Misaki Kondo, Zhongli Cai, Conrad Chan, Nubaira Forkan, and Raymond M. Reilly
EJNMMI Radiopharm Chem, 2023
Trastuzumab (Herceptin) has improved the outcome for patients with HER2-positive breast cancer (BC) but brain metastases (BM) remain a challenge due to poor uptake of trastuzumab into the brain. Radioimmunotherapy (RIT) with trastuzumab labeled with α-particle emitting, 225Ac may overcome this challenge by increasing the cytotoxic potency on HER2-positive BC cells. Our first aim was to synthesize and characterize [111In]In-DOTA-trastuzumab and [225Ac]Ac-DOTA-trastuzumab as a theranostic pair for imaging and RIT of HER2-positive BC, respectively. A second aim was to estimate the cellular dosimetry of [225Ac]Ac-DOTA-trastuzumab and determine its cytotoxicity in vitro on HER2-positive BC cells. A third aim was to study the tumour and normal tissue uptake of [225Ac]Ac-DOTA-trastuzumab using [111In]In-DOTA-trastuzumab as a radiotracer in vivo in NRG mice with s.c. 164/8-1B/H2N.luc+ human BC tumours that metastasize to the brain.
Trastuzumab was conjugated to 12.7 ± 1.2 DOTA chelators and labeled with 111In or 225Ac. [111In]In-DOTA-trastuzumab exhibited high affinity specific binding to HER2-positive SK-BR-3 human BC cells (KD = 1.2 ± 0.3 × 10–8 mol/L). Treatment with [225Ac]Ac-DOTA-trastuzumab decreased the surviving fraction (SF) of SK-BR-3 cells dependent on the specific activity (SA) with SF < 0.001 at SA = 0.74 kBq/µg. No surviving colonies were noted at SA = 1.10 kBq/µg or 1.665 kBq/µg. Multiple DNA double-strand breaks (DSBs) were detected in SK-BR-3 cells exposed to [225Ac]Ac-DOTA-trastuzumab by γ-H2AX immunofluorescence microscopy. The time-integrated activity of [111In]In-DOTA-trastuzumab in SK-BR-3 cells was measured and used to estimate the absorbed doses from [225Ac]Ac-DOTA-trastuzumab by Monte Carlo N-Particle simulation for correlation with the SF. The dose required to decrease the SF of SK-BR-3 cells to 0.10 (D10) was 1.10 Gy. Based on the D10 reported for γ-irradiation of SK-BR-3 cells, we estimate that the relative biological effectiveness of the α-particles emitted by 225Ac is 4.4. Biodistribution studies in NRG mice with s.c. 164/8-1B/H2N.luc+ human BC tumours at 48 h post-coinjection of [111In]In-DOTA-trastuzumab and [225Ac]Ac-DOTA-trastuzumab revealed HER2-specific tumour uptake (10.6 ± 0.6% ID/g) but spleen uptake was high (28.9 ± 7.4% ID/g). Tumours were well-visualized by SPECT/CT imaging using [111In]In-DOTA-trastuzumab.
We conclude that [225Ac]Ac-DOTA-trastuzumab exhibited potent and HER2-specific cytotoxicity on SK-BR-3 cells in vitro and HER2-specific uptake in s.c. 164/8-1B/H2N.luc+ human BC tumours in NRG mice, and these tumours were imaged by SPECT/CT with [111In]In-DOTA-trastuzumab. These results are promising for combining [111In]In-DOTA-trastuzumab and [225Ac]Ac-DOTA-trastuzumab as a theranostic pair for imaging and RIT of HER2-positive BC.
Results from nanoScan® SPECT/CT
Fig. 5 a Tumour and normal tissue uptake at 48 h p.i. of [111In]In-DOTA-trastuzumab (7–8 MBq) coinjected with [225Ac]Ac-DOTA-trastuzumab (4 kBq; total mass dose = 40 µg) in NRG mice with s.c. HER2-positive 164/8-1B/H2N.luc + human BC tumours. B: blood, Br: brain, H: heart, Lu: lungs, K: kidneys, Pn: pancreas, Sp: spleen, L: liver, S: stomach, I: intestine, Sk: skin, M: muscle, Bo: bone. Tissue activity was quantified by γ-counting of 111In. Values shown are the mean ± SD (n = 4–5). Significant differences (*P < 0.05) are indicated by asterisks. b Representative SPECT/CT images of tumour-bearing NRG mice at 48 h p.i. of [111In]In-DOTA-trastuzumab coinjected with [225Ac]Ac-DOTA-trastuzumab (left image) or [111In]In-DOTA-IgG1 coinjected with [225Ac]Ac-DOTA-IgG1 (right image). T: tumour, Sp: spleen. Scale for SPECT/CT images shows the percent injected dose/g (%ID/g). Also shown in grayscale is the scale for the CT image
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