RT Journal Article
SR Electronic
T1 1196 Streamlining T cell engager development with a diverse panel of fully human CD3-binding antibodies, bispecific engineering technology, and an integrated discovery engine
JF Journal for ImmunoTherapy of Cancer
JO J Immunother Cancer
FD BMJ Publishing Group Ltd
SP A1240
OP A1240
DO 10.1136/jitc-2022-SITC2022.1196
VO 10
IS Suppl 2
A1 DeVorkin, Lindsay
A1 Mai, Juntao (Matt)
A1 Caldwell, Kate
A1 Jacobs, Tim
A1 Tonikian, Raffi
A1 Herve, Karine
A1 Hwang, Yuri
A1 Faralla, Cristina
A1 Wei, Wei
A1 Lathouwers, Emma
A1 Chappell, Rhys
A1 Hannie, Stefan
A1 Lam, Katherine
A1 Dhupar, Harveer
A1 Tran, Tran
A1 Cid, Melissa
A1 Bolten, Lena
A1 Pinsky, Tova
A1 Xiang, Ping
A1 Lai, Courtenay
A1 Lee, Ahn
A1 Chan, Patrick
A1 Chin, Jasmine
A1 Yamniuk, Aaron
A1 Dalal, Kush
A1 Barnhart, Bryan
YR 2022
UL http://jitc.bmj.com/content/10/Suppl_2/A1240.abstract
AB Background CD3 T cell engagers have the potential to be a cornerstone of immuno-oncology. However, a limited pool of CD3-binding antibodies and technological challenges in engineering bispecifics have hindered development. Discovering effective T cell engagers requires two target-binding arms — a CD3 arm that fine-tunes T cell activation and a tumor arm with high specificity for cancer cells — optimized as a whole to work in concert with each other. Beginning with diverse panels of antibodies increases the probability of finding appropriately potent and developable T cell engagers and reduces the need for downstream engineering.Methods We used microfluidic technology to screen more than 3.5 million single cells from humanized mice and identified >200 CD3-specific antibodies. Using high-throughput assays, we determined affinity for CD3εδ and CD3εγ, cross-reactivity to human and cyno primary T cells, CD3 binding kinetics, and epitope bins. We assessed T cell activation by measuring CD25 and CD69 expression by flow cytometry. We then used our bispecific engineering platform, OrthoMabTM, to generate a proof-of-concept panel of CD3 x EGFR bispecific antibodies. Developability properties were assessed, including hydrophobicity (aHIC), self-association (AC-SINS), polyspecificity (BVP-ELISA), stability (nanoDSF), and aggregation (aSEC). CD3 T cell engager potencies were measured using an NFAT reporter T cell activation assay and an xCELLigence tumor cell killing assay, and cytokine release was assessed by FLEXMAP CD.Results We identified hundreds of fully human CD3-specific antibodies that are diverse, developable, and validated. The antibodies displayed a wide range of CD3 binding affinities (KD ~1 nM to 1 μM), binding kinetics, and T cell activation potencies (EC50 ~6 to 190 nM). Data on this novel panel includes epitope binning, which revealed human and cyno CD3-binders that are distinct from previously described cross-reactive antibodies. The antibodies were assessed using a range of biophysical assays and have favorable developability properties. In an expanded proof-of-concept study, we used OrthoMabTM to generate a panel of CD3 x EGFR bispecific antibodies. The resulting bispecifics had favorable developability properties, and displayed a wide range of antigen-dependent T cell activation (EC50 ~2 pM to 2 nM) and tumor cell killing potencies (EC50 ~0.01 to 1 nM). From this panel, we identified potent T cell engagers that achieved >90% tumor cell killing with low levels of cytokine release.Conclusions By integrating our panel of CD3-binding antibodies with our bispecific engineering and high-throughput antibody assessment capabilities, we identified developable CD3 T cell engagers with potent tumor cell-killing activity and minimal cytokine release.