Abstract 61: Sequence and structure-guided approach to identify functional mutations in G-protein coupled receptors
Authors: Sengupta Sohini , Ye Kai , Scott Adam D. , Niu Beifang , Bailey Matthew H. , McLellan Michael D. , Wendl Michael C. , Wyczalkowski Matthew A. , Ding Li 
Published: August 2015

Abstract
G-protein coupled receptors (GPCRs) account for about 4% of all encoded genes in the human genome with over 800 different types. They activate signal transduction pathways inside the cell that regulate a wide variety of cellular responses and physiological processes. GPCRs are known to play a role in disease progression and are the target of about 40% of drugs on market. However, the implication of GPCRs in tumor initiation and/or progression has not been extensively studied and remains unknown. Due to GPCRs being major drug targets, identifying functional mutations in GPCRs that may lead to tumorigenesis has large therapeutic implications. Nearly 20% of human tumors harbor mutations in GPCRs. However, individual GPCRs may harbor few recurrent mutations making it hard to detect potential functional mutations. Due to this lack of statistical power, it is beneficial to analyze the protein family as a whole and look at recurrent mutations at the same structural site. We can use sequence conservation, protein structure, and cancer mutation data to investigate the mutational landscape of GPCRs and identify conserved mutation hotspots that might be involved in tumorigenesis.
We use the positions of structural domains to guide our protein family alignment. Entropy scores, which are used to measure conservation, and somatic mutation densities are computed based on the structure-guided alignment for each position in each domain. Positions that exhibit high somatic mutation density and low entropy are prioritized as possible functional mutations. We analyzed GPCRs from 3,281 tumor samples in over 12 cancer types and have identified three residues located in highly conserved motifs: the DRY, NPxxY, and CWxP motifs that may be involved in the activation of oncogenic pathways. The arginine residue in the DRY motif serves as an “ionic lock”, which keeps GPCRs in an inactive state. Mutations at the arginine could disrupt the inactive conformation leading to a ligand-independent active form and constitutive activation. The NPxxY and CWxP motifs are known to control the equilibrium between the inactive and active states of GPCRs.
We plan to further develop our methodology by integrating specific amino acid changes and mutation expression levels during the prioritization process. We will generalize our methodology into a tool called AnaConDAS (Analysis of Conservation in Domain Alignments and Structure) for major druggable protein families. Our lab has created a new tool called HotSpot3D, which clusters mutations and associated drugs based on proximity on a 3D protein structure. We plan to integrate AnaConDAS and HotSpot3D by focusing on the hotpot residues identified by AnaConDAS and analyzing mutations that cluster around them. This approach is novel because it harnesses the statistical power of studying mutations across a protein family and uses 3D protein structure/proximity based analyses to uncover functional mutations in cancer.
Citation Format: Sohini Sengupta, Kai Ye, Adam D. Scott, Beifang Niu, Matthew H. Bailey, Michael D. McLellan, Michael C. Wendl, Matthew A. Wyczalkowski, Li Ding. Sequence and structure-guided approach to identify functional mutations in G-protein coupled receptors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 61. doi:10.1158/1538-7445.AM2015-61