Establishment and characterisation of tumour-bearing mouse models for evaluation of biodistribution of a radiopharmaceutical

Abstract

Introduction: To improve early detection of breast and ovarian cancer, characterised animal models of cancer are required for screening novel tumour-specific imaging radiopharmaceuticals. The purpose of this study was to establish and characterise allograft and xenograft tumour mouse models of breast and ovarian malignancies, respectively, for evaluation of 64Cu-GluCAB, a novel imaging radiopharmaceutical intended to target tumours through their high expression of glucose receptor (GLUT-1) and their increased vascularisation. Methods: The breast tumour allograft model was established by subcutaneous inoculation of E0771 cells suspended in Matrigel into the mammary fat pad of female C57BL/6 mice. The ovarian tumour xenograft model was established by subcutaneous inoculation of OVCAR-3 cells, with and without Matrigel, above the proximal tibia of the female athymic nude (nu/nu) mice. Tumour growth was monitored using a digital calliper and the tumours were excised after reaching the end-point tumour volume (≥300 mm3) to determine the tumour growth rate and confirm malignancy using haematoxylin and eosin (H &E) staining. To illustrate the application of the tumour models established, the E0771 derived allograft model was used for investigation of the ex vivo biodistribution and in vivo imaging of 64Cu-GluCAB. The mice were administered intravenously with 64Cu-GluCAB precursor (without albumin) and images acquired at 1, 2, 6 and 24 hours using microPET/CT. After 24 hours, blood, tumours and several organs and tissues were collected to determine the compound biodistribution using a gamma counter. Flow cytometry and immunofluorescence staining were conducted in order to evaluate the expression of the GLUT-1 receptor in E0771 cells and E0771 derived tumours. Results: Palpable tumours were detected within one-week post inoculation for the E0771 derived allograft model, with a tumour take rate of 100% (26/26) and average tumour growth rate of 0.03 g/day based on the final ex vivo tumour weight. For the OVCAR-3 derived xenograft model, tumours were palpable within approximately one month and two months with and without Matrigel, respectively, however, the tumour growth rate (based on the final ex vivo tumour weight) with or without Matrigel was statistically insignificant (p>0.05). Histological analysis revealed that the tumours of both models were malignant and actively proliferating. The biodistribution profile of 64Cu-GluCAB illustrated high accumulation of radioactivity in the plasma (4.07 ± 0.21%ID/g), confirming that 64Cu-GluCAB precursor (without albumin) bound to albumin in vivo thereby increasing the biological half-life of the compound. In correlation with the microPET/CT images, high uptake was observed in the liver (3.63 ± 0.80 %ID/g) and large intestine (2.82 ± 1.29 %ID/g), suggesting hepatobiliary excretion of the compound. In contrast, uptake of 64Cu-GluCAB by tumours (0.95 ± 0.30 %ID/g) and other organs was minimal. Moreover, the tumours could not be visualised using microPET/CT. Evaluation of GLUT-1 receptor expression in E0771 cells and E0771 derived tumour, yielded inconclusive results. Conclusion: The tumour-bearing mouse models of breast and ovarian cancers were successfully developed and characterised. Although the expression of the GLUT-1 receptor could not be confirmed, the biodistribution profile of 64Cu-GluCAB indicated a minimal amount of uptake by the tumour. The low radioactivity signal could however not be used for localisation and visualisation of the tumour by microPET/CT.