Imaging the distribution of an antibody-drug conjugate constituent targeting mesothelin with 89Zr and IRDye 800CW in mice bearing human pancreatic tumor xenografts.

Mesothelin is a tumor differentiation antigen expressed by epithelial tumors, including pancreatic cancer. Currently, mesothelin is being targeted with an antibody-drug conjugate (ADC) consisting of a mesothelin-specific antibody coupled to a highly potent chemotherapeutic drug. Considering the toxicity of the ADC and reduced accessibility of pancreatic tumors, non-invasive imaging could provide necessary information. We therefore developed a zirconium-89 (89Zr) labeled anti-mesothelin antibody (89Zr-AMA) to study its biodistribution in human pancreatic tumor bearing mice. Biodistribution and dose-finding of 89Zr-AMA were studied 144 h after tracer injection in mice with subcutaneously xenografted HPAC. MicroPET imaging was performed 24, 72 and 144 h after tracer injection in mice bearing HPAC or Capan-2. Tumor uptake and organ distribution of 89Zr-AMA were compared with nonspecific 111In-IgG. Biodistribution analyses revealed a dose-dependent 89Zr-AMA tumor uptake. Tumor uptake of 89Zr-AMA was higher than 111In-IgG using the lowest tracer dose. MicroPET showed increased tumor uptake over 6 days, whereas activity in blood pool and other tissues decreased. Immunohistochemistry showed that mesothelin was expressed by the HPAC and CAPAN-2 tumors and fluorescence microscopy revealed that AMA-800CW was present in tumor cell cytoplasm. 89Zr-AMA tumor uptake is antigen-specific in mesothelin-expressing tumors. 89Zr-AMA PET provides non-invasive, real-time information about AMA distribution and tumor targeting.

Visualising dual downregulation of insulin-like growth factor receptor-1 and vascular endothelial growth factor-A by heat shock protein 90 inhibition effect in triple negative breast cancer.

PURPOSE: Triple negative breast cancer (TNBC) is biologically characterised by heterogeneous presence of molecular pathways underlying it. Insulin-like growth factor receptor-1 (IGF-1R) expression and vascular endothelial growth factor-A (VEGF-A) have been identified as key factors in these pathways in TNBC. In this study, we aimed at in vivo PET imaging the effect of heat shock protein (Hsp) 90 inhibition by means of NVP-AUY922 on these pathways, with zirconium-89 ((89)Zr) labelled antibodies targeting IGF-1R and VEGF-A. MATERIALS AND METHODS: In vitro NVP-AUY922 effects on cellular IGF-1R expression and VEGF-A secretion were determined in MCF-7 and MDA-MB-231 cell lines. Moreover human TNBC bearing MDA-MB-231 mice received 50mg/kg NVP-AUY922 or vehicle q3d intraperitoneally for 21days. PET scans with (89)Zr-MAB391 and (89)Zr-bevacizumab for visualisation of IGF-1R and VEGF-A were performed before and during treatment. Ex vivo biodistribution and correlative tissue analyses were performed. RESULTS: NVP-AUY922 treatment reduced IGF-1R expression and VEGF-A excretion in both cell lines. Hsp90 inhibition lowered tumour uptake on (89)Zr-MAB391-PET by 37.3% (P<0.01) and on (89)Zr-bevacizumab-PET by 44.4% (P<0.01). This was confirmed by ex vivo biodistribution with a reduction of 41.3% injected dose (ID)/g for (89)Zr-MAB391 and 37.8% ID/g for (89)Zr-bevacizumab, while no differences were observed for other tissues. This coincided with reduced IGF-1R expression and mean vessel density in the NVP-AUY922 treated tumours. CONCLUSION: (89)Zr-MAB391 and (89)Zr-bevacizumab PET reflect effect of Hsp90 inhibitors and can therefore potentially be used to monitor therapeutic effects of Hsp90 inhibitor therapy in TNBC.

ImmunoPET and biodistribution with human epidermal growth factor receptor 3 targeting antibody 89Zr-RG7116.

