Original aim of SLIM was to study the effect of NPM1 mutations on the sensitivity of acute myeloid leukaemia(AML) cells to commonly used drugs and tool compounds. Methodologically it was envisioned to use CRISPR/Cas9 based genome editing to generate matched cell line pairs harbouring mutated or wild-type NPM1 genes whilst otherwise being identical. Further, automated high content microscopy would be used to study the differential effect of these drugs on the model cell lines. An integrated -omics approach would be used to understand differences in drug susceptibility in more detail and learn more about biology of NPM1. Due to unforeseen technical difficulties, the model system could not be established as planned. Rather an analogous research strategy was applied to study human immune processes. Whilst the original research aim could not be met, the training aims for the researcher could be achieved. The human immune system is a complex assembly of different cell types aimed at defending the host against invading pathogens. It is increasingly understood that the immune system plays an important role in controlling tumor growth. Tumors need to overcome the surveillance of the immune system to become macroscopically manifested and pharmacological interventions have been designed that can reactivate the immune system to treat cancer. When overactivated on the other hand, the human immune system can attack healthy cells of the body leading to autoimmune disease such as multiple sclerosis but also inflammatory diseases such as rheumatoid arthritis, ulcerative colitis etc. The societal cost of cancer and immune-related diseases combined is estimated to exceed EUR 200 Mio p.a. in the EU. The immune system is controlled in parts by soluble messenger factors (e.g. cytokines, chemokines) and by physical interactions of cells (“immunological synapses”,cell adherence processes etc.). So far the study of the immune system both in vitro and in animal models of disease have been limited to measuring soluble factors and events that are directly manifested in the expression of certain marker proteins on the surface of cells. The cell-cell interaction dimension until now has been very difficult to investigate. In the Superti-Furga lab a novel microscopy based approach has recently been developed that allows systematic measurement of cell-cell interactions in complex mixtures of immune cells ex vivo. This, for the first time, enables the study of the immune system directly in a cell-cell interaction dimension in high throughput in vitro. In a proof-of-concept study (Vladimer, Snijder et al, Nat.Chem.Biol.2017) it was shown that using the approach unexpected immunomodulatory properties of small molecule drugs could be identified. Specifically it was suggested that the TKI crizotinib could be combined with immunotherapy in a synergistic manner. For the purpose of the revised project the overall objective was in first place to better understand and systematically benchmark the novel cell-cell interaction readout by Vladimer, Snijder et al. We wanted to understand if certain stimuli elicited certain patterns of cell-cell interactions so that we could in reverse connect observed cell-cell interaction patterns of molecules of unknown function to known immune processes. In a second dimension, the objective was to understand if we could identify certain regulatory relationships based on our cell-cell interaction data. For example, if a certain gene is upregulated always together with another gene, the hypothesis can be set up that these genes are somehow functionally related. Similarly, we wanted to study if the occurrence of one cell-cell interaction could be functionally connected to the occurrence of another one. This could reveal important new insight into the wiring of the immune system.
