1H, 13C, and 15N assignments of the mRNA binding protein hnRNP A18.

Heterogeneous ribonuclear protein A18 (hnRNP A18) is an RNA binding protein (RBP) involved in the hypoxic cellular stress response and regulation of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression in melanoma, breast cancer, prostate cancer, and colon cancer solid tumors. hnRNP A18 is comprised of an N-terminal structured RNA recognition motif (RMM) and a C-terminal intrinsically disordered domain (IDD). Upon cellar stressors, such as UV and hypoxia, hnRNP A18 is phosphorylated by casein kinase 2 (CK2) and glycogen synthase kinase 3ß (GSK-3ß). After phosphorylation, hnRNP A18 translocates from the nucleus to the cytosol where it interacts with pro-survival mRNA transcripts for proteins such as hypoxia inducible factor 1α and CTLA-4. Both the hypoxic cellular response and modulation of immune checkpoints by cancer cells promote chemoradiation resistance and metastasis. In this study, the 1 H, 13 C, and 15 N backbone and sidechain resonances of the 172 amino acid hnRNP A18 were assigned sequence-specifically and provide a framework for future NMR-based drug discovery studies toward targeting hnRNP A18. These data will also enable the investigation of the dynamic structural changes within the IDD of hnRNP A18 upon phosphorylation by CK2 and GSK-3ß to provide critical insight into the structure and function of IDDs.

Small molecules inhibitors of the heterogeneous ribonuclear protein A18 (hnRNP A18): a regulator of protein translation and an immune checkpoint.

We have identified chemical probes that simultaneously inhibit cancer cell progression and an immune checkpoint. Using the computational Site Identification by Ligand Competitive Saturation (SILCS) technology, structural biology and cell-based assays, we identify small molecules that directly and selectively bind to the RNA Recognition Motif (RRM) of hnRNP A18, a regulator of protein translation in cancer cells. hnRNP A18 recognizes a specific RNA signature motif in the 3'UTR of transcripts associated with cancer cell progression (Trx, VEGF, RPA) and, as shown here, a tumor immune checkpoint (CTLA-4). Post-transcriptional regulation of immune checkpoints is a potential therapeutic strategy that remains to be exploited. The probes target hnRNP A18 RRM in vitro and in cells as evaluated by cellular target engagement. As single agents, the probes specifically disrupt hnRNP A18-RNA interactions, downregulate Trx and CTLA-4 protein levels and inhibit proliferation of several cancer cell lines without affecting the viability of normal epithelial cells. These first-in-class chemical probes will greatly facilitate the elucidation of the underexplored biological function of RNA Binding Proteins (RBPs) in cancer cells, including their effects on proliferation and immune checkpoint activation.