Monitoring melanoma immunotherapeutic efficacy through successive metabolic labeling and PD-L1-confined signal amplification based on membrane characteristics of the newly generated circulating tumor cells.

Unlike conventional detection, monitoring membrane characteristics of circulating tumor cells (CTCs) within a specific time frame can effectively indicate tumor progression, yet the challenge lies in selectively isolating CTCs generated during this period and precisely identifying subtle progression-related changes. This study focuses on CTCs newly generated during melanoma immunotherapy, utilizing a strategic combination of tumor-specific glycometabolic engineering, phenotypic protein-confined biotinylation and lanthanide luminescence to enrich and detect these spatiotemporally specific CTCs. First, Ac4ManNAz-associated cellular glycometabolic engineering selectively developes azide groups on the membranes of these CTCs, providing clickable artificial tags to distinguish them from pre-existing CTCs and blood cells. Subsequently, Fe3O4/lip-DBCO nanoparticles enrichs this CTC population through efficient click chemistry, achieving a capture efficiency exceeding 90 % for various types of CTCs, regardless of their phenotype, tumor type or species. Following this, a PD-L1-confined biotinylation is adopted to generate a significant number of biotin moieties in close proximity to the membrane PD-L1 of the captured CTCs, resulting in a 9-fold increase in the number of active sites available for introducing detection signal sources, compared to traditional immunofluorescent labeling methods. Further introduction of streptavidin-functionalized europium (Eu)-lanthanide nanoparticles (EuNPs-SA) quantifies the captured CTCs in a time-resolved manner and assesses CTC-related therapeutic response, while eliminating interferences from background biological substances. Specifically, significant advancements in detection performance have enabled the assessment of immunotherapeutic efficacy within melanoma model. Notably, substantial differences in lanthanide luminescent signals between the treated and untreated groups p = 0.000149), with the observed trends closely correlating with treatment outcomes.