ISIT-QA: In Silico Imaging Trial to Evaluate a Low-Count Quantitative SPECT Method Across Multiple Scanner-Collimator Configurations for 223Ra-Based Radiopharmaceutical Therapies.

Personalized dose-based treatment planning requires accurate and reproducible noninvasive measurements to ensure safety and effectiveness. Dose estimation using SPECT is possible but challenging for alpha (α)-particle-emitting radiopharmaceutical therapy (α-RPT) because of complex γ-emission spectra, extremely low counts, and various image-degrading artifacts across a plethora of scanner-collimator configurations. Through the incorporation of physics-based considerations and skipping of the potentially lossy voxel-based reconstruction step, a recently developed projection-domain low-count quantitative SPECT (LC-QSPECT) method has the potential to provide reproducible, accurate, and precise activity concentration and dose measures across multiple scanners, as is typically the case in multicenter settings. To assess this potential, we conducted an in silico imaging trial to evaluate the LC-QSPECT method for a 223Ra-based α-RPT, with the trial recapitulating patient and imaging system variabilities. Methods: A virtual imaging trial titled In Silico Imaging Trial for Quantitation Accuracy (ISIT-QA) was designed with the objectives of evaluating the performance of the LC-QSPECT method across multiple scanner-collimator configurations and comparing performance with a conventional reconstruction-based quantification method. In this trial, we simulated 280 realistic virtual patients with bone-metastatic castration-resistant prostate cancer treated with 223Ra-based α-RPT. The trial was conducted with 9 simulated SPECT scanner-collimator configurations. The primary objective of this trial was to evaluate the reproducibility of dose estimates across multiple scanner-collimator configurations using LC-QSPECT by calculating the intraclass correlation coefficient. Additionally, we compared the reproducibility and evaluated the accuracy of both considered quantification methods across multiple scanner-collimator configurations. Finally, the repeatability of the methods was evaluated in a test-retest study. Results: In this trial, data from 268 223RaCl2 treated virtual prostate cancer patients, with a total of 2,903 lesions, were used to evaluate LC-QSPECT. LC-QSPECT provided dose estimates with good reproducibility across the 9 scanner-collimator configurations (intraclass correlation coefficient > 0.75) and high accuracy (ensemble average values of recovery coefficients ranged from 1.00 to 1.02). Compared with conventional reconstruction-based quantification, LC-QSPECT yielded significantly improved reproducibility across scanner-collimator configurations, accuracy, and test-retest repeatability ([Formula: see text] Conclusion: LC-QSPECT provides reproducible, accurate, and repeatable dose estimations in 223Ra-based α-RPT as evaluated in ISIT-QA. These findings provide a strong impetus for multicenter clinical evaluations of LC-QSPECT in dose quantification for α-RPTs.

WIN-PDQ: A Wiener-estimator-based projection-domain quantitative SPECT method that accounts for intra-regional uptake heterogeneity.

SPECT can enable the quantification of activity uptake in lesions and at-risk organs in {\alpha}-particle-emitting radiopharmaceutical therapies ({\alpha}-RPTs). But this quantification is challenged by the low photon counts, complicated isotope physics, and the image-degrading effects in {\alpha}-RPT SPECT. Thus, strategies to optimize the SPECT system and protocol designs for the task of regional uptake quantification are needed. Objectively performing this task-based optimization requires a reliable (accurate and precise) regional uptake quantification method. Conventional reconstruction-based quantification (RBQ) methods have been observed to be erroneous for {\alpha}-RPT SPECT. Projection-domain quantification methods, which estimate regional uptake directly from SPECT projections, have demonstrated potential in providing reliable regional uptake estimates, but these methods assume constant uptake within the regions, an assumption that may not hold. To address these challenges, we propose WIN-PDQ, a Wiener-estimator-based projection-domain quantitative SPECT method. The method accounts for the heterogeneity within the regions of interest while estimating mean uptake. An early-stage evaluation of the method was conducted using 3D Monte Carlo-simulated SPECT of anthropomorphic phantoms with radium-223 uptake and lumpy-model-based intra-regional uptake heterogeneity. In this evaluation with phantoms of varying mean regional uptake and intra-regional uptake heterogeneity, the WIN-PDQ method yielded ensemble unbiased estimates and significantly outperformed both reconstruction-based and previously proposed projection-domain quantification methods. In conclusion, based on these preliminary findings, the proposed method is showing potential for estimating mean regional uptake in {\alpha}-RPTs and towards enabling the objective task-based optimization of SPECT system and protocol designs.

High-Dimensional Analyses Reveal IL15 Enhances Activation of Sipuleucel-T Lymphocyte Subsets and Reverses Immunoresistance.

