2022 Fiscal Year Research-status Report
Pathogenesis roles and therapeutic targets of PBRM1 mutation in intrahepatic cholangiocarcinoma
| Project/Area Number |
22K15336
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| Research Institution | Kanazawa University |
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Principal Investigator |
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| Project Period (FY) |
2022-04-01 – 2025-03-31
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| Keywords | Cholangiocarcinoma / PBRM1 / Organoid / CRISPR/Cas9 / TP53 |
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| Outline of Annual Research Achievements |
We have created 12 lines of intrahepatic cholangiocyte organoids (iCO)from non-tumor livers and optimized conditions for CHO and intrahepatic cholangiocarcinoma organoids (iCCAO). Normal iCOs were grown well in our culture condition with ROCK Inhibitor (Y-27632) and R-Spondin, without Wnt3a. We have created 03 iCCA cell lines with PBRM1 knockout (KO), performed step-by-step TP53 and PBRM1 KO on normal iCOs, and established TP53-KO iCO, and double TP53 and PBRM1-KO iCOs, using CRISPR/Cas9.
A novel TME classification of iCCA has been established, including four subtypes, showing distinct organizing patterns of T cells, myeloid cells, CAFs, an extracellular matrix such as collagen, and gene expression on spatial proteomic profiling, including CTLA4, PD-1, VISTA, LAG3, and TAGLN.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
We planned to collect and establish new organoids from human intrahepatic cholangiocarcinoma, but this has been dependent on primary tumor sample availability. It took a longer time for us to create gene-modified organoids from normal cholangiocytes.
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| Strategy for Future Research Activity |
We will analyze down-stream effects of TP53 and PBRM1 loss on iCOs in next experiments. We will also establish new organoids from human cholangiocarcinoma, develop a panel of immunohistochemistry makers to support the diagnosis of our histological-based TME classification of intrahepatic cholangiocarcinoma, evaluating clinical utility of the novel TME classification.
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