This article provides an update of the latest data and developments within the CATH protein structure classification database (http://www.cathdb.info). The resource provides two levels of release: CATH-B, a daily snapshot of the latest structural domain boundaries and superfamily assignments, and CATH+, which adds layers of derived data, such as predicted sequence domains, functional annotations and functional clustering (known as Functional Families or FunFams). The most recent CATH+ release (version 4.2) provides a huge update in the coverage of structural data. This release increases the number of fully- classified domains by over 40% (from 308 999 to 434 857 structural domains), corresponding to an almost two- fold increase in sequence data (from 53 million to over 95 million predicted domains) organised into 6119 superfamilies. The coverage of high-resolution, protein PDB chains that contain at least one assigned CATH domain is now 90.2% (increased from 82.3% in the previous release). A number of highly requested features have also been implemented in our web pages: allowing the user to view an alignment between their query sequence and a representative FunFam structure and providing tools that make it easier to view the full structural context (multi-domain architecture) of domains and chains.

Although several ATR inhibitors are in development, there are unresolved questions regarding their differential potency, molecular signatures of cancer patients for predicting activity and most effective therapeutic combinations. Here, we elucidate how to improve ATR-based chemotherapy with the newly developed ATR inhibitor, M4344 using in vitro and in vivo models. The potency of M4344 was compared with the clinically developed ATR inhibitors BAY1895344, berzosertib, and ceralasertib. The anticancer activity of M4344 was investigated as monotherapy and combination with clinical DNA damaging agents in multiple cancer cell lines, patient-derived tumor organoids and mouse xenograft models. We also elucidated the anticancer mechanisms and potential biomarkers for M4344. We demonstrate that M4344 is highly potent among the clinically developed ATR inhibitors. Replication stress (RepStress) and neuroendocrine (NE) gene expression signatures are significantly associated with a response to M4344 treatment. M4344 kills cancer cells by inducing cellular catastrophe and DNA damage. M4344 is highly synergistic with a broad range of DNA-targeting anticancer agents. It significantly synergizes with topotecan and irinotecan in patient-derived tumor organoids and xenograft models. Taken together, M4344 is a promising and highly potent ATR inhibitor. It enhances the activity of clinical DNA damaging agents commonly used in cancer treatment including topoisomerase inhibitors, gemcitabine, cisplatin and talazoparib. RepStress and NE gene expression signatures can be exploited as predictive markers for M4344.

Free fatty acid receptor 4 (FFA4), previously known as GPR120, is a G protein-coupled receptor that promotes numerous anti-inflammatory and antidiabetic effects upon its agonism by long chained unsaturated fatty acids. We have previously demonstrated that agonism of FFA4 with docosahexaenoic acid (DHA) and alpha-linoleic acid (ALA) facilitates rapid and transient phosphorylation of FFA4 expressed ectopically on the surface of HEK293 cells. However, the precise mechanisms that promote FFA4 phosphorylation remain elusive. In the current study, we examined the mechanisms behind both heterologous and homologous phosphorylation of FFA4 and set out to identify the foci of FFA4 phosphorylation. Our results demonstrate that basal and heterologous phosphorylation of FFA4 are mediated by protein kinase C (PKC), while G protein-coupled receptor kinase 6 (GRK6) plays the predominant role in DHA-mediated phosphorylation of FFA4. Furthermore, we identify Thr347, Ser350, and Ser357 in the C-terminal tail as major sites of FFA4 phosphorylation. Concurrent mutation of these three sites leads to a FFA4 receptor that seemingly affects Gαq/11 signaling in a positive manner as demonstrated by heightened intracellular Ca+2 responses following agonism with DHA. Importantly, this phosphodefective FFA4 mutant lacked the ability to promote β -arrestin-2 recruitment to the cell membrane. Since many of the functionally beneficial physiological effects of FFA4 are noted to be β -arrestin mediated, these findings could provide insight into the structural requirements for FFA4 function.