Sipuleucel-T (sip-T) is the only FDA-approved autologous cellular immunotherapy for metastatic castration-resistant prostate cancer (mCRPC). To elucidate parameters of the response profile to this therapy, we report high-dimensional analyses of sip-T using cytometry by time of flight (CyTOF) and show a lymphoid predominance, with CD3+ T cells constituting the highest proportion (median ∼60%) of sip-T, followed by B cells, and natural killer (NK) and NKT cells. We hypothesized that treatment of sip-T with homeostatic cytokines known to activate/expand effector lymphocytes could augment efficacy against prostate tumors. Of the cytokines tested, IL15 was the most effective at enhancing activation and proliferation of effector lymphocytes, as well as augmenting tumor cytotoxicity in vitro. Co-culture of sip-T with IL15 and control or prostate-relevant antigens showed substantial activation and expansion of CD8+ T cells and NKT cells in an antigen-specific manner. Adoptive transfer of IL15-treated sip-T into NSG mice resulted in more potent prostate tumor growth inhibition compared with control sip-T. Evaluation of tumor-infiltrating lymphocytes revealed a 2- to 14-fold higher influx of sip-T and a significant increase in IFNγ producing CD8+ T cells and NKT cells within the tumor microenvironment in the IL15 group. In conclusion, we put forward evidence that IL15 treatment can enhance the functional antitumor immunity of sip-T, providing rationale for combining IL15 or IL15 agonists with sip-T to treat patients with mCRPC.

Joint regional uptake quantification of Thorium-227 and Radium-223 using a multiple-energy-window projection-domain quantitative SPECT method.

Thorium-227-based alpha-particle radiopharmaceutical therapies ({\alpha}-RPTs) are being investigated in several clinical and pre-clinical studies. After administration, Thorium-227 decays to Radium-223, another alpha-particle-emitting isotope, which redistributes within the patient. Reliable dose quantification of both Thorium-227 and Radium-223 is clinically important, and SPECT may perform this quantification as these isotopes also emit X- and gamma-ray photons. However, reliable quantification is challenged by the orders-of-magnitude lower activity compared to conventional SPECT, resulting in a very low number of detected counts, the presence of multiple photopeaks, substantial overlap in the emission spectra of these isotopes, and the image-degrading effects in SPECT. To address these issues, we propose a multiple-energy-window projection-domain quantification (MEW-PDQ) method that jointly estimates the regional activity uptake of both Thorium-227 and Radium-223 directly using the SPECT projection from multiple energy windows. We evaluated the method with realistic simulation studies using anthropomorphic digital phantoms, in the context of imaging patients with bone metastases of prostate cancer and treated with Thorium-227-based {\alpha}-RPTs. The proposed method yielded reliable (accurate and precise) regional uptake estimates of both isotopes and outperformed state-of-the-art methods across different lesion sizes and contrasts, in a virtual imaging trial, as well as with moderate levels of intra-regional heterogeneous uptake and with moderate inaccuracies in the definitions of the support of various regions. Additionally, we demonstrated the effectiveness of using multiple energy windows and the variance of the estimated uptake using the proposed method approached the Cram\'er-Rao-lower-bound-defined theoretical limit.

Beyond Average: α-Particle Distribution and Dose Heterogeneity in Bone Metastatic Prostate Cancer.

α-particle emitters are emerging as a potent modality for disseminated cancer therapy because of their high linear energy transfer and localized absorbed dose profile. Despite great interest and pharmaceutical development, there is scant information on the distribution of these agents at the scale of the α-particle pathlength. We sought to determine the distribution of clinically approved [223Ra]RaCl2 in bone metastatic castration-resistant prostate cancer at this resolution, for the first time to our knowledge, to inform activity distribution and dose at the near-cell scale. Methods: Biopsy specimens and blood were collected from 7 patients 24 h after administration. 223Ra activity in each sample was recorded, and the microstructure of biopsy specimens was analyzed by micro-CT. Quantitative autoradiography and histopathology were segmented and registered with an automated procedure. Activity distributions by tissue compartment and dosimetry calculations based on the MIRD formalism were performed. Results: We revealed the activity distribution differences across and within patient samples at the macro- and microscopic scales. Microdistribution analysis confirmed localized high-activity regions in a background of low-activity tissue. We evaluated heterogeneous α-particle emission distribution concentrated at bone-tissue interfaces and calculated spatially nonuniform absorbed-dose profiles. Conclusion: Primary patient data of radiopharmaceutical therapy distribution at the small scale revealed that 223Ra uptake is nonuniform. Dose estimates present both opportunities and challenges to enhance patient outcomes and are a first step toward personalized treatment approaches and improved understanding of α-particle radiopharmaceutical therapies.

