ADAMTS3 restricts cancer invasion in models of early breast cancer progression through enhanced fibronectin degradation

•Metzincins display both pro- and anti-invasive effects in breast cancer.•Loss of myoepithelial ADAMTS3 expression drives invasion.•Degradomic analysis implicates fibronectin as a novel ADAMTS3 substrate.•ADAMTS3 restricts cell invasion through fibronectin degradation and reduced integrin α5β1 signalling. Proteases have long been associated with cancer progression, due to their ability to facilitate invasion upon matrix remodelling. However, proteases are not simply degraders of the matrix, but also play fundamental roles in modulating cellular behaviour through the proteolytic processing of specific substrates. Indeed, proteases can elicit both pro- and anti- tumorigenic effects depending on context. Using a heterocellular spheroid model of breast cancer progression, we demonstrate the repressive function of myoepithelial ADAMTS3, with its loss directing myoepithelial-led invasion of luminal cells through a physiologically relevant matrix. Degradomic analysis, using terminal amine isotopic labelling of substrates (TAILS), combined with functional assays, implicate ADAMTS3 as a mediator of fibronectin degradation. We show further that loss of ADAMTS3 enhances levels of fibronectin in the microenvironment, promoting invasion through canonical integrin α5β1 activation. Our data highlight a tumour suppressive role for ADAMTS3 in early stage breast cancer, and contribute to the growing evidence that proteases can restrain cancer progression.