| Project/Area Number |
22K15336
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| Research Category |
Grant-in-Aid for Early-Career Scientists
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| Allocation Type | Multi-year Fund |
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| Review Section |
Basic Section 47060:Clinical pharmacy-related
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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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| Project Status |
Granted (Fiscal Year 2023)
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| Budget Amount *help |
¥3,250,000 (Direct Cost: ¥2,500,000、Indirect Cost: ¥750,000)
Fiscal Year 2024: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2023: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
Fiscal Year 2022: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000)
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| Keywords | Cholangiocarcinoma / PBRM1 / Organoid / CRISPR/Cas9 / TP53 / IDH1 / KRAS |
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| Outline of Research at the Start |
Intrahepatic cholangiocarcinoma is a heterogeneous cancer with dismal prognosis. Somatic mutations of Polybromo-1(PBRM1)are often found in iCCA. This study aims to clarify the mechanism how PBRM1 loss drives iCCA progression, search for novel therapeutic targets for PBRM1-deficiency iCCA.
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| Outline of Annual Research Achievements |
We created four more lines of intrahepatic cholangiocyte organoids (iCO) from non-tumor livers, totaling 16 primary iCO lines, and one intrahepatic cholangiocarcinoma organoid (iCCAO). We constructed different vectors for PBRM1, IDH1, KRAS knockout, and optimized conditions for gene-edited organoids and cell lines. We collected 150 human intrahepatic cholangiocarcinomas (iCCA) and analyzed gene mutations, including PBRM1, P53, KRAS, IDH1/2, ARID1A. A panel of markers to support the classification of iCCA tumor microenvironment was established: SMA, TAGLN, CD276 were prominently found in activated stroma; CD163, CD66b, and VISTA for macrophages and myeloid-derived suppressor cells (MDSCs), prominently in early stroma; CD3, CD8, and PD1 for T cells in inflammatory stroma.
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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
Gene editing efficiency and the survival of gene-modified organoids from normal cholangiocytes were varied. Some lines of PBRM1-KO organoids were difficult to maintained.
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| Strategy for Future Research Activity |
We will perform CRISPR/Cas9 experiments on four human iCCA cell lines available in our lab, in addition to gene-modified iCO, and analyze how cell lines and iCO, iCCAO with PBRM1 loss respond to different treatments in the next experiments. We will also analyze human iCCAs, comparing tumor cell histological phenotypes and tumor microenvironment (TME) features, and TME class between PBRM1-mutant and PBRM1-wild type iCCAs.
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