Tumour initiation is a process of sequential genetic and biochemical alterations occurring over time in a three-dimensional environment. However, cooperation and competition of cellular populations within a living tissue are also fundamental, yet not fully understood, mechanisms underlying tumour initiation and promotion. Therefore, more accurate observations and modelling of these heterogeneous cellular populations are of strategic importance for the understanding of cancer and, consequently, to design future therapeutic interventions. Unfortunately, techniques typically used for in vivo cell tracing often present critical limitations: i) phototoxicity constrains time resolution and observational times, ii) the occurrence of oncogenic events in defined places and times within a tissue is extremely difficult to control and iii) how communication between different populations of cells contributes towards mutant cell competition is arduous to characterize.
OncoLive will integrate a three-dimensional organoid culturing system, cell genetics, optogenetic, biochemical imaging and light-sheet microscopy to control and visualize the early steps leading to oncogenic transformation. Our working hypothesis is that cooperation and competition between different mutant clonal populations contribute towards oncogenic tissue transformation and we will attempt to test this hypothesis by investigating clonal fitness, cellular cross-talk and cell fate dynamics within a three-dimensional culture controlling oncogenic events in space and time, as well as the cellular response to specific signalling cues.
We aim to provide the means to overcome existing methodological barriers and thus offer new insights about oncogenesis. The objectives of this collaborative efforts are:
1) Eliminating technology barriers with the development of a purpose-built light-sheet fluorescence microscope (LSFM), a three-dimensional imaging method proven for its low phototoxicity. With a unique integration of novel solid-state sensors and optogenetics, we will have the simultaneous control of light-inducible oncogenic events, recording of signalling pathways and cell lineage tracing.
2) Characterizing clonal dynamics and cross-talk between cellular populations in three-dimensional living organoid cultures during the controlled and sequential activation of specific oncogenic events by orthogonal (chemical and optical) means such as the light-inducible expression of the oncogenic mutants.
3) Modelling clonal dynamics with the implementation of novel methodologies for the mathematical modelling and Bayesian analysis in order to establish causal connections between oncogenic events and signalling cues at cell-autonomous or cell-to-cell level.
With outreach initiatives and by establishing strategic technologies, we envisage that OncoLive will impact how the wider community investigate the mutational and biochemical determinants of clonal dynamics leading to cancer, therefore providing more accurate information on how to target tumour prevention and therapeutic strategies.
OncoLive 