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Targeting canine bladder transitional cell carcinoma with a human bladder cancer-specific ligand.

Lin TY, Zhang H, Wang S, Xie L, Li B, Rodriguez CO, de Vere White R, Pan CX - Mol. Cancer (2011)

Bottom Line: In vivo tumor-specific homing/targeting property and biodistribution of PLZ4 was performed in a mouse xenograft model via tail vein injection and was confirmed with ex vivo imaging.No significant changes in cell viability or proliferation were observed upon incubation with PLZ4.The in vivo and ex vivo optical imaging study showed that, when linked with the near-infrared fluorescent dye Cy5.5, PLZ4 substantially accumulated at the canine bladder cancer foci in the mouse xenograft model as compared to the control.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Hematology and oncology, Department of Internal Medicine, University of California-Davis Cancer Center, Sacramento, CA 95817, USA.

ABSTRACT

Objective: To determine if a human bladder cancer-specific peptide named PLZ4 can target canine bladder cancer cells.

Experimental design: The binding of PLZ4 to five established canine invasive transitional cell carcinoma (TCC) cell lines and to normal canine bladder urothelial cells was determined using the whole cell binding assay and an affinitofluorescence assay. The WST-8 assay was performed to determine whether PLZ4 affected cell viability. In vivo tumor-specific homing/targeting property and biodistribution of PLZ4 was performed in a mouse xenograft model via tail vein injection and was confirmed with ex vivo imaging.

Results: PLZ4 exhibited high affinity and specific dose-dependent binding to canine bladder TCC cell lines, but not to normal canine urothelial cells. No significant changes in cell viability or proliferation were observed upon incubation with PLZ4. The in vivo and ex vivo optical imaging study showed that, when linked with the near-infrared fluorescent dye Cy5.5, PLZ4 substantially accumulated at the canine bladder cancer foci in the mouse xenograft model as compared to the control.

Conclusions and clinical relevance: PLZ4 can specifically bind to canine bladder cancer cells. This suggests that the preclinical studies of PLZ4 as a potential diagnostic and therapeutic agent can be performed in dogs with naturally occurring bladder cancer, and that PLZ4 can possibly be developed in the management of canine bladder cancer.

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Binding Affinity and biological effects of PLZ4 against canine TCC cell lines. A. Binding affinity of PLZ4 against K9TCC-PU and K9TCC-PU-In. Twenty thousands cells of K9TCC-PU and K9TCC-PU-In were seeded in 96 well plates. After culture for 24 hours, cells were fixed and incubated with different concentrations of PLZ4-biotin for 1.5 hours followed by SA-HRP for another 1 hour. Cells treated with SA-HRP alone served as background control. The color was developed using TMB substrate and read by ELISA readers. Three independent experiments conducted in triplicate were performed. The mean values of the 3 experiments are shown. B. Biological effects of PLZ4 on canine TCC cell lines. Ten thousand cells of K9TCC-PU-In or K9TCC-PU were seeded in the 96-well plates and treated with increasing concentrations of PLZ4 or PBS for 2 days. The cell proliferation assay was assessed by the WST-8 assay per manufacturer's protocol. Cells treated with PBS were used as 100% control. Each experiment was performed three times in triplicate. Mean values at each concentration are presented.
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Figure 2: Binding Affinity and biological effects of PLZ4 against canine TCC cell lines. A. Binding affinity of PLZ4 against K9TCC-PU and K9TCC-PU-In. Twenty thousands cells of K9TCC-PU and K9TCC-PU-In were seeded in 96 well plates. After culture for 24 hours, cells were fixed and incubated with different concentrations of PLZ4-biotin for 1.5 hours followed by SA-HRP for another 1 hour. Cells treated with SA-HRP alone served as background control. The color was developed using TMB substrate and read by ELISA readers. Three independent experiments conducted in triplicate were performed. The mean values of the 3 experiments are shown. B. Biological effects of PLZ4 on canine TCC cell lines. Ten thousand cells of K9TCC-PU-In or K9TCC-PU were seeded in the 96-well plates and treated with increasing concentrations of PLZ4 or PBS for 2 days. The cell proliferation assay was assessed by the WST-8 assay per manufacturer's protocol. Cells treated with PBS were used as 100% control. Each experiment was performed three times in triplicate. Mean values at each concentration are presented.

Mentions: To further quantify the binding affinity, K9TCC-PU and K9TCC-PU-In cells were seeded in 96-well plates, fixed with acetone, and incubated with increasing concentrations of PLZ4-biotin followed by SA-HRP. As shown in Figure 2A, PLZ4 exhibited a dose-dependent binding against canine TCC cell lines. The Kd50 values of PLZ4 for K9TCC-PU and K9TCC-PU-In (the concentration of PLZ4 to saturate 50% of cell surface receptor) were 21.3 and 10.3 μM, respectively, while the Kd50 value for human 5637 cells was 6.67 μM.


