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Transgenic plants as low-cost platform for chemotherapeutic drugs screening.

Vergara D, de Domenico S, Maffia M, Piro G, Di Sansebastiano GP - Int J Mol Sci (2015)

Bottom Line: Our results demonstrated that drugs tested alone or in combination differentially influenced the monitored cellular processes including cytoskeletal organization and endomembrane trafficking.In conclusion, we demonstrated that A. thaliana plants are sensitive to the action of human chemotherapeutics and can be used for preliminary screening of drugs efficacy.The cost-effective subcellular imaging in plant cell may contribute to better clarify drugs subcellular targets and their anticancer effects.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological and Environmental Sciences and Technologies (DiSTeBA), University of Salento, Campus ECOTEKNE, S.P. 6, Lecce-Monteroni, Lecce 73100, Italy. danielevergara@libero.it.

ABSTRACT
In this work we explored the possibility of using genetically modified Arabidopsis thaliana plants as a rapid and low-cost screening tool for evaluating human anticancer drugs action and efficacy. Here, four different inhibitors with a validated anticancer effect in humans and distinct mechanism of action were screened in the plant model for their ability to interfere with the cytoskeletal and endomembrane networks. We used plants expressing a green fluorescent protein (GFP) tagged microtubule-protein (TUA6-GFP), and three soluble GFPs differently sorted to reside in the endoplasmic reticulum (GFPKDEL) or to accumulate in the vacuole through a COPII dependent (AleuGFP) or independent (GFPChi) mechanism. Our results demonstrated that drugs tested alone or in combination differentially influenced the monitored cellular processes including cytoskeletal organization and endomembrane trafficking. In conclusion, we demonstrated that A. thaliana plants are sensitive to the action of human chemotherapeutics and can be used for preliminary screening of drugs efficacy. The cost-effective subcellular imaging in plant cell may contribute to better clarify drugs subcellular targets and their anticancer effects.

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Related in: MedlinePlus

Fluorescent patterns of GFPChi in transgenic Arabidopsis leaf epidermis treated with low doses of two combined drugs. Treatment with 0.2 μM Crizotinib had a moderate effect on GFPChi (A); but it was enhanced when combined with 10 μM Paclitaxel disturbing intermediate steps of its sorting (B) or causing mis-sorting to the apoplast when combined with 25 μM Y-27632 (C); combined treatment with 0.01 μM Sorafenib caused the increase of fluorescence in the central and small vacuoles (D). GFP fluorescence and chlorophyll autofluorescence are shown in green and blue, respectively. Scale bar = 20 μm.
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ijms-16-02174-f003: Fluorescent patterns of GFPChi in transgenic Arabidopsis leaf epidermis treated with low doses of two combined drugs. Treatment with 0.2 μM Crizotinib had a moderate effect on GFPChi (A); but it was enhanced when combined with 10 μM Paclitaxel disturbing intermediate steps of its sorting (B) or causing mis-sorting to the apoplast when combined with 25 μM Y-27632 (C); combined treatment with 0.01 μM Sorafenib caused the increase of fluorescence in the central and small vacuoles (D). GFP fluorescence and chlorophyll autofluorescence are shown in green and blue, respectively. Scale bar = 20 μm.

Mentions: Plantlets were treated with a low dose of Crizotinib (0.2 μM) in combination with Paclitaxel (10 μM), Y-27632 (25 μM), and Sorafenib (0.01 μM). It was hypothesized that the observed effect was influenced by a higher stress of tissue and for this reason the alterations taken into account had to be homogeneously visible throughout the tissue examined. With a longer treatment, up to 36–40 h, Crizotinib alone (0.2 μM) was already able to induce some of the symptoms typical of higher doses. Some cell kinds, possibly not fully differentiated (e.g., cells flanking the differentiating stomata) and not used for the screening, showed to be particularly sensitive (not shown). To evaluate synergistic effects, only stronger and extensive effects were taken into consideration. In doing so, some interesting interactions were evidenced. These effects were particularly evident on GFPChi because this marker sorting was already weakly affected by 0.2 μM Crizotinib (Figure 3A).


