Preclinical Assessment of [68Ga]Ga-Cell Death Indicator (CDI): A Novel hsp90 Ligand for Positron Emission Tomography of Cell Death.

BACKGROUND: 4-(N-(S-glutathionylacetyl)amino) phenylarsonous acid (GSAO) when conjugated with a bifunctional chelator 2,2'-(7-(1-carboxy-4-((2,5-dioxopyrrolidin-1-yl)oxy)-4- oxobutyl)-1,4,7-triazonane-1,4-diyl)diacetic acid (NODAGA) (hereafter referred to as Cell Death Indicator [CDI]), enters dead and dying cells and binds to 90kDa heat shock proteins (hsp90). OBJECTIVE: This study assesses stability, biodistribution, imaging, and radiation dosimetry of [68Ga]- Ga-CDI for positron emission tomography (PET). METHODS: Preparation of [68Ga]Ga-CDI was performed as previously described. Product stability and stability in plasma were assessed using high-performance liquid chromatography. Biodistribution and imaging were conducted in ten healthy male Lewis rats at 1 and 2 h following intravenous [68Ga]Ga-CDI injection. Human radiation dosimetry was estimated by extrapolation for a standard reference man and calculated with OLINDA/EXM 1.1. RESULTS: Radiochemical purity of [68Ga]Ga-CDI averaged 93.8% in the product and 86.7% in plasma at 4 h post-synthesis. The highest concentration of [68Ga]Ga-CDI is observed in the kidneys; [68Ga]Ga-CDI is excreted in the urine, and mean retained activity was 32.4% and 21.4% at 1 and 2 h post-injection. Lower concentrations of [68Ga]Ga-CDI were present in the small bowel and liver. PET CT was concordant and additionally demonstrated focal growth plate uptake. The effective dose for [68Ga]Ga-CDI is 2.16E-02 mSv/MBq, and the urinary bladder wall received the highest dose (1.65E-02 mSv/Mbq). CONCLUSION: [68Ga] Ga-CDI is stable and has favourable biodistribution, imaging, and radiation dosimetry for imaging of dead and dying cells. Human studies are underway.

Biodistribution and imaging of an hsp90 ligand labelled with 111In and 67Ga for imaging of cell death.

BACKGROUND: 4-(N-(S-glutathionylacetyl)amino) phenylarsonous acid (GSAO) when conjugated at the γ-glutamyl residue with fluorophores and radio-isotopes is able to image dead and dying cells in vitro and in vivo by binding to intracellular 90-kDa heat shock proteins (hsp90) when cell membrane integrity is compromised. The ability to image cell death has potential clinical impact especially for early treatment response assessment in oncology. This work aims to assess the biodistribution and tumour uptake of diethylene triamine pentaacetic acid GSAO labelled with 111In ([111In]In-DTPA-GSAO) and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid GSAO labelled with 67Ga ([67Ga]Ga-DOTA-GSAO) in a murine subcutaneous tumour xenograft model and estimate dosimetry of [67Ga]Ga-DOTA-GSAO. RESULTS: There was good tumour uptake of both [111In]In-DTPA-GSAO and [67Ga]Ga-DOTA-GSAO (2.44 ± 0.26% injected activity per gramme of tissue (%IA/g) and 2.75 ± 0.34 %IA/g, respectively) in Balb c nu/nu mice bearing subcutaneous tumour xenografts of a human metastatic prostate cancer cell line (PC3M-luc-c6). Peak tumour uptake occurred at 2.7 h post injection. [111In]In-DTPA-GSAO and [67Ga]Ga-DOTA-GSAO demonstrated increased uptake in the liver (4.40 ± 0.86 %IA/g and 1.72 ± 0.27 %IA/g, respectively), kidneys (16.54 ± 3.86 %IA/g and 8.16 ± 1.33 %IA/g) and spleen (6.44 ± 1.24 %IA/g and 1.85 ± 0.44 %IA/g); however, uptake in these organs was significantly lower with [67Ga]Ga-DOTA-GSAO (p = 0.006, p = 0.017 and p = 0.003, respectively). Uptake of [67Ga]Ga-DOTA-GSAO into tumour was higher than all organs except the kidneys. There was negligible uptake in the other organs. Excretion of [67Ga]Ga-DOTA-GSAO was more rapid than [111In]In-DTPA-GSAO. Estimated effective dose of [67Ga]Ga-DOTA-GSAO for an adult male human was 1.54 × 10- 2 mSv/MBq. CONCLUSIONS: [67Ga]Ga-DOTA-GSAO demonstrates higher specific uptake in dead and dying cells within tumours and lower uptake in normal organs than [111In]In-DTPA-GSAO. [67Ga]Ga-DOTA-GSAO may be potentially useful for imaging cell death in vivo. Dosimetry estimates for [67Ga]Ga-DOTA-GSAO are acceptable for future human studies. This work also prepares for development of 68Ga GSAO radiopharmaceuticals.

