Claudin-4 SPECT imaging allows detection of aplastic lesions in a mouse model of breast cancer

Abstract

The expression of claudin-4, a protein involved in tight junction complexes, is widely dysregulated in epithelial malignancies. Claudin-4 is overexpressed in several premalignant precursor lesions, including those of cancers of the breast, pancreas, and prostate, and is associated with poor survival. A noncytotoxic C-terminal fragment of Clostridium perfringens enterotoxin (cCPE) is a natural ligand for claudin-4. Here, we demonstrate whole-body quantitative SPECT imaging of preneoplastic breast cancer tissue using 111In-labeled cCPE. Methods: cCPE.GST or GST (GST is glutathione S-transferase) was conjugated to the metal ion chelator benzyl-diethylenetriaminepentaacetic acid to allow 111In radiolabeling. The affinity of radiolabeled cCPE.GST for claudin-4 was confirmed using claudin-4-expressing MDA-MB-468 and SQ20b cells, compared with claudin-4-negative HT1080 cells. In vivo SPECT imaging was performed using athymic mice bearing MDA-MB-468 or HT1080 xenografts and using genetically modified BALB/neuT mice, which spontaneously develop claudin-4-expressing breast cancer lesions. Results: The uptake of 111In-cCPE.GST in claudin-4-positive MDA-MB-468 xenograft tumors in athymic mice was significantly higher than in 111In-GST or claudin-4-negative HT1080 tumors (6.72 ± 0.18 vs. 3.88 ± 1.00 vs. 2.36 ± 1.25 percentage injected dose per gram [%ID/g]; P < 0.0001). No other significant differences were observed in any of the examined organs. BALB/neuT mice, expressing rat neuT under mmtv promotor control, spontaneously developed tumorous lesions within their mammary fat pads over the course of 130 d. Overt mammary tumors were claudin-4-positive, and 111In-cCPE.GST uptake was 3.2 ± 0.70%ID/g, significantly higher than 111In-GST (1.00 ± 0.60%ID/g; P < 0.05). Mammary fat pads in mice aged 80 d bore claudin-4-positive aplastic lesions and accumulated 111In-cCPE.GST (3.17 ± 0.51%ID/g) but not 111In-GST (0.99 ± 0.39%ID/g; P < 0.001). Conclusion: Taken together, 111In-cCPE.GST targets claudin-4 expression in frank tumors and preneoplastic tissue, and cCPE imaging may be used as an early detection tool for breast, prostate, and pancreatic cancer.