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CX-4945 (Silmitasertib) Induces Cell Death by Impairing Lysosomal Utilization in KRAS Mutant Cholangiocarcinoma Cell
DA Sol Lee1, Min Woo Han2, Yonghyun Kang1
1Asan Institute for Life Sciences, Asan Medical Center, Seoul, Republic of Korea.
Background/Aim:
Macropinocytosis is a non-selective form of endocytosis that facilitates the uptake of extracellular substances, such as nutrients and macromolecules, into the cells. In KRAS-driven cancers, including pancreatic ductal adenocarcinoma, macropinocytosis and subsequent lysosomal utilization are known to be enhanced to overcome metabolic stress. In this study, we investigated the role of Casein Kinase 2 (CK2) inhibition in macropinocytosis and subsequent metabolic processes in KRAS mutant cholangiocarcinoma (CCA) cell lines.
Materials And Methods:
The bovine serum albumin (BSA) uptake indicating macropinocytosis was performed by flow cytometry using the HuCCT1 KRAS mutant CCA cell line. To validate macropinosome, the Rab7 and LAMP2 were labeled and analyzed via immunocytochemistry and western blot. The CX-4945 (Silmitasertib), CK2 inhibitor, was used to investigate the role of CK2 in macropinocytosis and subsequent lysosomal metabolism.
Results:
The TFK-1, a KRAS wild-type CCA cell line, showed only apoptotic morphological changes. However, the HuCCT1 cell line showed macropinocytosis. Although CX-4945 induced morphological changes accompanied by the accumulation of intracellular vacuoles and cell death, the level of macropinocytosis did not change. These intracellular vacuoles were identified as late macropinosomes, representing Rab7+ vesicles before fusion with lysosomes. In addition, CX-4945 suppressed LAMP2 expression following the inhibition of the Akt-mTOR signaling pathway, which interrupts mature macropinosome and lysosomal metabolic utilization.
Conclusion:
Macropinocytosis is used as an energy source in the KRAS mutant CCA cell line HuCCT1. The inhibition of CK2 by CX-4945 leads to cell death in HuCCT1 cells through alteration of the lysosome-dependent metabolism.
Insights
In KRAS mutant cholangiocarcinoma (CCA), inhibiting Casein Kinase 2 (CK2) with CX-4945 causes cell death. This occurs by disrupting lysosome-dependent metabolism, even though macropinocytosis levels remain unchanged.
Area of Science:
- Cell Biology
- Cancer Metabolism
- Molecular Oncology
Background:
- Macropinocytosis is a cellular uptake mechanism crucial for nutrient acquisition, particularly in KRAS-driven cancers experiencing metabolic stress.
- KRAS mutant cholangiocarcinoma (CCA) exhibits enhanced macropinocytosis and lysosomal processing to meet metabolic demands.
Purpose of the Study:
- To investigate the impact of Casein Kinase 2 (CK2) inhibition on macropinocytosis and downstream metabolic pathways in KRAS mutant CCA cells.
- To determine if CK2 inhibition affects the utilization of macropinosomes for cellular metabolism.
Main Methods:
- Utilized the HuCCT1 KRAS mutant CCA cell line to assess bovine serum albumin (BSA) uptake via flow cytometry.
- Employed immunocytochemistry and western blot to validate macropinosomes (Rab7, LAMP2).
- Administered CX-4945 (Silmitasertib), a CK2 inhibitor, to evaluate its effects on macropinocytosis and lysosomal metabolism.
Main Results:
- KRAS mutant HuCCT1 cells demonstrated macropinocytosis, unlike KRAS wild-type TFK-1 cells which showed apoptosis.
- CX-4945 induced cell death and intracellular vacuole accumulation, identified as late macropinosomes, without altering macropinocytosis levels.
- CX-4945 suppressed LAMP2 expression and inhibited the Akt-mTOR pathway, disrupting lysosomal metabolic utilization.
Conclusions:
- Macropinocytosis serves as an energy source in KRAS mutant CCA (HuCCT1 cells).
- CK2 inhibition by CX-4945 induces cell death in HuCCT1 cells by impairing lysosome-dependent metabolism.
- Targeting CK2 represents a potential therapeutic strategy for KRAS mutant CCA by disrupting cellular metabolism.
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