“Zero-On” NIR-II Photoacoustic Organic Nanoprobes for Ultra-Accurate Companion Diagnostics of Cancer Immunotherapy Response

Companion diagnostics (CDx) employing biomarker-responsive “off-on” probes enable imaging-guided personalized cancer immunotherapy. However, CDx accuracy is compromised by even weak off-state probe background signals, which are amplified by intratumoral accumulation and can be indistinguishable from biomarker-activated signals. Here, we report a “zero-on” near-infrared-II (1000–1700 nm) photoacoustic semiconducting polymer nanoprobe (SPCDx) that switches from a nonconjugated, near-infrared-II-signal-silent “zero” state to a π-conjugated, signal-active “on” state upon exposure to immunoreactive biomarkers. Ingeniously nonconjugated polymer design achieves “zero” near-infrared-II probe background ( p > 0.05 vs water), eliminating probe background interference after intratumoral accumulation and ensuring detected signals in the tumor exclusively from true biomarker activation. In vivo, SPCDx accurately distinguishes immune drugs of varying efficacy and identifies low-efficacy immune response mice with 100% accuracy in a blind study, outperforming conventional “off-on” probes (56.3%). SPCDx further accurately detects early immune responses >42 h before clinical-standard flow cytometry biopsies and 6 days before observable tumor shrinkage. Our “zero-on” concept offers a molecular blueprint for ultra-accurate CDx of cancer immunotherapy response.