Radiopharmaceutical therapy using α -emitting 225 Ac is an emerging treatment for patients with advanced metastatic cancers. Measurement of the spatial dose distribution in organs and tumors is needed to inform treatment dose prescription and reduce off-target toxicity, at not only organ but also sub-organ scales. Digital autoradiography with α -sensitive detection devices can measure radioactivity distributions at 20-40 μ m resolution, but anatomical characterization is typically limited to 2D. We collected digital autoradiographs across whole tissues to generate 3D dose volumes and used them to evaluate the simultaneous tumor control and regional kidney dosimetry of a novel therapeutic radiopharmaceutical for prostate cancer, [225Ac]Ac-Macropa-PEG4-YS5, in mice. 22Rv1 xenograft-bearing mice treated with 18.5 kBq of [225Ac]Ac-Macropa-PEG4-YS5 were sacrificed at 24 h and 168 h post-injection for quantitative α -particle digital autoradiography and hematoxylin and eosin staining. Gamma-ray spectroscopy of biodistribution data was used to determine temporal dynamics and 213 Bi redistribution. Tumor control probability and sub-kidney dosimetry were assessed. Heterogeneous 225 Ac spatial distribution was observed in both tumors and kidneys. Tumor control was maintained despite heterogeneity if cold spots coincided with necrotic regions. 225 Ac dose-rate was highest in the cortex and renal vasculature. Extrapolation of tumor control suggested that kidney absorbed dose could be reduced by 41% while maintaining 90% TCP. The 3D dosimetry methods described allow for whole tumor and organ dose measurements following 225 Ac radiopharmaceutical therapy, which correlate to tumor control and toxicity outcomes.

Glypican-3 (GPC3) is overexpressed in hepatocellular carcinomas and hepatoblastomas and represents an important therapeutic target but the biologic importance of GPC3 in liver cancer is unclear. To date, there are limited data characterizing the biological implications of GPC3 knockout (KO) in liver cancers that intrinsically express this target. Here, we report on the development and characterization of GPC3-KO liver cancer cell lines and compare to them to parental lines. GPC3-KO variants were established in HepG2 and Hep3B liver cancer cell lines using a lentivirus-mediated CRISPR/Cas9 system. We assessed the effects of GPC3 deficiency on oncogenic properties in vitro and in murine xenograft models. Downstream cellular signaling pathway changes induced by GPC3 deficiency were examined by RNAseq and western blot. To confirm the usefulness of the models for GPC3-targeted drug development, we evaluated the target engagement of a GPC3-selective antibody, GC33, conjugated to the positron-emitting zirconium-89 (89Zr) in subcutaneous murine xenografts of wild type (WT) and KO liver cancer cell lines. Deletion of GPC3 significantly reduced liver cancer cell proliferation, migration, and invasion compared to the parental cell lines. Additionally, the tumor growth of GPC3-KO liver cancer xenografts was significantly slower compared with control xenografts. RNA sequencing analysis also showed GPC3-KO resulted in a reduction in the expression of genes associated with cell cycle regulation, invasion, and migration. Specifically, we observed the downregulation of components in the AKT/NFκB/WNT signaling pathways and of molecules related to cell cycle regulation with GPC3-KO. In contrast, pMAPK/ERK1/2 was upregulated, suggesting an adaptive compensatory response. KO lines demonstrated increased sensitivity to ERK (GDC09994), while AKT (MK2206) inhibition was more effective in WT lines. Using antibody-based positron emission tomography (immunoPET) imaging, we confirmed that 89Zr-GC33 accumulated exclusively in GPC3-expression xenografts but not in GPC3-KO xenografts with high tumor uptake and tumor-to-liver signal ratio. We show that GPC3-KO liver cancer cell lines exhibit decreased tumorigenicity and altered signaling pathways, including upregulated pMAPK/ERK1/2, compared to parental lines. Furthermore, we successfully distinguished between GPC3+ and GPC3- tumors using the GPC3-targeted immunoPET imaging agent, demonstrating the potential utility of these cell lines in facilitating GPC3-selective drug development.

