Vemurafenib resistance increases melanoma invasiveness and modulates the tumor microenvironment by MMP-2 upregulation

Abstract

The BRAF(V600E) mutation confers constitutive kinase activity and accounts for >90% of BRAF mutations in melanoma. This genetic alteration is a current therapeutic target; however, the antitumorigenic effects of the BRAF(V600E) inhibitor vemurafenib are short-lived and the majority of patients present tumor relapse in a short period after treatment. Characterization of vemurafenib resistance has been essential to the efficacy of next generation therapeutic strategies. Herein, we found that acute BRAF inhibition induced a decrease in active MMP-2, MT1-MMP and MMP-9, but did not modulate the metalloproteinase inhibitors TIMP-2 or RECK in naïve melanoma cells. In vemurafenib-resistant melanoma cells, we observed a lower growth rate and an increase in EGFR phosphorylation followed by the recovery of active MMP-2 expression, a mediator of cancer metastasis. Furthermore, we found a different profile of MMP inhibitor expression, characterized by TIMP-2 downregulation and RECK upregulation. In a 3D spheroid model, the invasion index of vemurafenib-resistant melanoma cells was more evident than in its non-resistant counterpart. We confirmed this pattern in a matrigel invasion assay and demonstrated that use of a matrix metalloproteinase inhibitor reduced the invasion of vemurafenib resistant melanoma cells but not drug naïve cells. Moreover, we did not observe a delimited group of cells invading the dermis in vemurafenib-resistant melanoma cells present in a reconstructed skin model. The same MMP-2 and RECK upregulation profile was found in this 3D skin model containing vemurafenib-resistant melanoma cells. Acute vemurafenib treatment induces the disorganization of collagen fibers and consequently, extracellular matrix remodeling, with this pattern observed even after the acquisition of resistance. Altogether, our data suggest that resistance to vemurafenib induces significant changes in the tumor microenvironment mainly by MMP-2 upregulation, with a corresponding increase in cell invasiveness.

Effects of treadmill exercise on cerebral angiogenesis and MT1‐MMP expression after cerebral ischemia in rats

“…In different cell lines, RECK may play different roles in regulation of MT1‐MMP (Silva et al., 2015. The activity of RECK and MT1‐MMP is essential for angiogenesis (Miki et al, 2010; Sandri et al, 2016). The increased RECK expression will inhibit angiogenesis, and vice versa (Golan, Vagima, & Goichberg, 2011).…”

Role of poly(ε-caprolactone) lipid-core nanocapsules on melanoma–neutrophil crosstalk

“…[1][2][3] Melanoma is extremely resistant to the current therapies such as chemotherapy, radiotherapy, immunotherapy and/or target therapy. [4][5][6] Therefore, the comprehension of the mechanisms underlying the disease is crucial for the establishment of novel and complementary therapeutic approaches.…”

Inhibition of proliferation and invasion in 2D and 3D models by 2-methoxyestradiol in human melanoma cells

“…Calcein and ethidium bromide staining were used as a live/dead cell marker, respectively. The images were imported to Definiens Tissue Studio v4.1 software to analyze the fluorescence as described by Sandri et al, (2016) (22) and the ratio was determined by live fluorescence divided by dead fluorescence.…”

“…The skin model was reconstructed based on the previous description by Sandri and collaborators (2016) with minor changes (22). The SK-Mel-28 melanoma cells were incorporated into the epidermis at the same time as keratinocytes and melanocytes.…”