Preclinical computational dosimetry of Nanobodies in targeted radionuclide therapy of HER-2 over-expressing cancers

Saldarriaga Vargas Clarita

Promoter

Caveliers Vicky, (VUB), vickycaveliers@scarlet.be

SCK•CEN Mentor

Saldarriaga Vargas Clarita
clarita.saldarriaga.vargas@sckcen.be
+32 14 33 88 14

SCK•CEN Co-mentor

Struelens Lara
lara.struelens@sckcen.be
+32 14 33 28 85

Expert group

Research in Dosimetric Applications

PhD started

2017-04-01

Short project description

Nanobodies are the smallest antigen-binding fragments (molecular weight < 15 kDa) derived from heavy chain-only antibodies naturally occurring in Camelidae. These proteins combine both rapid tumour penetration and rapid blood clearance. Additionally they are highly soluble in aqueous solution and highly robust. In a series of preclinical studies, the In Vivo Cellular and Molecular Imaging (ICMI) lab of the Vrije Universiteit Brussel demonstrated that in vivo, radiolabelled Nanobodies efficiently penetrate tumours and tissues and bind very fast to biomarkers expressed on cells. Unbound circulating Nanobody is rapidly cleared from blood and non-target organs through kidneys. However, the cleared radioactivity is partially trapped in the kidney cortex. This can become a major dosimetric concern in terms of renal tissue toxicity when translating this development to a clinical application. However, a recent preclinical study in collaboration with SCK·CEN, showed that an i.v. injected 177Lu-labelled anti-HER-2 Nanobody efficiently targeted HER-2 over-expressing tumours, while radioactivity levels in non-target organs were low. Kidney retention could be minimized by removing the c-terminal hexahistidine tail and with a coinfusion of 177Lu-Nanobody with gelofusin. No signs of toxicity were noted by histological analysis of kidneys of treated animals. Based on the preliminary biodistribution studies and dosimetric calculations 177Lu-labelled Nanobodies delivered a similar radioactive dose to both tumour and kidneys. Despite the fact that signs of renal inflammation, apoptosis or necrosis were absent in histopathology studies, a further reduction of renal retention might be more optimal for clinical translation. Therefore, the radiolabelling of anti-HER-2 Nanobodies with 131I was initiated, using the prostetic group N-succinimidyl 4-guanidinomethyl-3-iodobenzoate (SGMIB). SGMIB stabilises 131I and maximizes retention in tumour cells. In kidneys on the other hand, the 131I-labelled Nanobodies are metabolised and cleared to the bladder, leading to a significant reduction of long-term kidney retention observed in in vivo biodistribution studies, while tumour targeting is maintained.

These promising results initiated the need for further investigation of the potential use of the alpha-emitting radionuclide 211At for radiolabelling Nanobodies. The combination of their specific targeting capacity and the localised energy deposition of alpha particles at high LET would allow the delivery of a highly specific toxic load to residual or micrometastatic cancer cells, while minimising impairment to healthy cells. The relatively short half-life of 211At (7.2 h) would also be beneficial in terms of dosimetry and would match the relatively short biological half-life of Nanobodies. Moreover Choi et al. studied already, in collaboration with ICMI, the labelling anti-HER-2 Nanobodies with N-succinimidyl 3-[211At]astato-4-guanidinomethylbenzoate (SAGMB) as an analogue for SGMIB and reported encouraging results.

So we believe that radionuclide therapy with both alpha- and bèta-emitting Nanobodies has great potential, but there is a need to integrate more accurate dose estimates of different target and non-target organs in the ongoing preclinical mice studies using radiolabelled Nanobodies to treat HER-2 over-expressing cancers.

Accurate dose estimates in preclinical models (like mice) have become indispensable to support and explain the results of histological analysis of targeted and untargeted tissues and the outcome of renal function studies. In this respect, there is still a lot of improvement possible regarding the preclinical internal dosimetry of radiolabelled Nanobodies. Detailed preclinical computational dosimetry is therefore one of the cornerstones of strategies in future clinical trials with 131I- and 211At-labelled Nanobodies.

Objective

The global aim of the project will be to perform detailed computational dosimetry of targeted and untargeted tissues after targeted radionuclide therapy with 131I- and 211At-labelled anti-HER-2 Nanobodies with a major focus on kidney dosimetry.

General internal dosimetry tools following the MIRD schema use biokinetics of radiopharmaceuticals to calculate estimates of the radiation dose to all organs and the entire body. This organ-level or single-region dosimetry is based on the assumption of a uniform distribution of radioactivity across the entire organ. The average absorbed dose to the entire organ predicted by such models can misrepresent local regional doses to specific substructures (6). Variations in absorbed dose can be of great importance in the case of radiolabelled Nanobodies which are partially trapped in the cortical region of the kidneys.

Moreover, this importance will increase when Nanobodies are labelled with alpha-emitters for which smaller scale dosimetry methods should be applied. For alpha-particle emitters, accurate dosimetry calculations require knowledge of the activity distribution as a function of time at the cellular and even subcellular levels. To tackle the problem of preclinical dosimetry of 131I- and 211At-labelled anti-HER-2 Nanobodies several steps will be taken:

In a first phase, in vivo quantitative imaging at different time intervals of mice treated with 131I-labelled Nanobodies will be performed using the MILabs micro SPECT/PET/CT/MR imaging platform, recently installed at ICMI-Lab VUB. This platform allows imaging of anatomical tissues and of the biodistribution of 131I-labelled Nanobodies with a sub-mm spatial resolution. Measured Time-Activity Curves (TACs) will be used to obtain total cumulated activities in different tissues of interest per unit of administered activity of radiopharmaceutical. Additional spatial information at higher resolution will be obtained by parallel performed ex vivo autoradiography. Image segmentation and processing will allow generating a series of voxel phantoms of different targeted and untargeted tissues which will be used for Monte Carlo simulations to calculate S-factors for specific substructures of individual organs. This multiregional approach will be compared to histological analysis and renal function studies, which will be performed in parallel at ICMI.

In the next step of the project, the time and spatial biodistribution of the 131I-labelled Nanobodies can be used to perform multiregional computational dosimetry after a simulated radionuclide exchange of 131I by its alpha-emitting surrogate 211At. Validation of this proof of principle method should be done by studying the feasibility of in vivo quantitative imaging using micro SPECT/CT of mice after injection with 211At-labelled Nanobodies and ex vivo autoradiography. Again, this multiregional approach for 211At-labelled Nanobodies will be compared to histological analysis and renal function studies parallel performed at ICMI.

Finally the focus of the project will shift to smaller scale dosimetry of 211At-labelled Nanobodies. The input for this part will be delivered by a parallel ongoing study in which time-activity distributions of 211At-labelled Nanobodies in cell clusters of targeted and untargeted organs will be determined through ex vivo imaging using the Alpha Camera technology at the University of Gothenburg. This technology allows ex vivo imaging with a spatial resolution of 35 µm. Detailed information of the spatial distribution of 211At-labelled Nanobodies at this small scale should be used as input for dedicated computational dosimetry codes enabling to calculate absorbed doses to isolated cells and cell clusters of 211At-labelled Nanobodies for a simulated location at the cell surface or in the cytoplasm.

Results of the smaller scale and cell-level dosimetry approach will be evaluated against the more macroscopic multiregional approach.