Periodic Reporting for period 4 - PanCaT (Next-generation in vivo models for improved pancreatic cancer therapies) Okres sprawozdawczy: 2020-08-01 do 2021-12-31 Podsumowanie kontekstu i ogólnych celów projektu Cancer is a disease with a major socio-economic impact and the 2nd leading cause of death in Europe. Pancreatic ductal adenocarcinoma (PDAC), which is driven by oncogenic KRAS, is almost universally fatal and the 3rd leading cause of cancer death in the western world. The incidence of PDAC is constantly rising. Despite modern treatment regimens, the prognosis of PDAC did not change significantly in the last 30 years with less than 1% of patients surviving 10 years. This contrasts other solid tumor entities such as breast and colon cancer, where death rates have decreased >50%. Due to the increasing incidence and the lack of efficient therapies, PDAC is predicted to become the 2nd leading cause of cancer death in the next decade. Therefore, PDAC poses a major challenge in Europe and novel therapeutic approaches are urgently needed.Though mortality rates have remained stagnant for more than 30 years, recent advances provide reason for true optimism. Breakthroughs in various fields of cancer research, such as genomic analysis and next-generation sequencing, high-throughput drug and genetic screening and computational approaches are providing a wealth of resources that hold the promise of advancing treatment strategies for PDAC patients. Thus, the field is primed for major advances as never before.What has been lacking so far to take full advantage of these developments is a means of modeling and manipulating the developing tumor entity and its tumor microenvironment (TME) in a realistic manner on a genome-wide scale in vivo. This is vital for identifying and assessing candidate therapeutic targets and mechanisms of resistance, as well as revealing basic principles of tumor biology. Therefore, there is an urgent need for a next generation of improved model systems, which will pave the way to novel therapies. The overarching goal of this ERC funded project is to develop next generation models and technologies for PDAC research, to investigate why it resists conventional and targeted therapies and to pave the way to novel individualized treatment strategies.We invented new models and research tools such as dual- and triple recombinase systems (DRS and TRS) and combined them with novel sophisticated technologies, such as single cell sequencing, computational modelling and advanced genetic and drug screening. Together, these novel models and tools allowed us for the first time to control gene expression in a spatial and temporal manner in autochthonous tumors in vivo and provided unparalleled access to the native biology of cancer cells and their hosting stroma, as well as the rigorous validation of candidate therapeutic targets and novel therapeutic strategies. By the application of cutting edge genetic engineering and interfering technologies we were able to address long-standing biological questions in a rigorous manner that could not be addressed before. The project thus opened new horizons for the functional understanding of PDAC biology and the development of novel therapies. Prace wykonane od początku projektu do końca okresu sprawozdawczego oraz najważniejsze dotychczasowe rezultaty First, we generated DRS and TRS models and developed novel non-genetic strategies to deliver site-specific recombinases to the pancreas via electroporation strategies as well as viral AAV based gene transfer for somatic gene targeting and CRISPR/Cas9 based genetic engineering in vivo. This opened new possibilities to investigate PDAC mechanistically in vivo and to identify novel cancer genes and therapeutic targets. To exploit the full potential of the DRS and TRS based PDAC models, we systematically investigated subpopulations of cells of the tumor microenvironment, which support tumor growth and developed systems to target cell subtypes, which are not precisely characterized by a single marker, but only by marker combinations. These results have been published in Nature Communications, Nature Protocols, Nature Reviews Cancer and EMBO Molecular Medicine.Next, we applied our novel PDAC models to address therapeutic questions of high clinical relevance for PDAC patients. We used the models to 1) uncover evolutionary routes of PDAC development and understand their impact on tumor phenotypes (published in Nature, 2018), 2) identify cancer genes and candidate therapeutic targets in PDAC functionally (published in Cancer Discovery 2021, Gastroenterology 2021, Nature Communications 2016), 3) perform synthetic lethal CRISPR/Cas-based genetic loss and gain of function screens to identify genes which induce cancer cell death (unpublished), and to 4) show that the molecular makeup of the primary PDAC dictates if a cell in the tumor microenviroment promotes or restains tumor development. This indicates an extraordinary context specificity of PDAC stroma crosstalk, which is of fundamental relevance for establishing novel therapeutic strategies. Indeed, the combined targeting of PDAC cells and their immunosuppressive stroma sensitizes them to immunotherapy (published in Nature Cancer 2022).Finally, we investigated primary and secondary resistance towards targeted PDAC therapies, which is a major clinical problem. We used our novel models and tools to uncover resistance at the genetic, epigenetic and transcriptional level and identified several pathways and modes of resistance, such as Myc amplification, which provide novel and unexpected vulnerabilities and opportunities for combination therapies (unpublished). We exploited and disseminated our discoveries by various means, such as 1) exchange of models, technologies and data with collaborators nationally and internationally, 2) international scientific meetings and symposia, 3) educational programs in Germany and Europe, 4) seminars, workshops and international conferences in Munich, 5) national and international research societies for oncology and pancreas research, 6) publications, reviews, and overview articles in high profile journals, such as Nature Reviews Cancer, and 7) knowledge transfer and public relations via social media (e.g. Twitter channel with >350 followers, interviews in the German magazines, patient involvement, press releases and public events). Innowacyjność oraz oczekiwany potencjalny wpływ (w tym dotychczasowe znaczenie społeczno-gospodarcze i szersze implikacje społeczne projektu) Our new DRS/TRS based PDAC models provide novel and unique opportunities to model, manipulate, investigate and understand diverse aspects of malignant tumors. This includes rigorous mechanistic analysis of i) genetic alterations in catastrophic & multi-step carcinogenesis and tumor heterogeneity, ii) tumor cell subpopulations such as cancer stem cells, iii) the tumor microenvironment, iv) immune cell subpopulations, v) the metastatic niche of host organs, vi) therapeutic targets and vii) resistance mechanisms. The next-generation models of human PDAC, which we developed in the project, are highly flexible and of ground breaking importance to the field, because they allowed us to address fundamental biological and therapeutic questions at the genetic level in established tumors in vivo that could not be addressed before. Therefore, they set a new global standard for basic cancer research and drug target discovery and validation. In addition, they are of great clinical relevance because they contributed directly to the development of novel therapeutic strategies, such as novel cancer therapies that are capable of reprogramming the immunosuppressive TME and sensitise PDAC towards immunotherapy. Based on the results of the ERC funded project, we are currently initiating fist clinical trials to test our novel concepts in the clinic. A combination therapy sensitises PDAC to T cell attack
