Abstract
BackgroundThe Musashi (MSI) family of RNA-binding proteins is best known for the role in post-transcriptional regulation of target mRNAs. Elevated MSI1 levels in a variety of human cancer are associated with up-regulation of Notch/Wnt signaling. MSI1 binds to and negatively regulates translation of Numb and APC (adenomatous polyposis coli), negative regulators of Notch and Wnt signaling respectively.MethodsPreviously, we have shown that the natural product (−)-gossypol as the first known small molecule inhibitor of MSI1 that down-regulates Notch/Wnt signaling and inhibits tumor xenograft growth in vivo. Using a fluorescence polarization (FP) competition assay, we identified gossypolone (Gn) with a > 20-fold increase in Ki value compared to (−)-gossypol. We validated Gn binding to MSI1 using surface plasmon resonance, nuclear magnetic resonance, and cellular thermal shift assay, and tested the effects of Gn on colon cancer cells and colon cancer DLD-1 xenografts in nude mice.ResultsIn colon cancer cells, Gn reduced Notch/Wnt signaling and induced apoptosis. Compared to (−)-gossypol, the same concentration of Gn is less active in all the cell assays tested. To increase Gn bioavailability, we used PEGylated liposomes in our in vivo studies. Gn-lip via tail vein injection inhibited the growth of human colon cancer DLD-1 xenografts in nude mice, as compared to the untreated control (P < 0.01, n = 10).ConclusionOur data suggest that PEGylation improved the bioavailability of Gn as well as achieved tumor-targeted delivery and controlled release of Gn, which enhanced its overall biocompatibility and drug efficacy in vivo. This provides proof of concept for the development of Gn-lip as a molecular therapy for colon cancer with MSI1/MSI2 overexpression.
Highlights
The Musashi (MSI) family of RNA-binding proteins is best known for the role in post-transcriptional regulation of target mRNAs
Ki values were calculated based on the Kd and the dose-response curves. b Surface plasmon resonance (SPR) analyses of Gn binding to immobilized GB1-tagged MSI1-RNA-Binding Domain 1 (RBD1)
Four RNA binding residues undergo line broadening upon addition of Gn indicating that they are involved in binding to Gn. d Docked model of MSI1 RBD1 bound to gossypolone
Summary
The Musashi (MSI) family of RNA-binding proteins is best known for the role in post-transcriptional regulation of target mRNAs. Elevated MSI1 levels in a variety of human cancer are associated with up-regulation of Notch/Wnt signaling. The expression of the RNA-binding protein Musashi-1 (MSI1) is elevated in a variety of human cancers, including glioblastoma, breast, colon and lung cancers [1,2,3,4,5,6,7,8,9,10], with higher levels corresponding to poor prognosis [3,4,5, 10,11,12]. Our previous study identified (−)-gossypol as a small molecule inhibitor of MSI1 that reduced cancer cell proliferation and xenograft growth [27]. Gn can potentially be used as a MSI1/2 dual inhibitor
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