Theranostic 64Cu-DOTHA2-PSMA allows low toxicity radioligand therapy in mice prostate cancer model

Introduction We have previously shown that copper-64 (64Cu)-DOTHA2-PSMA can be used for positron emission tomography (PET) imaging of prostate cancer. Owing to the long-lasting, high tumoral uptake of 64Cu-DOTHA2-PSMA, the objective of the current study was to evaluate the therapeutic potential of 64Cu-DOTHA2-PSMA in vivo. Methods LNCaP tumor-bearing NOD-Rag1nullIL2rgnull (NRG) mice were treated with an intraveinous single-dose of 64Cu-DOTHA2-PSMA at maximal tolerated injected activity, natCu-DOTHA2-PSMA at equimolar amount (control) or lutetium-177 (177Lu)-PSMA-617 at 120 MBq to assess their impact on survival. Weight, well-being and tumor size were followed until mice reached 62 days post-injection or ethical limits. Toxicity was assessed through weight, red blood cells (RBCs) counts, pathology and dosimetry calculations. Results Survival was longer with 64Cu-DOTHA2-PSMA than with natCu-DOTHA2-PSMA (p < 0.001). Likewise, survival was also longer when compared to 177Lu-PSMA-617, although it did not reach statistical significance (p = 0.09). RBCs counts remained within normal range for the 64Cu-DOTHA2-PSMA group. 64Cu-DOTHA2-PSMA treated mice showed non-pathological fibrosis and no other signs of radiation injury. Human extrapolation of dosimetry yielded an effective dose of 3.14 × 10-2 mSv/MBq, with highest organs doses to gastrointestinal tract and liver. Discussion Collectively, our data showed that 64Cu-DOTHA2-PSMA-directed radioligand therapy was effective for the treatment of LNCaP tumor-bearing NRG mice with acceptable toxicity and dosimetry. The main potential challenge is the hepatic and gastrointestinal irradiation.


Dosimetry calculations 1) Data acquisition
For healthy organs, proportions of injected activity per gram (%IA/g) is obtained by biodistributions (results previously published, (2)). Data is corrected for physical decay to injection time. For tumor, %IA/cc is obtained from positron emission imaging.

2) Generation of time-activity curve and calculation of area under the curve
Physical decay is reapplied to %IA/g to represent time of death of the animal (1 h, 2 h, 4 h, 24 h or 48 h p.i.). A curve is plotted for each organ in Graphpad Prism 8 (X: time (h), Y: %IA/g. Area under the curve (AUC) is calculated by geometrical method from 0 h to 48 h or 0 to 24 h in tumor. To calculate the area under the curve from 48 h to infinity (24h to infinity for tumor), we assumed physical decay only using %IA/g at 48h (24h for tumor) as a starting point. Both values are added to form a total AUC, representing the total fraction of disintegrations per injected dose per gram of organ.

3) Mouse and tumor dosimetry calculation
Tumor kinetics was obtained by multiplying tumor AUC to the average tumor mass at time of radiotherapy and imaging (estimating from measured volume, assuming 1 g = 1 cm 3 ). In OLINDA/EXM 2.2.3, sphere model was used. Whole-body dosimetry for mouse was obtained by multiplying AUC by 25 g mouse model organs weight. Remainder of the body kinetic value was calculated from the muscle and biodistributed organs not included in OLINDA/EXM model. For muscle, lean mass weight of 19.2 g minus all known organs weight was used (9.46 g), therefore assuming muscle AUC to for organs with an unknown number of disintegrations.

4) Extrapolation to human
Using organ masses provided by OLINDA/EXM for the human male model IRCP 89, we extrapolated mice AUC to human proportions using Sparks method (1): Where "AUC" means "area under the curve", "w" means "weight" for either a single human tissue (e.g., liver), mice total weight (25 g) or human total weight (73 kg). Skin, fat, muscle and seminal vesicles were not available in the OLINDA model. Average masses from the literature were used and their estimated human AUC were added to form "Remainder of the body" (or "Total body"), scaled for their human weight. Values used for model unknowns were: average fat mass of 19.4 kg, average muscle mass of 25.6 kg, average skin mass of 12.2 kg, and average seminal vesicles mass of 4.57 g.

5) Human dosimetry calculation
OLINDA/EXM 2.2.3 software (Hermes Medical Solution) was used to calculate doses. To calculate 64 Cu dose factor, OLINDA/EXM creator used emissions that contributed to 0.01% or more of the total decay scheme based on data from Brookhaven National Laboratory National Nuclear Data Center.
The average adult human male model IRCP 89 is used for healthy organ dosimetry. Blood AUC value was used for "red marrow". Stomach AUC value was used for "stomach contents". Whole-heart AUC value was used for "heart wall" and blood AUC scaled to the heart-blood volume was used for "heart contents". Bowel AUC value was an average value and is therefore scaled to the size of every section and used for the small intestine, the right colon, the left colon and the rectum. Bone AUC value was used for cortical and trabecular compartments and "value in bone volume" was selected. Gallbladder wall and urinary bladder wall values were unavailable due to difficulty with sampling. Other unavailable AUCs: eyes, prostate, thymus, esophagus.

6) Confidence intervals
95% confidence intervals (CI) was obtained on biodistribution data. Dosimetry was then calculated following the previously described method with the maximum and minimum from this interval for every organ.  12160.0 9.0077E-01 (6.0869E-01 -1.1924E00) Remainder of the body (sum of the above) 2.8505E00 (2.1236E00 -3.5756E00) * Values exceptionally presented with up to four decimals to facilitate reproduction.