Longitudinal cancer monitoring is crucial to clinical implementation of precision medicine. There is growing evidence indicating important functions of extracellular vesicles (EVs) in tumor progression and metastasis, including matrix remodeling via transporting matrix metalloproteases (MMPs). However, the clinical relevance of EVs remains largely undetermined, partially owing to challenges in EV analysis. Distinct from existing technologies mostly focused on characterizing molecular constituents of EVs, here we report a nanoengineered lab-on-a-chip system that enables integrative functional and molecular phenotyping of tumor-associated EVs. A generalized, high-resolution colloidal inkjet printing method was developed to allow robust and scalable manufacturing of three-dimensional (3D) nanopatterned devices. With this nanochip platform, we demonstrated integrative analysis of the expression and proteolytic activity of MMP14 on EVs to detect in vitro cell invasiveness and monitor in vivo tumor metastasis, using cancer cell lines and mouse models. Analysis of clinical plasma specimen showed that our technology could be used for cancer detection including accurate classification of age-matched controls and patients with ductal carcinoma in situ, invasive ductal carcinoma, or locally metastatic breast cancer in a training cohort (n = 30, 96.7% accuracy) and an independent validation cohort (n = 70, 92.9% accuracy). With clinical validation, our technology could provide a useful liquid biopsy tool to improve cancer diagnostics and real-time surveillance of tumor evolution in patients to inform personalized therapy.

Musashi‐1 (MSI1) is an RNA‐binding protein that acts as a translation activator or repressor of target mRNAs. The best‐characterized MSI1 target is Numb mRNA, whose encoded protein negatively regulates Notch signaling. Additional MSI1 targets include the mRNAs for the tumor suppressor protein APC that regulates Wnt signaling and the cyclin‐dependent kinase inhibitor P21WAF−1. We hypothesized that increased expression of NUMB, P21 and APC, through inhibition of MSI1 RNA‐binding activity might be an effective way to simultaneously downregulate Wnt and Notch signaling, thus blocking the growth of a broad range of cancer cells. We used a fluorescence polarization assay to screen for small molecules that disrupt the binding of MSI1 to its consensus RNA binding site. One of the top hits was (−)‐gossypol (Ki = 476 ± 273 nM), a natural product from cottonseed, known to have potent anti‐tumor activity and which has recently completed Phase IIb clinical trials for prostate cancer. Surface plasmon resonance and nuclear magnetic resonance studies demonstrate a direct interaction of (−)‐gossypol with the RNA binding pocket of MSI1. We further showed that (−)‐gossypol reduces Notch/Wnt signaling in several colon cancer cell lines having high levels of MSI1, with reduced SURVIVIN expression and increased apoptosis/autophagy. Finally, we showed that orally administered (−)‐gossypol inhibits colon cancer growth in a mouse xenograft model. Our study identifies (−)‐gossypol as a potential small molecule inhibitor of MSI1‐RNA interaction, and suggests that inhibition of MSI1's RNA binding activity may be an effective anti‐cancer strategy.