Specific molecular imaging of BALB/c model mice with Graves' ophthalmopathy based on high expression of insulin-like growth factor 1 receptor

Abstract

Objective: At present, most of the targeted imaging based on insulin-like growth factor 1 receptor (IGF-1R) is for tumor research, and there is no IGF-1R-targeted imaging for Graves' ophthalmopathy(GO). This study aims to develop a peptide probe, ^99mTc-Z_IGF1R:4551-GGGC, targeting the IGF-1R, and to achieve specific imaging in Graves' disease (GD) animal models exhibiting GO.

Methods: 99mTc-ZIGF1R:4551-GGGC probe was synthesized using a direct labeling method and its labeling efficiency assessed via instant thin-layer chromatography (ITLC). Western blot analysis confirmed the overexpression of IGF-1R in malignant melanoma B16F10 cells. Subsequent SPECT/CT whole-body imaging of B16F10 tumor-bearing mice evaluated the probe's targeting accuracy. In addition, a GO model was established using an electroporation immunoassay, followed by serological and histopathological examinations. The GO models then underwent 99mTc-ZIGF1R:4551-GGGC SPECT/CT imaging to assess eye-targeted imaging capabilities.

Results: The peptide probe exhibited a labeling efficiency exceeding 90%. Both GD and GO models were effectively created via electroporation immunoassay. Imaging results indicated significant accumulation and retention of the peptide probes in the tumors of B16F10 tumor-bearing mice. In the GO models, probe uptake was predominantly observed in retrobulbar tissues, contrasting with primary accumulation in the lungs and gastrointestinal tract in normal mice, where only minimal tracer was observed in retrobulbar tissues. Notably, GO mice demonstrated higher probe uptake and prolonged retention.

Conclusion: This study successfully established GD and GO models, reducing the duration of the immune cycle. Moreover, a peptide probe targeting IGF-1R was synthesized, enabling specific imaging of retrobulbar tissues in GO models.

Keywords: Graves’ disease; Graves’ ophthalmopathy; IGF-1R; Peptide probe; TSHR; ZIGF1R:4551-GGGC.

Fig. 1

Results of IGF-1R expression in B16F10 and Cal62 cells

Fig. 2

Grouping and related operations of BALB/c mice

Fig. 3

Examples of ITLC for ^99mTc-Z_IGF1R:4551-GGGC and ^99mTcO₄⁻

Fig. 4

Imaging of tumor-bearing mice in the experimental group and control group. A and C SPECT/CT images of the experimental group 0.5 h and 4 h after injection of ^99mTc-Z_IGF1R:4551-GGGC, respectively. B and D SPECT/CT images of the control group 0.5 h and 4 h after injection of ^99mTcO₄.⁻, respectively. (In the red box is the tumor)

Fig. 5

Weight and serological changes in each group

Fig. 6

Thyroid SPECT/CT imaging. A Thyroid imaging before and after immunization in group A. B Thyroid imaging before and after immunization in group B. C Thyroid imaging at weeks 0 and 18 of group C (red box represents thyroid)

Fig. 7

Positive sagittal position of ocular MRI T2. A Group A imaging before and after immunization: a eye muscle tissue with slightly low-signal, b optic nerve surrounded by cerebrospinal fluid with high signal, c harderian glands with high signal; B group B imaging before and after immunization

Fig. 8

^99mTc-Z_IGF1R:4551-GGGC imaging of groups A and B and ^99mTcO₄⁻ imaging of group A

Fig. 9

Immunohistochemical analysis of thyroid and retrobulbar tissue. A The expression of TSHR in thyroid tissues of each group; B the expression of IGF-1R in thyroid tissues of each group; C the expression of TSHR in retrobulbar tissues of each group; D the expression of IGF-1R in retrobulbar tissues of each group