Pre-clinical assessment of phosphokinase activity by an anti-phosphor-tyrosine antibody labeled with In-111

Objectives It is crucial to predict the selection of patients who may benefit from targeted treatment using non-invasive imaging technique. This study was aimed to determine whether Bcr-Abl non-receptor tyrosine kinase could be assessed by using Indium-111 labeled anti-phospho-tyrosine antibody (APT).

Methods Ethylenedicysteine (EC) was conjugated to either APT or bovine serum albumin (BSA) using sulfo-N-hydroxysuccinimide and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide HCl as coupling agents. After dialysis, EC-APT was labeled with Indium (¹¹¹In). To determine if ¹¹¹In-EC-APT could be used to assess a non-receptor tyrosine kinase, in human CML K562 xenografts. Human chronic myelogenous leukemia cell line K562 was selected because it expresses a high level of the non-receptor tyrosine kinase Bcr-Abl. Gamma scintigraphy of the tumor-bearing mice administered with ¹¹¹In-EC-APT (a positive control) and ¹¹¹In-EC-BSA (a negative control) (100 μCi/mouse, iv) before and after Imatinib treatment was obtained at 24 and 48 hrs. Changes in the expression of phosphorylated Bcr-Abl of tumor specimen were determined by western blot analysis.

Results ¹¹¹In-EC-APT is preferentially taken up by tumor cells when compared to ¹¹¹In-EC-BSA control. The optimal imaging time was at 24 and 48 hours after injection of ¹¹¹In-EC-APT. Imatinib treatment resulted in decreased tumor volume and decreased expression of phosphorylated Bcr-Abl by western blot analysis. Bcr-Abl positive tumor uptake changes after Imantinib treatment could be assessed by ¹¹¹In-EC-APT. The imaging findings did correlate with tumor response.

Conclusions The results inidcate that ¹¹¹In-EC-APT could assess tumor phosphokinase activity in our animal model. ¹¹¹In-EC-APT may provide early evidence of response in phosphokinase levels after treatment with a kinase inhibitor