2018 Fiscal Year Annual Research Report
|Research Institution||Okinawa Institute of Science and Technology Graduate University |
Hoh Hong Huat 沖縄科学技術大学院大学, 科学技術研究科, 特別研究員(DC1)
|Project Period (FY)
2018-04-25 – 2021-03-31
|Keywords||Pancreatic cancer / 3D / Organoids|
|Outline of Annual Research Achievements
This study focuses on how pancreatic cancer cell-to-cell interaction in 3D tumor microenvironment. Five pairs of normal and cancerous pancreatic ductal organoids had been established from wild-type C57BL/6 mice and pancreatic cancer mouse models (KPC mice), respectively. The harvested cells developed into 3D structures in Matrigel in vitro. Multiple lines of pancreatic organoids were expanded for experiments and some were frozen for future use. CRISPR-Cas9 mediated gene editing for KRAS activating point mutation was attempted in wild-type pancreatic organoids to generate early stage pancreatic intraepithelial neoplasia (PanIN). KRAS-activated organoids presented with abnormal ductal structures in 3D culture as compared to normal, wild-type organoids. Together with the normal organoids and tumor organoids derived from KPC mice that resembles late-stage, aggressive pancreatic ductal adenocarcinoma (PDAC), I have established a series of mouse organoids representing normal, early-stage and late-stage pancreatic cancer in 3D. These organoids were verified for their gene expression for pancreatic ducts as well as aberrant expression of mutated genes in the tumor organoids by qPCR before they were used for upcoming experiments.
|Current Status of Research Progress
Current Status of Research Progress
3: Progress in research has been slightly delayed.
Due to the late arrival of tumor samples from the collaborators, there was a significant delay in the progress of the establishment of 3D organoid model which has a consequence on the subsequent experimental plan. However, once the tumor samples were received, immediate establishment and expansion of the organoids were carried out and used for experiments.
|Strategy for Future Research Activity
Future work involves genotypic and phenotypic verification of the KRAS-mutant organoids using cellular imaging, molecular profiling including RNA sequencing and exosome profiling. Other mutations including point mutation of p53 will also be induced using genomic engineering method to create a more advanced model of PDAC in 3D for experiments looking into the molecular evolution and exosome characteristic at advanced stage of PDAC. Exosome profiling will be conducted by nanoparticle tracking analysis and RNA sequencing following extraction using differential ultracentrifugation. Normal pancreatic epithelial organoids will be used as a control for comparison.
Research Products (2 results)