Noninvasive Detection of HER2 Expression in Gastric Cancer by ⁶⁴Cu-NOTA-Trastuzumab in PDX Mouse Model and in Patients

Abstract

The purpose of this study was to establish the quality control and quantify the novel 64Cu-NOTA-Trastuzumab in gastric cancer patient-derived xenografts (PDX) mice models and patients by applying the molecular imaging technique. Trastuzumab was labeled with 64Cu using NCS-Bz-NOTA as bifunctional chelator, and hIgG1 was labeled with the same procedures as a negative control agent. HER2-positive (case 176, n = 12) and HER2-negative (case 168, n = 3) PDX models were established and validated by Western blot, DNA amplification, and immunohistochemistry (IHC). Both models were conducted for micro-PET imaging by tail injection of 18.5 MBq of 64Cu-NOTA-Trastuzumab or 64Cu-NOTA-hIgG1. Radioprobe uptake in tumor and main organs was quantified by region of interested (ROI) analysis of the micro-PET images and autoradiography. Finally, gastric cancer patients were enrolled in preliminary 64Cu-NOTA-Trastuzumab PET/CT scans. NOTA-Trastuzumab was efficiently radiolabeled with 64Cu over a 99% radiochemical purity and 17.5 GBq/μmol specific activity. The immune activity was preserved as the nonmodified antibody, and the radiopharmaceutical proved to be stable for up to 5 half-decay lives of 64Cu both in vitro and in vivo. Two serials of PDX gastric cancer models were successfully established: case 176 for HER2 positive and case 168 for HER2 negative. In micro-PET imaging studies, 64Cu-NOTA-Trastuzumab exhibits a significant higher tumor uptake (11.45 ± 0.42 ID%/g) compared with 64Cu-NOTA-IgG1 (3.25 ± 0.28 ID%/g, n = 5, p = 0.0004) at 36 h after intravenous injection. Lower level uptake of 64Cu-NOTA-Trastuzumab (6.35 ± 0.48 ID%/g) in HER2-negative PDX tumor models further confirmed specific binding of the radioprobe. Interestingly, the coinjection of 2.0 mg of Trastuzumab (15.52 ± 1.97 ID%/g) or 2.0 mg of hIgG1 (15.64 ± 3.54 ID%/g) increased the 64Cu-NOTA-Trastuzumab tumor uptake in PDX tumor (HER2+) models compared with 64Cu-NOTA-Trastuzumab alone (p < 0.05) at 36 h postinjection. There were good correlations between micro-PET images and IHC (n = 4) and autoradiography in PDX (HER2+) tumor tissues. Therefore, 64Cu-NOTA-Trastuzumab successfully translated to clinical PET imaging, and 64Cu-NOTA-Trastuzumab PET/CT scan in gastric cancer patients showed good detection ability. In conclusion, we reported quality control and application of novel 64Cu-NOTA-Trastuzumab for HER2 expression in PDX gastric cancer mice models and gastric cancer patients. Moreover, 64Cu-NOTA-Trastuzumab holds great potential for noninvasive PET detection, staging, and follow-up of HER2 expression in gastric cancer.

Antibody-Targeted Imaging of Gastric Cancer

“…ImmunoPET and immunoSPECT have been used for the non-invasive imaging of gastric cancer in both preclinical and clinical studies. The first section of the review will discuss the use of immunoPET in GC targeting the antigens carcinoma-associated antigen (MG7) [ 14 ], programmed death-1 (PD-1) [ 16 ], cadherin-17 (CDH17) [ 15 ], human epidermal growth factor receptors 2 and 3 (HER2 [ 5 , 9 , 21 , 22 , 23 , 24 ] and HER3 [ 12 ]), hepatocyte growth factor (HGF [ 11 ]), and the mesenchymal-epithelial transition factor (MET) [ 10 ]. As shown in Table 1 , FDA-approved or newly developed antibodies targeting membrane antigens were radiolabeled with gallium-68 ( 68 Ga), technetium-99m ( 99m Tc), indium-111 ( 111 In), copper-64 ( 64 Cu), zirconium-89 ( 89 Zr), and bromine-76 ( 76 Br) and used for PET or SPECT imaging of gastric tumors.…”

“…Indeed, HER2 heterogeneity exists not only between primary tumors and metastases, but also in the primary tumor alone, and within the same lesion before and after treatment [ 6 ]. In this context, PET and SPECT using radiolabeled anti-HER2 antibodies have demonstrated great potential to non-invasively image and monitor HER2 expression in both preclinical models and patients with GC [ 5 , 9 , 21 , 22 , 23 , 24 ].…”

“…In addition to zirconium-89, trastuzumab was radiolabeled with the shorter-lived radioisotope copper-64 [ 9 ]. At 36 h after intravenous injection of the radioconjugate in PDX GC models, 64 Cu-NOTA-trastuzumab demonstrated higher tumor uptake than the negative control 64 Cu-NOTA-hIgG1.…”

“…Additionally, radiolabeled trastuzumab accumulation was higher in HER2-expressing PDXs than in tumors containing low levels of the protein. Clinical studies further validated the ability of 64 Cu-NOTA-trastuzumab to image GC [ 9 ].…”

“…The use of FDG-PET is associated with false-negative and false-positive images that could misdirect therapy planning and decrease diagnostic accuracy. In this context, PET has evolved into immunoPET, wherein antibodies with high specificity for antigens overexpressed or uniquely expressed in tumor cells are labeled with PET radiometals [ 5 , 9 , 10 , 11 , 12 , 13 , 14 ]. In addition to PET, antibodies radiolabeled with single photon emission computed tomography (SPECT) radiometals allow noninvasive, highly sensitive imaging of GC [ 15 , 16 ].…”