Preparation of a Dithiol-Reactive Probe for PET Imaging of Cell Death.

Conjugates of 4-(N-(S-glutathionylacetyl)amino)phenylarsonous acid (GSAO) with optical or radionuclide probes are able to image cell death in vivo. GSAO conjugates are retained in the cytosol of dying and dead cells via the formation of covalent bonds between the As(III) ion and the thiol groups of proximal cysteine residues. Here we describe the method for preparing a NODAGA-GSAO conjugate and its radiolabeling with gallium-68 (68Ga-NODAGA-GSAO) for positron-emission tomography (PET) imaging of cell death.

Comparison of PSMA-HBED and PSMA-I&T as diagnostic agents in prostate carcinoma.

PURPOSE: Gallium(68)-labelled prostate-specific membrane antigen (PSMA) radiopharmaceuticals can be used to detect prostate cancer (PCa) cells due the their over expression of PSMA. The 68Ga HBED-PSMA (PSMA-HBED) ligand has been most widely used and can be considered the current gold standard agent. Further PSMA ligands based on the DOTAGA and DOTA conjugates have more recently been developed. These agents (PSMA-I&T and PSMA-617) have potential theranostic capabilities as they can be conjugated with therapeutic radioisotopes. In this study, we examine whether PSMA-I&T has comparative efficacy, such that it could replace PSMA-HBED as a diagnostic agent in prostate carcinoma. METHODS: 19 patients with PCa referred for 68Ga-PSMA imaging were imaged with PSMA-HBED and PSMA-I&T PET-CT imaging within a 2-week period. The two pharmaceuticals were synthesised using click chemistry. Imaging was performed using the same standardised methodology on a Siemens Biograph mCT. All sites of PSMA binding thought to represent PCa (probable or definite) were included in a lesion analysis that examined lesion concordance and lesional binding efficiency (SUVpeak) between the two radiopharmaceuticals. For each patient, SUVmean of the LV cavity blood pool, bone, muscle and liver were determined as image background measures. RESULTS: Across all patients, PSMA uptake was observed in 47 lesions (10 bone lesions, 19 nodal lesions, 18 high-grade intraprostatic binding). Lesions were concordant between the agents in all except for two small (<4 mm) nodal lesions which were not visualised with PSMA-I&T. SUVpeak assessment showed significantly greater overall lesion binding with HBED (paired t test, p = 0.0001). LV blood pool and bone marrow SUVmean were significantly higher for I&T than HBED (paired t test, blood pool p < 1 × 10-5, bone marrow p < 0.005). CONCLUSION: Intra-patient comparative imaging demonstrates higher lesional PSMA-HBED binding than PSMA-I&T and that the HBED agent is likely to have better lesion contrast. While there was concordance in 96% of lesions, 2 small nodal lesions were appreciated with PSMA-HBED imaging while considered normal with PSMA-I&T. These findings suggest that HBED-PSMA has a slightly higher diagnostic accuracy in comparison to PSMA-I&T.

Clinical benefits of two different dosing schedules of recombinant human erythropoietin in anemic patients with advanced head and neck cancer.

A total of 100 patients with stage III or IV head or neck cancer, a performance status of 0-1, and anemia with hemoglobin (Hb) < 10 g/dL at baseline who where to receive chemotherapy concomitantly or sequentially with radiotherapy were randomized to receive either epoetin beta 10,000 IU thrice weekly (TW) (n = 52) and oral iron starting 10-15 days before the start of treatment or epoetin beta 30,000 IU once weekly (OW) (n = 48) and oral iron before the start of treatment. The mean Hb in patients on the thrice weekly (11.96 g/dL) and once weekly (12.50 g/dL) dosing schedules increased significantly (p < 0.01) at the end of the treatment in comparison to respective baseline values of 9.38 g/dL and 9.41 g/dL; levels were 1.2-fold higher, which was significant (p < 0.01), for patients on the once weekly schedule. That said, there was significant improvement (p < 0.01) in mean linear analog scale assessment (LASA) scores for energy level (EL), ability to perform daily activities (AL), and overall quality of life (QOL) for patients on both dosing schedules but these improvements did not differ significantly between schedules (p > 0.05). The 2-year overall survival for patients on both dosing schedules did not differ significantly (p > 0.05). Epoetin beta therapy was found to be equally beneficial and well tolerated for patients on both thrice weekly and once weekly dosing schedules.

