Gamma-Delta (γδ) CAR-T Cells targeting CD33 and lacking the CD3ζ signaling domain enhance killing of AML cells without toxicity to normal hematopoietic cells

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Presentation Time: 03:15 PM - 04:30 PM
Poster Board Number: B958
Abstract ID: 7472

Presenting Author:
Lawrence Lamb , Chief Scientific Officer at IN8Bio

Abstract:

Extending CAR-T therapy to myeloid malignancies has proven challenging due to co-expression of targetable antigens on hematopoietic progenitors. We leveraged the enhanced targeting conferred by CAR-T technology with the broad stress-associated antigen recognition and killing capabilities of γδ T cells by employing a non-signaling CAR (nsCAR) that excludes the CD3ζ domain, thereby facilitating targeted tumor recognition while sparing healthy tissues. Jurkat cells expressing a CD3ζ+ CD33+ CAR showed significant upregulation of CD69 in co-culture with KG1a AML cells but nsCAR-expressing Jurkat cells did not upregulate CD69. A time-dependent reduction (43%) in the CD3ζ+CD33CAR+ Jurkat population was also noted over a 7-day coculture while the nsCAR+ Jurkat population remained stable, suggesting improved durability of the nsCAR modified Jurkat cells. nsCAR-expressing expanded and activated γδ T cells from healthy donors exhibited enhanced killing against AML lines HL-60 (1.3x)  and K-562 (1.6x) compared to unmodified γδ T cells (UTD) in a 24-hour cytotoxicity assay. Incorporation of sIL-15 also increased killing (1.8x for HL-60, 2.6x for KG-1a, 2.0x for MOLM-13 2.0x for K-562). Minimal cytotoxicity (<10%) was observed for nsCARs or UTD γδ T cells against CD34+ HPCs. Our findings suggest that combining nsCAR constructs with γδ T cells may widen the therapeutic window to enable of CAR-T therapy for cancers with target antigen expression on healthy tissues.