The glucoincretin hormone glucagon-like peptide-1 (GLP-1) increases pancreatic ␤-cell proliferation and survival through sequential activation of the epidermal growth factor receptor (EGFR), phosphatidylinositol-3 kinase (PI 3-kinase), and Akt. We investigated the role of transcription factor

Identification of low-dose, low-molecular-weight, drug-like inhibitors of protein–protein interactions (PPIs) is a challenging area of research. Despite the challenges, the therapeutic potential of PPI inhibition has driven significant efforts toward this goal. Adding to recent success in this area, we describe herein our efforts to optimize a novel purine carboxylic acid-derived inhibitor of the HDM2–p53 PPI into a series of low-projected dose inhibitors with overall favorable pharmacokinetic and physical properties. Ultimately, a strategy focused on leveraging known binding hot spots coupled with biostructural information to guide the design of conformationally constrained analogs and a focus on efficiency metrics led to the discovery of MK-4688 (compound 56), a highly potent, selective, and low-molecular-weight inhibitor suitable for clinical investigation.

Two novel methods were developed to stimulate B cells in rat whole blood, that have the potential to be used to support drug discovery efforts in the therapeutic targeting of B cells. Furthermore, we pharmacologically validated these whole blood assays using highly selective inhibitors of BTK, SYK, and PI3Kδ, signaling kinases which are downstream of the B cell receptor.

Natural killer cells are innate immune cells whose activity is gated by activating and inhibitory receptors that bind ligands on potential target cells. NK cell gene signatures are associated with improved survival in several indications, and NK cell therapies have shown some encouraging results in the clinic. These data highlight the relevance of this cell type in anti-tumor immunity and has given rise to interest in amplifying NK responses to cancer as a complementary therapeutic strategy to current T cell based immunotherapies. The goal of this project is to identify tumor cell intrinsic negative regulators of NK cell-mediated tumor cell killing through in vivo CRISPR/Cas9 screening, with identification of both PD-1 blockade-dependent and -independent regulators. The syngeneic B16F10 model in C57Bl6 mice was used with or without NK depletion and with or without anti-PD1 treatment, with a custom library of 900 genes split into four subpools (one guide per gene per subpool). B16F10 cells were transduced in vitro and inoculated into mice ten days after spinfection. On day 14 after inoculation, tumors were harvested and subsequently the gDNA was isolated and sequenced. QC analysis indicated good guide recovery and screen performance. There were several hits from this screen that were depleted in the NK+ context but not in the NK depleted context, indicating their potential role in regulating NK mediated cell killing. Of the hits, several expected genes scored including B2M which is a known negative regulator of NK cell killing. Validation efforts include confirmation of dropout in other models as well as in vitro co-culture assays to assess enhanced NK killing of single gene KO cells. The hits identified in the screen will increase our understanding of anti-tumor immunity and serve as potential novel Oncology targets. Citation Format: Kayla Duval, Elsa Krall, Federica Piccioni, Maria Monberg, Marianne Spatz, Tobias Ehrenberger, Nicole Follmer. In vivo functional genomics screen to uncover tumor intrinsic negative regulators of NK cell-mediated tumor cell killing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 227.