Targeting of RNA-binding protein FXR1 in HNSCC

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Abstract Head and neck squamous cell carcinoma (HNSCC) is the 6th most common cancer worldwide and accounts for more than 90% of head and neck cancers, with a 5-year survival rate of approximately 50%. Despite specific effective and novel therapies, most patients remain unresponsive to current treatment modalities and develop drug resistance. Therefore, identifying new molecules that target cancer pathways, alone or in combination, is critical for developing novel cancer therapies. In HNSCC, dysregulated RNA-binding proteins (RBPs) controls co- and post-transcriptional events. Approaches targeting RBPs are not well established to regulate the gene expression in cancers. Thus, new therapeutic strategies to control gene expression and tumor growth are necessary. Our published findings demonstrate that FXR1 directly binds the RNA G-quadruplex (G4) regions of mRNA CDKN1A and a non-coding RNA TERC (Telomerase RNA component) and promotes the growth and proliferation of HNSCC cells. Herein, our preliminary findings indicate that methylated arginine amino acids at the NES (nuclear export signal) and RGG (arginine-glycine rich) domain of FXR1 are responsible for G4-RNA binding and the growth of cancer cells. However, mutation of specific arginine residues in the NES/RGG domain abolished FXR1 binding with CDKN1A and TERC and reduced the growth and proliferation of oral cancer cells. Thus, targeting FXR1's RGG domain with specific inhibitors is considered an appealing drug target for oral cancer treatment. We plan to use nucleic acid-based aptamers that directly bind to the arginine amino acids present in NES/RGG domain of FXR1 and disrupt its RNA binding activities. Hence, the overall goal of this R21 proposal is to utilize structural and biochemical assays to screen and identify aptamers specific to the FXR1 NES/RGG domain to control its gene expression activities and the growth of oral cancer cells. Two specific aims are proposed to test this model. In specific aim1, we will determine how arginine methylation impacts FXR1 RNA- binding functions and oral tumor progression using in vitro RNA binding assays and in vivo tumor models. In specific aim2, we will develop an aptamer that directly binds and interfere with methylated FXR1-RNA complex and show anti-tumor activity in our preclinical animal models of HNSCC. Our present work is an innovative, straightforward, and robust approach to target the dysregulated RBP-mediated gene expression in HNSCC. We will use these mechanistic findings to develop novel strategies to treat aggressive HNSCC tumors.