Abstract A226: ImmunoPET with 89Zr-DFO-trastuzumab: Monitoring the pharmacodynamic effects on HER2/neu expression by Hsp90 inhibition in vivo

Abstract

Abstract Objectives: The positron-emitting radionuclide 89Zr (t1/2 = 3.17 days) has been isolated in high specific-activity and was used to prepare 89Zr-radiolabeled monoclonal antibodies (mAbs) for use as immunoPET imaging agents. The work presented describes the preparation of 89Zr-DFO-trastuzumab for use as a radiotracer for delineating HER2/neu positive tumors for the non-invasive imaging of breast cancer in vivo. In addition, pharmacokinetic studies on HER2/neu expression levels in response to administration of therapeutic doses of PU-H71 (a specific inhibitor of heat-shock protein 90 [Hsp90]) were conducted. Methods: 89Zr was prepared via the 89Y(p, n)89Zr transmutation with high radiochemical yields (1.52±0.11 mCi/ Ah) and purity (>99.99%). Trastuzumab was functionalized with the tris-hydroxamate chelate, desferrioxamine B (DFO) and radiolabeled with [89Zr]Zr-oxalate at room temperature. ImmunoPET imaging experiments in female, athymic nu/nu mice bearing sub-cutaneous BT-474 (HER2/neu positive) and/or MDA-MB-468 (HER2/neu negative) were conducted. The change in 89Zr-DFO-trastuzumab tissue uptake in response to high- and low-specific-activity formulations and co-administration of PU-H71 (a Hsp90 chaperone inhibitor) was measured by using both acute biodistribution studies and Western blot analysis. Results: 89Zr-DFO-trastuzumab radiolabeling proceeded in high radiochemical yield and specific-activity of 2.82±0.05 mCi/mg. In vitro assays demonstrated >99% radiochemical purity with an immunoreactive fraction of 0.87±7. In vivo biodistribution experiments revealed high and specific tumor uptake after 24, 48 and 72 h (64.68 ± 13.06 %ID/g; 71.71 ± 10.35 %ID/g and 85.18±11.10 %ID/g, respectively) with retention of activity for over 120 h. Pre-treatment with PU-H71 was followed by quantitative biodistribution and 89Zr-DFO-trastuzumab immunoPET imaging. Expression levels of HER2/neu were modulated during the first 24 and 48 h post-administration (29.75 ± 4.43 %ID/g and 41.42 ± 3.64 %ID/g, respectively). By 72 h radiotracer uptake studies (73.64 ± 12.17 %ID/g) and Western blot analysis demonstrated that HER2/neu expression recovered to baseline levels. Conclusions: The results indicate that 89Zr-DFO-trastuzumab provides quantitative and highly-specific delineation of HER2/neu positive tumors and may be used to monitor treatment with Hsp90 inhibitors. Efforts towards the “current Good Manufacturing Process” (cGMP) production and clinical translation of 89Zr-DFO-trastuzumab are underway. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A226.