Introduction

Gastric chief cells can re-enter the cell cycle after injury through paligenosis, a conserved regenerative program regulated by mTORC1. This process involves (1) cellular degradation, (2) induction of progenitor genes (spasmolytic polypeptide-expressing metaplasia, SPEM), and (3) cell cycle re-entry. Dysregulated paligenosis contributes to gastric cancer, but mechanisms governing this reprogramming remain incompletely understood. Here, we investigated the role and regulation of YAP1, a key effector of the Hippo pathway, during paligenosis.

Aims & Methods

Our objectives were to determine whether YAP1 is activated during paligenosis, define its role, and identify upstream regulators. Using a mouse model of injury-induced paligenosis (via high-dose tamoxifen, HDTAM, intraperitoneal injection), we analyzed YAP1 activation and Hippo pathway dynamics via proteomics, single-cell RNA sequencing, western blot, immunohistochemistry, and immunofluorescence. To dissect upstream signaling, we employed a series of genetically engineered mouse models targeting Hippo pathway components.

Results

YAP1 is rapidly activated (via dephosphorylation) during the early stage of paligenosis. Functional assays revealed that YAP1 (and its paralog, TAZ) are necessary and sufficient to drive chief cell reprogramming. Among upstream kinases, only STK38 activity mirrored YAP1 activation patterns. STK38 is essential for YAP1 activation and is degraded via autophagy during transition to stage 2. Notably, STK38 harbors a UFM1/LC3-interacting region (LIR) domain and is targeted to autophagosomes. We found that MIST1 induces UFM1, which protects STK38 from autophagic degradation. In Mist1^−/− mice, reduced UFM1 correlates with heightened basal autophagy, STK38 loss, and constitutive YAP1 activation. Furthermore, STK38 interacts with NF2, similar to canonical Hippo kinases, modulating YAP1 regulation.

Conclusion

We identify a novel MIST1–UFM1–STK38 axis that regulates YAP1 activity in an autophagy-dependent manner during paligenosis. STK38 acts as a non-canonical Hippo kinase whose stability dictates YAP1 activation. These findings uncover new layers of Hippo pathway regulation and highlight the dynamic control of YAP1 during gastric epithelial regeneration.

Introduction

The adhesion G protein-coupled receptor F5 (ADGRF5) belongs to seven-transmembrane receptor family which action seems to be crucial for various cancers and immune response [1-3]. Nevertheless, limited efforts have been made to explore ADGRF5 significance in gastrointestinal cancers.

Aims & Methods

The aim of the study was to assess the significance of ADGRF5 for macrophages function and its impact on gastrointestinal cancer progression. Bone marrow of wild type (WT) and ADGRF5 knockout (KO) mice was used to isolate macrophage population employing differentiation medium. M1 and M2 macrophage phenotype was obtained by polarization of naïve macrophages with a mixture of lipopolysaccharide (LPS) with interferon γ (IFN-γ) and interleukin (IL) 4, respectively. Macrophage phenotype was confirmed using real-time PCR. In vivo studies employing MC38 and PK5L1940 cell-derived allograft murine model of colon and pancreas cancers, respectively, were performed. Multiplex analysis was used to explore secretome in media obtained from tumors.

Results

In murine model of colon cancer, animals with ADGRF5 KO developed significantly smaller tumors which was accompanied by enhanced secretion of macrophage-related cytokines (such as macrophage colony-stimulating factor) in colon tumors compared to WT animals. Ex vivo analysis revealed overexpression of IL-12, nitric oxidase synthase 2 and tumor necrosis factor α in M1 macrophages with ADGRF5 KO compared with WT macrophages under LPS with IFN-γ treatment. In contrast, lower expression of arginase 1 was noted in M2 macrophages with ADGRF5 KO compared with WT macrophages treated with IL-4. In vivo studies employing allograft model of colon and pancreas cancers revealed that the administration of ADGRF5 KO macrophages led to a reduction in tumor size when compared to WT macrophages.

