Contributed Talk

Effect of fibroblast contractility on immune infiltration in 3D co-culture spheroids of colorectal cancer

Eleni Dalaka¹, Ruchi Goswami², Salvatore Girardo², Ana Henriques³, Eduard Batlle^3,4,5, Jochen Guck^2,6, Xavier Trepat^1,5,7,8

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Treating cancer has been a major focus in cancer research, with several therapies being developed over the years. Immunotherapies – exploiting patient’s immune system to trigger immune responses against tumours – has provided several encouraging results against tumour progression and recurrence [1,2,3,4]. However, only 20-40% of cancer patients are eligible for immunotherapies and less than 15% of them respond to their treatment [5]. When immunotherapies fail, tumours exhibit a certain phenotype, with immune cells being trapped inside the tumour microenvironment (TME) and thus being unable to migrate towards the tumour.

In this study, we investigate the role of mechanical forces that arise in the TME on tumour immune infiltration. We hypothesize that fibroblasts – a major cellular component of TME – exert high mechanical forces, which create a tense and compact physical barrier around the tumour, inhibiting immune cells from navigating their environment and thus migrate towards the tumour. By reducing fibroblast contractility, relaxation of the apparent TME tension is expected, accompanied by the formation of physical paths among fibroblasts that immune cells can use to migrate through this complex environment. To confirm this hypothesis, we developed an in vitro model of liver metastasis of colorectal cancer by forming 3D co-culture spheroids using colorectal tumour organoids enveloped by liver fibroblasts. These 3D multicellular structures are then embedded in alginate matrices together with primary T-cells, which migrate in different directions, and they reach the 3D co-culture spheroids over time. This system allows us to study mechanics, migration and physical interactions of cells in a 3D manner.

First, we investigated how immune cells interact with the different cell types in the co-culture spheroids over time and we quantified immune infiltration in the fibroblast layer and the cancer core, separately. We followed and tracked immune cells as they adhere, enter and migrate inside co-culture spheroids. Next, we observed that upon TGF-b inhibition, immune cells can bypass the fibroblast layer and infiltrate into cancer areas that reside at the core of the co-culture spheroids, confirming previous in vivo studies [6,7]. By using soft polyacrylamide (PAAm) beads [8], we measured the mechanical forces developed in the co-culture spheroids, and we observed a decrease in the mechanical force exertion upon TGF-b inhibition. These data suggest that fibroblast contractility plays a key role in tumour immune infiltration. Overall, this study aims at identifying – for the first time – the mechanical components of TME that control tumour immune infiltration in colorectal cancer.