title: Parallel in vivo and in vitro selection using phage display identifies protease-dependent tumor-targeting peptides.
abstract: We recently developed activatable cell-penetrating peptides (ACPPs) that target contrast agents to in vivo sites of matrix metalloproteinase activity, such as tumors. Here we use parallel in vivo and in vitro selection with phage display to identify novel tumor-homing ACPPs with no bias for primary sequence or target protease. Specifically, phage displaying a library of ACPPs were either injected into tumor-bearing mice, followed by isolation of cleaved phage from dissected tumor, or isolated based on selective cleavage by extracts of tumor versus normal tissue. Selected sequences were synthesized as fluorescently labeled peptides, and tumor-specific cleavage was confirmed by digestion with tissue extracts. The most efficiently cleaved peptide contained the substrate sequence RLQLKL and labeled tumors and metastases from several cancer models with up to 5-fold contrast. This uniquely identified ACPP was not cleaved by matrix metalloproteinases or various coagulation factors but was efficiently cleaved by plasmin and elastases, both of which have been shown to be aberrantly overexpressed in tumors. The identification of an ACPP that targets tumor expressed proteases without rational design highlights the value of unbiased selection schemes for the development of potential therapeutic agents.
authors.affiliation: Department of Pharmacology, University of California at San Diego, La Jolla, California 92093, USA.
authors.forename: Mike
authors.initials: M
authors.lastname: Whitney
authors.forename: Jessica L
authors.initials: JL
authors.lastname: Crisp
authors.forename: Emilia S
authors.initials: ES
authors.lastname: Olson
authors.forename: Todd A
authors.initials: TA
authors.lastname: Aguilera
authors.forename: Larry A
authors.initials: LA
authors.lastname: Gross
authors.forename: Lesley G
authors.initials: LG
authors.lastname: Ellies
authors.forename: Roger Y
authors.initials: RY
authors.lastname: Tsien
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016503:Drug Delivery Systems / Q000379:methods*; D051379:Mice; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000378:metabolism*; D010447:Peptide Hydrolases / Q000378:metabolism*; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D014020:Tissue Extracts
chemical_list: D019151:Peptide Library; D010455:Peptides; D014020:Tissue Extracts; D010447:Peptide Hydrolases
references.citation: Int J Cancer. 2006 Dec 1;119(11):2539-45
references.pmid: 16929516
references.citation: Br J Cancer. 2004 Apr 5;90(7):1414-21
references.pmid: 15054465
references.citation: Nano Lett. 2006 Sep;6(9):1988-92
references.pmid: 16968013
references.citation: Cancer. 1996 Jul 15;78(2):237-43
references.pmid: 8673998
references.citation: Proc Natl Acad Sci U S A. 2010 Mar 2;107(9):4311-6
references.pmid: 20160077
references.citation: Int J Cancer. 1991 Feb 20;47(4):592-6
references.pmid: 1995487
references.citation: Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):7978-82
references.pmid: 1896445
references.citation: Cancer Res. 2003 Jan 15;63(2):337-41
references.pmid: 12543785
references.citation: Integr Biol (Camb). 2009 Jun;1(5-6):382-93
references.pmid: 20023745
references.citation: J Med Chem. 2000 Apr 6;43(7):1253-6
references.pmid: 10753462
references.citation: Mol Biol Rep. 2004 Jun;31(2):121-9
references.pmid: 15293788
references.citation: Clin Cancer Res. 2004 Jul 15;10(14):4761-8
references.pmid: 15269150
references.citation: Curr Protein Pept Sci. 2003 Apr;4(2):105-24
references.pmid: 12678850
references.citation: Bioconjug Chem. 2003 Mar-Apr;14(2):368-76
references.pmid: 12643747
references.citation: Bioconjug Chem. 2006 May-Jun;17(3):787-96
references.pmid: 16704219
references.citation: Lab Invest. 2002 Nov;82(11):1583-90
references.pmid: 12429818
references.citation: J Cell Biol. 1997 May 19;137(4):953-63
references.pmid: 9151696
references.citation: J Biochem. 1999 Jun;125(6):1067-76
references.pmid: 10348909
references.citation: Br J Cancer. 2003 Oct 6;89(7):1270-5
references.pmid: 14520459
references.citation: Thromb Haemost. 2005 Apr;93(4):676-81
references.pmid: 15841311
references.citation: Br J Cancer. 1994 Jan;69(1):72-6
references.pmid: 8286213
references.citation: J Med Chem. 2002 Aug 15;45(17):3612-8
references.pmid: 12166934
references.citation: Radiology. 2001 Nov;221(2):523-9
references.pmid: 11687699
references.citation: Ann Surg. 1994 Feb;219(2):131-4
references.pmid: 8129483
references.citation: Nat Med. 2001 Jun;7(6):743-8
references.pmid: 11385514
references.citation: Integr Biol (Camb). 2009 Jun;1(5-6):371-81
references.pmid: 20023744
references.citation: Proc Natl Acad Sci U S A. 1999 Aug 17;96(17):9497-502
references.pmid: 10449721
references.citation: Matrix Biol. 2003 Mar;22(1):55-61
references.pmid: 12714042
references.citation: Biomol Eng. 2001 Sep;18(2):57-63
references.pmid: 11535417
references.citation: J Biol Chem. 2002 Jun 28;277(26):23788-93
references.pmid: 11959855
references.citation: Am J Pathol. 2003 Nov;163(5):2113-26
references.pmid: 14578209
references.citation: Int J Cancer. 2008 Jan 1;122(1):1-4
references.pmid: 17960586
references.citation: Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10335-40
references.pmid: 12149457
references.citation: J Nucl Med. 2004 Sep;45(9):1431-4
references.pmid: 15347707
references.citation: Mol Ther. 2003 Aug;8(2):295-305
references.pmid: 12907152
references.citation: Proc Natl Acad Sci U S A. 2004 Dec 21;101(51):17867-72
references.pmid: 15601762
references.citation: Proc Natl Acad Sci U S A. 2004 Jun 22;101(25):9381-6
references.pmid: 15197262
references.citation: J Med Chem. 2002 Dec 5;45(25):5523-33
references.pmid: 12459020
references.citation: Mol Cell Biol. 1992 Mar;12(3):954-61
references.pmid: 1312220
references.citation: Oncogene. 2006 Aug 17;25(36):4998-5010
references.pmid: 16568086
references.citation: Cancer Res. 2001 Nov 15;61(22):8298-305
references.pmid: 11719463
references.citation: Am J Pathol. 1998 Sep;153(3):937-44
references.pmid: 9736042