Image-based dosimetry-guided radiopharmaceutical therapy has the potential to personalize treatment by limiting toxicity to organs at risk and maximizing the therapeutic effect. The 177Lu dosimetry challenge of the Society of Nuclear Medicine and Molecular Imaging consisted of 5 tasks assessing the variability in the dosimetry workflow. The fifth task investigated the variability associated with the last step, dose conversion, of the dosimetry workflow on which this study is based. Methods: Reference variability was assessed by 2 medical physicists using different software, methods, and all possible combinations of input segmentation formats and time points as provided in the challenge. General descriptive statistics for absorbed dose values from the global submissions from participants were calculated, and variability was measured using the quartile coefficient of dispersion. Results: For the liver, which included lesions with high uptake, variabilities of up to 36% were found. The baseline analysis showed a variability of 29% in absorbed dose results for the liver from datasets where lesions included and excluded were grouped, indicating that variation in how lesions in normal liver were treated was a significant source of variability. For other organs and lesions, variability was within 7%, independently of software used except for the local deposition method. Conclusion: The choice of dosimetry method or software had a small contribution to the overall variability of dose estimates.

OBJECTIVE: Treatment planning through the diagnostic dimension of theranostics provides insights into predicting the absorbed dose of RPT, with the potential to individualize radiation doses for enhancing treatment efficacy. However, existing studies focusing on dose prediction from diagnostic data often rely on organ-level estimations, overlooking intra-organ variations. This study aims to characterize the intra-organ theranostic heterogeneity and utilize artificial intelligence techniques to localize them, i.e. to predict voxel-wise absorbed dose map based on pre-therapy PET.
METHODS: 23 patients with metastatic castration-resistant prostate cancer treated with [177Lu]Lu-PSMA I&T RPT were retrospectively included. 48 treatment cycles with pre-treatment PET imaging and at least 3 post-therapeutic SPECT/CT imaging were selected. The distribution of PET tracer and RPT dose was compared for kidney, liver and spleen, characterizing intra-organ heterogeneity differences. Pharmacokinetic simulations were performed to enhance the understanding of the correlation. Two strategies were explored for pre-therapy voxel-wise dosimetry prediction: (1) organ-dose guided direct projection; (2) deep learning (DL)-based distribution prediction. Physical metrics, dose volume histogram (DVH) analysis, and identity plots were applied to investigate the predicted absorbed dose map.
RESULTS: Inconsistent intra-organ patterns emerged between PET imaging and dose map, with moderate correlations existing in the kidney (r = 0.77), liver (r = 0.5), and spleen (r = 0.58) (P < 0.025). Simulation results indicated the intra-organ pharmacokinetic heterogeneity might explain this inconsistency. The DL-based method achieved a lower average voxel-wise normalized root mean squared error of 0.79 ± 0.27%, regarding to ground-truth dose map, outperforming the organ-dose guided projection (1.11 ± 0.57%) (P < 0.05). DVH analysis demonstrated good prediction accuracy (R2 = 0.92 for kidney). The DL model improved the mean slope of fitting lines in identity plots (199% for liver), when compared to the theoretical optimal results of the organ-dose approach.
CONCLUSIONS: Our results demonstrated the intra-organ heterogeneity of pharmacokinetics may complicate pre-therapy dosimetry prediction. DL has the potential to bridge this gap for pre-therapy prediction of voxel-wise heterogeneous dose map.

UNASSIGNED: Computational models yield valuable insights into biological interactions not fully elucidated by experimental approaches. This study investigates an innovative spatiotemporal model for simulating the controlled release and dispersion of radiopharmaceutical therapy (RPT) using 177Lu-PSMA, a prostate-specific membrane antigen (PSMA) targeted radiopharmaceutical, within solid tumors via a dual-release implantable delivery system. Local delivery of anticancer agents presents a strategic approach to mitigate adverse effects while optimizing therapeutic outcomes.
UNASSIGNED: This study evaluates various factors impacting RPT efficacy, including hypoxia region extension, binding affinity, and initial drug dosage, employing a novel 3-dimensional computational model. Analysis gauges the influence of these factors on radiopharmaceutical agent concentration within the tumor microenvironment. Furthermore, spatial and temporal radiopharmaceutical distribution within both the tumor and surrounding tissue is explored.
UNASSIGNED: Analysis indicates a significantly higher total concentration area under the curve within the tumor region compared to surrounding normal tissue. Moreover, drug distribution exhibits notably superior efficacy compared to the radiation source. Additionally, low microvascular density in extended hypoxia regions enhances drug availability, facilitating improved binding to PSMA receptors and enhancing therapeutic effectiveness. Reductions in the dissociation constant (KD) lead to heightened binding affinity and increased internalized drug concentration. Evaluation of initial radioactivities (7.1×107, 7.1×108, and 7.1×109 [Bq]) indicates that an activity of 7.1×108 [Bq] offers a favorable balance between tumor cell elimination and minimal impact on normal tissues.
UNASSIGNED: These findings underscore the potential of localized radiopharmaceutical delivery strategies and emphasize the crucial role of released drugs relative to the radiation source (implant) in effective tumor treatment. Decreasing the proximity of the drug to the microvascular network and enhancing its distribution within the tumor promote a more effective therapeutic outcome. The study furnishes valuable insights for future experimental investigations and clinical trials, aiming to refine medication protocols and minimize reliance on in vivo testing.

