DOI: 10.5061/dryad.6j018
title: Data from: Development and evaluation of a fluorescent antibody-drug conjugate for molecular imaging and targeted therapy of pancreatic cancer

description: Antibodies are widely available and cost-effective research tools in life
 science, and antibody conjugates are now extensively used for targeted
 therapy, immunohistochemical staining, or in vivo diagnostic imaging of
 cancer. Significant advances in site-specific antibody labeling
 technologies have enabled the production of highly characterized and
 homogenous conjugates for biomedical purposes, and some recent studies
 have utilized site-specific labeling to synthesize bifunctional antibody
 conjugates with both imaging and drug delivery properties. While these
 advances are important for the clinical safety and efficacy of such
 biologics, these techniques can also be difficult, expensive, and
 time-consuming. Furthermore, antibody-drug conjugates (ADCs) used for
 tumor treatment generally remain distinct from conjugates used for
 diagnosis. Thus, there exists a need to develop simple dual-labeling
 methods for efficient therapeutic and diagnostic evaluation of antibody
 conjugates in pre-clinical model systems. Here, we present a rapid and
 simple method utilizing commercially available reagents for synthesizing a
 dual-labeled fluorescent ADC. Further, we demonstrate the fluorescent
 ADC’s utility for simultaneous targeted therapy and molecular imaging of
 cancer both in vitro and in vivo. Employing non-site-specific,
 amine-reactive chemistry, our novel biopharmaceutical theranostic is a
 monoclonal antibody specific for a carcinoembryonic antigen (CEA)
 biomarker conjugated to both paclitaxel and a near-infrared (NIR),
 polyethylene glycol modified (PEGylated) fluorophore (DyLight™ 680-4xPEG).
 Using in vitro systems, we demonstrate that this fluorescent ADC
 selectively binds a CEA-positive pancreatic cancer cell line (BxPC-3) in
 immunofluorescent staining and flow cytometry, exhibits efficient
 internalization kinetics, and is cytotoxic. Model studies using a
 xenograft of BxPC-3 cells in athymic mice also show the fluorescent ADC’s
 efficacy in detecting tumors in vivo and inhibiting tumor growth more
 effectively than equimolar amounts of unconjugated drug. Overall, our
 results demonstrate that non-selective, amine-targeting chemistry is an
 effective dual-labeling method for synthesizing and evaluating a
 bifunctional fluorescent antibody-drug conjugate, allowing concurrent
 detection, monitoring and treatment of cancer.
description: Conjugate
 Characterization - Data and GraphsThis file contains HPLC
 chromatograms and compiled numerical data concerning the characterization
 (drug:ab and fluorophore:ab ratios) of conjugates synthesized in this
 study (Fig 1 and Table 1).Conjugate
 Characterization - Linker StabilityThis file contains all
 HPLC chromatograms, graphs and computations for determining antibody-drug
 linker stability of the fluorescent antibody drug conjugate as described
 in the paper (Fig 2).In Vitro Fluorescent
 Evalution - QuantitationThis file contains all
 of the raw and analyzed fluorescent quantitation data for assessing
 localization of conjugates with various cell lines (Fig.
 3).Immunofluorescence
 Sample Image BxPC-3 CEA 680 PTXThis file is a raw jpeg
 of an immunofluorescent image used in Fig 3, panel A.Immunofluorescence
 Sample Image BxPC-3 CEA 680This file is a raw jpeg
 of an immunofluorescent image used in Fig 3, panel A.Immunofluorescence
 Sample Image BxPC3 IgG 680 PTXThis file is a raw jpeg
 of an immunofluorescent image used in Fig 3, panel A.Immunofluorescence
 Sample Image BxPC3 IgG 680This file is a raw jpeg
 of an immunofluorescent image used in Fig 3, panel A.In Vitro Fluorescent
 Evaluation - BxPC3 Flow DataThis file contains the
 raw flow cytometry data and overlayed images (“analyze” tab) which were
 arranged for Fig 4.In Vitro Fluorescent
 Evaluation - HeLa Flow DataThis file contains the
 raw flow cytometry data and overlayed images (“analyze” tab) which were
 arranged for Fig 4.In Vitro Fluorescent
 Evaluation - HepG2 Flow DataThis file contains the
 raw flow cytometry data and overlayed images (“analyze” tab) which were
 arranged for Fig 4.In Vitro Fluorescent
 Evaluation - MCF7 Flow DataThis file contains the
 raw flow cytometry data and overlayed images (“analyze” tab) which were
 arranged for Fig 4.In Vitro Fluorescent
 Evaluation - CEA Internalization DataThis file contains the
 compiled and processed data surrounding the quantitative estimation of CEA
 internalization (Fig 5, panel B).In Vitro Cytotoxicity
 Evaluation - MTT Assay Compiled DataThis excel file contains
 all of the compiled MTT assay data, as well as graphs (Fig 6) and linear
 regression analysis for estimating IC50 (Table 2).In Vitro Cytotoxic
 Evaluation - Flow Cytometry Cell Cycle AnalysisThis file contains the
 raw flow cytometry cell cycle analysis data and analyzed images that were
 arranged for Fig 7. This file was originally a template that was included
 with the BD cell cycle analysis kit as described in the
 paper.In Vivo Evaluation -
 Tumor Growth and Auto Exposure DataThis excel file contains
 all raw tumor growth measurements and graphs for in vivo efficacy
 evaluation (Fig 8). This file also contains the compiled exposure times
 generated for Fig 10.In Vivo Imaging Sample
 Image - Day 7 - White Light - RawThis file is a raw jpeg
 generated for Fig 9. It depicts imaging of a mouse in group 3 imaged under
 white light as described in the methods section.In Vivo Imaging Sample
 Image - Day 7 - Infrared - RawThis file is a raw jpeg
 generated for Fig 9. It depicts a mouse in group 3 imaged under an
 infrared filter set as described in the methods
 section.In Vivo Imaging Sample
 Image - Day 7 - Infrared - False ColorThis file is a jpeg
 generated for Fig 9. It depicts a mouse in group 3 imaged under an
 infrared filter as described in the methods section. Red false color has
 been added with Pxlr editor as described in the methods
 section.In Vivo Evaluation -
 ANOVA AnalysisThis file contains the
 statistical analysis of the data generated for Fig 8.