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Conditioned Medium From Azurin-Expressing Human Mesenchymal Stromal Cells Demonstrates Antitumor Activity Against Breast and Lung Cancer Cell Lines.

ABSTRACT: Recently, cell-based therapies have been explored as a strategy to enhance the specificity of anticancer therapeutic agents. In this perspective, human mesenchymal stromal cells (MSC) hold a promising future as cell delivery systems for anticancer proteins due to their unique biological features. In this study, we engineered human MSC to secrete a human codon-optimized version of azurin (hazu), a bacterial protein that has demonstrated anticancer activity toward different cancer models both in vitro and in vivo. To this end, microporation was used to deliver plasmid DNA encoding azurin into MSC derived from bone marrow (BM) and umbilical cord matrix (UCM), leading to expression and secretion of hazu to the conditioned medium (CM). Engineered hazu-MSC were shown to preserve tumor tropism toward breast (MCF-7) and lung (A549) cancer cell lines, comparable to non-modified MSC. Azurin was detected in the CM of transfected MSC and, upon treatment with hazu-MSC-CM, we observed a decrease in cancer cell proliferation, migration, and invasion, and an increase in cell death for both cancer cell lines. Moreover, expression of azurin caused no changes in MSC expression profile of cytokines relevant in the context of cancer progression, thus suggesting that the antitumoral effects induced by hazu-MSC secretome might be due to the presence of azurin independently. In conclusion, data shown herein indicate that MSC-produced azurin in a CM configuration elicits an anticancer effect.

SUBMITTER: Silva M

PROVIDER: S-EPMC7363770 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Conditioned Medium From Azurin-Expressing Human Mesenchymal Stromal Cells Demonstrates Antitumor Activity Against Breast and Lung Cancer Cell Lines.

Silva Marília M Monteiro Gabriel Amaro GA Fialho Arsenio M AM Bernardes Nuno N da Silva Cláudia Lobato CL

Frontiers in cell and developmental biology 20200709

Recently, cell-based therapies have been explored as a strategy to enhance the specificity of anticancer therapeutic agents. In this perspective, human mesenchymal stromal cells (MSC) hold a promising future as cell delivery systems for anticancer proteins due to their unique biological features. In this study, we engineered human MSC to secrete a human codon-optimized version of azurin (hazu), a bacterial protein that has demonstrated anticancer activity toward different cancer models both <i>i ...[more]

PMID: 32733876

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Project description:BackgroundWhen genes responsible for normal embryonic development are abnormally expressed in adults, it can lead to tumor development. This can suggest that the same mechanism that controls embryonic differentiation can also control tumor differentiation. We hypothesize that the malignant phenotype of lung cancer cells could acquire benign characteristics when in contact with an embryonic lung microenvironment. We cultured two lung cancer cell lines in embryonic lung mesenchyme-conditioned medium and evaluated morphological, functional and molecular changes.MethodsThe human embryonic mesenchymal lung-conditioned medium (hEML-CM) was obtained by culturing lung cells from embryos in the pseudoglandular stage of development. The NSCLC cell lines A549 and H1299 we cultured in the hEML-CM and in a tumor-conditioned medium. Morphological changes were analyzed with optical and transmission electron microscopy. To evaluate the functional effect of conditioned medium in tumor cells, we analyzed cell proliferation, migration, colony formation capacity in 2D and 3D and in vivo tumor growth capacity. The expression of the pluripotency genes OSKM, the adenocarcinoma marker NKX2-1, the lung surfactant proteins SFTP, the myofibroblast marker MYH and DNMT3A/3B was analyzed with qRT-PCR and the presence of the myofibroblast markers vimentin and α-SMA with immunofluorescence. Transcriptomic analysis was performed using Affymetrix arrays.ResultsThe A549 and H1299 cells cultured in hEML-CM lost their epithelial morphology, acquired mesodermal characteristics, and decreased proliferation, migration, and colony formation capacity in 2D and 3D, as well as reduced its capacity to growth in vivo. The expression of OSKM, NKX2-1 and SFTP decreased, while that of DNMT3A/3B, vimentin, α-SMA and MYH increased. Distant matrix analysis based on transcriptomic profile showed that conditioned cells were closer to myoblast and human lung fibroblast than to normal epithelial immortalized lung cells. A total of 1631 for A549 and 866 for H1299 differentially expressed genes between control and conditioned cells were identified.ConclusionsTo the best of our knowledge, this is the first study to report that stimuli from the embryonic lung can modulate the malignant phenotype of lung cancer cells, control their growth capacity and activate their differentiation into myofibroblasts. These findings could lead to new strategies for lung cancer management.

Project description:The endometrium contains a distinct population of immune cells consisting of 70% natural killer (NK) cells that undergo cyclic changes during the menstrual cycle. However, how these uterine NK (uNK) cells interact with uterine stromal cells (SC) remains unclear. We therefore investigated the paracrine effect of medium conditioned by uNK cells on the gene expression profile of endometrial SC in-vitro using a cDNA Microarray. Our results, verified by real-time PCR and ELISA, reveal that soluble factors from uNK cells substantially alter endometrial SC gene expression. The largest group of up-regulated genes found were chemokines and cytokines, including IL-15 and IL-15RÎ±. The latter could produce a niche for uNK cells allowing proliferation within and recruitment into the uterus, as seen in bone marrow. In addition, the most abundantly up-regulated genes, including IL-8, CCL8 and CXCL1 have also been shown to be stimulated by contact of SC with trophoblast, suggesting that uNK cells work synergistically to support the initial trophoblast migration during implantation. Overall this study demonstrates for the first time the paracrine communication between uNK cells and uterine SC, and adds to the understanding of how the uterine immune system contributes to the changes seen within the cycling endometrium. Keywords: Response of endometrial stromal cells to uNK conditioned medium This study was designed to identify the response of non-decidualised stromal cells from the endometrium, to soluble factors secreted by uterine NK cells. Endometrial stromal cells were isolated from 7 patients and treated with control medium or medium conditioned by uterine Nk cells. THE 'REF' COLUMN ON EACH ARRAY IS THE SIGNAL PRODUCED BY A COMMON REFERNCE RNA SAMPLE THAT WAS LABELLED AS A SINGLE BATCH SAMPLE AND HYBRIDISED TO ALL THE ARRAYS- IT IS A 'COMMON REFERENCE'. THE 'TEST' SAMPLE COMPRISES EACH INDIVIDUAL SAMPLE OF CELLS TREATED AS DESCRIBED IN THE SERIES SUBMISSION. FOR EXAMPLE; SAMPLE Y1 (gsm2435820) IS RNA FROM PATIENT 1 TREATED WITH CONTROL MEDIUM, SAMPLE G1 (GSM245371) IS RNA FROM PATIENT 1 TREATED WITH NK CONDITIONED MEDIUM THESE TWO SAMPLES THEREFORE FORM A PAIR- IE CELLS FROM THE SAME PATIENT TREATED WITH CONTROL OR NK-CONDITIONED MIDIUM. Y2,G2 ARE FROM PATIENT 2, ETC

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Conditioned Medium From Azurin-Expressing Human Mesenchymal Stromal Cells Demonstrates Antitumor Activity Against Breast and Lung Cancer Cell Lines.

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Conditioned Medium From Azurin-Expressing Human Mesenchymal Stromal Cells Demonstrates Antitumor Activity Against Breast and Lung Cancer Cell Lines.

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