Molecular Imaging of ACE2 Expression in Infectious Disease and Cancer.

Angiotensin-converting enzyme 2 (ACE2) is a cell-surface receptor that plays a critical role in the pathogenesis of SARS-CoV-2 infection. Through the use of ligands engineered for the receptor, ACE2 imaging has emerged as a valuable tool for preclinical and clinical research. These can be used to visualize the expression and distribution of ACE2 in tissues and cells. A variety of techniques including optical, magnetic resonance, and nuclear medicine contrast agents have been developed and employed in the preclinical setting. Positron-emitting radiotracers for highly sensitive and quantitative tomography have also been translated in the context of SARS-CoV-2-infected and control patients. Together this information can be used to better understand the mechanisms of SARS-CoV-2 infection, the potential roles of ACE2 in homeostasis and disease, and to identify potential therapeutic modulators in infectious disease and cancer. This review summarizes the tools and techniques to detect and delineate ACE2 in this rapidly expanding field.

A list-mode multi-energy window low-count SPECT reconstruction method for isotopes with multiple emission peaks.

SPECT provides a mechanism to perform absorbed-dose quantification tasks for $\alpha$-particle radiopharmaceutical therapies ($\alpha$-RPTs). However, quantitative SPECT for $\alpha$-RPT is challenging due to the low number of detected counts, the complex emission spectrum, and other image-degrading artifacts. Towards addressing these challenges, we propose a low-count quantitative SPECT reconstruction method for isotopes with multiple emission peaks. Given the low-count setting, it is important that the reconstruction method extract the maximal possible information from each detected photon. Processing data over multiple energy windows and in list-mode (LM) format provide mechanisms to achieve that objective. Towards this goal, we propose a list-mode multi-energy window (LM-MEW) OSEM-based SPECT reconstruction method that uses data from multiple energy windows in LM format, and includes the energy attribute of each detected photon. For computational efficiency, we developed a multi-GPU-based implementation of this method. The method was evaluated using 2-D SPECT simulation studies in a single-scatter setting conducted in the context of imaging [$^{223}$Ra]RaCl${_2}$. The proposed method yielded improved performance on the task of estimating activity uptake within known regions of interest in comparison to approaches that use a single energy window or use binned data. The improved performance was observed in terms of both accuracy and precision and for different sizes of the region of interest. Results of our studies show that the use of multiple energy windows and processing data in LM format with the proposed LM-MEW method led to improved quantification performance in low-count SPECT of isotopes with multiple emission peaks. These results motivate further development and validation of the LM-MEW method for such imaging applications, including for $\alpha$-RPT SPECT.

A list-mode multi-energy window low-count SPECT reconstruction method for isotopes with multiple emission peaks.

BACKGROUND: Single-photon emission computed tomography (SPECT) provides a mechanism to perform absorbed-dose quantification tasks for [Formula: see text]-particle radiopharmaceutical therapies ([Formula: see text]-RPTs). However, quantitative SPECT for [Formula: see text]-RPT is challenging due to the low number of detected counts, the complex emission spectrum, and other image-degrading artifacts. Towards addressing these challenges, we propose a low-count quantitative SPECT reconstruction method for isotopes with multiple emission peaks. METHODS: Given the low-count setting, it is important that the reconstruction method extracts the maximal possible information from each detected photon. Processing data over multiple energy windows and in list-mode (LM) format provide mechanisms to achieve that objective. Towards this goal, we propose a list-mode multi energy window (LM-MEW) ordered-subsets expectation-maximization-based SPECT reconstruction method that uses data from multiple energy windows in LM format and include the energy attribute of each detected photon. For computational efficiency, we developed a multi-GPU-based implementation of this method. The method was evaluated using 2-D SPECT simulation studies in a single-scatter setting conducted in the context of imaging [[Formula: see text]Ra]RaCl[Formula: see text], an FDA-approved RPT for metastatic prostate cancer. RESULTS: The proposed method yielded improved performance on the task of estimating activity uptake within known regions of interest in comparison to approaches that use a single energy window or use binned data. The improved performance was observed in terms of both accuracy and precision and for different sizes of the region of interest. CONCLUSIONS: Results of our studies show that the use of multiple energy windows and processing data in LM format with the proposed LM-MEW method led to improved quantification performance in low-count SPECT of isotopes with multiple emission peaks. These results motivate further development and validation of the LM-MEW method for such imaging applications, including for [Formula: see text]-RPT SPECT.

A Projection-Domain Low-Count Quantitative SPECT Method for α-Particle-Emitting Radiopharmaceutical Therapy.

