111In-1,4,7,10-Tetraazacyclododecane-1,4,7,10-tetraacetic acid-anti-insulin-like growth factor 1 receptor Affibody ZIGF1R:4551

111In-DOTA-ZIGF1R:4551

Leung K.

Publication Details

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In vitro Rodents

Background

[PubMed]

Insulin-like growth factor 1 (IGF-1, 70 amino acids) is secreted by the liver and target tissues in response to growth hormone stimulation (1). IGF-1 plays an important role in cell growth and development. IGF-1 binds to IGF-1 receptor (IGF-1R) with ~1,000-fold higher affinity than to insulin receptor. IGF-1R is a transmembrane tyrosine kinase receptor that is highly expressed in many human cancers (2). IGF-1R plays an important role in tumor proliferation, apoptosis, angiogenesis, and metastasis (3, 4). Triple-negative breast tumors are negative for estrogen receptors, progesterone receptors, and epidermal growth factor receptor-2 (HER2). Patients with triple-negative breast tumors have a lower survival rate than patients with estrogen receptor- or HER2-positive breast tumors (5). IGF-1R is overexpressed in 36% of all triple-negative breast carcinomas (6).Therefore, monitoring of IGF-1R expression in breast tumor lesions with a noninvasive imaging modality is important for development of therapeutic treatments of these patients with anti-IGF-1R monoclonal antibodies (7). The human anti-IGF-1R monoclonal antibody R1507 is directed against the extracellular domain of human IGF-1R (8). However, the pharmacokinetics of intact radiolabeled mAbs, with high liver uptake and slow blood elimination, are generally not ideal for imaging. Smaller antibody fragments, such as Fab or F(ab')2, have better imaging pharmacokinetics because they are rapidly excreted by the kidneys. A novel class of recombinant affinity ligands (Affibody molecules) for HER2 was constructed based on the 58-amino-acid Z-domain residues from one of the IgG-binding domains of staphylococcal protein A (9). Affibody molecules exhibit high binding affinity to HER2 with Kd values of <1 nM. Various radiolabeled Affibody molecules have been studied in terms of their ability to image HER2 in tumors [PubMed]. A new Affibody with site-specific cysteine at the C-terminus for labeling, ZIGF1R:4551, has been identified to bind to IGF-1R with high affinity (10). 1,4,7,10-Tetraazacyclododecane-1,4,7-tris-acetic acid-10-maleimidoethylacetamide (MMA-DOTA) chelator was used for 111In labeling of ZIGF1R:4551. 111In-[MMA-DOTA-Cys]-ZIGF-1R:4551 (111In-DOTA-ZIGF-1R:4551) was evaluated in nude mice bearing human prostate DU-145 tumors.

Synthesis

[PubMed]

Affibody ZIGF-1R:4551 was reduced with dithiothreitol in 0.02 M ascorbic acid for 18 h at 37°C (10). The reduced ZIGF-1R:4551 was incubated with MMA-DOTA in 1:1 molar ratio in ammonium acetate buffer (pH, 5.5) for 18 h at 37°C. MMA-DOTA-ZIGF-1R:4551 was isolated with column chromatography. MMA-DOTA-ZIGF-1R:4551 (10 nmol) was mixed with 60 MBq (1.6 mCi) 111InCl3 and incubated for 30 min at 90°C. The labeling efficiency of 111In incorporation was >99%. Specific activity of the preparation was ~6 GBq/µmol (162 mCi/µmol). 111In-DOTA-ZIGF-1R:4551 was found to be stable after incubation for 4 h in the presence of 1,000-fold molar excess of EDTA.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

Tolmachev et al. (10) reported that ZIGF-1R:4551 exhibited an affinity constant (Kd) of 500 ± 70 pM with MCF-7 cells using flow cytometry. 111In-DOTA-ZIGF-1R:4551 saturation binding studies with DU-145 cells and DOTA-ZIGF-1R:4551 (displacement ligand) showed that there were 25,000 ± 3,000 receptor sites per cells. Cell binding and internalization of 111In-DOTA-ZIGF-1R:4551 (1.5 nM) were determined at 1–4 h of incubation with DU-145 cells at 37°C. Cell-associated radioactivity reached a plateau after 1 h, whereas internalization of 111In-DOTA-ZIGF-1R:4551 gradually increased with time of incubation from 25% at 1 h to 35% of total cellular radioactivity at 4 h.

Animal Studies

Rodents

[PubMed]

Tolmachev et al. (10) performed ex vivo biodistribution studies in normal mice (n = 4/group) at 4 h after intravenous injection of 0.025 MBq (0.00067 mCi) 111In-DOTA-ZIGF-1R:4551. The kidney showed the highest accumulation of radioactivity with 40% injected dose/gram (ID/g), followed by the liver (4% ID/g), salivary gland (3% ID/g), lung (2% ID/g), spleen (<2% ID/g), pancreas (<2% ID/g), and stomach (<1% ID/g). The blood radioactivity level was 0.5% ID/g. Co-injection of DOTA-ZIGF-1R:4551 (1.6 nmol) lowered the radioactivity levels in the pancreas, lung, stomach, and salivary gland (P < 0.05), organs known to be rich in IGF-1R.

Tolmachev et al. (10) performed ex vivo biodistribution studies in nude mice (n = 4/group) bearing DU-145 xenografts at 1–24 h after injection of 0.025 MBq (0.00067 mCi) 111In-DOTA-ZIGF-1R:4551. The maximum accumulation of radioactivity in all tissues studied was observed at 1 h after injection. There was a rapid clearance from the blood and lung, whereas the radioactivity levels remained almost constant in the kidney, muscle, and bone. There was a slight washout from the tumors, pancreas, salivary gland, spleen, and stomach. The tumor/blood ratios were 1.2, 2.5, and 5.2 at 1, 4, and 24 h after injection, respectively. Co-injection of 50 nmol DOTA-ZIGF-1R:4551 decreased the tumor/blood ratio to ~1 at 4 h after injection. Planar gamma imaging showed a clear visualization of the tumors with the tumor/contralateral site ratio of 2.9 at 8 h after injection. The kidneys, liver, and salivary gland were also visualized.

Other Non-Primate Mammals

[PubMed]

No publication is currently available.

Non-Human Primates

[PubMed]

No publication is currently available.

Human Studies

[PubMed]

No publication is currently available.

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