In vivoDiagnosis of Epidermal Growth Factor Receptor Expression using Molecular Imaging with a Cocktail of Optically Labeled Monoclonal Antibodies

Abstract

Purpose: Epidermal growth factor receptors (EGFR) play an important role in tumorigenesis and, therefore, have become targets for new molecular therapies. Here, we use a “cocktail” of optically labeled monoclonal antibodies directed against EGFR-1 (HER1) and EGFR-2 (HER2) to distinguish tumors by their cell surface expression profiles. Experimental Design: In vivo imaging experiments were done in tumor-bearing mice following s.c. injection of A431 (overexpressing HER1), NIH3T3/HER2+ (overexpressing HER2), and Balb3T3/DsRed (non-expression control) cell lines. After tumor establishment, a cocktail of optically labeled antibodies: Cy5.5-labeled cetuximab (anti-HER1) and Cy7-labeled trastuzumab (anti-HER2) was i.v. injected. In vivo and ex vivo fluorescence imaging was done. For comparison with radionuclide imaging, experiments were undertaken using 111Indium-labeled antibodies. Additionally, a “blinded” diagnostic study was done for mice bearing one tumor type. Results: In vivo spectral fluorescent molecular imaging of 14 mice with three tumor types clearly differentiated tumors using the cocktail of optically labeled antibodies both in vivo and ex vivo. Twenty-four hours after injection, A431 and NIH3T3/HER2+ tumors were detected distinctly by their peak on Cy5.5 and Cy7 spectral images, respectively; radionuclide imaging was unable to clearly distinguish tumors at this time point. In blinded single tumor experiments, investigators were able to correctly diagnose a total of 40 tumors. Conclusion: An in vivo imaging technique using an antibody cocktail simultaneously differentiated two tumors expressing distinct EGFRs and enabled an accurate characterization of each subtype.

Affibody Molecules for In vivo Characterization of HER2-Positive Tumors by Near-Infrared Imaging

“…Pharmacokinetics of trastuzumab conjugate was consistent with previous findings by other groups using different fluorophores, thus verifying our imaging system (43,44). On the contrary, Affibody monomer Z HER2:342 -Alexa Fluor 750 conjugate failed to show clear signs of tumor accumulation and fast clearance.…”

A comparative study of affibody, panitumumab, and EGF for near-infrared fluorescence imaging of EGFR- and EGFRvIII-expressing tumors

“…Examples include the small molecule 2-deoxyglucose (2-DG), 23,24 small peptide RGD, 25 large peptides annexin V, 26 EGF, 5,6,27,28 affibody, 15,29 and large antibodies. [30][31][32] Quenched probes that only produce fluorescence signals in the presence of tumor-specific proteinases have also been developed. 33 Cell surface receptors, such as EGFR, that are overexpressed in cancers can be reliable targets for molecular imaging.…”

Towards detecting the HER-2 receptor and metabolic changes induced by HER-2-targeted therapies using medical imaging

“…Coincident administration of different coloured probes that target several cell surface determinants can further facilitate tissue discrimination. Pre-clinical studies [35][36][37] have shown that cocktails of antibodies labelled with differently coloured fluorescent tags could differentiate between different tumour types. Thus, Longmire et al [35] recently demonstrated that the use of conjugates consisting of trastuzumab-rhodamine green to target HER-2 could discriminate HER-2-overexpressing cells from SHIN3-RFP (red fluorescent protein) tumour cells, which are engineered to express red fluorescence.…”

“…Thus, Longmire et al [35] recently demonstrated that the use of conjugates consisting of trastuzumab-rhodamine green to target HER-2 could discriminate HER-2-overexpressing cells from SHIN3-RFP (red fluorescent protein) tumour cells, which are engineered to express red fluorescence. Barrett et al [36] were able to detect and differentiate tumours overexpressing HER-1 and HER-2 24 h after administration of a mixture of Cy5.5-labelled cetuximab and Cy7-labelld trastuzumab. Koyama [37] used multifilter spectrally resolved optical imaging to detect tumours overexpressing HER-1, HER-2 and interleukin-2 receptor alpha-subunit receptor (IL-2Ra) grown as xenografts using cetuximab-Cy5 (targeting HER-1), trastuzumab-Cy7 (HER-2), and daclizumab-Alexa-Fluor-700 (IL-2Ra) labelled antibodies, respectively, in mice.…”