287 Mass spectrometry-based secretome analysis of melanocyte-keratinocyte-fibroblast co-culture systems

Abstract

Melanoma is an aggressive form of skin cancer in the later stages of the disease. Despite recent advances in targeted and immunotherapies the relapse rate among melanoma patients remains significant. The TME has been shown in other cancers and melanoma to provide a protective niche where the malady can reside and resist treatment. It has been shown that the increase in secreted fibroblasts growth factors can in turn lead to melanoma cell growth and further para/autocrine signaling, leading to increased survival of the tumour. This altered TME can have a negative effect on the efficacy of treatment and may be a contributing factor to the relapse rate amongst melanoma patients. A better understanding of the interplay between the secreted factors of the melanoma TME would be beneficial to therapy. Here, using transwell inserts we have simulated the melanoma microenvironment by allowing the cells to physically interact or be separated from other skin cells while still being able to communicate via secreted signals. Melanoma cells and keratinocytes were seeded in a transwell insert and normal human dermal fibroblasts or patient derived cancer associated fibroblasts were grown below separated by a semipermeable membrane. Different combinations were also explored to represent a move advance in vivo setting, and combinations of pigmented and unpigmented melanoma cell lines. Our preliminary data suggests that the proteins secreted from the CAF coculture, the NHDF coculture and melanoma cells alone are fundamentally different. Amongst the CAF coculture there are more factors associated with ECM organization and signaling cell fate than the normal fibroblast secretome. This MS approach in combination with simulation of the TME in transwells allows us to identify and exploit novel factors involved in melanoma signaling and cell fate.