Periodic Reporting for period 1 - PRECODE (PancREatic Cancer OrganoiDs rEsearch Network)
Reporting period: 2019-10-01 to 2021-09-30
The PRECODE Network (PancREatic Cancer OrganoiDs rEsearch) central mission is to establish pancreatic organoid research firmly in the European Union, by training the next generation of creative and innovative researchers in pancreatic cancer. This will be done through a shared doctorate program of excellence that fulfils the three “I”: international, interdisciplinary and intersectorial. Organoids can be viewed as small micro-organs which can recapitulate the organization and the function thereof. As micro-organs organoids can be used to reduce animal experiments and help to reach 3R goals. Organoids from pancreatic cancer will help us to learn which drugs might help in the therapy of this terrible disease. They can be isolated from a variety of clinical sources with minimal material needed. This makes them a perfect tool for precision medicine. Sequencing, metabolomics and high throughput in vitro experiments to deduct the best possible drug combination fit hand in glove with this approach. Training of ESRs will be achieved by e-learning, workshops and secondments. They will earn to cultivate and manipulate organoids as well as cutting edge molecular biology techniques. Therefore our work group consists of 15 beneficiaries and 7 partner organizations (including three patient advocate groups from Sweden, Italy and Germany) to develop those technologies and to train ESRs in their application. PRECODE will therefore connect the concepts of investigation, education and innovation establishing a capacitation for the ESRs that goes beyond the state of the art and that will allow them to improve their competences and their professional career at the same time that we advance towards innovation in the field of pancreatic cancer research.
The early activities of PRECODE have focused on the production of a robust and reliable protocol for the derivation of organoid cultures. The protocol has been produced in the form of Standard Operating Procedures and includes the description of methods for preparation of nucleic acids and protein lysates from the models. In addition to the protocol, PRECODE has also generated a flowchart for the qualification of organoids before their distribution which included: (1) expansion of the organoids for a minimum of five consecutive passages; (2) “resuscitation” of the culture following freeze/thaw cycle; (3) genetic evidence of cancer based on the detection of KRAS mutation and/or alteration of other common PaCa genes. To date, 26 organoid cultures have passed the first 2 QC steps, 15 of which have also been genetically verified, and 20 are in the derivation pipeline. We have also defined a minimum medium condition that enables long-term organoid propagation. We have also defined Common Data Elements to be recorded from the source patients by the clinical collaborators. Tissues and models related information are anonymized and collected at each contributing site before submission into a RedCap database that has been established at Umea University. PRECODE has been able to establish organoids from IPMN (a precursor lesion) patients which is a rare histology of pancreatic tissue. We have also successfully developed a protocol for isolation of cancer-associated fibroblasts, and have established normal fibroblasts and cancer-associated fibroblast cultures. To identify metastases associated genes we have established genome wide gene editing in organoids from different sources. Moreover, we have established about 20 wildtype and eight Zeb1 knockout organoid lines from the KPC PaCa GEMM in order to block the EMT program by inactivating one central transcription factor. We have optimized media conditions for high throughput drug screening of PDAC organoid monocultures and initial drug screens with standard of care drugs have been performed including treatment schedules mimicking clinical applications. Co-cultures of PDAC organoids and platelets have been established and initial drug treatments with standard of care and nanodrugs were performed. We have established a centralised sequencing and informatic platform with our partners at SciLifeLab Sweden, that will expedite both the genomic and transcriptomic analyses of PRECODE organoids. PRECODE has trained the ESR via eLearning and in three meetings in the area of organoid production, bioinformatics and CRISPR/Cas9 based genome editing. ESR training also included several courses on transferable skills.
Within the reporting period we achieved several results. First, we introduced a standard operating procedure (SOP) for the establishment of organoid lines and the extraction of RNA, DNA and proteins, which is currently also used in the planned clinical trials of ESPAC-6 and ESPAC-7. The participation in PRECODE permitted the beneficiary Peter Bailey from UG to become the head of translational research for the two clinical trials mentioned above. Second, we were able to publish a collection of peer reviewed reviews in the journal “Frontiers in cell and developmental biology” (IF:6.514) in which we presented the anticipated work of PRECODE. Third, we have agreed on a SOP for high throughput screening for the produced organoid lines, so that we achieve comparable results between different beneficiaries. Our efforts in dissemination of PRECODE has enabled us to include organoid lines from two other European groups (Hanna Seppänen from Helsinki and Michael Linnebacher from Rostock) into the PRECODE pipeline indicating the amplifying effect of the MSCA action.
Within the reporting period we achieved several results. First, we introduced a standard operating procedure (SOP) for the establishment of organoid lines and the extraction of RNA, DNA and proteins, which is currently also used in the planned clinical trials of ESPAC-6 and ESPAC-7. The participation in PRECODE permitted the beneficiary Peter Bailey from UG to become the head of translational research for the two clinical trials mentioned above. Second, we were able to publish a collection of peer reviewed reviews in the journal “Frontiers in cell and developmental biology” (IF:6.514) in which we presented the anticipated work of PRECODE. Third, we have agreed on a SOP for high throughput screening for the produced organoid lines, so that we achieve comparable results between different beneficiaries. Our efforts in dissemination of PRECODE has enabled us to include organoid lines from two other European groups (Hanna Seppänen from Helsinki and Michael Linnebacher from Rostock) into the PRECODE pipeline indicating the amplifying effect of the MSCA action.
We expect to create a resource of human organoid lines, which are fully characterised and available for other European researchers. By this PRECODE will increase its impact since such models are highly sought after. We also expect to identify genes crucial for the early development of metastasis in pancreatic cancer. These validated target genes will serve as a resource for further investigation and European drug development. Finally, PRECODE will have further impact on the European research by educating 15 ESR in state-of-the-art technologies, including organoid production, next generation sequencing and genome editing. These researchers of the future will drive the European research landscape by becoming the next generation research leader.