Single-photon emission-computed tomography (SPECT) provides a mechanism to estimate regional isotope uptake in lesions and at-risk organs after administration of α-particle-emitting radiopharmaceutical therapies (α-RPTs). However, this estimation task is challenging due to the complex emission spectra, the very low number of detected counts (~20 times lower than in conventional SPECT), the impact of stray-radiation-related noise at these low counts, and the multiple image-degrading processes in SPECT. The conventional reconstruction-based quantification methods are observed to be erroneous for α-RPT SPECT. To address these challenges, we developed a low-count quantitative SPECT (LC-QSPECT) method that directly estimates the regional activity uptake from the projection data (obviating the reconstruction step), compensates for stray-radiation-related noise, and accounts for the radioisotope and SPECT physics, including the isotope spectra, scatter, attenuation, and collimator-detector response, using a Monte Carlo-based approach. The method was validated in the context of 3-D SPECT with 223Ra, a commonly used radionuclide for α-RPT. Validation was performed using both realistic simulation studies, including a virtual clinical trial, and synthetic and 3-D-printed anthropomorphic physical-phantom studies. Across all studies, the LC-QSPECT method yielded reliable regional-uptake estimates and outperformed the conventional ordered subset expectation-maximization (OSEM)-based reconstruction and geometric transfer matrix (GTM)-based post-reconstruction partial-volume compensation methods. Furthermore, the method yielded reliable uptake across different lesion sizes, contrasts, and different levels of intralesion heterogeneity. Additionally, the variance of the estimated uptake approached the Cramér-Rao bound-defined theoretical limit. In conclusion, the proposed LC-QSPECT method demonstrated the ability to perform reliable quantification for α-RPT SPECT.

Evaluation of Candidate Theranostics for 227Th/89Zr Paired Radioimmunotherapy of Lymphoma.

227Th is a promising radioisotope for targeted α-particle therapy. It produces 5 α-particles through its decay, with the clinically approved 223Ra as its first daughter. There is an ample supply of 227Th, allowing for clinical use; however, the chemical challenges of chelating this large tetravalent f-block cation are considerable. Using the CD20-targeting antibody ofatumumab, we evaluated chelation of 227Th4+ for α-particle-emitting and radiotheranostic applications. Methods: We compared 4 bifunctional chelators for thorium radiopharmaceutical preparation: S-2-(4-Isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane tetraacetic acid (p-SCN-Bn-DOTA), 2-(4-isothicyanatobenzyl)-1,2,7,10,13-hexaazacyclooctadecane-1,4,7,10,13,16-hexaacetic acid (p-SCN-Bn-HEHA), p-isothiacyanatophenyl-1-hydroxy-2-oxopiperidine-desferrioxamine (DFOcyclo*-p-Phe-NCS), and macrocyclic 1,2-HOPO N-hydroxysuccinimide (L804-NHS). Immunoconstructs were evaluated for yield, purity, and stability in vitro and in vivo. Tumor targeting of the lead 227Th-labeled compound in vivo was performed in CD20-expressing models and compared with a companion 89Zr-labeled PET agent. Results: 227Th-labeled ofatumumab-chelator constructs were synthesized to a radiochemical purity of more than 95%, excepting HEHA. 227Th-HEHA-ofatumumab showed moderate in vitro stability. 227Th-DFOcyclo*-ofatumumab presented excellent 227Th labeling efficiency; however, high liver and spleen uptake was revealed in vivo, indicative of aggregation. 227Th-DOTA-ofatumumab labeled poorly, yielding no more than 5%, with low specific activity (0.08 GBq/g) and modest long-term in vitro stability (<80%). 227Th-L804-ofatumumab coordinated 227Th rapidly and efficiently at high yields, purity, and specific activity (8 GBq/g) and demonstrated extended stability. In vivo tumor targeting confirmed the utility of this chelator, and the diagnostic analog, 89Zr-L804-ofatumumab, showed organ distribution matching that of 227Th to delineate SU-DHL-6 tumors. Conclusion: Commercially available and novel chelators for 227Th showed a range of performances. The L804 chelator can be used with potent radiotheranostic capabilities for 89Zr/227Th quantitative imaging and α-particle therapy.

Interpretable spatial cell learning enhances the characterization of patient tissue microenvironments with highly multiplexed imaging data.

Multiplexed imaging technologies enable highly resolved spatial characterization of cellular environments. However, exploiting these rich spatial cell datasets for biological insight is a considerable analytical challenge. In particular, effective approaches to define disease-specific microenvironments on the basis of clinical outcomes is a complex problem with immediate pathological value. Here we present InterSTELLAR, a geometric deep learning framework for multiplexed imaging data, to directly link tissue subtypes with corresponding cell communities that have clinical relevance. Using a publicly available breast cancer imaging mass cytometry dataset, InterSTELLAR allows simultaneous tissue type prediction and interested community detection, with improved performance over conventional methods. Downstream analyses demonstrate InterSTELLAR is able to capture specific pathological features from different clinical cancer subtypes. The method is able to reveal potential relationships between these regions and patient prognosis. InterSTELLAR represents an application of geometric deep learning with direct benefits for extracting enhanced microenvironment characterization for multiplexed imaging of patient samples.

