Efficient methods for generation and expansion of, and gene delivery to 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. By comparing several media types and conditions, 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. We also identified the serotypes (2.5 and D-J) and conditions necessary for efficient transduction of NHP B cells with AAV vectors for the purposes of producing a secreted protein (BAFF). We hope that this work will accelerate proof-of-concept in vivo studies using engineered protein-secreting B cells in an NHP model.

In vivotracking ofex vivogenerated89Zr-oxine labeled plasma cells by PET in a non-human primate model

“…RM peripheral blood B cells have been successfully expanded and differentiated ex vivo into class switched antibody secreting cells using commercially available B cell expansion media. 43,44 We optimized these culture conditions for ex vivo B cell RM expansion and differentiation and were able to consistently achieve 10- to 15-fold cell expansion over 7 days, with preserved viability (80-90%) starting with cryopreserved peripheral blood B cells from multiple subjects ( Figure 1B ). After 7-8 days of culture, B cell expansion slowed, and viability began to decline to below 80%.…”

Advancing Cell Therapies: Single-Cell Profiling, Generation, Expansion, and Gene Delivery in Rhesus Macaque Plasma B Cells

“…In this study, we refined the reagents and methods we previously developed 12 to establish a comprehensive workflow for expanding and engineering NHP PCs. This included the identification of a range of reagents for isolation, transduction with AAV/lentivirus, expansion, and differentiation of B cells sourced from peripheral rhesus macaque blood mononuclear cells.…”