Dual role of Transforming Growth Factor Beta1&2 during Tumor Promotion and Metastasis in Primary Liver Cancer

Background:

Transforming Growth Factor Beta (TGF-β) belongs to a superfamily of cytokines that induces pleiotropic effects on different processes and cell types in the liver. While TGF-β signaling exerts tumor suppressive functions at pre-neoplastic and early tumor stages, cytostatic effects of TGF-β are often lost in progressed stages due to (epi-) genetic disruption of several members of the signaling pathway. Consequently, cancer cells display an epithelial-mesenchymal-transition (EMT) phenotype and acquire pro-metastatic properties.

Aims:To evaluate the effect of the TGF-β1 and TGF-β2 on (i) proliferative (ii) migratory and pro-metastatic properties of primary and established PLC cell lines.

Method:

Primary patient-derived (HCC & CCA) and established cell lines (PLC & HuCCT-1) were treated with TGF-β1 and TGF-β2 (1 ng/ml) for 72 hr. The effect of TGF-β1&2 on proliferation was determined by colony and sphere formation assays. Invasive and migratory properties were determined using the wound healing invasion assays. Next Generation Sequencing and RPPA (reverse phase protein array) was performed to explore differential transcriptomic and protein expression patterns across treatments.

Results:

Treatment with TGF-β1 and TGF-β2 led to a significant reduction in colony and spheroid forming ability in all investigated cell lines. Interestingly, a significant downregulation of epithelial marker E-cadherin and concomitant upregulation of mesenchymal markers such as vimentin and SNAIL was exclusively observed after TGF-β1 treatment. In addition, transcriptome profiling confirmed activation of gene sets involved in Cell Cycle:G1/S Checkpoint in response to both treatments (TGF-β1&β2) whereas enrichment in signaling pathways known to be involved in pro-metastatic properties resembling P13K, MAPK, MMPs and Hippo signaling pathway were predominantly associated with the TGF-β1 response.

Conclusions:

In conclusion, the cytostatic effect of TGF- β1 and TGF- β2 is reflected by a reduction in proliferation in both HCC and iCCA. Further, TFG-β1 seems to be an important regulator of EMT as well as invasive properties in progressed PLCs. Transcriptome profiling and Proteomics data indicates an increase in p21 that induces cell cycle arrest upon treatment of TGF-β1&2, while an increase in EMT related properties is associated only with the TGF-β1.These context-dependent dichotomic effects should be considered in TGF-β based therapeutic approaches.