Conclusion

Collectively, ADGRF5 appears to be crucial regulator of gastrointestinal cancer progression through regulating macrophage function. Our study also noted that ADGRF5 depletion reprograms macrophages into M1 phenotype which is responsible for anti-tumor activity.

References

[1] Jacenik D., Hikisz P., Beswick E.J., Fichna J. The clinical relevance of the adhesion G protein-coupled receptor F5 for human diseases and cancers. Biochim Biophys Acta Mol Basis Dis. 2023; 1869: 166683. doi:10.1016/j.bbadis.2023.166683
[2] Kubo F., Ariestanti D.M., Oki S., Fukuzawa T., Demizu R., Sato T., Sabirin R. M., Hirose S., Nakamura N. Loss of the adhesion G-protein coupled receptor ADGRF5 in mice induces airway inflammation and the expression of CCL2 in lung endothelial cells. Respir Res. 2019; 20: 11. doi:10.1186/s12931-019-0973-6
[3] Tang X., Jin R., Qu G., Wang X., Li Z., Yuan Z., Zhao C., Siwko S., Shi T., Wang P., Xiao J., Liu M., Luo J. GPR116, an adhesion G-protein-coupled receptor, promotes breast cancer metastasis via the Gαq-p63RhoGEF-Rho GTPase pathway. Cancer Res, 2013; 73: 6206–6218. doi:10.1158/0008-5472.CAN-13-1049

Introduction

Familial adenomatous polyposis (FAP) is an autosomal dominant syndrome caused by mutations in the APC gene, a negative regulator of the Wnt signaling pathway. It is characterized by the development of hundreds to thousands of Wnt-driven adenomatous polyps in the colon. To prevent the inevitable progression to colorectal cancer, patients with FAP typically undergo total proctocolectomy in early adulthood. Currently, no effective medical therapy exists for FAP. Casein kinase I alpha (CKIα), another negative regulator of Wnt signaling, has emerged as a potential therapeutic target. Prior work in our lab demonstrated that, unlike APC loss, epithelial deletion of Csnk1a1 (CKIα) in the gut does not induce tumorigenesis. Instead, it activates a non-canonical DNA damage response, leading to robust p53 stabilization and suppression of tumorigenic changes. Given that TP53 mutations are typically late events in the adenoma–carcinoma sequence, early-stage lesions in FAP often retain wild-type p53. Unlike sporadic adenomas, which can be resected during colonoscopy, the high polyp burden in FAP precludes endoscopic management, underscoring the need for effective chemoprevention.

Aims & Methods

Our aim in this work was to investigated CKIα as a therapeutic target in preclinical models of FAP.
We employed Csnk1a1^fl/fl, APC^fl/fl, Villin-CreERT2, and CDX2-CreERT2 transgenic mice for in vivo studies, with tamoxifen-inducible gene ablation. Intestinal organoids derived from normal and tumor tissues were used for ex vivo experiments.

Results

In vivo, simultaneous deletion of APC and CKIα in intestinal epithelium did not further enhance Wnt target gene expression compared to APC loss alone. Instead, dual knockout (DKO) stabilized p53, promoted apoptosis, and blocked the dysplastic progression normally observed in APC-deficient gut tissue. In organoid models, treatment with the novel CKIα degrader BTX1050 selectively inhibited tumor-derived organoid proliferation, but did not fully eliminate tumor-initiating cells. We therefore tested A51, a dual CKIα and CDK7/9 inhibitor developed in our lab, which both stabilized and activated p53 and suppressed transcription of super-enhancer-driven oncogenes. A51 treatment significantly downregulated Myc and Mcl1, increased apoptosis in tumor organoids, but only partially suppressed tumor growth in vivo. Notably, A51 induced MAPK pathway activation, as indicated by increased pERK levels. This prompted combination therapy with A51 and the MEK inhibitor trametinib, which demonstrated potent synergistic anti-tumor activity in both organoid and mouse models.

Conclusion

CKIα depletion exerts anti-tumor effects in intestinal tumor models with wild-type p53. Our findings support the combination of A51 and MEK inhibition as a promising "chemical polypectomy" approach for FAP patients.

Disclosure

n/a