Hematologic toxicity, although often transient, is the most common limiting adverse effect during somatostatin peptide receptor radionuclide therapy. This study investigated the association between Monte Carlo-derived absorbed dose to the red marrow (RM) and hematologic toxicity in patients being treated for their neuroendocrine tumors. Methods: Twenty patients each receiving 4 treatment cycles of [177Lu]Lu-DOTATATE were included. Multiple-time-point 177Lu SPECT/CT imaging-based RM dosimetry was performed using an artificial intelligence-driven workflow to segment vertebral spongiosa within the field of view (FOV). This workflow was coupled with an in-house macroscale/microscale Monte Carlo code that incorporates a spongiosa microstructure model. Absorbed dose estimates to RM in lumbar and thoracic vertebrae within the FOV, considered as representations of the whole-body RM absorbed dose, were correlated with hematologic toxicity markers at about 8 wk after each cycle and at 3- and 6-mo follow-up after completion of all cycles. Results: The median of absorbed dose to RM in lumbar and thoracic vertebrae within the FOV (D median,vertebrae) ranged from 0.019 to 0.11 Gy/GBq. The median of cumulative absorbed dose across all 4 cycles was 1.3 Gy (range, 0.6-2.5 Gy). Hematologic toxicity was generally mild, with no grade 2 or higher toxicity for platelets, neutrophils, or hemoglobin. However, there was a decline in blood counts over time, with a fractional value relative to baseline at 6 mo of 74%, 97%, 57%, and 97%, for platelets, neutrophils, lymphocytes, and hemoglobin, respectively. Statistically significant correlations were found between a subset of hematologic toxicity markers and RM absorbed doses, both during treatment and at 3- and 6-mo follow-up. This included a correlation between the platelet count relative to baseline at 6-mo follow up: D median,vertebrae (r = -0.64, P = 0.015), D median,lumbar (r = -0.72, P = 0.0038), D median,thoracic (r = -0.58, P = 0.029), and D average,vertebrae (r = -0.66, P = 0.010), where D median,lumbar and D median,thoracic are median absorbed dose to the RM in the lumbar and thoracic vertebrae, respectively, within the FOV and D average,vertebrae is the mass-weighted average absorbed dose of all vertebrae. Conclusion: This study found a significant correlation between image-derived absorbed dose to the RM and hematologic toxicity, including a relative reduction of platelets at 6-mo follow up. These findings indicate that absorbed dose to the RM can potentially be used to understand and manage hematologic toxicity in peptide receptor radionuclide therapy.

