Mol Oncol. 2015 Aug;9(7):1406-20. doi: 10.1016/j.molonc.2015.03.014.

Natural product (-)-gossypol inhibits colon cancer cell growth by targeting RNA-binding protein Musashi-1.

 

Lan L1, Appelman C1, Smith AR1, Yu J2, Larsen S1, Marquez RT1, Liu H1, Wu X1, Gao P3, Roy A4, Anbanandam A5, Gowthaman R6, Karanicolas J6, De Guzman RN1, Rogers S7, Aubé J8, Ji M9, Cohen RS10, Neufeld KL1, Xu L11.
  • 1Department of Molecular Biosciences, The University of Kansas, Lawrence, KS, USA.
  • 2Department of Molecular Biosciences, The University of Kansas, Lawrence, KS, USA; School of Chemistry and Chemical Engineering, Southeast University, Nanjing, China.
  • 3COBRE Protein Production Group, The University of Kansas, Lawrence, KS, USA.
  • 4High Throughput Screening Laboratory, The University of Kansas, Lawrence, KS, USA.
  • 5Bio-NMR Core Facility, The University of Kansas, Lawrence, KS, USA.
  • 6Department of Molecular Biosciences, The University of Kansas, Lawrence, KS, USA; Center for Bioinformatics, The University of Kansas, Lawrence, KS, USA.
  • 7Center of Biomedical Research Excellence, Center for Cancer Experimental Therapeutics, The University of Kansas, Lawrence, KS, USA.
  • 8Center of Biomedical Research Excellence, Center for Cancer Experimental Therapeutics, The University of Kansas, Lawrence, KS, USA; Specialized Chemistry Center, The University of Kansas, Lawrence, KS, USA; Center for Chemical Methodologies and Library Development, The University of Kansas, Lawrence, KS, USA; Department of Medicinal Chemistry, The University of Kansas, Lawrence, KS, USA.
  • 9School of Chemistry and Chemical Engineering, Southeast University, Nanjing, China.
  • 10Department of Biological Sciences, Clemson University, Clemson, SC, USA.
  • 11Department of Molecular Biosciences, The University of Kansas, Lawrence, KS, USA; Department of Radiation Oncology, The University of Kansas Cancer Center, Kansas City, KS, USA. Electronic address: xul@ku.edu.

 

Abstract

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 P21(WAF-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.

KEYWORDS: Colon cancer; Musashi-1; Notch; RNA binding protein; Wnt

PMID: 25933687

 

Supplements

Cancer is among the top leading causes of mortality in the world and the second leading cause of death in the United States. Finding the right cancer therapies is imperative. With the development of next-generation gene sequencing and tissue array, targeted molecular therapy will be developed to fit the needs of individual patient. Precision medicine, or targeted molecular therapy will be more effective and have fewer side effects compared to traditional therapy because it deals with a common cause that drives cancer growth in that particular population.

Our goal is to find solutions for cancers with Musashi-1 (MSI-1) over-expression. MSI-1, a RNA-binding protein (RBP) that plays key roles in posttranscriptional gene regulation, is overexpressed in several different cancers. MSI-1 promotes Wnt and Notch signaling pathways by binding and inhibiting the translation of APC and NUMB, negative regulators of Wnt and Notch pathway, respectively (Figure 1). Our hypothesis is: if we block the RNA-binding function of MSI-1 with a small molecule, we can release APC and NUMB from their translation repression. Increased APC and NUMB protein will inhibit Wnt and Notch signaling pathway, thus inhibit tumor growth (Figure 2, (–)-gossypol as an example of a small molecule.).

 

Figure 1

Figure 1: In cells with MSI1 overexpression, MSI1 blocks the translation of NUMB and APC mRNA, which leads to the up-regulation of Notch/Wnt signaling. Thus MSI1 indirectly promotes proliferation/survival of the cells and inhibits cell death.(from the original paper, with permission from the publisher)

 

The significance of our study lies in two aspects. First, we screened and validated small molecular inhibitors of an RBP, and we are the first to report their anti-cancer activities associated with MSI-1 in vivo. Although there are many examples of compounds that specifically interfere with protein-protein interactions, there is limited success of drug discoveries for protein-RNA interactions, especially for RBPs that are classified as “undruggable”, potentially due to the lack of defined binding pockets. Our strategy may apply to many other important RBPs and will potentially revolutionize the drug discovery field by identifying and validating novel small molecule inhibitors targeting the so far undruggable RBPs.

Second, we discovered a new target of an existing compound (–)-gossypol, thus potentially expanding its scope of treatment. Discovering new therapeutic uses for existing molecules will shorten the time between bench to bedside. (–)-Gossypol has already completed phase IIb clinical trial for treating prostate cancer and several other cancers with over-expression of anti-apoptotic Bcl-2 family proteins. Our study suggested that (–)-gossypol would be beneficial for cancer patients with MSI1 over-expression.

 

Figure 2

Figure 2: In the presence of (–)-gossypol, (–)-gossypol binds to the RBD1 of MSI1, presumably releasing NUMB, APC and P21 mRNA from their translational repression. Increased level of Numb and APC protein will block Notch and Wnt signaling respectively. Increased P21 will block cell cycle progression. Thus (–)-gossypol-MSI1 complex indirectly blocks proliferation/survival of the cells. (from the original paper, with permission from the publisher)

 

 

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