The Hedgehog-GLI (HH-GLI) signalling pathway is an important regulator of embryonic development, while it is mostly inactive in the adult organism. It only remains active in somatic stem cells. Aberrant activation of the pathway contributes to tumorigenesis in many tumor types. The GLI1, GLI2 and GLI3 transcription factors regulate gene expression of targets involved in proliferation, metastasis and cancer stem cell renewal. In our previous research we were the first to analyse the transcriptomes of all three GLI proteins in melanoma, and we have identified a large number of target genes, some of them previously known from literature data, but also some novel. In this project, we wish to investigate: (1) which growth factors are directly regulated by HH-GLI signalling on cell models of ovarian, prostate and head and neck cancers, (2) how are the signals transmitted between cells, (3) how do the growth factors affect tumor and stromal cells, (4) does the ratio of tumor to stromal cells affect gene expression in the two populations, (5) what is the effect of HH-GLI and growth factor inhibition (individually or combined) on both populations in vitro.

To answer these questions, we will use cell culture models. The cells will be grown in 2D, but also in 3D cultures, as spheroids. In both setups, co-culture of tumor and stromal cells will be used as well. The results from the first three years of the project will then be validated on patient-derived archive material: tumor RNA, paraffin-embedded tissue slides and primary cell cultures of both tumor and stromal origin. We will be using many standard molecular biology techniques (qPCR, Western blot, immunofluorescence), but also some advanced techniques such as protein antibody arrays, RNA-seq and live cell imaging.

With this project we will demonstrate the role of HH-GLI signalling in tumor-stroma communication, which could lead to advances in combined tumor therapy and to discovery of new prognostic markers.