Cure of Disseminated Human Lymphoma with [225Ac]Ac-Ofatumumab in a Preclinical Model.

Immunotherapies that target the CD20 protein expressed on most non-Hodgkin lymphoma cells have improved clinical outcomes, but relapse is common. We prepared 225Ac-labeled anti-CD20 ofatumumab and evaluated its in vitro characteristics and therapeutic efficacy in a murine model of disseminated human lymphoma. Methods: 225Ac was chelated by DOTA-ofatumumab, and radiochemical yield, purity, immunoreactivity, stability, and chelate number were determined. In vitro cell killing of CD20-positive, human B-cell lymphoma Raji-Luc cells was assayed. Biodistribution was determined as percentage injected activity per gram (%IA/g) in mice with subcutaneous Raji-cell tumors (n = 4). [225Ac]Ac-ofatumumab biodistribution in C57BL/6N mice was performed to estimate projected human dosimetry. Therapeutic efficacy was tested in mice with systemically disseminated Raji-Luc cells, tracking survival, bioluminescence, and animal weight for a targeted 200 d, with single-dose therapy initiated 8, 12, or 16 d after cell injection, comparing no treatment, ofatumumab, and low (3.7 kBq/mouse) and high (9.25 kBq/mouse) doses of [225Ac]Ac-IgG and [225Ac]Ac-ofatumumab (n = 8-10/cohort). Results: Radiochemical yield and purity were 32% ± 9% and more than 95%, respectively. Specific activity was more than 5 MBq/mg. Immunoreactivity was preserved, and more than 90% of the 225Ac remained chelated after 10 d in serum. Raji-Luc cell killing in vitro was significant, specific, and dose-dependent. In tumor-bearing mice, [225Ac]Ac-ofatumumab displayed low liver (7 %IA/g) and high tumor (28 %IA/g) uptake. Dosimetry estimates indicated that bone marrow is likely the dose-limiting organ. When therapy was initiated 8 d after cell injection, untreated mice and mice treated with cold ofatumumab or low- or high-dose [225Ac]Ac-IgG showed indistinguishable median survivals of 20-24 d, with extensive cancer-cell burden before death. Low- and high-dose [225Ac]Ac-ofatumumab profoundly (P < 0.05) extended median survival to 190 d and more than 200 d (median not determinable), with 5 and 9 of 10 mice, respectively, surviving at study termination with no detectable cancer cells. Surviving mice treated with high-dose [225Ac]Ac-ofatumumab showed reduced weight gain versus naïve mice. When therapy was initiated 12 d, but not 16 d, after cell injection, high-dose [225Ac]Ac-ofatumumab significantly extended median survival to 40 d but was not curative. Conclusion: In an aggressive disseminated tumor model, [225Ac]Ac-ofatumumab was effective at cancer-cell killing and curative when administered 8 d after cell injection. [225Ac]Ac-ofatumumab has substantial potential for clinical translation as a next-generation therapeutic for treatment of patients with non-Hodgkin lymphoma.

IMC-Denoise: a content aware denoising pipeline to enhance Imaging Mass Cytometry.

Imaging Mass Cytometry (IMC) is an emerging multiplexed imaging technology for analyzing complex microenvironments using more than 40 molecularly-specific channels. However, this modality has unique data processing requirements, particularly for patient tissue specimens where signal-to-noise ratios for markers can be low, despite optimization, and pixel intensity artifacts can deteriorate image quality and downstream analysis. Here we demonstrate an automated content-aware pipeline, IMC-Denoise, to restore IMC images deploying a differential intensity map-based restoration (DIMR) algorithm for removing hot pixels and a self-supervised deep learning algorithm for shot noise image filtering (DeepSNiF). IMC-Denoise outperforms existing methods for adaptive hot pixel and background noise removal, with significant image quality improvement in modeled data and datasets from multiple pathologies. This includes in technically challenging human bone marrow; we achieve noise level reduction of 87% for a 5.6-fold higher contrast-to-noise ratio, and more accurate background noise removal with approximately 2 × improved F1 score. Our approach enhances manual gating and automated phenotyping with cell-scale downstream analyses. Verified by manual annotations, spatial and density analysis for targeted cell groups reveal subtle but significant differences of cell populations in diseased bone marrow. We anticipate that IMC-Denoise will provide similar benefits across mass cytometric applications to more deeply characterize complex tissue microenvironments.

