Discovery and Characterization of PROTACs Targeting Tissue Transglutaminase (TG2).

Tissue transglutaminase (TG2) is a multifunctional enzyme involved in the cross-linking of extracellular matrix proteins, formation of complexes with fibronectin (FN) and integrins, and GTP hydrolysis. TG2 is activated in several pathological conditions, including cancer. We recently described a novel series of ligands that bind to TG2 and inhibit its interaction with FN. Because TG2 acts via multiple mechanisms, we set out to pursue a targeted protein degradation strategy to abolish TG2's myriad functions. Here, we report the synthesis and characterization of a series of VHL-based degraders that reduce TG2 in ovarian cancer cells in a proteasome-dependent manner. Degradation of TG2 resulted in significantly reduced cancer cell adhesion and migration in vitro in scratch-wound and migration assays. These results strongly indicate that further development of more potent and in vivo efficient TG2 degraders could be a new strategy for reducing the dissemination of ovarian and other cancers.

Transcriptional Repression by FoxM1 Suppresses Tumor Differentiation and Promotes Metastasis of Breast Cancer.

The transcription factor Forkhead box M1 (FoxM1) is overexpressed in breast cancers and correlates with poor prognosis. Mechanistically, FoxM1 associates with CBP to activate transcription and with Rb to repress transcription. Although the activating function of FoxM1 in breast cancer has been well documented, the significance of its repressive activity is poorly understood. Using CRISPR-Cas9 engineering, we generated a mouse model that expresses FoxM1-harboring point mutations that block binding to Rb while retaining its ability to bind CBP. Unlike FoxM1-null mice, mice harboring Rb-binding mutant FoxM1 did not exhibit significant developmental defects. The mutant mouse line developed PyMT-driven mammary tumors that were deficient in lung metastasis, which was tumor cell-intrinsic. Single-cell RNA-seq of the tumors revealed a deficiency in prometastatic tumor cells and an expansion of differentiated alveolar type tumor cells, and further investigation identified that loss of the FoxM1/Rb interaction caused enhancement of the mammary alveolar differentiation program. The FoxM1 mutant tumors also showed increased Pten expression, and FoxM1/Rb was found to activate Akt signaling by repressing Pten. In human breast cancers, expression of FoxM1 negatively correlated with Pten mRNA. Furthermore, the lack of tumor-infiltrating cells in FoxM1 mutant tumors appeared related to decreases in pro-metastatic tumor cells that express factors required for infiltration. These observations demonstrate that the FoxM1/Rb-regulated transcriptome is critical for the plasticity of breast cancer cells that drive metastasis, identifying a prometastatic role of Rb when bound to FoxM1. SIGNIFICANCE: This work provides new insights into how the interaction between FoxM1 and Rb facilitates the evolution of metastatic breast cancer cells by altering the transcriptome.