Human plasma cells engineered to secrete bispecifics drive effectivein vivoleukemia killing

Abstract

Bispecific antibodies are an important tool for the management and treatment of acute leukemias. Advances in genome-engineering have enabled the generation of human plasma cells that secrete therapeutic proteins and are capable of long-termin vivoengraftment in humanized mouse models. As a next step towards clinical translation of engineered plasma cells (ePCs) towards cancer therapy, here we describe approaches for the expression and secretion of bispecific antibodies by human plasma cells. We show that human ePCs expressing either fragment crystallizable domain deficient anti-CD19 x anti-CD3 (blinatumomab) or anti-CD33 x anti-CD3 bispecific antibodies mediate T cell activation and direct T cell killing of specific primary human cell subsets and B-acute lymphoblastic leukemia or acute myeloid leukemia cell linesin vitro. We demonstrate that knockout of the self-expressed antigen, CD19, boosts anti-CD19 bispecific secretion by ePCs and prevents self-targeting. Further, anti-CD19 bispecific-ePCs elicited tumor eradicationin vivofollowing local delivery in flank-implanted Raji lymphoma cells. Finally, immunodeficient mice engrafted with anti-CD19 bispecific-ePCs and autologous T cells potently preventedin vivogrowth of CD19+acute lymphoblastic leukemia in patient-derived xenografts. Collectively, these findings support further development of ePCs for use as a durable, local delivery system for the treatment of acute leukemias, and potentially other cancers.Key pointsUsing gene editing, we engineered human plasma cells that secrete functional bispecifics to target leukemia cells expressing CD19 or CD33Engineered plasma cells secreting bispecifics suppress patient-derived leukemia in immunodeficient mice

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

“…Challenges to B cell medicine approaches include inefficient transducOon, achieving protein producOon at a level likely to be therapeuOc, and manufacturing at scale. 20 However, recent breakthroughs in B cell ediOng 14,[21][22][23][24] using CRISPR/Cas9 coupled with adeno-associated virus mediated delivery of a homology directed repair template enables the efficient introducOon of transgenes into safe-harbor loci (e.g., CCR5) or the immunoglobulin locus. [22][23][24][25] Advances have also been seen with lenOviral systems.…”

“…20 However, recent breakthroughs in B cell ediOng 14,[21][22][23][24] using CRISPR/Cas9 coupled with adeno-associated virus mediated delivery of a homology directed repair template enables the efficient introducOon of transgenes into safe-harbor loci (e.g., CCR5) or the immunoglobulin locus. [22][23][24][25] Advances have also been seen with lenOviral systems. 21 These techniques produce stable expression at levels predicted to be clinically significant.…”

“…[25][26][27] Furthermore, xenotransplant models confirm that edited human plasma cells derived from ex vivo expanded B cells can be detected for up to a year following infusion and conOnue to produce human immunoglobulins detectable in the plasma. 14 As a result of these discoveries, work is underway to engineer B cells to produce cloong factors, 23 immunotherapeuOcs, 22 pathogen-specific anObodies for HIV and RSV, 21,25,26 and viral inhibitors. 25 TradiOonally, the mouse is the plaaorm of choice for pre-clinical experimentaOon, including studies to date on the adopOve transfers of engineered B lineage cells, due to ease of manipulaOon and wellcharacterized geneOc background.…”

“…25–27 Furthermore, xenotransplant models confirm that edited human plasma cells derived from ex vivo expanded B cells can be detected for up to a year following infusion and continue to produce human immunoglobulins detectable in the plasma. 14 As a result of these discoveries, work is underway to engineer B cells to produce clotting factors, 23 immunotherapeutics, 22 pathogen-specific antibodies for HIV and RSV, 21,25,26 and viral inhibitors. 25…”

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

“…Recently, we developed a cell-based method to deliver protein drugs, which we have successfully tested in immune-deficient mice. To do this, we generated ex vivo differentiated human plasma cells (PCs) and engineered them to produce protein drugs (including bispecific antibodies), and in immuno-deficient mice, we observed long-lasting antibody secretion for a year and potent tumor killing [1][2][3][4] .…”