Small molecule LATS kinase inhibitors block the Hippo signaling pathway and promote cell growth under 3D culture conditions

by Selleckchem | Nov 09 2023

Although 3D cell culture models are considered to reflect the physiological microenvironment and exhibit high concordance with in vivo conditions, one disadvantage has been that cell proliferation is slower in 3D culture as compared to 2D culture. However, the signaling differences that lead to this slower proliferation are unclear. Here, we conducted a cell-based high-throughput screening study and identified novel small molecules that promote cell proliferation, particularly under 3D conditions. We found that one of these molecules, designated GA-017, increases the number and size of spheroids of various cell-types in both scaffold-based and scaffold-independent cultures. In addition, GA-017 also enhances the ex vivo formation of mouse intestinal organoids. Importantly, we demonstrate that GA-017 inhibits the serine/threonine protein kinases large tumor suppressor kinase 1/2, which phosphorylate Yes-associated protein and transcriptional coactivator with PDZ-binding motif , key effectors of the growth- and proliferation-regulating Hippo signaling pathway. We showed that GA-017 facilitates the growth of spheroids and organoids by stabilizing and translocating Yes-associated protein and transcriptional coactivator with PDZ-binding motif into the cell nucleus. Another chemical analog of GA-017 obtained in this screening also exhibited similar activities and functions. We conclude that experiments with these small molecule large tumor suppressor kinase inhibitors will contribute to further development of efficient 3D culture systems for the ex vivo expansion of spheroids and organoids.

Comments:

The research you've described highlights an important breakthrough in the field of 3D cell culture models and their potential applications in various areas of biology and medicine. Here's a summary of the key findings and implications of the study:

1. **Slower Cell Proliferation in 3D Culture:** It's well-known that 3D cell culture models more closely mimic the physiological microenvironment compared to traditional 2D cultures. However, a drawback of 3D culture has been slower cell proliferation, which has limited their use in some applications.

2. **Identification of GA-017:** The study conducted a high-throughput screening to identify small molecules that can promote cell proliferation, especially in 3D culture conditions. One of the identified molecules, GA-017, was found to be particularly effective.

3. **Promotion of Spheroid Growth:** GA-017 was shown to increase the number and size of spheroids formed by various cell types in both scaffold-based and scaffold-independent 3D cultures. This suggests that GA-017 has a broad range of applications in 3D cell culture systems.

4. **Enhancement of Organoid Formation:** GA-017 was also found to enhance the ex vivo formation of mouse intestinal organoids. Organoids are 3D structures that mimic the structure and function of organs, making them valuable tools for studying organ development and disease.

5. **Inhibition of Large Tumor Suppressor Kinases:** The study revealed that GA-017 inhibits the activity of serine/threonine protein kinases known as large tumor suppressor kinase 1/2. These kinases are involved in the Hippo signaling pathway, which regulates cell growth and proliferation.

6. **Translocation of Key Effectors:** GA-017 was found to stabilize and translocate key effectors of the Hippo signaling pathway, including Yes-associated protein and transcriptional coactivator with PDZ-binding motif, into the cell nucleus. This translocation likely plays a role in promoting cell growth.

7. **Potential for 3D Culture Systems:** The research suggests that small molecule inhibitors of large tumor suppressor kinases, like GA-017, can be valuable tools for developing more efficient 3D cell culture systems for the ex vivo expansion of spheroids and organoids.

Overall, this study has important implications for advancing 3D cell culture techniques and improving their efficiency, which can have far-reaching applications in drug development, disease modeling, and regenerative medicine. Further research and development in this area could lead to more physiologically relevant in vitro models for studying cellular behavior and tissue development.