Engineering a modular 44Ti/44Sc generator: eluate evaluation in preclinical models and estimation of human radiation dosimetry.

BACKGROUND: 44Sc/47Sc is an attractive theranostic pair for targeted in vivo positron emission tomographic (PET) imaging and beta-particle treatment of cancer. The 44Ti/44Sc generator allows daily onsite production of this diagnostic isotope, which may provide an attractive alternative for PET facilities that lack in-house irradiation capabilities. Early animal and patient studies have demonstrated the utility of 44Sc. In our current study, we built and evaluated a novel clinical-scale 44Ti/44Sc generator, explored the pharmacokinetic profiles of 44ScCl3, [44Sc]-citrate and [44Sc]-NODAGA (1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid) in naïve mice, and estimated the radiation burden of 44ScCl3 in humans. METHODS: 44Ti/44Sc (101.2 MBq) in 6 M HCl solution was utilized to assemble a modular ZR resin containing generator. After assembly, 44Sc was eluted with 0.05 M HCl for further PET imaging and biodistribution studies in female Swiss Webster mice. Based on the biodistribution data, absorbed doses of 44/47ScCl3 in human adults were calculated for 18 organs and tissues using the IDAC-Dose software. RESULTS: 44Ti in 6 M HCl was loaded onto the organic resin generator with a yield of 99.97%. After loading and initial stabilization, 44ScCl3 was eluted with 0.05 M HCl in typical yields of 82.9 ± 5.3% (N = 16), which was normalized to the estimated generator capacity. Estimated generator capacity was computed based on elution time interval and the total amount of 44Ti loaded on the generator. Run in forward and reverse directions, the 44Sc/44Ti ratio from a primary column was significantly improved from 1038 ± 440 to 3557 ± 680 (Bq/Bq) when a secondary, replaceable, ZR resin cartridge was employed at the flow outlet. In vivo imaging and ex vivo distribution studies of the reversible modular generator for 44ScCl3, [44Sc]-citrate and [44Sc]-NODAGA show that free 44Sc remained in the circulation significantly longer than the chelated 44Sc. The dose estimation of 44ScCl3 reveals that the radiation burden is 0.146 mSv/MBq for a 70 kg adult male and 0.179 mSv/MBq for a 57 kg adult female. Liver, spleen and heart wall will receive the highest absorbed dose: 0.524, 0.502, and 0.303 mGy/MBq, respectively, for the adult male. CONCLUSIONS: A clinical-scale 44Ti/44Sc generator system with a modular design was developed to supply 44ScCl3 in 0.05 M HCl, which is suitable for further radiolabeling and in vivo use. Our data demonstrated that free 44ScCl3 remained in the circulation for extended periods, which resulted in approximately 10 times greater radiation burden than stably chelated 44Sc. Stable 44Sc/47Sc-complexation will be more favorable for in vivo use and for clinical utility.

Cure of Disseminated Human Lymphoma with [177Lu]Lu-Ofatumumab in a Preclinical Model.

Although immunotherapies that target CD20 on most non-Hodgkin lymphoma (NHL) cells have improved patient outcomes, current therapies are inadequate because many cases are, or become, refractory or undergo relapse. Here, we labelled the third-generation human anti-CD20 antibody ofatumumab with 177Lu, determined the in vitro characteristics of [177Lu]Lu-ofatumumab, estimated human dosimetry, and assayed tumor targeting and therapeutic efficacy in a murine model of disseminated NHL. Methods: CHX-A″-diethylenetriaminepentaacetic acid-[177Lu]Lu-ofatumumab was prepared. We evaluated radiochemical yield, purity, in vitro immunoreactivity, stability, (n = 7), affinity, and killing of CD20-expressing Raji cells (n = 3). Human dosimetry was estimated from biodistribution studies as percentage injected activity per gram using C57BL/6N mice. Tissue and organ biodistribution was determined in R2G2 immunodeficient mice with subcutaneous Raji-cell tumors. Therapy studies used R2G2 mice with disseminated human Raji-luc tumor cells (n = 10 mice/group). Four days after cell injection, the mice were left untreated or were treated with ofatumumab, 8.51 MBq of [177Lu]Lu-IgG, or 0.74 or 8.51 MBq of [177Lu]Lu-ofatumumab. Survival, weight, and bioluminescence were tracked. Results: Radiochemical yield was 93% ± 2%, radiochemical purity was 99% ± 1%, and specific activity was 401 ± 17 MBq/mg. Immunoreactivity was substantially preserved, and more than 75% of 177Lu remained chelated after 7 d in serum. [177Lu]Lu-ofatumumab specifically killed Raji-luc cells in vitro (P < 0.05). Dosimetry estimated that an effective dose for human administration is 0.36 mSv/MBq and that marrow may be the dose-limiting organ. Biodistribution in subcutaneous tumors 1, 3, and 7 d after [177Lu]Lu-ofatumumab injection was 11, 15, and 14 percentage injected activity per gram, respectively. In the therapy study, median survival of untreated mice was 19 d, not statistically different from mice treated with 8.51 MBq of [177Lu]Lu-IgG (25 d). Unlabeled ofatumumab increased survival to 46 d, similar to 0.74 MBq of [177Lu]Lu-ofatumumab (59 d), with both being superior to no treatment (P < 0.0003). Weight loss and increased tumor burden preceded death or killing of the animal for cause. In contrast, treatment with 8.51 MBq of [177Lu]Lu-ofatumumab dramatically increased median survival (>221 d), permitted weight gain, eliminated detectable tumors, and was curative in 9 of 10 mice. Conclusion: [177Lu]Lu-ofatumumab shows favorable in vitro characteristics, localizes to tumor, and demonstrates curative therapeutic efficacy in a disseminated lymphoma model, showing potential for clinical translation to treat NHL.

