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Modified bleomycin disaccharides exhibiting improved tumor cell targeting.

Madathil MM, Bhattacharya C, Yu Z, Paul R, Rishel MJ, Hecht SM - Biochemistry (2014)

Bottom Line: In earlier studies, we have demonstrated that the tumor cell selectivity resides in the mannose carbamoyl moiety of the BLM saccharide and that both the BLM disaccharide and monosaccharide containing the carbamoyl moiety were capable of the delivery/uptake of a conjugated cyanine dye into cultured cancer cell lines.A library of seven disaccharide-Cy5** dye conjugates was prepared that are structural analogues of the BLM disaccharide.These differed from the natural BLM disaccharide in the position, orientation, and substitution of the carbamoyl group.

View Article: PubMed Central - PubMed

Affiliation: Center for Bioenergetics, Biodesign Institute, and Department of Chemistry and Biochemistry, Arizona State University , Tempe, Arizona 85287, United States.

ABSTRACT
The bleomycins (BLMs) are a family of antitumor antibiotics used clinically for anticancer chemotherapy. Their antitumor selectivity derives at least in part from their ability to target tumor cells, a property that resides in the carbohydrate moiety of the antitumor agent. In earlier studies, we have demonstrated that the tumor cell selectivity resides in the mannose carbamoyl moiety of the BLM saccharide and that both the BLM disaccharide and monosaccharide containing the carbamoyl moiety were capable of the delivery/uptake of a conjugated cyanine dye into cultured cancer cell lines. Presently, the nature of the participation of the carbamoyl moiety has been explored further to provide compounds of utility for defining the nature of the mechanism of tumor cell recognition and uptake by BLM saccharides and in the hope that more efficient compounds could be identified. A library of seven disaccharide-Cy5** dye conjugates was prepared that are structural analogues of the BLM disaccharide. These differed from the natural BLM disaccharide in the position, orientation, and substitution of the carbamoyl group. Studies of these compounds in four matched sets of tumor and normal cell lines revealed a few that were both tumor cell selective and internalized 2-4-fold more efficiently than the natural BLM disaccharide.

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(Top)Comparison of binding/uptake of disaccharide–Cy5**conjugates 1, 2, 4, 5, and 9 in BxPC-3 and SVR A221a cell lines. The cellswere treated with 25 μM disaccharide–Cy5** conjugatesat 37 °C for 1 h, washed with PBS, and fixed with 4% paraformaldehyde.The cell nuclei were stained with 2-(4-amidinophenyl)-6-indolecarbamidine(DAPI). Fluorescence imaging was carried out with a 2 s exposure time.(Bottom) Quantification of the binding/uptake of disaccharide–Cy5**conjugates 1–9 in BxPC-3 and SVRA221a cell lines. The cells were treated with 25 μM dye conjugates,irradiated for 2 s prior to imaging, and then analyzed using a ZeissAxiovert 200M inverted microscope with a 40× oil objective.
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fig3: (Top)Comparison of binding/uptake of disaccharide–Cy5**conjugates 1, 2, 4, 5, and 9 in BxPC-3 and SVR A221a cell lines. The cellswere treated with 25 μM disaccharide–Cy5** conjugatesat 37 °C for 1 h, washed with PBS, and fixed with 4% paraformaldehyde.The cell nuclei were stained with 2-(4-amidinophenyl)-6-indolecarbamidine(DAPI). Fluorescence imaging was carried out with a 2 s exposure time.(Bottom) Quantification of the binding/uptake of disaccharide–Cy5**conjugates 1–9 in BxPC-3 and SVRA221a cell lines. The cells were treated with 25 μM dye conjugates,irradiated for 2 s prior to imaging, and then analyzed using a ZeissAxiovert 200M inverted microscope with a 40× oil objective.

Mentions: The cell binding/uptakeof BLM disaccharide–Cy5** conjugate(1), decarbamoyl BLM disaccharide–Cy5** conjugate(2), and the newly synthesized disaccharide–Cy5**conjugates 3–9 by BxPC-3 pancreaticcarcinoma cells and SVR A221a normal pancreatic cells (Figures 3 and S1), by A549 lungcarcinoma cells and WI-38 normal lung cells (Figures 4 and S2), by DU-145 prostate carcinomacells and PZ-HPV-7 normal prostate cells (Figures 5 and S3), and by A498 kidney carcinomacells and CCD-1105 KIDTr normal kidney cells (Figures 6 and S4) was quantified by fluorescenceimaging. As shown (Figures 3–6), in all tumor and normal cell lines tested, thedecarbamoyl BLM disaccharide exhibited very low binding and uptake,highlighting the importance of the carbamoyl moiety to effective cellbinding and uptake. This is in agreement with earlier studies reportedby our laboratory.15,21,22


Modified bleomycin disaccharides exhibiting improved tumor cell targeting.