BACKGROUND: With recent interest in patient-specific dosimetry for radiopharmaceutical therapy (RPT) and selective internal radiation therapy (SIRT), an increasing number of voxel-based algorithms are being evaluated. Monte Carlo (MC) radiation transport, generally considered to be the most accurate among different methods for voxel-level absorbed dose estimation, can be computationally inefficient for routine clinical use.
OBJECTIVE: This work demonstrates a recently implemented grid-based linear Boltzmann transport equation (LBTE) solver for fast and accurate voxel-based dosimetry in RPT and SIRT and benchmarks it against MC.
METHODS: A deterministic LBTE solver (Acuros MRT) was implemented within a commercial RPT dosimetry package (Velocity 4.1). The LBTE is directly discretized using an adaptive mesh refined grid and then the coupled photon-electron radiation transport is iteratively solved inside specified volumes to estimate radiation doses from both photons and charged particles in heterogeneous media. To evaluate the performance of the LBTE solver for RPT and SIRT applications, 177Lu SPECT/CT, 90Y PET/CT, and 131I SPECT/CT images of phantoms and patients were used. Multiple lesions (2-1052 mL) and normal organs were delineated for each study. Voxel dosimetry was performed with the LBTE solver, dose voxel kernel (DVK) convolution with density correction, and a validated in-house MC code using the same time-integrated activity and density maps as input to the different dose engines. The resulting dose maps, difference maps, and dose-volume-histogram (DVH) metrics were compared, to assess the voxel-level agreement. Evaluation of mean absorbed dose included comparison with structure-level estimates from OLINDA.
RESULTS: In the phantom inserts/compartments, the LBTE solver versus MC and DVK convolution demonstrated good agreement with mean absorbed dose and DVH metrics agreeing to within 5% except for the D90 and D70 metrics of a very low activity concentration insert of 90Y where the agreement was within 15%. In the patient studies (five patients imaged after 177Lu DOTATATE RPT, five after 90Y SIRT, and two after 131I radioimmunotherapy), in general, there was better agreement between the LBTE solver and MC than between LBTE solver and DVK convolution for mean absorbed dose and voxel-level evaluations. Across all patients for all three radionuclides, for soft tissue structures (kidney, liver, lesions), the mean absorbed dose estimates from the LBTE solver were in good agreement with those from MC (median difference < 1%, maximum 9%) and those from DVK (median difference < 5%, maximum 9%). The LBTE and OLINDA estimates for mean absorbed dose in kidneys and liver agreed to within 10%, but differences for lesions were larger with a maximum 14% for 177Lu, 23% for 90Y, and 26% for 131I. For bone regions, the agreement in mean absorbed doses between LBTE and both MC and DVK were similar (median < 11%, max 11%) while for lung the agreement between LBTE and MC (median < 1%, max 8%) was substantially better than between LBTE and DVK (median < 16%, max 33%). Voxel level estimates for soft tissue structures also showed good agreement between the LBTE solver and both MC and DVK with a median difference < 5% (maximum < 13%) for the DVH metrics with all three radionuclides. The largest difference in DVH metrics was for the D90 and D70 metric in lung and bone where the uptake was low. Here, the difference between LBTE and MC had a median value < 14% (maximum 23%) for bone and < 4% (maximum 37%) for lung, while the corresponding differences between LBTE and DVK were < 23% (maximum 31%) and < 67% (maximum 313%), respectively. For a typical patient with a matrix size of 166 × 166 × 129 (voxel size 3 × 3 × 3 mm3), voxel dosimetry using the LBTE solver was as fast as ∼2 min on a desktop computer.
CONCLUSIONS: Having established good agreement between the LBTE solver and MC for RPT and SIRT applications, the LBTE solver is a viable option for voxel dosimetry that can be faster than MC. Further analysis is being performed to encompass the broad range of radionuclides and conditions encountered clinically.

A comprehensive investigation of the Hg2+ coordination chemistry and 197m/gHg radiolabeling capabilities of cyclen-based commercial chelators, namely, DOTA and DOTAM (aka TCMC), along with their bifunctional counterparts, p-SCN-Bn-DOTA and p-SCN-Bn-TCMC, was conducted to assess the suitability of these frameworks as bifunctional chelators for the 197m/gHg2+ theranostic pair. Radiolabeling studies revealed that TCMC and DOTA exhibited low radiochemical yields (0%-6%), even when subjected to harsh conditions (80°C) and high ligand concentrations (10-4 M). In contrast, p-SCN-Bn-TCMC and p-SCN-Bn-DOTA demonstrated significantly higher 197m/gHg radiochemical yields (100% ± 0.0% and 70.9% ± 1.1%, respectively) under the same conditions. The [197 m/gHg]Hg-p-SCN-Bn-TCMC complex was kinetically inert when challenged against human serum and glutathione. To understand the differences in labeling between the commercial chelators and their bifunctional counterparts, non-radioactive natHg2+ complexes were assessed using NMR spectroscopy and DFT calculations. The NMR spectra of Hg-TCMC and Hg-p-SCN-Bn-TCMC suggested binding of the Hg2+ ion through the cyclen backbone framework. DFT studies indicated that binding of the Hg2+ ion within the backbone forms a thermodynamically stable product. However, competition can form between isothiocyanate binding and binding through the macrocycle, which was experimentally observed. The isothiocyanate bound coordination product was dominant at the radiochemical scale as, in comparison, the macrocycle bound product was seen at the NMR scale, agreeing with the DFT result. Furthermore, a bioconjugate of TCMC (TCMC-PSMA) targeting prostate-specific membrane antigen was synthesized and radiolabeled, resulting in an apparent molar activity of 0.089 MBq/nmol. However, the complex demonstrated significant degradation over 24 h when exposed to human serum and glutathione. Subsequently, cell binding assays were conducted, revealing a Ki value ranging from 19.0 to 19.6 nM. This research provides crucial insight into the effectiveness of current commercial chelators in the context of 197m/gHg2+ radiolabeling. It underscores the necessity for the development of specific and customized chelators to these unique \"soft\" radiometals to advance 197m/gHg2+ radiopharmaceuticals.