Radiochemical Quality Control Methods for Radium-223 and Thorium-227 Radiotherapies.

Background: The majority of radiopharmaceuticals for use in disease detection and targeted treatment undergo a single radioactive transition (decay) to reach a stable ground state. Complex emitters, which produce a series of daughter radionuclides, are emerging as novel radiopharmaceuticals. The need for validation of chemical and radiopurity with such agents using common quality control instrumentation is an area of active investigation. Here, we demonstrate novel methods to characterize 227Th and 223Ra. Materials and Methods: A radio-TLC scanner and a γ-counter, two common and widely accessible technologies, as well as a solid-state α-particle spectral imaging camera were evaluated for their ability to characterize and distinguish 227Th and 223Ra. We verified these results through purity evaluation of a novel 227Th-labeled protein construct. Results: The γ-counter and α-camera distinguished 227Th from 223Ra, enabling rapid and quantitative determination of radionuclidic purity. The radio-TLC showed limited ability to describe purity, although use under α-particle-specific settings enhanced resolution. All three methods were able to distinguish a pure from impure 227Th-labeled protein. Conclusions: The presented quality control evaluation for 227Th and 223Ra on three different instruments can be applied to both research and clinical settings as new alpha particle therapies are developed.

[18F]-Labeled PARP-1 PET imaging of PSMA targeted alpha particle radiotherapy response.

The growing interest and clinical translation of alpha particle (α) therapies brings with it new challenges to assess target cell engagement and to monitor therapeutic effect. Noninvasive imaging has great potential to guide α-treatment and to harness the potential of these agents in the complex environment of disseminated disease. Poly(ADP) ribose polymerase 1 (PARP-1) is among the most abundantly expressed DNA repair enzymes with key roles in multiple repair pathways-such as those induced by irradiation. Here, we used a third-generation PARP1-specific radiotracer, [18F]-PARPZ, to delineate castrate resistant prostate cancer xenografts. Following treatment with the clinically applied [225Ac]-PSMA-617, positron emission tomography was performed and correlative autoradiography and histology acquired. [18F]-PARPZ was able to distinguish treated from control (saline) xenografts by increased uptake. Kinetic analysis of tracer accumulation also suggests that the localization of the agent to sites of increased PARP-1 expression is a consequence of DNA damage response. Together, these data support expanded investigation of [18F]-PARPZ to facilitate clinical translation in the ⍺-therapy space.

Radiopharmaceutical Quality Control Considerations for Accelerator-Produced Actinium Therapies.

Background: Alpha-particle-emitting radiotherapies are of great interest for the treatment of disseminated cancer. Actinium-225 (225Ac) produces four α-particles through its decay and is among the most attractive radionuclides for use in targeted radiotherapy applications. However, supply issues for this isotope have limited availability and increased cost for research and translation. Efforts have focused on accelerator-based methods that produce 225Ac in addition to long-lived 227Ac. Objective: The authors investigated the impact of 225Ac/227Ac material in the radiolabeling and radiopharmaceutical quality control evaluation of a DOTA chelate-conjugated peptide under good manufacturing practices. The authors use an automated module under identical conditions with either generator or accelerator-produced actinium radiolabeling. Methods: The authors have performed characterization of the radiolabeled products, including thin-layer chromatography, high-pressure liquid chromatography, gamma counting, and high-energy resolution gamma spectroscopy. Results: Peptide was radiolabeled and assessed at >95% radiochemical purity with high yields for generator produced 225Ac. The radiolabeling results produced material with subtle but detectable differences when using 225Ac/227Ac. Gamma spectroscopy was able to identify peptide initially labeled with 227Th, and at 100 d for quantification of 225Ac-bearing peptide. Conclusion: Peptides produced using 225Ac/227Ac material may be suitable for translation, but raise new issues that include processing times, logistics, and contaminant detection.

