Comparison of the Anti-Tumour Activity of the Somatostatin Receptor (SST) Antagonist [177Lu]Lu-Satoreotide Tetraxetan and the Agonist [177Lu]Lu-DOTA-TATE in Mice Bearing AR42J SST2-Positive Tumours

Abstract

Limited experiments have compared the treatment effects of repetitive cycles of radiolabelled somatostatin (SST) analogues. In vitro and in vivo experiments were conducted in an AR42J cancer cell model, comparing the antagonist [¹⁷⁷Lu]Lu-satoreotide tetraxetan with the agonist [¹⁷⁷Lu]Lu-DOTA-TATE in terms of their binding properties, biodistribution, anti-tumour activity and toxicity. Histopathological and immunohistochemical examinations were performed at different timepoints. In the in vitro assays, [¹⁷⁷Lu]Lu-satoreotide tetraxetan recognised twice as many SST₂ binding sites as [¹⁷⁷Lu]Lu-DOTA-TATE. In mice treated once a week for four consecutive weeks, [¹⁷⁷Lu]Lu-satoreotide tetraxetan (15 MBq) revealed a significantly greater median time taken to reach a tumour volume of 850 mm³ (68 days) compared to [¹⁷⁷Lu]Lu-DOTA-TATE at 15 MBq (43 days) or 30 MBq (48 days). This was associated with a higher tumour uptake, enhanced DNA damage and no or mild effects on body weight, haematological toxicity, or renal toxicity with [¹⁷⁷Lu]Lu-satoreotide tetraxetan (15 MBq). At the end of the study, complete tumour senescence was noted in 20% of animals treated with [¹⁷⁷Lu]Lu-satoreotide tetraxetan, in 13% of those treated with [¹⁷⁷Lu]Lu-DOTA-TATE at 30 MBq, and in none of those treated with [¹⁷⁷Lu]Lu-DOTA-TATE at 15 MBq. In conclusion, repeated administrations of [¹⁷⁷Lu]Lu-satoreotide tetraxetan were able to potentiate peptide receptor radionuclide therapy with a higher tumour uptake, longer median survival, and enhanced DNA damage, with a favourable efficacy/safety profile compared to [¹⁷⁷Lu]Lu-DOTA-TATE.

Keywords: AR42J; [177Lu]Lu-DOTA-TATE; [177Lu]Lu-satoreotide tetraxetan; peptide receptor radionuclide therapy; somatostatin receptor subtype 2.

Figure 1

Biodistribution of [¹⁷⁷Lu]Lu-DOTA-TATE at 15 and 30 MBq and [¹⁷⁷Lu]Lu-satoreotide tetraxetan at 15 MBq at 96 h following the fourth injection of the radiopharmaceuticals. (a) Tumour, kidney, and femur radioactivity uptake expressed as mean ± standard error of the mean. * denotes p ≤ 0.05, ** ≤0.01, *** ≤0.001, and ns, non-significant. (b) Tumour-to-organ ratios expressed as median (range).

Figure 2

Anti-tumour effects of [¹⁷⁷Lu]Lu-DOTA-TATE at 15 and 30 MBq and [¹⁷⁷Lu]Lu-satoreotide tetraxetan at 15 MBq. (a) Tumour growth over time. Data are mean ± standard error of the mean. (b) Kaplan-Meier curves of Time to Reach Volume of 850 mm³ (TTRV850).

Figure 3

Somatostatin receptor subtype 2 (SST₂) expression in tumours. The basal high SST₂ expression (a) was not modified by the administered treatments either with [¹⁷⁷Lu]Lu-DOTA-TATE (b) or [¹⁷⁷Lu]Lu-satoreotide tetraxetan (c) 96h post last administration. All images were taken using an objective 40.

Figure 4

Histopathological and immunohistochemical analyses at 96 h post-administration of last treatment. (a,b) are representative images of haematoxylin and eosin (H&E) tumour staining. (a) The arrowheads point to mitoses in a control/untreated tumour. (b) In a [¹⁷⁷Lu]Lu-satoreotide tetraxetan-treated tumour, mitoses were absent. The arrowheads point to degenerating cells. (c) Mean ± standard error of the mean (SEM) mitotic count (ranging from 22 to 66 at objective 40 in control tumours). (d,e) are representative images of sirius red staining. (d) In a control/untreated tumour, fibrosis was absent. (e) Fibrosis grade 1 (minimal) was noted in a [¹⁷⁷Lu]Lu-satoreotide tetraxetan-treated tumour. (f) Mean ± SEM fibrosis score, ranging from 0 to 4. (g,h) are representative images of pH2AX tumour staining. (g) In a control/untreated tumour, there were low levels of staining in the nuclei of cancer cells (arrowheads) unlike in a [¹⁷⁷Lu]Lu-satoreotide tetraxetan-treated tumour (h). (i) Mean ± SEM pH2AX H-score, ranging from 0 to 300. Figure (a,b,d,e) were taken at the objective 40, and figures (g,h) at the objective 100.

Figure 5

Histopathological and immunohistochemical analyses at the end of the study. Two distinct tumor phenotypes were noted: a re-growing/relapsing phenotype (a–c), and a senescent phenotype (d–f). In re-growing/relapsing tumours, mitotic figures were commonly seen (a, arrowheads) and associated with Ki67-positive staining (b). pH2AX expression was in some re-growing/relapsing tumours (c), which were treated with [¹⁷⁷Lu]Lu-satoreotide tetraxetan (h, H-score > 200). In senescent tumours, there was no presence of mitoses of degenerating cancer (d, arrowhead) and a lack of Ki67 (e) and pH2AX (f) staining in cancer cells. All images were taken at the objective 100. (g) illustrates the proportion of mice with different tumour phenotypes in each treated group. (h) illustrates the mean ± standard error of the mean pH2AX H-score at the end of the study in re-growing/relapsing tumours.

Figure 6

High expression of SST² in relapsing tumours (a) and senescent tumours (b). The isotype control antibody confirmed the specificity of the staining (c). All images were taken using the objective 40.

Figure 7

SST₂ expression in the (a) adrenal medulla, (b) colon (including the enteric nervous system and epithelial crypt cells, arrowhead), (c) spleen (rare lymphoid cells, arrowheads), and (d) bone marrow osteoblasts (arrowheads). The isotype control antibody confirmed the specificity of the staining in the different tissues, as in the bone marrow (e). SSTR2 was not detected in the kidneys (f). Figures (a–c,f) were taken using the objective 40, and figures (d,e) were taken using the objective 100.

Figure 8

Histopathological analyses of the bone marrow at 96 h post administration of last treatment and at the end of study. Compared to a control bone marrow (a), similar treatment-related toxicity was observed in all three treated groups, which was mainly characterized by an increased myeloid/erythroid ratio (b) at 96 h after last administration. An increased myeloid/erythroid ratio was also noted at the end of the study in all study groups, with the presence of rare foci of eosinophilic vacuolated cells in a limited number of animals treated with either [¹⁷⁷Lu]Lu-satoreotide tetraxetan at 15 MBq or [¹⁷⁷Lu]Lu-DOTA-TATE at 30 MBq (c, arrowheads). Figures (a–c) were taken using the objective 100.