A Biocompatible Cu(I)-Catalyzed Click Chemistry Framework for Long-Term Cell Tracking

Cellular immunotherapies have transformed oncologic treatment in recent years, yet the inability to monitor cell migration post-transfusion remains a significant barrier to their development. Here, we explore the feasibility of a non-invasive cell-tracking method utilizing Cu(I)-catalyzed click chemistry in which cells are first labeled with alkynes and, after 1-2 weeks, are selectively targeted via an azide-conjugated fluorescent probe. We developed Click1, a novel tris-Cu-chelating azide capable of a rapid, robust click reaction directly on live cell surfaces with minimal cytotoxicity. When paired with Click1, two distinct alkyne-labeling reagents, N-(4-pentynoyl)-mannosamine-tetraacylated (Ac4ManNAl) and propargyl choline, were capable of detecting human T cells for over two weeks. Our findings elucidate the metabolic turnover of biochemical handles in primary immune cells, providing a potential framework for long-term cell tracking in immunotherapies.