Is (99m)Tc-labelled pamidronate a better agent than (99m)Tc-medronate for bone imaging?

OBJECTIVE: To evaluate the potential of (99m)Tc-pamidronate ((99m)Tc-APD) against (99m)Tc-medronate ((99m)Tc-MDP) as a new bone-seeking agent using intact bone and fractured femur in a rat model. METHODS: (99m)Tc-APD was prepared by the stannous reduction method. Scintigraphic images were obtained at 2 h and 24 h after intravenous injection of (99m)Tc-APD or (99m)Tc-MDP in rats, then they were culled to estimate activities in various organs. Bone uptake (as percent injected dose/gram weight) was estimated in an intact femur and in 1 week post-fracture model. The urinary excretion dose (as percent injected dose) was also estimated. RESULTS: The bone uptake of (99m)Tc-APD was significantly higher (P<0.05) than (99m)Tc-MDP at 2 h and 24 h post-injection studies. (99m)Tc-APD uptake was further increased (P<0.05) in the fracture model than the intact femur. (99m)Tc-APD uptake in the soft tissues including liver and the kidneys was lower than (99m)Tc-MDP. Renal excretion was faster and the ratios of bone to blood and bone to soft tissues were higher with APD than MDP. APD dose was selected at 1% of MDP, to obviate therapeutic effect, as the former compound is 100 times more potent than MDP. CONCLUSIONS: Our results suggest that (99m)Tc-APD uptake by intact bone and fractured bone was significantly higher than (99m)Tc-MDP. The renal clearance of (99m)Tc-APD was faster and soft tissue uptake was lower than (99m)Tc-MDP. These results suggest that APD has the potential to become an excellent bone-imaging agent.

Evaluation of biodistribution by local versus systemic administration of 99mTc-labeled pamidronate.

BACKGROUND: There is an emerging interest in utilizing local and systemic administration of bisphosphonates in orthopedics. The primary objective of this study was to use (99m)Tc-pamidronate ((99m)Tc-PAM) as a tool and compare bone and tissue uptake by local versus systemic administration. METHODS: (99m)Tc-PAM was administered intravenously (i.v.), subcutaneously (s.c.) and by direct application (d.a.) on a surgically exposed and fractured femur (d.a.#f). The animals were imaged at 2 h and 24 h after administration and then killed. Organs were harvested, and their radioactivity was estimated. Specific uptake in the right femur was compared between groups, as was systemic exposure to (99m)Tc PAM. RESULTS: Bone uptake of (99m)Tc-PAM in the i.v. and s.c. groups was 2.2 +/- 0.15 and 0.65 +/- 0.07% ID/g, respectively, at the 2 h time point. Uptake by surgically exposed right femur (d.a) was 5.15 +/- 0.26% ID/g, 134% higher than the femoral uptake by the i.v. method (P < 0.05). In the presence of exposed bone when the femur was fractured (d.a.#f), the uptake was 7.89 +/- 0.46% ID/g, a further 50% increase (P < 0.05). The uptake of (99m)Tc-PAM increased after 24 h of application to 2.4 +/- 0.15, 1.53 +/- 0.09, 7.94 +/- 0.99, and 13.2 +/- 0.80% ID/g) for i.v., s.c., d.a., and d.a.#f methods, respectively. The increases in uptake for the d.a. methods were significantly higher than for the local methods at the 24-h time point (P < 0.05). Although renal uptake was comparable with the i.v. and s.c. methods (0.22 +/- 0.03 and 0.22 +/- 0.04% ID/g), it was significantly lower with the d.a. methods (0.05 +/- 0.07 and 0.16 +/- 0.07% ID/g) (P < 0.05). The corresponding urinary excretion was 55%, 45%, 36%, and 35% of the injected dose at 24 h. CONCLUSIONS: The results indicate that the bone uptake of (99m)Tc-PAM was significantly higher (P = 0.001) and the kidney uptake significantly lower (P = 0.004) with the d.a. methods than with the i.v. or s.c. method. The findings indicate the need for further study into the potential of local administration of bisphosphonates in the presence of orthopedic indications.