Transforming growth factor-beta-induced protein using human 3D organoids: a novel immune check-point in colorectal cancer?

Introduction: The transforming growth factor—beta-induced (TGF-I) protein was found significantly upregulated in colorectal cancer (CRC) secretome (secreted proteome) as compared with the non-tumor. TGFBI is an RGD-containing extracellular matrix protein that binds to type I, II and IV collagens, serves as a ligand recognition sequence for several integrins, and inhibits cell adhesion. Its release from primary tumors has been associated with increased tumor proliferation/migration/metastasis, but its role as secreted immune check-points (sICs) has not been fully investigated. Methods: A LC/Mass-spectometry (Orbitrap)-based platform was set up to identify the secretome in conditioned medium (CM) from fresh tumor and non-tumor surgery samples (20 patients). By this approach, we selected a multitude of secreted proteins that were upregulated in CRC secretome as compared to the non-tumor, in order to identify those potentially acting as sIC . Aim: Their discovery may represent a tremendous resource for tumor specific drug targets, potentially acting as sIC inhibitors in both cold and hot tumors, unlike current IC inhibitors (e.g., IpilumumAb and NivolumAb). Results: We first validated by Elisa that TGFBI was overexpressed in CM tissue samples and in serum from CRC patients (as compared with non-tumor patients), and positively correlated with the tumor stage. Interestingly, tissue-IHC and confocal microscopy revealed that TGFBI was overexpressed by tumor cells, T cells, monocytes and plasma cells in tumors in a significantly higher extent than in non-tumor, suggesting a massive involvement of the tumor microenvironment (TME) in secreting it. These data are also confirming at the level of the same cell populations isolated from tumor tissues. Importantly, the recombinant form of TGFBI, as well as the tumor CM containing high levels of native TGFBI, significantly inhibited various functions (IFN- and TNF- production, GZB and T-bet expression…) of anti-CD3/CD28-activated CD4 and CD8 T cells, which could be restored by the addition of the neutralizing anti-TGFBI mAb in vitro. Finally, we are validating that TGFBI can act as a sIC by using human 3D CRC organoids as a surrogate of animal models in vivo. Human 3D-organoids generated from various tumor tissues allow to determine the interaction between tumor and immune system, the response (activation, cytokine production, killing…) by autologous CD8 and CD4 T cells derived from cancer patients, the role of sICs in inhibiting anti-tumor T cell response, the role of related sIC inhibitors in unlashing the anti-tumor T cell response. Conclusions: TGFBI with human 3D CRC organoids can act as a tremendous effective sIC inhibitors in CRC patients. Furthermore, human-based models, such as human organoids, can offer effective ways “to accelerate transition to a research system that does not involve testing on animals”, as the European Parliament has recently declared (see go.nature.com/3hzprhj).