title: Design optimization and characterization of Her2/neu-targeted immunotoxins: comparative in vitro and in vivo efficacy studies.
abstract: Targeted therapeutics are potential therapeutic agents because of their selectivity and efficacy against tumors resistant to conventional therapy. The goal of this study was to determine the comparative activity of monovalent, engineered anti-Her2/neu immunotoxins fused to recombinant gelonin (rGel) to the activity of bivalent IgG-containing immunoconjugates. Utilizing Herceptin and its derived humanized single-chain antibody (single-chain fragment variable, designated 4D5), we generated bivalent chemical Herceptin/rGel conjugate, and the corresponding monovalent recombinant immunotoxins in two orientations, 4D5/rGel and rGel/4D5. All the constructs showed similar affinity to Her2/neu-overexpressing cancer cells, but significantly different antitumor activities. The rGel/4D5 orientation construct and Herceptin/rGel conjugate were superior to 4D5/rGel construct in in vitro and in vivo efficacy. The enhanced activity was attributed to improved intracellular toxin uptake into target cells and efficient downregulation of Her2/neu-related signaling pathways. The Her2/neu-targeted immunotoxins effectively targeted cells with Her2/neu expression level >1.5 × 10(5) sites per cell. Cells resistant to Herceptin or chemotherapeutic agents were not cross-resistant to rGel-based immunotoxins. Against SK-OV-3 tumor xenografts, the rGel/4D5 construct with excellent tumor penetration showed impressive tumor inhibition. Although Herceptin/rGel conjugate demonstrated comparatively longer serum half-life, the in vivo efficacy of the conjugate was similar to the rGel/4D5 fusion. These comparative studies demonstrate that the monovalent, engineered rGel/4D5 construct displayed comparable in vitro and in vivo antitumor efficacy as bivalent Herceptin/rGel conjugate. Immunotoxin orientation can significantly impact the overall functionality and performance of these agents. The recombinant rGel/4D5 construct with excellent tumor penetration and rapid blood clearance may reduce the unwanted toxicity when administrating to patients, and warrants consideration for further clinical evaluation.
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: Y
authors: Y
authors: Cao
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: J W
authors: JW
authors: Marks
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: Z
authors: Z
authors: Liu
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: L H
authors: LH
authors: Cheung
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: W N
authors: WN
authors: Hittelman
authors: Immunopharmacology and Targeted Therapy Laboratory, Department of Experimental Therapeutics, M.D. Anderson Cancer Center, Houston, TX, USA.
authors: M G
authors: MG
authors: Rosenblum
mesh_terms: D000818:Animals; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000737:chemistry / Q000494:pharmacology*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006801:Humans; D018796:Immunoconjugates / Q000737:chemistry / Q000494:pharmacology*; D007150:Immunohistochemistry; D000922:Immunotoxins / Q000737:chemistry / Q000494:pharmacology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D058990:Molecular Targeted Therapy / Q000379:methods*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D054789:Ribosome Inactivating Proteins, Type 1 / Q000494:pharmacology; D000068878:Trastuzumab; D023041:Xenograft Model Antitumor Assays
chemical_list: D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D018796:Immunoconjugates; D000922:Immunotoxins; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D000068878:Trastuzumab; C025674:GEL protein, Gelonium multiflorum; C508053:ERBB2 protein, human
references: Curr Opin Mol Ther. 2003 Feb;5(1):44-51
references: 12669470
references: Cancer Res. 1992 Jun 15;52(12):3402-8
references: 1596900
references: Cancer. 2007 Jan 15;109(2):170-9
references: 17154393
references: Am J Obstet Gynecol. 1993 Jan;168(1 Pt 1):228-32
references: 8420332
references: Clin Cancer Res. 1999 Sep;5(9):2311-5
references: 10499598
references: Cancer Res. 1992 Nov 15;52(22):6310-7
references: 1358432
references: Proc Natl Acad Sci U S A. 1992 May 15;89(10):4285-9
references: 1350088
references: Clin Cancer Res. 1999 Apr;5(4):865-74
references: 10213223
references: J Clin Oncol. 2010 Sep 1;28(25):3910-6
references: 20679614
references: Clin Cancer Res. 2006 Mar 1;12(5):1599-605
references: 16533787
references: Oncogene. 2001 Nov 8;20(51):7551-62
references: 11709727
references: Mol Cancer Ther. 2012 Jan;11(1):143-53
references: 22090420
references: Ann Oncol. 2001;12 Suppl 1:S81-7
references: 11521727
references: Adv Cancer Res. 2001;81:93-124
references: 11430597
references: Clin Cancer Res. 1995 Sep;1(9):1023-9
references: 9816075
references: Clin Cancer Res. 2009 Dec 15;15(24):7479-7491
references: 20008848
references: Recent Pat Anticancer Drug Discov. 2009 Jan;4(1):9-18
references: 19149684
references: Methods. 2003 Jul;30(3):256-68
references: 12798140
references: Breast Cancer Res Treat. 2003 Dec;82(3):155-64
references: 14703062
references: Microbiol Immunol. 1997;41(7):563-70
references: 9272702
references: Anticancer Res. 2006 Jan-Feb;26(1B):715-22
references: 16739343
references: J Mol Biol. 1998 Aug 21;281(3):475-83
references: 9698563
references: Clin Cancer Res. 2007 Mar 1;13(5):1374-82
references: 17309914
references: Cancer Lett. 2007 Nov 8;257(1):124-35
references: 17698286
references: Immunol Res. 2002;25(2):177-91
references: 11999171
references: Cancer Res. 1991 Oct 1;51(19):5361-9
references: 1680547
references: Oncogene. 2003 May 22;22(21):3205-12
references: 12761490
references: Cancer Res. 2008 Nov 15;68(22):9280-90
references: 19010901
references: Eur J Nucl Med Mol Imaging. 2009 Jan;36(1):81-93
references: 18712381
references: Cancer Res. 2003 Jul 15;63(14):3995-4002
references: 12873997
references: Cancer Res. 1999 Aug 15;59(16):4018-22
references: 10463601
references: Cancer Res. 1988 Dec 15;48(24 Pt 1):7022-32
references: 3191477
references: Br J Cancer. 1998 May;77(9):1405-12
references: 9652755
references: Cancer Res. 2009 Dec 1;69(23):8987-95
references: 19934334
references: Protein Expr Purif. 2006 Mar;46(1):73-84
references: 16230023
references: Cancer Res. 1998 Jun 15;58(12):2661-6
references: 9635594
references: In Vivo. 2000 Jan-Feb;14(1):21-7
references: 10757057
references: Breast Dis. 2000;11:113-24
references: 15687597
references: Expert Opin Biol Ther. 2001 Mar;1(2):291-300
references: 11727536
references: Oncology (Williston Park). 2010 Apr 30;24(5):410-5
references: 20480738
references: Breast Cancer Res. 2011;13(6):R123
references: 22129105