Cathepsin D Specifically Cleaves the Chemokines Macrophage Inflammatory Protein-1α, Macrophage Inflammatory Protein-1β, and SLC That Are Expressed in Human Breast Cancer

Abstract

Cathepsin D (Cath-D) expression in human primary breast cancer has been associated with a poor prognosis. In search of a better understanding of the Cath-D substrates possibly involved in cancer invasiveness and metastasis, we investigated the potential interactions between this protease and chemokines. Here we report that purified Cath-D, as well as culture supernatants from the human breast carcinoma cell lines MCF-7 and T47D, selectively degrade macrophage inflammatory protein (MIP)-1 alpha (CCL3), MIP-1 beta (CCL4), and SLC (CCL21). Proteolysis was totally blocked by the protease inhibitor pepstatin A, and specificity of Cath-D cleavage was demonstrated using a large chemokine panel. Whereas MIP-1 alpha and MIP-1 beta degradation was rapid and complete, cleavage of SLC was slow and not complete. Mass spectrometry analysis showed that Cath-D cleaves the Leu(58) to Trp(59) bond of SLC producing two functionally inactive fragments. Analysis of Cath-D proteolysis of a series of monocyte chemoattractant protein-3/MIP-1 beta hybrids indicated that processing of MIP-1 beta might start by cleaving off amino acids located in the C-terminal domain. In situ hybridization studies revealed MIP-1 alpha, MIP-1 beta, and Cath-D gene expression mainly in the stromal compartment of breast cancers whereas SLC transcripts were found in endothelial cells of capillaries and venules within the neoplastic tissues. Cath-D production in the breast carcinoma cell lines MCF-7 and T47D, as assessed by enzyme-linked immunosorbent assay of culture supernatants and cell lysates, was not affected by stimulation with chemokines such as interleukin-8 (CXCL8), SDF-1 (CXCL12), and SLC. These data suggest that inactivation of chemokines by Cath-D possibly influences regulatory mechanisms in the tumoral extracellular microenvironment that in turn may affect the generation of the antitumoral immune response, the migration of cancer cells, or both processes.

Native matrix-based human lung alveolar tissue model in vitro: studies of the reparatory actions of mesenchymal stem cells

“…MMP-9, and their up-regulation reflects similar tendencies reported by other authors (Braber et al 2011;Overbeek et al 2013). In general, expression of MMPs in lung tissue is described as a characteristic of tissue remodeling associated with various biological processes, such as development, inflammation, migration of cells, wound repair and other (Legrand et al 1999;Itoh et al 2002;Barnes et al 2003;Wolf et al 2003;Greenlee et al 2007;Crosby and Waters 2010). Secretion of pro-inflammatory chemokine GROa was increased by CSE in our native matrix model in contrast to some other reports (Victoni et al 2014) suggesting that study model and matrix origin might be important.…”

Proteases as modulators of tumor–stromal interaction: Primary tumors to bone metastases

“…Cathepsin D potentially modifies the microenvironment and tumor-stromal interactions in a way that affects immune response and migration of cancer cells [34]. Cathepsin D has been shown to cleave and inactivate macrophage inflammatory protein (MIP)-1α (CCL3), MIP-1β (CCL4), and SLC (CCL21) [34].…”

“…Cathepsin D potentially modifies the microenvironment and tumor-stromal interactions in a way that affects immune response and migration of cancer cells [34]. Cathepsin D has been shown to cleave and inactivate macrophage inflammatory protein (MIP)-1α (CCL3), MIP-1β (CCL4), and SLC (CCL21) [34]. CCL3 and CCL4 are typically expressed by the stroma of breast cancers as demonstrated by in situ hybridization, while CCL21 is typically expressed by endothelial cells of capillaries and venules within the tumor tissue [34].…”

“…Cathepsin D has been shown to cleave and inactivate macrophage inflammatory protein (MIP)-1α (CCL3), MIP-1β (CCL4), and SLC (CCL21) [34]. CCL3 and CCL4 are typically expressed by the stroma of breast cancers as demonstrated by in situ hybridization, while CCL21 is typically expressed by endothelial cells of capillaries and venules within the tumor tissue [34]. These chemokines are potentially capable of generating an immune response to the newly formed tumor.…”

“…These chemokines are potentially capable of generating an immune response to the newly formed tumor. By degrading chemokines, Cathepsin D modifies the relationship of the tumor to the stroma in a manner that allows tumor cells to escape the immune system and thrive unchecked (Figure 1) [34].…”