Integral membrane proteins remain difficult antibody targets because they often lack soluble, native antigens for screening. This study addresses that hurdle by coupling in vivo immunization (humanized transgenic or wild-type mice and llamas) with Adimab’s yeast-based platform to discover antibodies against multiple membrane-obligate targets (MOTs), yielding large, clonally diverse panels of high-affinity IgGs and HCAbs.

Approach and outcomes

• The Adimab platform discovered antibodies against multiple MOT classes, including GPCRs and tetraspanins, as shown by cell staining on target-expressing CHO cells versus empty vector controls.
• Using CCR8 as a case study, we demonstrated efficient recovery of 57 CCR8-specific antibodies and confirmed cell binding on human donor tumor-infiltrating T cells at 100 nM.
• The CCR8 antibodies were functionally active, antagonizing CCL1 signaling in a GPCR biosensor BRET assay and mediating antibody-dependent cellular cytotoxicity in a luciferase reporter system.
• After humanization, the CCR8-specific antibodies demonstrated favorable developability profiles, with consistent SEC, high Tm, short HIC retention times, and low PSR binding when benchmarked against clinical-stage controls.
• Integrating immunized llama VHH repertoires enabled a yeast immune library that yielded 98 unique HCAbs to a model GPCR, including triple cross-reactivity to human, cynomolgus monkey, and mouse, with good developability profiles and output diversity.

The data show that pairing in vivo diversities with a yeast-based discovery engine overcomes the soluble-antigen barrier for MOTs by delivering functional CCR8 antagonists in one case study and cross-reactive HCAbs to a model GPCR in another. Finally, this workflow allows the scientist control over the selection of developability profiles suitable for therapeutic leads. 

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