Background: Breach in central and peripheral tolerance mechanisms generates autoreactive B-cells that contribute to the pathogenesis of autoimmune rheumatological diseases like systemic lupus erythematosus (SLE), myositis and others. Recently, promising remission rates compared to conventional therapies have been demonstrated in early clinical trials with bispecific CD3 T-cell engagers in refractory rheumatic indications. Toxicities including ICANS and CRS, often observed with CD3 T-cell engagers (TCE) in oncology patients, may limit therapeutic potential for such treatments. AZD5492 is an asymmetric, tri-specific, IgG1-based antibody which harbors two Fab binding domains to CD20, one VHH binding domain to TCR, and one VHH binding domain to CD8. AZD5492 is currently in Ph1 clinical trial in NHL and CLL patients as well as SLE and Myositis patients. We are reporting pre-clinical data in patient samples demonstrating the robust depletion potency of AZD5492 along with specific engagement and activation of CD8 T-cells while sparing CD4 T-cells provides an attractive opportunity for treatment of immune-mediated rheumatological diseases.

Objectives: We aimed to assess the AZD5492-mediated depletion of B cells in patient-derived PBMCs with preferential activation of CD8 T cells.

Methods: PBMCs isolated from healthy donors (HDs) and patients with SLE/myositis/RA were used for in vitro assays to determine B-cell cytolysis and T-cell (both CD8 and CD4) activation 72 hours post-AZD5492 treatment. Cytokine release was measured from supernatants collected from the treated PBMCs using multiplex technology (MesoScale Discovery). CD20xCD3 targeting bi-specific TCE was used as a comparator for in vitro assays. For in vivo experiments, humanized NOD scid gamma (NSG) mice, which had been engrafted with CD34+ hematopoietic stem cells, were treated with a single dose of AZD5492 for 72 hours. B-cell depletion was measured via flow cytometry in the blood and tissues (bone marrow, spleen, thymus, kidney, and lymph nodes) collected after dosing.

Results: In PBMCs samples from HDs and patients (SLE, myositis and RA), specific B-cell cytolysis was observed with AZD5492 and demonstrated superior potency compared to comparator TCE. T-cell activation was specifically biased towards CD8+ T-cells after treatment with AZD5492, while minimal CD4+ T-cell activation was observed. In contrast, both CD8+ and CD4+ T-cell activation was observed with comparator TCE-mediated depletion of B-cells. In addition, treatment with AZD5492 induced minimal cytokine release and T-cell activation in a non-antigen specific T-cell activation assay as compared to comparator TCE. The in vivo studies demonstrated that AZD5492 induced robust depletion of B cells in the peripheral blood and tissues.

Conclusions: Compared to conventional CD20xCD3 TCEs, which equally engage and activate both CD4+ and CD8+ T cells, AZD5492 drives potent cytolysis of B cells in a CD8-T cell-biased manner. Sparing the activation of CD4+ T-cells and associated secretion of cytokines could potentially lead to a better safety profile; based on current knowledge, AZD5492 is the only TCE molecule to do so in autoimmune disease indications. AZD5492 represents a best-in-class TCE of its kind in autoimmune indications with opportunity for tolerability, efficacy, and safety compared to its competitors. Phase 1 clinical evaluation in SLE, myositis and RA patients is ongoing (NCT06916806).

REFERENCES: NIL.

Acknowledgments: NIL.

Disclosure of Interests: Naresh Kumar Astrazeneca, Astrazeneca, Becca Ursin Astrazeneca, Astrazeneca, Karin Golan Astrazeneca, Astrazeneca, Yuanyue Zhang Astrazeneca, Astrazeneca, Brian Naiman Astrazeneca, Astrazeneca, Zakia Goodwin Astrazeneca, Astrazeneca, Ariful Qadri Astrazeneca, Astrazeneca, Supinya Iamsawat Astrazeneca, Astrazeneca, Alex Estrada Astrazeneca, Astrazeneca, Shonda Hawkins Astrazeneca, Astrazeneca, Francoise Meylan Astrazeneca, Astrazeneca, Tilbe Creigh-pulatmen Astrazeneca, Astrazeneca, Abigail Lara Astrazeneca, Astrazeneca, Nazli Jafarzadeh Astrazeneca, Astrazeneca, Thomas Ciucci Astrazeneca, Astrazeneca, Corinne Cayatte Astrazeneca, Astrazeneca, David Close Astrazeneca, Astrazeneca, David Rowlands Astrazeneca, Astrazeneca, Tatiana Ort Astrazeneca, Astrazeneca.