0994: UTAA91: An Off-the-Shelf Allogeneic CAR-Vδ1T Therapy Leveraging CD19 Nanoantibody for B-cell Depletion and PD-1 Agonist Nanoantibody for Pathogenic T-cell Elimination in Refractory Autoimmune Diseases

Background/Purpose: Current therapies inadequately restore immune homeostasis due to incomplete pathogenic cell clearance and limited multi-lineage targeting. UTAA91 integrates:

1) PD-1 agonist-targeted cytotoxicity: The PD-1 nanoantibody module directs Vδ1T cells to selectively eliminate PD-1+ pathogenic T cells via their potent killing capacity: includng follicular helper T cells (Tfh), peripheral helper T cells (Tph) and effectors;

2) CD19-targeted B-cell clearance: Concurrent CD19 nanoantibody-mediated B-cell depletion disrupts T-B cell collaboration;

3) Vδ1T cells for deep tissue penetration.

Methods: Manufacturing: Vδ1T cells from donor PBMCs were enriched via MACS, engineered with CD19/PD1-CAR (Chimeric antigen receptor), and Fas-knockout via CRISPR-Cas9 to mitigate HvGR;

In vitro: PD-1+ T-cell/B-cell depletion quantified in SLE/RA/SS patient PBMCs;

In vivo: Humanized NPG mice with CD19+ tumors assessed for HvGR suppression, PD1+ T-cell clearance, tumor inhibition (flow cytometry/IVIS), and safety.

Results: The cGMP-compliant manufacturing process for UTAA91 demonstrates exceptional industrial feasibility, achieving the following critical quality attributes: Vδ1T cell purity >90%, stable CAR molecule expression rate >30%, Fas gene-specific knockout efficiency >90%, expansion capability exceeding 10,000-fold, Tnaive subset proportion >90%, and sustained high expression of NKG2D/DNAM1 and chemokine receptors, establishing a robust cellular foundation for long-term immune regulation.

In vitro functional validation confirmed UTAA91’s dual-targeted cytotoxicity against CD19+ and PD1+ cells, maintaining >90% specific killing efficiency through multiple rounds of antigen stimulation, which highlights its superior functional persistence. In humanized mouse models, UTAA91 effectively overcame allogeneic therapy barriers by significantly suppressing host-versus-graft reaction (HvGR), clearing peripheral PD1+ T cells, and inhibiting tumor growth, with no treatment-related toxicity observed.

Of significant translational value, UTAA91 exhibited precise and potent depletion of pathogenic cells in PBMCs derived from autoimmune disease patients (including SLE, RA, and SS), specifically eliminating CD19+ B cells and Tfh/Tph populations while selectively removing hyperactivated CD4+PD1+and CD8+PD1+T cells. Notably, functional PD1-T cell subsets were preserved post-treatment. This marks the first preclinical demonstration of UTAA91’s unique capability to simultaneously target bidirectional T-B cell pathogenic pathways, offering a transformative therapeutic strategy for autoimmune diseases.

Conclusion: UTAA91, the first dual-targeting allogeneic UCAR-Vδ1T therapy, pioneers a "dual depletion" mechanism to eliminate B and Tfh/Tph cells, overcoming T-B interaction limitations and offering a groundbreaking solution for autoimmune diseases.