Preclinical evaluation of a monoclonal antibody targeting the epidermal growth factor receptor as a radioimmunodiagnostic and radioimmunotherapeutic agent

Abstract

Background and purpose: The studies described here are the first to evaluate the in vitro and in vivo properties of (111)In-CHX-A''-panitumumab for radioimmunotherapy (alpha- and beta(-)-emitters) and radioimmunoimaging (single photon emission computed tomography and positron emission tomography).

Experimental approach: Twenty-seven human carcinoma cell lines were analysed for expression of epidermal growth factor receptors by flow cytometry. Panitumumab was conjugated with CHX-A''-DTPA (diethylenetriamine-pentaacetic acid) and radiolabelled with (111)In. Immunoreactivity of the CHX-A''-DTPA-panitumumab and (111)In-CHX-A''-DTPA-panitumumab was evaluated by radioimmunoassays. Tumour targeting was determined in vivo by direct quantitation of tumour and normal tissues and by gamma-scintigraphy.

Key results: For 26 of 27 human tumour cell lines, 95% of the cells expressed epidermal growth factor receptors over a range of intensity. Immunoreactivity of panitumumab was retained after modification with CHX-A''-DTPA. Radiolabelling of the immunoconjugate with (111)In was efficient with a specific activity of 19.5 +/- 8.9 mCi.mg(-1) obtained. Immunoreactivity and specificity of binding of the (111)In-panitumumab was shown with A431 cells. Tumour targeting by (111)In-panitumumab was demonstrated in athymic mice bearing A431, HT-29, LS-174T, SHAW or SKOV-3 s.c. xenografts with little uptake observed in normal tissues. The (111)In-panitumumab was also evaluated in non-tumour-bearing mice. Pharmacokinetic studies compared the plasma retention time of the (111)In-panitumumab in both non-tumour-bearing and A431 tumour-bearing mice. Tumour targeting was also visualized by gamma-scintigraphy.

Conclusions and implications: Panitumumab can be efficiently radiolabelled with (111)In with high labelling yields. Based on the efficiency in tumour targeting and low normal tissue uptake, panitumumab may be an effective targeting component for radioimmunodiagnostic and radioimmunotherapeutic applications.

Figure 1

Evaluation of panitumumab immunoreactivity in a competition radioimmunoassay. The immunoreactivity of panitumumab-CHX-A″-DTPA (diethylenetriamine-pentaacetic acid) for purified epidermal growth factor receptor was compared with that of unmodified panitumumab. The anti-CD33 monoclonal antibody, HuM195 was used as a non-specific control.

Figure 2

Blood pharmacokinetics of radiolabelled panitumumab. Mice (n= 5) bearing s.c. A431 xenografts as well as mice without tumours were given i.v. injections of ¹¹¹In-CHX-A″-panitumumab (∼7.5 µCi) via the tail vein. Blood samples (10 µL) were collected over a 1 week period as described in Methods. The average %ID·mL⁻¹ (percentage of injected dose per millilitre) with the standard deviations is plotted. Panel A presents the first 20 h while Panel B presents the complete study period of 168 h.

Figure 3

Biodistribution of ¹¹¹In-CHX-A″-panitumumab in tumour xenografts and normal tissues. Mice (n= 5) without tumour (Panel A) or bearing s.c. A431 xenografts (Panel B) were injected i.v. with ¹¹¹In-CHX-A″-panitumumab (∼7.5 µCi). Mice were killed at the indicated times, when blood, tumour and tissues were removed, weighed and the radioactivity measured. The mean %ID·g⁻¹ (percentage of injected dose per gram) with standard deviations are plotted.

Figure 4

γ-Scintigraphy of mice bearing tumour xenografts. Following i.v. injection with 80–100 µCi of ¹¹¹In-CHX-A″-panitumumab, mice bearing LS-174T (upper row) xenografts were imaged over a 1 week period. γ-Scintigraphy was also conducted with other tumour xenograft models with representative scintigraphs taken at 48 h presented in the lower panel.