Stable Chelation of the Uranyl Ion by Acyclic Hexadentate Ligands: Potential Applications for 230U Targeted α-Therapy.

Uranium-230 is an α-emitting radionuclide with favorable properties for use in targeted α-therapy (TAT), a type of nuclear medicine that harnesses α particles to eradicate cancer cells. To successfully implement this radionuclide for TAT, a bifunctional chelator that can stably bind uranium in vivo is required. To address this need, we investigated the acyclic ligands H2dedpa, H2CHXdedpa, H2hox, and H2CHXhox as uranium chelators. The stability constants of these ligands with UO22+ were measured via spectrophotometric titrations, revealing log ßML values that are greater than 18 and 26 for the "pa" and "hox" chelators, respectively, signifying that the resulting complexes are exceedingly stable. In addition, the UO22+ complexes were structurally characterized by NMR spectroscopy and X-ray crystallography. Crystallographic studies reveal that all six donor atoms of the four ligands span the equatorial plane of the UO22+ ion, giving rise to coordinatively saturated complexes that exclude solvent molecules. To further understand the enhanced thermodynamic stabilities of the "hox" chelators over the "pa" chelators, density functional theory (DFT) calculations were employed. The use of the quantum theory of atoms in molecules revealed that the extent of covalency between all four ligands and UO22+ was similar. Analysis of the DFT-computed ligand strain energy suggested that this factor was the major driving force for the higher thermodynamic stability of the "hox" ligands. To assess the suitability of these ligands for use with 230U TAT in vivo, their kinetic stabilities were probed by challenging the UO22+ complexes with the bone model hydroxyapatite (HAP) and human plasma. All four complexes were >95% stable in human plasma for 14 days, whereas in the presence of HAP, only the complexes of H2CHXdedpa and H2hox remained >80% intact over the same period. As a final validation of the suitability of these ligands for radiotherapy applications, the in vivo biodistribution of their UO22+ complexes was determined in mice in comparison to unchelated [UO2(NO3)2]. In contrast to [UO2(NO3)2], which displays significant bone uptake, all four ligand complexes do not accumulate in the skeletal system, indicating that they remain stable in vivo. Collectively, these studies suggest that the equatorial-spanning ligands H2dedpa, H2CHXdedpa, H2hox, and H2CHXhox are highly promising candidates for use in 230U TAT.

Blind Image Restoration Enhances Digital Autoradiographic Imaging of Radiopharmaceutical Tissue Distribution.

Digital autoradiography (DAR) is a powerful tool to quantitatively determine the distribution of a radiopharmaceutical within a tissue section and is widely used in drug discovery and development. However, the low image resolution and significant background noise can result in poor correlation, even errors, between radiotracer distribution, anatomic structure, and molecular expression profiles. Differing from conventional optical systems, the point-spread function in DAR is determined by properties of radioisotope decay, phosphor, and digitizer. Calibration of an experimental point-spread function a priori is difficult, prone to error, and impractical. We have developed a content-adaptive restoration algorithm to address these problems. Methods: We model the DAR imaging process using a mixed Poisson-gaussian model and blindly restore the image by a penalized maximum-likelihood expectation-maximization algorithm (PG-PEM). PG-PEM implements a patch-based estimation algorithm with density-based spatial clustering of applications with noise to estimate noise parameters and uses L2 and Hessian Frebonius norms as regularization functions to improve performance. Results: First, PG-PEM outperformed other restoration algorithms at the denoising task (P < 0.01). Next, we implemented PG-PEM on preclinical DAR images (18F-FDG, treated mouse tumor and heart; 18F-NaF, treated mouse femur) and clinical DAR images (bone biopsy sections from 223RaCl2-treated castration-resistant prostate cancer patients). DAR images restored by PG-PEM of all samples achieved a significantly higher effective resolution and contrast-to-noise ratio and a lower SD of background (P < 0.0001). Additionally, by comparing the registration results between the clinical DAR images and the segmented bone masks from the corresponding histologic images, we found that the radiopharmaceutical distribution was significantly improved (P < 0.0001). Conclusion: PG-PEM is able to increase resolution and contrast while robustly accounting for DAR noise and demonstrates the capacity to be widely implemented to improve preclinical and clinical DAR imaging of radiopharmaceutical distribution.