Detection of Cell-Cell Interactions via Photocatalytic Cell Tagging

The growing appreciation of immune cell-cell interactions within disease environments has led to significant efforts to develop highly effective protein-, and cell-based immunotherapies.  However, characterizing these complex cell-cell interactions in high resolution remains challenging.  Thus, technologies that leverage therapeutic-based modalities for profiling intercellular environments can provide unique advantages towards understanding these cellular interactions at molecular-level detail.  To address this, we introduce photocatalytic cell tagging (PhoTag), a platform for profiling cell-cell interactions that utilizes a single domain antibody (VHH) conjugated to a photoactivatable flavin-based cofactor.  Upon irradiation with visible light, the tethered flavin photocatalyst generates phenoxy radical tags for targeted labeling within cell-cell contact environments.  Using anti-PD-1 or anti-PD-L1 VHH flavin conjugates, we demonstrate that PhoTag achieves highly selective synaptic labeling in antigen presenting cell-T cell co-culture systems. By combining the high resolution transcellular biotinylation capability of PhoTag with multi-omics single cell sequencing, we interrogated transient interactions between Peripheral blood mononuclear cell (PBMC) populations and Raji PD-L1 B cells and discovered that specific T cell subtypes can transiently interact more efficiently than others.   We envision that the spatio-temporal and modular nature of PhoTag will enable its broad utilization for detailed profiling of intercellular interactions across different biological systems. For the single cell sequencing aspect of this study, 3 donors were used with samples from each donor split into the different treatment groups (Isotype, PD-L1, and LFA-1 inhibitor). Isotype labeling serves as the reference samples.