PB Immune Therapeutics, Inc

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder with a complex immune-mediated pathogenesis. The efficacy of human-specific cellular immunotherapies and biological medications cannot be accurately evaluated using traditional murine IBD models. Therefore, a humanized mouse model of IBD is necessary. Regulatory T cells (Tregs) are critical for maintaining intestinal immune homeostasis and may have therapeutic potential for treating IBD.

Donor peripheral blood mononuclear cells (PBMCs) were used to reconstitute the human immune system in NOG mice and for Treg isolation. T cells were sorted and stimulated with anti-CD3 and anti-CD28 in the presence of irradiated feeder cells to prepare Treg cells. Two weeks after PBMC reconstitution in NOG mice, colitis was induced with dextran sodium sulfate (DSS). The expanded Treg cells were administered intravenously. Ozanimod was used as a positive control.

After expansion, 65.4% of the live CD4+ cells were Foxp3+CD25+ Treg cells and 14.5% were non-Treg cells. The mean human leukocyte (hCD45+) engraftment rate in the humanized mice was 56.5% ± 4.5%. Autologous Treg-cell therapy significantly reduced the disease activity index by 78% on day 7. Colonic length was preserved, and colonic inflammation was reduced in mice treated with Treg cells. Immunohistology revealed reduced human T-cell infiltration in Treg-treated mice.

Autologous Treg therapy ameliorated the symptoms of DSS-induced colitis in a humanized mouse model. The autologous PBMC-humanized DSS-induced colitis model may serve as a robust preclinical platform for evaluating the efficacy of personalized Treg cell therapy for IBD.

Porcine‐to‐human islet xenotransplantation offers a promising solution to type 1 diabetes. This study investigates the use of Belimumab, a human monoclonal antibody that inhibits B cell activating factor (BAFF) as part of an immunosuppressive regimen in porcine‐to‐non‐human primate xenotransplantation.

Porcine islets were transplanted into Rhesus monkeys with an immunosuppressant regimen. Blood samples were collected from the monkeys pre‐ and post‐treatment, and single‐cell RNA sequencing was performed to investigate immune cell landscape changes, focusing on B cells.

UMAP clustering of B cells identified five distinct subsets. Notably, Belimumab treatment significantly reduced the proportion of both CXCR5 + and CXCR5 − atypical memory B cells, which possess the potential to differentiate into antibody‐secreting cells. Furthermore, differentially expressed gene (DEG) analysis revealed a comprehensive functional impairment, along with significant downregulation of activation markers such as CD69 and CD83, across all B cell subsets.

Our study elucidates the mechanism of action of Belimumab in non‐human primate models, serving as preclinical subjects for B‐cell–targeted therapy research in a xenotransplantation context. Significantly, our data indicate that Belimumab not only reduces the proportion of antibody‐secreting atypical memory B cells but also induces functional impairment across all B cell subsets. Given the potential pathogenic roles of atypical memory B cells in autoimmunity and other related settings, their reduction by Belimumab could play a crucial role in regulating B‐cell–mediated immune responses in pig‐to‐non‐human primate pancreatic islet xenotransplantation. Thus, our findings highlight the prospective utility of Belimumab as a B cell suppressant in future clinical xenotransplantation applications.

Single-cell RNA sequencing (scRNA-seq) has improved our ability to characterize rare cell populations. In practice, cells from different tissues or donors are simultaneously loaded onto the instrument (multiplexed) at the recommended (standard loading) or higher (superloading) numbers to save time and money. Although cost-effective, superloading can stymie computational analyses owing to high multiplet rates and sample complexity.

We compared the effects of superloading on multiplexed single-cell gene expression and T cell receptor (TCR) data generated from human thymus and blood samples from different donors.

Minimal transcriptomic differences were observed between the data generated by either standard or superloading. Irrespective of the loading cell number, we found that over 50% of the T cells expressing multiple TCR chains were doublets.

Multiple samples can be run simultaneously without compromising data quality and subsequent analyses. However, an additional doublet removal step based on TCR configuration may improve the accuracy of T cell analysis.