Molecular and cellular characterization of different cancer cell lines in vitro following irradiation with beams of different qualities: the role of Hedgehog signaling pathway

Radiation therapy seeks to eliminate cancer cells by irradiation-induced DNA damage. Paradoxically, recent research findings have implicated X-ray radiotherapy in tumor recurrence, metastasis and radio-resistance. This has been linked in part, to an X-irradiation-induced up-regulation of the Hedgehog (Hh) signalling pathway. Several studies have associated this pathway with the development and progression of various cancer types into aggressive and radio-resistant forms. The clinical adaptation of charged particles in radiation therapy has increased in the past few years. This is owed to their enhanced relative biological effectiveness and superior dose distribution which spares non-target healthy tissue from exposure to a radiation dose. It is not known how charged particles influence the expression of the Hh signalling pathway genes. We carried out a comparative molecular and cellular characterization of PC3, DAOY and MCF-7 cancer cell lines in vitro following irradiation with varying doses of X-rays, protons or carbon ions. We investigated the effect of ionising radiation on cancer cell survival, migration and expression of Hh signalling pathway genes. We observed a decrease in clonogenic capacity corresponding to higher irradiation doses, irrespective of radiation type in all the three cell types used. This decrease was more pronounced in cells irradiated with higher doses of protons compared to cells irradiated with similar X-ray doses. Wound-healing assay results indicated that in vitro, photon irradiation may amplify migration of PC3 cancer cells and hence need for more research to build up information explaining this observation. In combination with X-rays, inhibiting the Hh pathway with GANT61 did not augment the radiosensitivity of the three cancer cell lines. Our gene expression analyses revealed that different radiation qualities have distinct effects on Hh pathway gene expression and that carbon ions (C-ions) induce more pronounced changes compared to X-rays. Our results provide insights into how X-rays and C-ion irradiation affect the Hh pathway. Given that we observed an increase in expression of some key pathway activating genes corresponding to specific radiation qualities, it is of importance to include more studies both in vitro and in vivo to increase the knowledge on long term effects of ionising irradiation on cancer cells for the optimization and adoption of especially, particle radiation therapy.