illness, but their association with recovery or death from COVID-19 has not been characterized. Methods: In a cohort study of 178 COVID-19 patients, 73 non-hospitalized and 105 hospitalized patients, mucosal swabs and plasma samples were collected at hospital enrollment and up to 3 months post-enrollment (MPE) to measure virus RNA, cytokines/chemokines, binding antibodies, ACE2 binding inhibition, and Fc effector antibody responses against SARS-CoV-2. The association of demographic variables and more than 20 serological antibody measures with intubation or death due to COVID-19 was determined using machine learning algorithms. Results: Predictive models reveal that IgG binding and ACE2 binding inhibition responses at 1 MPE are positively and anti-Spike antibody-mediated complement activation at enrollment is negatively associated with an increased probability of intubation or death from COVID-19 within 3 MPE. Conclusions: At enrollment, serological antibody measures are more predictive than demographic variables of subsequent intubation or death among hospitalized COVID-19 patients.

A defining feature of systemic lupus erythematosus (SLE) is loss of tolerance to self-DNA, and deficiency of DNASE1L3, the main enzyme responsible for chromatin degradation in blood, is also associated with SLE. This association can be found in an ultrarare population of pediatric patients with DNASE1L3 deficiency who develop SLE, adult patients with loss-of-function variants of DNASE1L3 who are at a higher risk for SLE, and patients with sporadic SLE who have neutralizing autoantibodies against DNASE1L3. To mitigate the pathogenic effects of inherited and acquired DNASE1L3 deficiencies, we engineered a long-acting enzyme biologic with dual DNASE1/DNASE1L3 activity that is resistant to DNASE1 and DNASE1L3 inhibitors. Notably, we found that the biologic prevented the development of lupus in Dnase1^–/–Dnase1L3^–/– double-knockout mice and rescued animals from death in pristane-induced lupus. Finally, we confirmed that the human isoform of the enzyme biologic was not recognized by autoantibodies in SLE and efficiently degraded genomic and mitochondrial cell–free DNA, as well as microparticle DNA, in SLE plasma. Our findings suggest that autoimmune diseases characterized by aberrant DNA accumulation, such as SLE, can be effectively treated with a replacement DNASE tailored to bypass pathogenic mechanisms, both genetic and acquired, that restrict DNASE1L3 activity.

The risk of autoimmunity linked to female sex is substantially larger than any susceptibility gene discovered to date (1). The mechanisms underlying sex bias in autoimmune diseases, however, remain poorly understood. By approaching different hypotheses, two recent studies reached to the conclusion that X-inactive-specific transcript (XIST)a X chromosome-encoded long noncoding RNA (lncRNA)may explain sex bias in systemic lupus erythematosus (SLE) and potentially other femaleassociated autoimmune diseases (2, 3). Because the function of XIST is to inactivate one of the two X chromosomes in females (4), it is not expressed in males, explaining why XIST is a strong candidate for underlying female predominance in autoimmune diseases. Interestingly, far from its function in X chromosome inactivation (XCI), both studies suggest that XIST rather induces autoimmunity by activating innate or adaptive immune responses (2, 3). While both studies complement one other, the proposed mechanisms are different, which merit an examination of their discrepancies. Because gain-of-function of toll-like receptor 7 (TLR7), a cellular sensor of viral infection activated by RNA, is a cause of SLE (one of the most female sex-biased autoimmune diseases) (5), Crawford et al., used unbiased approaches to search for endogenous female-specific TLR7 ligands (2). Using differential expression analysis of peripheral blood between female and male donors, as well as transcriptional data from SLE tissues (i.e., blood, spleen, and kidney), they ranked transcripts based on 4 criteria: female expression bias, total UU count, maximum UU richness, and expression. Remarkably, this approach led to the discovery of XIST as the strongest candidate carrying female-specific TLR7 ligands in the entire human genome. Further studies demonstrated that XIST, but not other RNAs in the cellular transcriptome, activates TLR7 triggering the production of interferon (IFN)-a by plasmacytoid dendritic cells, which is a hallmark in SLE (6). Notably, XIST expression was found to be increased in leukocytes from women with SLE, which correlated with disease activity and the IFN signature. Moreover, they showed that XIST is not IFN inducible, implying that XIST is a cause rather than a result of IFN production in SLE. In contrast to the work by Crawford et. al., which indicates that XIST drives autoimmunity by directly acting as a DAMP (Damage-Associated Molecular Pattern) (2), Dou et al., proposed that XIST plays an indirect role in autoimmunity by providing protein autoantigens to the adaptive immune system (3). Since XIST is a ribonucleoprotein Frontiers in Immunology frontiersin.org 01

Critically ill people with COVID-19 have greater antibody titers than those with mild to moderate illness, but their association with recovery or death from COVID-19 has not been characterized. In 178 COVID-19 patients, 73 non-hospitalized and 105 hospitalized patients, mucosal swabs and plasma samples were collected at hospital enrollment and up to 3 months post-enrollment (MPE) to measure virus RNA, cytokines/chemokines, binding antibodies, ACE2 binding inhibition, and Fc effector antibody responses against SARS-CoV-2. The association of demographic variables and >20 serological antibody measures with intubation or death due to COVID-19 was determined using machine learning algorithms. Predictive models revealed that IgG binding and ACE2 binding inhibition responses at 1 MPE were positively and C1q complement activity at enrollment was negatively associated with an increased probability of intubation or death from COVID-19 within 3 MPE. Serological antibody measures were more predictive than demographic variables of intubation or death among COVID-19 patients.