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Optimizing Glioma Detection Using an EGFR-Targeted Fluorescent Affibody.

ABSTRACT: Since many types of cancers overexpress EGFR, this surface receptor has been used as a target for therapy or diagnosis of malignant disease. Uptake kinetics of EGFR-targeted fluorescent Affibody (ABY-029) were studied with a view toward optimizing efficacy of tumor detection in a glioma as a function of both delivered dose and concurrent administration of unlabeled cetuximab (an EGFR antagonist). U251 glioma cells were inoculated in brain of nude rats, and the fluorescence from each brain was analyzed after the administration of ABY-029. Although cetuximab was able to systematically block ABY-029 binding to EGFR in a dose-dependent manner in cell culture, no influence on the tumor-to-normal brain contrast was seen when unlabeled cetuximab was administered prior to ABY-029. Ex vivo imaging of ABY-029 fluorescence showed increasing values of the tumor-to-normal brain ratio with an increasing injected dose. A saturation value was obtained at a dose of 245 ?g kg-1 which represents a 10-fold increase over a "microdose" value. According to FDA, the microdose of protein products is considered ?30 nanomoles due to its difference in molecular weight as compared to synthetic drugs. This observation indicates that glioma detection will be optimal if the ABY-029 dose exceeds the "microdose" value.

SUBMITTER: Ribeiro de Souza AL

PROVIDER: S-EPMC6234080 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Optimizing Glioma Detection Using an EGFR-Targeted Fluorescent Affibody.

Ribeiro de Souza Ana Luiza AL Marra Kayla K Gunn Jason J Samkoe Kimberley S KS Hull Sally S Paulsen Keith D KD Pogue Brian W BW

Photochemistry and photobiology 20180928 6

Since many types of cancers overexpress EGFR, this surface receptor has been used as a target for therapy or diagnosis of malignant disease. Uptake kinetics of EGFR-targeted fluorescent Affibody (ABY-029) were studied with a view toward optimizing efficacy of tumor detection in a glioma as a function of both delivered dose and concurrent administration of unlabeled cetuximab (an EGFR antagonist). U251 glioma cells were inoculated in brain of nude rats, and the fluorescence from each brain was an ...[more]

PMID: 30129069

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Project description:In head and neck squamous cell cancer, the human epidermal growth factor receptor 1 (EGFR) is the dominant signaling molecule among all members of the family. So far, cetuximab is the only approved anti-EGFR monoclonal antibody used for the treatment of head and neck squamous cell cancer, but despite the benefits of adding it to standard treatment regimens, attempts to define a predictive biomarker to stratify patients for cetuximab treatment have been unsuccessful. We hypothesized that imaging with EGFR-specific radioligands may facilitate noninvasive measurement of EGFR expression across the entire tumor burden and allow for dynamic monitoring of cetuximab-mediated changes in receptor expression. Methods: EGFR-specific Affibody molecule (ZEGFR:03115) was radiolabeled with 89Zr and 18F. The radioligands were characterized in vitro and in mice bearing subcutaneous tumors with varying levels of EGFR expression. The protein dose for imaging studies was assessed by injecting 89Zr-deferoxamine-ZEGFR:03115 (2.4-3.6 MBq, 2 μg) either together with or 30 min after increasing amounts of unlabeled ZEGFR:03115 (1, 5, 10, 15, and 20 μg). PET images were acquired at 3, 24, and 48 h after injection, and the image quantification data were correlated with the biodistribution results. The EGFR expression and biodistribution of the tracer were assessed ex vivo by immunohistochemistry, Western blot, and autoradiography. To downregulate the EGFR level, treatment with cetuximab was performed, and 18F-aluminium fluoride-NOTA-ZEGFR:03115 (12 μg, 1.5-2 MBq/mouse) was used to monitor receptor changes. Results: In vivo studies demonstrated that coinjecting 10 μg of nonlabeled molecules with 89Zr-deferoxamine-ZEGFR:03115 allows for clear tumor visualization 3 h after injection. The radioconjugate tumor accumulation was EGFR-specific, and PET imaging data showed a clear differentiation between xenografts with varying EGFR expression levels. A strong correlation was observed between PET analysis, ex vivo estimates of tracer concentration, and receptor expression in tumor tissues. Additionally, 18F-aluminium fluoride-NOTA-ZEGFR:03115 could measure receptor downregulation in response to EGFR inhibition. Conclusion: ZEGFR:03115-based radioconjugates can assess different levels of EGFR level in vivo and measure receptor expression changes in response to cetuximab, indicating a potential for assessment of adequate treatment dosing with anti-EGFR antibodies.

Project description:UnlabelledEpidermal growth factor receptor (EGFR) is often overexpressed in a variety of human cancers, and its expression is associated with poor prognosis for many cancer types. However, an accurate technique to noninvasively image EGFR expression in vivo is not available in the clinical setting. In this research, an Affibody analog, anti-EGFR Ac-Cys-Z(EGFR:1907), was successfully site-specifically (18)F-labeled for PET of EGFR expression.MethodsThe prosthetic group N-[2-(4-(18)F-fluorobenzamido) ethyl] maleimide ((18)F-FBEM) was conjugated to Ac-Cys-Z(EGFR:1907) under mild conditions (pH 7) to produce the probe (18)F-FBEM-Cys-Z(EGFR:1907). The binding affinity and specificity tests of (18)F-FBEM-Cys-Z(EGFR:1907) to EGFR were conducted using A431 cancer cells. Small-animal PET and biodistribution studies were conducted on various mice tumor xenograft models with EGFR overexpression (6 types) after injection of approximately 2.0 MBq of (18)F-FBEM-Cys-Z(EGFR:1907) with or without coinjection of unlabeled Ac-Cys-Z(EGFR:1907) for up to 3 h after injection. A correlation study between (18)F-FBEM-Cys-Z(EGFR:1907) small- animal PET quantification and ex vivo Western blot analysis of tumor EGFR expression was conducted in those 6 types of tumor models.Results(18)F-FBEM-Cys-Z(EGFR:1907) binds to EGFR with low nanomolar affinity (37 nM) in A431 cells. (18)F-FBEM-Cys-Z(EGFR:1907) rapidly accumulated in the tumor and cleared from most of the normal organs except the liver and kidneys at 3 h after injection, allowing excellent tumor-to-normal tissue contrast to be obtained. In the A431 tumor xenograft model, coinjection of the PET probe with 45 μg of Ac-Cys-Z(EGFR:1907) was able to improve the tumor uptake (3.9 vs. 8.1 percentage of the injected radioactive dose per gram of tissue, at 3 h after injection) and tumor imaging contrast, whereas coinjection with 500 μg of Ac-Cys-Z(EGFR:1907) successfully blocked the tumor uptake significantly (8.1 vs. 1.0 percentage of the injected radioactive dose per gram of tissue, at 3 h after injection, 88% inhibition, P < 0.05). Moderate correlation was found between the tumor tracer uptake at 3 h after injection quantified by PET and EGFR expression levels measured by Western blot assay (P = 0.007, R = 0.59).Conclusion(18)F-FBEM-Cys-Z(EGFR:1907) is a novel protein scaffold-based PET probe for imaging EGFR overexpression of tumors, and its ability to differentiate tumors with high and low EGFR expression in vivo holds promise for future clinical translation.

Dataset Information

Optimizing Glioma Detection Using an EGFR-Targeted Fluorescent Affibody.

Publications

Optimizing Glioma Detection Using an EGFR-Targeted Fluorescent Affibody.

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