Exploiting Zebrafish Xenografts for Testing the in vivo Antitumorigenic Activity of Microcin E492 Against Human Colorectal Cancer Cells

Abstract

One of the approaches to address cancer treatment is to develop new drugs not only to obtain compounds with less side effects, but also to have a broader set of alternatives to tackle the resistant forms of this pathology. In this regard, growing evidence supports the use of bacteria-derived peptides such as bacteriocins, which have emerged as promising anti-cancer molecules. In addition to test the activity of these molecules on cancer cells in culture, their in vivo antitumorigenic properties must be validated in animal models. Although the standard approach for such assays employs experiments in nude mice, at the initial stages of testing, the use of high-throughput animal models would permit rapid proof-of-concept experiments, screening a high number of compounds, and thus increasing the possibilities of finding new anti-cancer molecules. A validated and promising alternative animal model are zebrafish larvae harboring xenografts of human cancer cells. Here, we addressed the anti-cancer properties of the antibacterial peptide microcin E492 (MccE492), a bacteriocin produced by Klebsiella pneumoniae, showing that this peptide has a marked cytotoxic effect on human colorectal cancer cells in vitro. Furthermore, we developed a zebrafish xenograft model using these cells to test the antitumor effect of MccE492 in vivo, demonstrating that intratumor injection of this peptide significantly reduced the tumor cell mass. Our results provide, for the first time, evidence of the in vivo antitumoral properties of a bacteriocin tested in an animal model. This evidence strongly supports the potential of this bacteriocin for the development of novel anti-cancer therapies.

Bacteriocins: An Overview of Antimicrobial, Toxicity, and Biosafety Assessment by in vivo Models

“…On the other hand, Acuña et al (2012) were the first to design chimeric bacteriocins that retained the properties to kill both Gram-positive and Gram-negative bacteria. Moreover, other authors have preferred to repurpose the bacteriocins by exploiting their capability against tumor cells ( Varas et al, 2020 ) or by exploring synergistic effects of bacteriocins while combining with renowned antibiotics ( Mathur et al, 2017 ) or with other bacteriocins ( Hanchi et al, 2017 ).…”

Zebrafish as a Model for Anticancer Nanomedicine Studies

“…Therefore, using 2 dpf embryos could allow the spread, metastasis, and survival of cancer cells without the need for a prior immunosuppressive treatment [ 16 ]. Several types of primary tumor cells or cancer cell lines have been transplanted to generate several different cancer xenograft zebrafish models such as, melanoma [ 48 ], leukemia [ 49 ], colorectal [ 50 ], kidney [ 51 ], breast [ 52 ], ovarian [ 53 ], prostate [ 54 ], pancreatic [ 55 ], oral [ 56 ] and lung [ 57 ] cancers’ models.…”

Colorectal cancer treatment using bacteria: focus on molecular mechanisms

“…Nisin is a positive charged molecule that causes pore development in the cell membrane of the target organism and thereby results in cytoplasmic membrane depolarization [ 31 ]. Nisin is synthesized and secreted by Lactococcus lactis appeared to have a considerable cytotoxic effect against CRC cell lines such as Caco-2 and HT-29 colorectal cancer cell lines [ 32 ]. In a study that the apoptotic effect of nisin on the SW480 cells was investigated, cell viability and Bax/Bcl-2 ratio at both mRNA and protein levels were considered.…”

“…For in vivo investigation, SW620 zebrafish larvae xenografts were developed. The intratumoral injection of microcin E492 reduced tumor growth significantly [ 32 ]. Studies indicated microcin depolarizes cell membrane potential by pore-forming ability, and also DNA fragmentation, phosphatidylserine release, caspase activity, and releasing of intracellular calcium ions are the principle mechanisms of apoptotic cell death by inducing microcin [ 39 ].…”

“…Cancer cell studies demonstrated numerous vacuoles and autolysosomes and autophagosomes structures in the cells. Entap has anti-proliferative activity against prostatic carcinoma (22Rv1), mammary gland adenocarcinoma (MDA-MB-231), human gastric adenocarcinoma (AGS), uterine cervix adenocarcinoma (HeLa), and colorectal adenocarcinoma (HT-29) cell lines [ 32 ].…”