Transgenic plants as low-cost platform for chemotherapeutic drugs screening.

Vergara D, de Domenico S, Maffia M, Piro G, Di Sansebastiano GP - Int J Mol Sci (2015)

Fluorescent patterns of GFPChi in transgenic Arabidopsis leaf epidermis treated with low doses of two combined drugs. Treatment with 0.2 μM Crizotinib had a moderate effect on GFPChi (A); but it was enhanced when combined with 10 μM Paclitaxel disturbing intermediate steps of its sorting (B) or causing mis-sorting to the apoplast when combined with 25 μM Y-27632 (C); combined treatment with 0.01 μM Sorafenib caused the increase of fluorescence in the central and small vacuoles (D). GFP fluorescence and chlorophyll autofluorescence are shown in green and blue, respectively. Scale bar = 20 μm.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4307356&req=5

ijms-16-02174-f003: Fluorescent patterns of GFPChi in transgenic Arabidopsis leaf epidermis treated with low doses of two combined drugs. Treatment with 0.2 μM Crizotinib had a moderate effect on GFPChi (A); but it was enhanced when combined with 10 μM Paclitaxel disturbing intermediate steps of its sorting (B) or causing mis-sorting to the apoplast when combined with 25 μM Y-27632 (C); combined treatment with 0.01 μM Sorafenib caused the increase of fluorescence in the central and small vacuoles (D). GFP fluorescence and chlorophyll autofluorescence are shown in green and blue, respectively. Scale bar = 20 μm.
Mentions: Plantlets were treated with a low dose of Crizotinib (0.2 μM) in combination with Paclitaxel (10 μM), Y-27632 (25 μM), and Sorafenib (0.01 μM). It was hypothesized that the observed effect was influenced by a higher stress of tissue and for this reason the alterations taken into account had to be homogeneously visible throughout the tissue examined. With a longer treatment, up to 36–40 h, Crizotinib alone (0.2 μM) was already able to induce some of the symptoms typical of higher doses. Some cell kinds, possibly not fully differentiated (e.g., cells flanking the differentiating stomata) and not used for the screening, showed to be particularly sensitive (not shown). To evaluate synergistic effects, only stronger and extensive effects were taken into consideration. In doing so, some interesting interactions were evidenced. These effects were particularly evident on GFPChi because this marker sorting was already weakly affected by 0.2 μM Crizotinib (Figure 3A).

Bottom Line: Our results demonstrated that drugs tested alone or in combination differentially influenced the monitored cellular processes including cytoskeletal organization and endomembrane trafficking.In conclusion, we demonstrated that A. thaliana plants are sensitive to the action of human chemotherapeutics and can be used for preliminary screening of drugs efficacy.The cost-effective subcellular imaging in plant cell may contribute to better clarify drugs subcellular targets and their anticancer effects.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological and Environmental Sciences and Technologies (DiSTeBA), University of Salento, Campus ECOTEKNE, S.P. 6, Lecce-Monteroni, Lecce 73100, Italy. danielevergara@libero.it.

ABSTRACT
In this work we explored the possibility of using genetically modified Arabidopsis thaliana plants as a rapid and low-cost screening tool for evaluating human anticancer drugs action and efficacy. Here, four different inhibitors with a validated anticancer effect in humans and distinct mechanism of action were screened in the plant model for their ability to interfere with the cytoskeletal and endomembrane networks. We used plants expressing a green fluorescent protein (GFP) tagged microtubule-protein (TUA6-GFP), and three soluble GFPs differently sorted to reside in the endoplasmic reticulum (GFPKDEL) or to accumulate in the vacuole through a COPII dependent (AleuGFP) or independent (GFPChi) mechanism. Our results demonstrated that drugs tested alone or in combination differentially influenced the monitored cellular processes including cytoskeletal organization and endomembrane trafficking. In conclusion, we demonstrated that A. thaliana plants are sensitive to the action of human chemotherapeutics and can be used for preliminary screening of drugs efficacy. The cost-effective subcellular imaging in plant cell may contribute to better clarify drugs subcellular targets and their anticancer effects.

Show MeSH
Related in: MedlinePlus