Cancer stem cells orchestrate immune evasion through extracellular vesicle-mediated non-canonical signaling pathways

Cancer stem cells (CSCs) within tumors exhibit a remarkable capacity to
evade immune surveillance, thereby escaping elimination by the host immune
system, yet the molecular mechanisms underlying this immune-evasive
plasticity remain incompletely defined. Compared to other breast cancer
subtypes, triple-negative breast cancer (TNBC) is particularly enriched
with CSCs exhibiting basal-like molecular features characterized by
elevated stemness indices, positioning it as an exemplary model for
investigating bidirectional CSCs-TME crosstalk. Here, we mapped the
spatial immunometabolic landscape of TNBC using high-dimensional flow
cytometry to simultaneously interrogate 50 TME-associated proteins at
single-cell resolution, including CSC markers tetraspanin-8 (TSPAN8) and
CD44. A total of 161 treatment-naïve TNBC specimens were analyzed,
stratified into CSC-high (CSCHi) and CSC-low (CSCLo) cohorts based on
TSPAN8 and CD44 protein expression thresholds. TNBCs with elevated
stemness indices drive effector-to-regulatory T cell (Treg) phenotypic
conversion, establishing immunosuppressive niches. Blocking EV-associated
TSPAN8 using an anti-TSPAN8 monoclonal antibody synergized with PD-1
checkpoint inhibition, providing preclinical validation for dual targeting
of CSC plasticity and immune checkpoint pathways. The raw and
processed data from high-dimensional single-cell proteomics analysis
performed on the BD FACSymphony Flow Cytometry system, as shown in Figures
1B, 2B, and 2G, have been deposited.