Targeting canine bladder transitional cell carcinoma with a human bladder cancer-specific ligand.

Lin TY, Zhang H, Wang S, Xie L, Li B, Rodriguez CO, de Vere White R, Pan CX - Mol. Cancer (2011)

Binding Affinity and biological effects of PLZ4 against canine TCC cell lines. A. Binding affinity of PLZ4 against K9TCC-PU and K9TCC-PU-In. Twenty thousands cells of K9TCC-PU and K9TCC-PU-In were seeded in 96 well plates. After culture for 24 hours, cells were fixed and incubated with different concentrations of PLZ4-biotin for 1.5 hours followed by SA-HRP for another 1 hour. Cells treated with SA-HRP alone served as background control. The color was developed using TMB substrate and read by ELISA readers. Three independent experiments conducted in triplicate were performed. The mean values of the 3 experiments are shown. B. Biological effects of PLZ4 on canine TCC cell lines. Ten thousand cells of K9TCC-PU-In or K9TCC-PU were seeded in the 96-well plates and treated with increasing concentrations of PLZ4 or PBS for 2 days. The cell proliferation assay was assessed by the WST-8 assay per manufacturer's protocol. Cells treated with PBS were used as 100% control. Each experiment was performed three times in triplicate. Mean values at each concentration are presented.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3040722&req=5

Figure 2: Binding Affinity and biological effects of PLZ4 against canine TCC cell lines. A. Binding affinity of PLZ4 against K9TCC-PU and K9TCC-PU-In. Twenty thousands cells of K9TCC-PU and K9TCC-PU-In were seeded in 96 well plates. After culture for 24 hours, cells were fixed and incubated with different concentrations of PLZ4-biotin for 1.5 hours followed by SA-HRP for another 1 hour. Cells treated with SA-HRP alone served as background control. The color was developed using TMB substrate and read by ELISA readers. Three independent experiments conducted in triplicate were performed. The mean values of the 3 experiments are shown. B. Biological effects of PLZ4 on canine TCC cell lines. Ten thousand cells of K9TCC-PU-In or K9TCC-PU were seeded in the 96-well plates and treated with increasing concentrations of PLZ4 or PBS for 2 days. The cell proliferation assay was assessed by the WST-8 assay per manufacturer's protocol. Cells treated with PBS were used as 100% control. Each experiment was performed three times in triplicate. Mean values at each concentration are presented.
Mentions: To further quantify the binding affinity, K9TCC-PU and K9TCC-PU-In cells were seeded in 96-well plates, fixed with acetone, and incubated with increasing concentrations of PLZ4-biotin followed by SA-HRP. As shown in Figure 2A, PLZ4 exhibited a dose-dependent binding against canine TCC cell lines. The Kd50 values of PLZ4 for K9TCC-PU and K9TCC-PU-In (the concentration of PLZ4 to saturate 50% of cell surface receptor) were 21.3 and 10.3 μM, respectively, while the Kd50 value for human 5637 cells was 6.67 μM.

Bottom Line: In vivo tumor-specific homing/targeting property and biodistribution of PLZ4 was performed in a mouse xenograft model via tail vein injection and was confirmed with ex vivo imaging.No significant changes in cell viability or proliferation were observed upon incubation with PLZ4.The in vivo and ex vivo optical imaging study showed that, when linked with the near-infrared fluorescent dye Cy5.5, PLZ4 substantially accumulated at the canine bladder cancer foci in the mouse xenograft model as compared to the control.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Hematology and oncology, Department of Internal Medicine, University of California-Davis Cancer Center, Sacramento, CA 95817, USA.

ABSTRACT

Objective: To determine if a human bladder cancer-specific peptide named PLZ4 can target canine bladder cancer cells.

Experimental design: The binding of PLZ4 to five established canine invasive transitional cell carcinoma (TCC) cell lines and to normal canine bladder urothelial cells was determined using the whole cell binding assay and an affinitofluorescence assay. The WST-8 assay was performed to determine whether PLZ4 affected cell viability. In vivo tumor-specific homing/targeting property and biodistribution of PLZ4 was performed in a mouse xenograft model via tail vein injection and was confirmed with ex vivo imaging.

Results: PLZ4 exhibited high affinity and specific dose-dependent binding to canine bladder TCC cell lines, but not to normal canine urothelial cells. No significant changes in cell viability or proliferation were observed upon incubation with PLZ4. The in vivo and ex vivo optical imaging study showed that, when linked with the near-infrared fluorescent dye Cy5.5, PLZ4 substantially accumulated at the canine bladder cancer foci in the mouse xenograft model as compared to the control.

Conclusions and clinical relevance: PLZ4 can specifically bind to canine bladder cancer cells. This suggests that the preclinical studies of PLZ4 as a potential diagnostic and therapeutic agent can be performed in dogs with naturally occurring bladder cancer, and that PLZ4 can possibly be developed in the management of canine bladder cancer.

Show MeSH
Related in: MedlinePlus