Madathil MM, Bhattacharya C, Yu Z, Paul R, Rishel MJ, Hecht SM - Biochemistry (2014)

(Top)Comparison of binding/uptake of disaccharide–Cy5**conjugates 1, 2, 4, 5, and 9 in BxPC-3 and SVR A221a cell lines. The cellswere treated with 25 μM disaccharide–Cy5** conjugatesat 37 °C for 1 h, washed with PBS, and fixed with 4% paraformaldehyde.The cell nuclei were stained with 2-(4-amidinophenyl)-6-indolecarbamidine(DAPI). Fluorescence imaging was carried out with a 2 s exposure time.(Bottom) Quantification of the binding/uptake of disaccharide–Cy5**conjugates 1–9 in BxPC-3 and SVRA221a cell lines. The cells were treated with 25 μM dye conjugates,irradiated for 2 s prior to imaging, and then analyzed using a ZeissAxiovert 200M inverted microscope with a 40× oil objective.
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Related In: Results  -  Collection

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fig3: (Top)Comparison of binding/uptake of disaccharide–Cy5**conjugates 1, 2, 4, 5, and 9 in BxPC-3 and SVR A221a cell lines. The cellswere treated with 25 μM disaccharide–Cy5** conjugatesat 37 °C for 1 h, washed with PBS, and fixed with 4% paraformaldehyde.The cell nuclei were stained with 2-(4-amidinophenyl)-6-indolecarbamidine(DAPI). Fluorescence imaging was carried out with a 2 s exposure time.(Bottom) Quantification of the binding/uptake of disaccharide–Cy5**conjugates 1–9 in BxPC-3 and SVRA221a cell lines. The cells were treated with 25 μM dye conjugates,irradiated for 2 s prior to imaging, and then analyzed using a ZeissAxiovert 200M inverted microscope with a 40× oil objective.
Mentions: The cell binding/uptakeof BLM disaccharide–Cy5** conjugate(1), decarbamoyl BLM disaccharide–Cy5** conjugate(2), and the newly synthesized disaccharide–Cy5**conjugates 3–9 by BxPC-3 pancreaticcarcinoma cells and SVR A221a normal pancreatic cells (Figures 3 and S1), by A549 lungcarcinoma cells and WI-38 normal lung cells (Figures 4 and S2), by DU-145 prostate carcinomacells and PZ-HPV-7 normal prostate cells (Figures 5 and S3), and by A498 kidney carcinomacells and CCD-1105 KIDTr normal kidney cells (Figures 6 and S4) was quantified by fluorescenceimaging. As shown (Figures 3–6), in all tumor and normal cell lines tested, thedecarbamoyl BLM disaccharide exhibited very low binding and uptake,highlighting the importance of the carbamoyl moiety to effective cellbinding and uptake. This is in agreement with earlier studies reportedby our laboratory.15,21,22

Bottom Line: In earlier studies, we have demonstrated that the tumor cell selectivity resides in the mannose carbamoyl moiety of the BLM saccharide and that both the BLM disaccharide and monosaccharide containing the carbamoyl moiety were capable of the delivery/uptake of a conjugated cyanine dye into cultured cancer cell lines.A library of seven disaccharide-Cy5** dye conjugates was prepared that are structural analogues of the BLM disaccharide.These differed from the natural BLM disaccharide in the position, orientation, and substitution of the carbamoyl group.

View Article: PubMed Central - PubMed

Affiliation: Center for Bioenergetics, Biodesign Institute, and Department of Chemistry and Biochemistry, Arizona State University , Tempe, Arizona 85287, United States.

ABSTRACT
The bleomycins (BLMs) are a family of antitumor antibiotics used clinically for anticancer chemotherapy. Their antitumor selectivity derives at least in part from their ability to target tumor cells, a property that resides in the carbohydrate moiety of the antitumor agent. In earlier studies, we have demonstrated that the tumor cell selectivity resides in the mannose carbamoyl moiety of the BLM saccharide and that both the BLM disaccharide and monosaccharide containing the carbamoyl moiety were capable of the delivery/uptake of a conjugated cyanine dye into cultured cancer cell lines. Presently, the nature of the participation of the carbamoyl moiety has been explored further to provide compounds of utility for defining the nature of the mechanism of tumor cell recognition and uptake by BLM saccharides and in the hope that more efficient compounds could be identified. A library of seven disaccharide-Cy5** dye conjugates was prepared that are structural analogues of the BLM disaccharide. These differed from the natural BLM disaccharide in the position, orientation, and substitution of the carbamoyl group. Studies of these compounds in four matched sets of tumor and normal cell lines revealed a few that were both tumor cell selective and internalized 2-4-fold more efficiently than the natural BLM disaccharide.

Show MeSH
Related in: MedlinePlus