Cytotoxicity of purified listeriolysin O on mouse and human leukocytes and leukaemia cells

Stachowiak, Radosław; Lyzniak, M.; Grabowska, Maja; Roeske, Katarzyna; Jagielski, Tomasz; Bielecki, Jacek; Budziszewska, Bożena Katarzyna; Hoser, Grazyna; Kawiak, Jerzy

doi:10.1186/1472-6750-14-77

Cited by 5 publications

(5 citation statements)

References 18 publications

(22 reference statements)

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“…The anticancer properties of this factor have been previously studied in various cell lines, such as Jurkat cells or human peripheral blood mononuclear cells. Similar to other CDCs, LLO may display some specificity [ 34 , 36 ]. However, it is necessary to further modify this toxin to ensure selectiveness towards cancer cells.…”

Section: Resultsmentioning

confidence: 99%

“…The LLO, LLO-GFP and GFP-LLO proteins were purified from the bacterial cell lysates using Ni-NTA resin columns via affinity chromatography and concentrated with a centrifugal concentrator. Construction and purity was confirmed by SDS PAGE and western blot, and activity was assessed using the hemolytic test [ 34 ]. The final concentration 0.6 μg/ml was estimated by NanoDrop spectrophotometer.…”

Section: Methodsmentioning

confidence: 99%

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Stabilized nanosystem of nanocarriers with an immobilized biological factor for anti-tumor therapy

et al. 2017

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ObjectiveThe inadequate efficiency of existing therapeutic anti-cancer regiments and the increase in the multidrug resistance of cancer cells underscore the need to investigate novel anticancer strategies. The induction of apoptosis in tumors by cytotoxic agents produced by pathogenic microorganisms is an example of such an approach. Nevertheless, even the most effective drug should be delivered directly to targeted sites to reduce any negative impact on other cells. Accordingly, the stabilized nanosystem (SNS) for active agent delivery to cancer cells was designed for further application in local anti-tumor therapy. A product of genetically modified Escherichia coli, listeriolysin O (LLO), was immobilized within the polyelectrolyte membrane (poly(ethylenimine)|hyaluronic acid) shells of ‘LLO nanocarriers’ coupled with the stabilizing element of natural origin.Methods and resultsThe impact of LLO was evaluated in human leukemia cell lines in vitro. Correspondingly, the influence of the SNS and its elements was assessed in vitro. The viability of targeted cells was evaluated by flow cytometry. Visualization of the system structure was performed using confocal microscopy. The membrane shell applied to the nanocarriers was analyzed using atomic force microscopy and Fourier transform infrared spectroscopy techniques. Furthermore, the presence of a polyelectrolyte layer on the nanocarrier surface and/or in the cell was confirmed by flow cytometry. Finally, the structural integrity of the SNS and the corresponding release of the fluorescent solute listeriolysin were investigated.ConclusionThe construction of a stabilized system offers LLO release with a lethal impact on model eukaryotic cells. The applied platform design may be recommended for local anti-tumor treatment purposes.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Stabilized nanosystem of nanocarriers with an immobilized biological factor for anti-tumor therapy

et al. 2017

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“…To our knowledge, this is the first study in which wt LLO or its mutants have been used on urothelial cells. To date, wt LLO activity has been studied in a variety of other cell types, including colon epithelial cells Caco-2 and HT-29/B6, embryonic kidney cells HEK 293, human brain endothelial cells, and Jurkat cells, each without comparing the response of wt LLO in cancer vs. normal cells [ 34 , 35 , 36 , 37 , 38 ]. Herein, we initially planned to use LLO Y406A, designed for activity at a narrow pH optimum of 5–6 [ 11 ] to prevent pore-forming activity at the plasma membrane with usually neutral pH surrounding, which should induce selective pore-forming activity in acidic endosomes.…”

Section: Discussionmentioning

confidence: 99%

Cytotoxic Activity of LLO Y406A Is Targeted to the Plasma Membrane of Cancer Urothelial Cells

