Background: Recent clinical data suggests that autologous CAR-T cells are emerging as promising treatments for B-cell mediated autoimmune diseases, having demonstrated durable drug-free remissions in patients. Off-the-shelf allogeneic cell therapies promise to build on the efficacy of their autologous counterparts, while providing patients with a well-defined, scalable, and cost-efficient drug product that can be administered on-demand. Due to their ability to self-renew, induced pluripotent stem cells (iPSCs) provide an ideal platform to develop allogeneic cell therapies, allowing precision editing with desired characteristics that can be expanded, characterized, and banked for future differentiation to any desired cell type. Successful development requires complex engineering to prevent GvHD, rejection by the patient’s immune system, and armoring the cells with an antigen targeting moiety (such as a chimeric antigen receptor). Here, we describe preclinical development of iPSC-derived γδ CAR-T, αβ CAR-T, and CAR-NK cell platforms for use in autoimmune diseases.

Objectives: Our objective is to demonstrate the potency and efficacy of Century Therapeutic’s different iPSC-derived platforms (γδ CAR-T, αβ CAR-T, and CAR-NK) by in vitro and in vivo B cell aplasia.

Methods: iPSCs were reprogrammed from peripheral γδ T cells, peripheral blood mononuclear cells, or human umbilical cord blood CD34+ cells. iPSCs were engineered using either MAD7 or CRISPR/Cas9 to express a CD19-targeting (FMC63) CAR and Allo-Evasion ^TM edits. Engineered iPSCs were differentiated into hematopoietic progenitor cells and then subsequently differentiated into γδ CAR-T cells, αβ CAR-T cells, and CAR-NK cells. PBMCs were isolated from leukopaks donated by normal healthy donors from which B cells were subsequently isolated for in vitro assays. Acute potency of iPSC-derived γδ CAR-T cells, αβ CAR-T cells, and CAR-NK cells toward healthy B cells and Nalm6 tumor cells was assessed in single-round killing assays. Effector cells and target cells were plated at multiple effector-to-target (E:T) ratios, and incubated for 24 hours, at which point cells were stained and analyzed by flow cytometry for target cell cytolysis. A PBMC-humanized NOD SCID Gamma (NSG) mouse model was utilized to test the B cell killing capacity of iPSC-derived γδ CAR-T cells, αβ CAR-T cells, and CAR-NK cells. Mice were intravenously engrafted with PBMCs, and 14 days later received a single intravenous injection of γδ CAR-T cells, αβ CAR-T cells, and CAR-NK cells. One week later, mice were euthanized, and blood and tissues were collected for flow cytometry detection of B cells.

Results: iPSC-derived γδ CAR-T, αβ CAR-T, and CAR-NK cells showed strong potency against both healthy donor B cells and Nalm6 target cells with significant killing observed across all platforms. Mouse blood and tissues were evaluated by FACS for the presence of human B cells following CD19 CAR effector cell treatment. Compared to untreated humanized mice, B cell depletion was observed in blood and tissues of γδ CAR-T, αβ CAR-T, and CAR-NK treated mice, at levels equivalent to that observed in primary CD19 CAR-T treated mice. Splenic B cell depletion was incomplete but nearly equivalent across all treatment groups, including primary CAR-T cells.

Conclusion: Here, we demonstrate iPSC-derived γδ CAR-T, αβ CAR-T, and CAR-NK cells are potent cell therapies against CD19-expressing cells, each displaying antigen-dependent cytotoxicity. The data presented provide support for the use of these platforms for the development of allogeneic CAR cell medicines against B-cell mediated autoimmune diseases. The unique attributes of each platform (γδ CAR-T, αβ CAR-T, and CAR-NK) could potentially be harnessed for different disease indications (e.g., tissue distribution, expansion, and persistence).

REFERENCES: NIL.

Acknowledgements: NIL.

Disclosure of Interests: Jonathan Kurtz Century Therapeutics, Daniel Perry Century Therapeutics, Dar Heinze Century Therapeutics, Liam Campion Century Therapeutics, Mark Mendonca Century Therapeutics, Diana Chin Century Therapeutics, Buddha Gurung Century Therapeutics, Chen-Yuan Kao Century Therapeutics, Arina Perez Century Therapeutics, Hyam Levitsky Century Therapeutics, Chad Cowan Century Therapeutics, G. Grant Welstead Century Therapeutics.

© The Authors 2025. This abstract is an open access article published in Annals of Rheumatic Diseases under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ ). Neither EULAR nor the publisher make any representation as to the accuracy of the content. The authors are solely responsible for the content in their abstract including accuracy of the facts, statements, results, conclusion, citing resources etc.