TLR4-mediated immunomodulatory properties of the bacterial metalloprotease arazyme in preclinical tumor models

Abstract

Despite the recent approval of new agents for metastatic melanoma, its treatment remains challenging. Moreover, few available immunotherapies induce a strong cellular immune response, and selection of the correct immunoadjuvant is crucial for overcoming this obstacle. Here, we studied the immunomodulatory properties of arazyme, a bacterial metalloprotease, which was previously shown to control metastasis in a murine melanoma B16F10-Nex2 model. The antitumor activity of arazyme was independent of its proteolytic activity, since heat-inactivated protease showed comparable properties to the active enzyme; however, the effect was dependent on an intact immune system, as antitumor properties were lost in immunodeficient mice. The protective response was IFNγ-dependent, and CD8(+) T lymphocytes were the main effector antitumor population, although B and CD4(+) T lymphocytes were also induced. Macrophages and dendritic cells were involved in the induction of the antitumor response, as arazyme activation of these cells increased both the expression of surface activation markers and proinflammatory cytokine secretion through TLR4-MyD88-TRIF-dependent, but also MAPK-dependent pathways. Arazyme was also effective in the murine breast adenocarcinoma 4T1 model, reducing primary and metastatic tumor development, and prolonging survival. To our knowledge, this is the first report of a bacterial metalloprotease interaction with TLR4 and subsequent receptor activation that promotes a proinflammatory and tumor protective response. Our results show that arazyme has immunomodulatory properties, and could be a promising novel alternative for metastatic melanoma treatment.

Combination Treatment of Arazyme and Soy Leaf Extract Attenuates Hyperglycemia and Hepatic Steatosis in High-Fat Diet-Fed C57BL/6J Mice

“…Previous studies have shown that Arazyme exerts hepatoprotective effects by improving CCl 4 -induced acute hepatic injury, HFD-induced chronic hepatic injury, and tumor development [ 16 , 17 , 22 , 23 ]. In addition, Arazyme treatment inhibits the expression of inflammatory cytokines, chemokines, and reactive oxygen species in lipopolysaccharide-induced human endothelial cells [ 24 ].…”

Ethanolic Extract of Senna velutina Roots: Chemical Composition, In Vitro and In Vivo Antitumor Effects, and B16F10-Nex2 Melanoma Cell Death Mechanisms

“…Accordingly, several anticancer drugs of natural origin are available on the market [31]. In this context, scientific studies have shown that Brazilian biodiversity due to its various biomes provides various natural compounds with anticancer potential both in vitro [32, 33] and in vivo [30, 34]. In the present study, we assessed the anticancer effects of the ethanolic extract of the roots of S. velutina , a plant species native to Brazil whose phytochemical composition and potential pharmacological applications have been poorly studied.…”

“…The experiment was conducted according to Pereira et al in 2016 [30] with minor modifications. Thus, 5 × 10 5 B16F10-Nex2 melanoma cells were injected through the caudal vein into C57Bl/6 mice (five animals per group).…”

A comparative study of the arazyme-based fusion proteins with various ligands for more effective targeting cancer therapy: an in-silico analysis

“…Arazyme (AraA), an extracellular metalloproteinase secreted by Serratiaproteamaculans, has strong proteolytic activity on various protein substrates such as albumin, elastin, creatinine, and collagen in a wide pH and temperature range( 8 9 ). Several studies have reported the strong anticancer activity of AraA, especially its antimetastatic effect on various cancers such as colon, ovarian, and melanoma cancer cell lines( 8 10 11 ). Ghadaksaz et al have investigated a fusion protein consisting of transforming growth factor alpha third loop (TGFαL3)-targeted AraA for breast cancer therapy( 12 ).…”