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J Urol. 2003 Apr;169(4):1535-40.

Targeting urothelium: ex vivo assay standardization and selection of internalizing ligands.

Author information

1
Department of Genitourinary Medical Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, 77030, USA.

Abstract

PURPOSE:

With the goal of targeting the human bladder using phage display technology we designed and tested a tissue binding assay on intact urothelium ex vivo. This approach may form the molecular basis for clinical development of peptide or peptidomimetic guided intravesical compounds.

MATERIALS AND METHODS:

We screened 2 phage display random peptide libraries on human urothelium. Select peptides were tested for their binding ability to human urothelium, 2 human transitional cell carcinoma cell lines and a nontransitional cell carcinoma cell line. Next we standardized an ex vivo binding assay, validated binding of selected phage to whole urothelium, and evaluated whether receptor mediated internalization into urothelium derived cells occurred. Finally we tested if the presence of the glycosaminoglycan layer had any effect on the binding of the urothelium targeted phage.

RESULTS:

Phage selected and recovered in the screening were isolated and sequenced. Displayed peptide sequences were searched against online protein databases. Five classes of peptide motifs were characterized based on their ability to bind to normal urothelium but not to control cell lines. Remarkable consistency and reproducibility were observed in the ex vivo binding assays. Two classes of peptide motifs sharing the sequence Ile/Leu-Ser-Gly-Leu bound to normal urothelium and to 2 transitional cell carcinoma cells but not to nontransitional cell carcinoma cells in a glycosaminoglycan independent manner and mediated internalization into cells of urothelial origin.

CONCLUSIONS:

We introduce a strategy for screening combinatorial peptide libraries on bladder mucosa, a standard model for ex vivo intact urothelium binding assays and a panel of urothelium binding peptides that may be suitable for translation into targeted intravesical therapy applications.

[Indexed for MEDLINE]

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