Uniting dual-modality of fluorescence and photoacoustic (PA) imaging into theranostic nanoprobes is imperative for spatio-temporally tracking of drug delivery,distribution,and release.Herein,we present a rational design strategy of molecularly precise amphiphilic prodrugs BPn-Cy-S-CPT (n=0,5,and 20,refers to the degree of polyethylene glycol (PEG) polymerization;CPT=camptothecin) to tune their self-assembly behaviour,innovatively integrating dual-modal PA and near-infrared (NIR)fluorescence imaging in a single-molecular framework.Among these elaborately designed prodrugs,it is found that only BP20-Cy-S-CPT could form uniform and highly stable self-assemblies,especially in showing synergistically enhanced PA and dual-channel NIR signals.In detail,PA signal is employed to trace the in vivo delivery with high spatial resolution,meanwhile the glutathione (GSH)-triggered dual-channel fluorescence response could real-timely monitor drug distribution and release without"blind spot".The results of in vivo dual-modal PA/NIR imaging have verified that BP2o-Cy-S-CPT displayed synergistic targeting (including passive,active,and activatable targeting) for tumor-specific delivery,and thereby executed CPT release in the tumor site.Consequently,our molecularly precise BP20-Cy-S-CPT self-assemblies could make a breakthrough to spatio-temporally track the in vivo drug release profile,expanding the intelligent theranostic toolbox for precise cancer treatment.