The humanized monoclonal antibody with high affinity for the human epidermal growth factor receptor (HER) 3, RG7116, is a glycoengineered, IgG1 class antibody. By labeling RG7116 with zirconium-89 ((89)Zr) we aimed to visualize in vivo HER3 expression and study the biodistribution of this antibody in human tumor-bearing mice. Biodistribution of (89)Zr-RG7116 was studied in subcutaneously xenografted FaDu tumor cells (HER3-positive). Dose-dependency of (89)Zr-RG7116 organ distribution and specific tumor uptake was assessed by administering doses ranging from 0.05 to 10 mg/kg RG7116 to SCID/Beige mice. Biodistribution was analyzed at 24 and 144 h after injection. MicroPET imaging was performed at 1, 3, and 6 days after injection of 1.0 mg/kg (89)Zr-RG7116 in the FaDu, H441, QG-56 and Calu-1 xenografts with varying HER3 expression. The excised tumors were analyzed for HER3 expression. Biodistribution analyses showed a dose- and time-dependent (89)Zr-RG7116 tumor uptake in FaDu tumors. The highest tumor uptake of (89)Zr-RG7116 was observed in the 0.05 mg/kg dose group with 27.5%ID/g at 144 h after tracer injection. MicroPET imaging revealed specific tumor uptake of (89)Zr-RG7116 in FaDu and H441 models with an increase in tumor uptake over time. Biodistribution data was consistent with the microPET findings in FaDu, H441, QG56 and Calu-1 xenografts, which correlated with HER3 expression levels. In conclusion, (89)Zr-RG7116 specifically accumulates in HER3 expressing tumors. PET imaging with this tracer provides real-time non-invasive information about RG7116 distribution, tumor targeting and tumor HER3 expression levels.

89Zr-bevacizumab PET imaging in primary breast cancer.

UNLABELLED: Vascular endothelial growth factor (VEGF)-A is overexpressed in most malignant and premalignant breast lesions. VEGF-A can be visualized noninvasively with PET imaging and using the tracer (89)Zr-labeled bevacizumab. In this clinical feasibility study, we assessed whether VEGF-A in primary breast cancer can be visualized by (89)Zr-bevacizumab PET. METHODS: Before surgery, breast cancer patients underwent a PET/CT scan of the breasts and axillary regions 4 d after intravenous administration of 37 MBq of (89)Zr-bevacizumab per 5 mg. PET images were compared with standard imaging modalities. (89)Zr-bevacizumab uptake was quantified as the maximum standardized uptake value (SUV max). VEGF-A levels in tumor and normal breast tissues were assessed with enzyme-linked immunosorbent assay. Data are presented as mean ± SD. RESULTS: Twenty-five of 26 breast tumors (mean size ± SD, 25.1 ± 19.8 mm; range, 4-80 mm) in 23 patients were visualized. SUV max was higher in tumors (1.85 ± 1.22; range, 0.52-5.64) than in normal breasts (0.59 ± 0.37; range, 0.27-1.69; P < 0.001). The only tumor not detected on PET was 10 mm in diameter. Lymph node metastases were present in 10 axillary regions; 4 could be detected with PET (SUV max, 2.66 ± 2.03; range, 1.32-5.68). VEGF-A levels in the 17 assessable tumors were higher than in normal breast tissue in all cases (VEGF-A/mg protein, 184 ± 169 pg vs. 10 ± 21 pg; P = 0.001), whereas (89)Zr-bevacizumab tumor uptake correlated with VEGF-A tumor levels (r = 0.49). CONCLUSION: VEGF-A in primary breast cancer can be visualized by means of (89)Zr-bevacizumab PET.

Measurement of tumor VEGF-A levels with 89Zr-bevacizumab PET as an early biomarker for the antiangiogenic effect of everolimus treatment in an ovarian cancer xenograft model.

PURPOSE: The mTOR pathway is frequently activated in ovarian cancers. mTOR inhibitors, such as everolimus, can reduce VEGF-A production by cancer cells. We investigated whether early everolimus treatment effects could be monitored by positron emission tomography (PET) with (89)Zr-bevacizumab. EXPERIMENTAL DESIGN: The effect of everolimus on VEGF-A secretion was determined in a panel of human ovarian cancer cell lines and in A2780(luc+) ovarian cancer cells xenografted subcutaneously in BALB/c mice. Mice received daily 10 mg/kg everolimus intraperitoneally (i.p.) for 14 days. PET scans with the tracer (89)Zr-labeled bevacizumab were conducted before and after treatment. Ex vivo (89)Zr-bevacizumab biodistribution and correlative tissue analyses were conducted. Tumor VEGF-A levels were measured with ELISA and mean vascular density (MVD) was determined with immunohistochemistry. RESULTS: Everolimus treatment reduced VEGF-A levels in the supernatant of all cell lines. Everolimus lowered (89)Zr-bevacizumab tumor uptake by 21.7% ± 4.0% [mean standardized uptake value (SUV(mean)) 2.3 ± 0.2 vs. 2.9 ± 0.2, P < 0.01]. Ex vivo biodistribution also showed lower tracer uptake in the tumors of treated as compared with control animals (7.8 ± 0.8%ID/g vs. 14.0 ± 1.7%ID/g, P < 0.01), whereas no differences were observed for other tissues. This coincided with lower VEGF-A protein levels in tumor lysates in treated versus untreated tumors (P = 0.04) and reduced MVD (P < 0.01). CONCLUSION: Tumor VEGF-A levels are decreased by everolimus. (89)Zr-bevacizumab PET could be used to monitor tumor VEGF-A levels as an early biomarker of the antiangiogenic effect of mTOR inhibitor therapy.