BACKGROUND: In radiopharmaceutical therapies (RPT) involving beta emitters, absorbed dose (Dabs) calculations often employ the use of dose voxel kernels (DVK). Such methods are faster and easier to implement than Monte Carlo (MC) simulations. Using DVK methods implies a non-stochastic distribution of particles. This is a valid assumption for betas where thousands to tens of thousands of particles traversing the cell nucleus are required to achieve cell kill. However, alpha particles have linear energy transfers (LET) that are ∼500 times higher than LETs of betas. This results in a significant probability of killing a cell from even a single traversal through its nucleus. Consequently, the activity used for therapy involving alphas is very low, and the use of DVKs for estimating Dabs will generate results that may be erroneous.
OBJECTIVE: This work aims at illustrating how use of DVKs affect the resulting Dabs in small tumors when irradiated with clinically relevant amounts of beta- and alpha-emitters. The results are compared with those from using a Monte Carlo method where the energy deposition from individual tracks is simulated.
METHODS: To illustrate the issues associated with DVK for alpha radiopharmaceutical therapies at the microscale, a tumor cluster model was used to compare beta (177Lu) and alphas (211At, 225Ac, and 227Th) irradiations. We used 103 beta particles and 20 alpha particles per cell, which is within the range of the required number of particle traversals through its nucleus to sterilize a cell. Results from using both methods were presented with Dabs histograms, dose volume histograms, and Dabs error maps.
RESULTS: For beta-emitter (177Lu) irradiating the modeled tumor cluster, resulting Dabs was similar for both DVK and MC methods. For all alpha emitters, the use of DVK led to an overestimation of Dabs when compared to results generated using a MC approach.
CONCLUSIONS: Our results demonstrate that the use of DVK methods for alpha emitters can lead to an overestimation in the calculated Dabs. The use of DVKs for therapies involving alpha emitters may therefore not be appropriate when only referring to the mean Dabs metric.

OBJECTIVE: 177Lutetium [Lu] Ludotadipep is a novel prostate-specific membrane antigen targeting therapeutic agent with an albumin motif added to increase uptake in the tumors. We assessed the biodistribution and dosimetry of [177Lu]Ludotadipep in patients with metastatic castration-resistant prostate cancer (mCRPC).
METHODS: Data from 25 patients (median age, 73 years; range, 60-90) with mCRPC from a phase I study with activity escalation design of single administration of [177Lu]Ludotadipep (1.85, 2.78, 3.70, 4.63, and 5.55 GBq) were assessed. Activity in the salivary glands, lungs, liver, kidneys, and spleen was estimated from whole-body scan and abdominal SPECT/CT images acquired at 2, 24, 48, 72, and 168 h after administration of [177Lu]Ludotadipep. Red marrow activity was calculated from blood samples obtained at 3, 10, 30, 60, and 180 min, and at 24, 48, and 72 h after administration. Organ- and tumor-based absorbed dose calculations were performed using IDAC-Dose 2.1.
RESULTS: Absorbed dose coefficient (mean ± standard deviation) of normal organs was 1.17 ± 0.81 Gy/GBq for salivary glands, 0.05 ± 0.02 Gy/GBq for lungs, 0.14 ± 0.06 Gy/GBq for liver, 0.77 ± 0.28 Gy/GBq for kidneys, 0.12 ± 0.06 Gy/GBq for spleen, and 0.07 ± 0.02 Gy/GBq for red marrow. The absorbed dose coefficient of the tumors was 10.43 ± 7.77 Gy/GBq.
CONCLUSIONS: [177Lu]Ludotadipep is expected to be safe at the dose of 3.7 GBq times 6 cycles planned for a phase II clinical trial with kidneys and bone marrow being the critical organs, and shows a high tumor absorbed dose.