Resnik

Tratnjek

Kreft

et al. 2021

IJMS

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Identification of novel agents for bladder cancer treatment is highly desirable due to the high incidence of tumor recurrence and the risk of progression to muscle-invasive disease. The key feature of the cholesterol-dependent toxin listeriolysin O mutant (LLO Y406A) is its preferential activity at pH 5.7, which could be exploited either directly for selective targeting of cancer cells or the release of accumulated therapeutics from acidic endosomes. Therefore, our goal was to compare the cytotoxic effect of LLO Y406A on cancer cells (RT4) and normal urothelial cells (NPU), and to identify which cell membranes are the primary target of LLO Y406A by viability assays, life-cell imaging, fluorescence, and electron microscopy. LLO Y406A decreased viability, altered cell morphology, provoked membrane blebbing, and induced apoptosis in RT4 cells, while it did not affect NPU cells. LLO Y406A did not cause endosomal escape in RT4 cells, while the plasma membrane of RT4 cells was revealed as the primary target of LLO Y406A. It has been concluded that LLO Y406A has the ability to selectively eliminate cancer urothelial cells through pore-forming activity at the plasma membrane, without cytotoxic effects on normal urothelial cells. This promising selective activity merits further testing as an anti-cancer agent.

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“…Also for a long time therapies have been applied in which antibodies administered to patients were linked to isotopes (Ibritumomab). Attempts were also made to use bacterial toxins or to amplify with them effects of antibodies [42,133].…”

Section: Traditional Approach To Tumour Treatment: Chemotherapy Radimentioning

confidence: 99%

Problems of Cancer Treatment. Part I. Theory of Treatment Based on Known Mechanisms of Anticancer Immunological Responses

Kawiak

Hoser

Domagała‐Kulawik

2017

Advances in Cell Biology

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Summary: Various processes, taking place both in cells and in their environment, are linked to carcinogenesis. This paper aims at recalling the complex mechanisms of oncogenesis, with particular attention paid to responses of the immune system. In development of solid tumours, leukaemias and lymphomas several common stages can be noted. A neoplastic disease cannot be understood considering only phenomena of genetic mutations. Neoplastic cells are characterised by an extensive antigenic variability and resistance to apoptosis. The cells create around them a microenvironment which protects them from defensive activity of the host. In the paper we present the recognised mechanisms of anti-neoplastic defense as well as several elements allowing the solid tumours and leukaemias to escape from the immune surveillance. The generally accepted treatment of tumours aims at reducing numbers of tumour cells. Following resection of a tumour, radiotherapy or chemotherapy, the parallel or consecutive stage of treatment was found to involve an increase in number of clones of immune system cells. One of the ways in which the immune system can be activated involves autovaccination of the host with own neoplastic cells in an apoptosis. However, attempts of such a therapy frequently brought no expected results due to blocked activity of cytotoxic cells. Therefore, the subsequent stage in activation of the immune system should involve elimination of the tumor-mobilized blockade of the system. Attempts toward this aim include neutralization of the tumour-blocked cytotoxic properties of defensive cells, first of all T lymphocytes. The recognized mechanisms of blocking T cells activity in the PD-1/PD-L1 system or due to inhibition of activation by CTLA-4 molecule provided rationale for development of effective tumour immunotherapy approaches. Keywords

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Cytotoxicity of purified listeriolysin O on mouse and human leukocytes and leukaemia cells

Cited by 5 publications

References 18 publications

Stabilized nanosystem of nanocarriers with an immobilized biological factor for anti-tumor therapy

Stabilized nanosystem of nanocarriers with an immobilized biological factor for anti-tumor therapy

Cytotoxic Activity of LLO Y406A Is Targeted to the Plasma Membrane of Cancer Urothelial Cells

Problems of Cancer Treatment. Part I. Theory of Treatment Based on Known Mechanisms of Anticancer Immunological Responses

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