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Advancing Cell Therapies: Single-Cell Profiling in Rhesus Macaque Plasma B Cells

ABSTRACT: Engineered long lived plasma cells have the potential to be a new area of cell therapy. A key step in developing this cell therapy is testing in a model with an intact immune system similar to humans. To that end, we have developed methods to purify, expand, and differentiate non-human primate (NHP; rhesus macaque) B cells ex vivo. We consistently achieved 10-fold expansion of NHP B cells using a readily available commercial supplement. After only seven days in culture, large percentages of cells in NHP B cell cultures were differentiated. These cells expressed surface markers found in human antibody secreting cells (CD38 and CD138) and secreted immunoglobulin G. From single cell transcriptome analysis of NHP, we verified the presence of plasma cell markers commonly shared with humans, and have unearthed less recognized markers such as CD59 and CD79A. In addition, we identified unique NHP plasma cell markers that are absent in humans including the immune checkpoint molecule CD274 (PD-L1, Programmed Death-Ligand 1). Furthermore, we found that MHC class I molecules were upregulated in NHP plasma cells, in contrast to the pattern observed in humans. Lastly, we also identified the serotypes (AAVD-J) and established the conditions for efficient transduction of NHP B cells with AAV vectors, achieving an editing rate of approximately 60%. We envision that this work will accelerate proof-of-concept in vivo studies using engineered protein-secreting B cells in the NHP model.

ORGANISM(S): Macaca mulatta

PROVIDER: GSE247023 | GEO | 2023/11/14

REPOSITORIES: GEO

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Project description:Normal human prostate (NHP) epithelial cells undergo senescence in vitro and in vivo, but the underlying molecular mechanisms remain incompletely understood. Here we first show that multiple primary strains of NHP cells are all immunophenotyped as CK5+/CK18+CD44+α2β1+p63+hTERT+ progenitor cells, which gradually lose progenitor markers as they lose proliferative capacity. NHP cell senescence involves loss of telomerase expression, upregulation of p16, and activation of p53. Using genetically defined manipulations of these three signaling pathways, we show that p16 is the primary determinant of the NHP cell proliferative capacity and hTERT is required for unlimited proliferative lifespan. Hence, suppression of 16 alone significantly extends NHP cell lifespan but both p16 inhibition and hTERT are required to immortalize NHP cells. Importantly, the immortalized NHP cells are normal progenitors that possess the ability to differentiate into functional prostatic glands containing both basal and luminal cells and, frequently, neuroendocrine cells. The immortalized NHP cells possess gene expression profiles characteristic of proliferating progenitor cells. Our studies shed light on the molecular mechanisms regulating the proliferative lifespan of NHP progenitor cells and provide direct evidence that basal-like progenitor cells can regenerate the entire prostatic glands in vivo. The availability of these cells should facilitate future studies of prostate cancer development. Further processed data linked below as Supplementary files. TestvsControl: corresponding GSM# 3vs1: GSM266720, 721, 722 4vs1: GSM266727, 730, 731 5vs1: GSM266732, 733, 734 7avs1: GSM266735, 736, 737 9vs1: GSM266772, 773, 812 11vs1: GSM266813, 816, 823 13vs1: GSM266824, 826, 828 Note: The 'TestvsControl' comparisons report the top 100 up- or down-regulated genes. The numeric designations (1 to 13) are used in the corresponding manuscript. The 'SignificantGenes' lists were used in clustering analysis. Keywords: Cell type comparison (senscent and immortalized NHP cells compared to young NHP progenitors)

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Advancing Cell Therapies: Single-Cell Profiling in Rhesus Macaque Plasma B Cells

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