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An Automated Radiosynthesis of [68Ga]Ga-FAPI-46 for Routine Clinical Use
In This Article
Summary
This work describes the automated production of up to 1.7 GBq of [68Ga]Ga-FAPI-46 on the iPHASE MultiSyn synthesizer for PET imaging of fibroblast activation protein.
Abstract
[68Ga]Ga-FAPI-46 is a promising new tracer for the imaging of fibroblast activation protein (FAP) by positron emission tomography (PET). Labeled FAP inhibitors (FAPIs) have demonstrated uptake in various types of cancers, including breast, lung, prostate, pancreatic and colorectal cancer. FAPI-PET also possesses a practical advantage over FDG-PET as fasting and resting are not required. [68Ga]Ga-FAPI-46 exhibits enhanced pharmacokinetic properties, improved tumor retention, and higher contrast images than the earlier presented [68Ga]Ga-FAPI-02 and [68Ga]Ga-FAPI-04. Although a manual synthesis protocol for [68Ga]Ga-FAPI-46 was initially described, in recent years, automated methods using different commercial synthesizers have been reported.
In this work, we describe the development of the automated synthesis of [68Ga]Ga-FAPI-46 using the iPHASE MultiSyn synthesizer for clinical applications. Initially, optimization of the reaction time and comparison of the performance of four different solid phase extraction (SPE) cartridges for final product purification were investigated. Then, the development and validation of the production of 0.6-1.7 GBq of [68Ga]Ga-FAPI-46 were conducted using these optimized parameters. The product was synthesized in 89.8 ± 4.8% decay corrected yield (n = 6) over 25 min. The final product met all recommended quality control specifications and was stable up to 3 h post synthesis.
Introduction
Fibroblast activation protein (FAP) has become a prominent target for cancer imaging and therapy1,2. FAP is a specific marker of cancer-associated fibroblasts (CAFs), a stromal cell type constituting much of the microenvironment of solid cancers. CAFs play a key role in tumor growth, invasion, and metastasis3. They are found in most solid tumors, including breast, prostate, and pancreatic cancers1. By targeting FAP with small molecule inhibitors labeled with diagnostic or therapeutic radionuclides, selective non-invasive imaging and therapy of these cancers may b....
Protocol
CAUTION: This protocol involves the handling of radioactive materials. All personnel undertaking this work must be adequately trained in working with unsealed radioisotopes and have the approval of their institution's radiation safety officer. The automated synthesizer must be located in a dedicated shielded hot cell. Manual experiments must be performed in a shielded hot cell or behind radiation shielding. Preliminary experiments for the optimization of the reaction time and testing of various SPE cartridges are des.......
Representative Results
The radiolabeling efficiency assessed between 5 and 20 min of reaction at 95 °C is reported in Table 2. The postpurification SPE cartridges HLB 30 mg, Strata X 60 mg, and Sep-Pak C18 Plus Short 360 mg showed very similar recoveries (94.3 ± 0.5% decay-corrected [DC]) (Table 3). In our hands, the recovery off the HLB 225 mg was much lower (63.8 ± 3.5% DC).
Table 2: Summary of the radiochemical conversion of the crude reaction measured by T.......
Discussion
This work describes the reliable and high-yielding production of [68Ga]Ga-FAPI-46 on the MS synthesizer for clinical applications. The preliminary workup of this protocol tested, in the same series of experiments, different reaction times for the synthesis of [68Ga]Ga-FAPI-46 as well as four different SPE cartridges for the purification of the final product. In order to (i) reduce the radiation dose to the operator and (ii) re-create the conditions in which the routine production of [68Ga.......
Acknowledgements
The authors acknowledge and thank SOFIE Biosciences Inc. for supplying the FAPI-46 chemical precursor and [natGa]Ga-FAPI-46 standard, the Charlies Foundation for Research for financial support, Stan Poniger from iPHASE Pty Ltd, and the Radiopharmaceuticals Production team (RAPID) at the Medical Technology and Physics Department at Sir Charles Gairdner Hospital for their scientific and technical support. The authors also acknowledge the assistance of the WA National Imaging Facility Node, which is supported by infrastructure funding from the Western Australian State Government in partnership with the Australian Federal Government, through the National Collab....
Materials
| Name | Company | Catalog Number | Comments |
| 0.1 M Hydrochloric acid (HCl) ultra pure | ABX advanced biochemical compound (Radberg, Germany) | HCl-103-G | Used for generator(s) elution |
| Ammonium acetate | Sigma Aldrich Pty Ltd (NSW, Australia) | A1542-250G | Used to make iTLC mobile phase |
| C18 SepPak Plus short (360 mg) | Waters | WAT020515 | Post-purification silica SPE |
| Chromolith Performance RP-18 endcapped 100-4.6 monolithic | Merck Pty Ltd (Victoria (Australia) | 1021290001 | HPLC RP-18 endcapped column, used for HPLC quality control |
| Dose calibrator | Capintec | CRC-15PET | Used to calibrate and measure 68Ga activity |
| Dual scan-RAM | LabLogic Limited (VA, USA) | SR-1A | Radio-TLC scanner to analysise the iTLC paper |
| FAPI-46 precursor (GMP) | ABX advanced biochemical compound (Radberg, Germany) | 3601.0000.050 | Peptide precursor |
| Fill ease Sterile vacuum vial (10 mL) | HUAYI iosotopes | SVV-10C | Used for sterility and retention samples |
| Fill ease Sterile vacuum vial (25 mL) | HUAYI iosotopes | SVV-25A | Used for final product |
| Ga68 peptide radiolabelling with generator pre-purification | iPHASE Technologies (Melbourne, Australia) | MSR-120G-(RK-3296) | Reagent set |
| Ga68 radiolabeling with generator prepurification | iPHASE Technologies (Melbourne, Australia) | MSH-120 | Hardware Cassette + ancillaries set |
| Gas chromatography (GC) system | Agilent technologies (Vic, Australia) | G2630A | Used to measure residual solvent |
| GS Standard source (Ba133) | Global Medical Solutions Pty Ltd (Australia) | D-102-19 | Used to calibrate the Gamma Spectometer |
| GS Standard source (Co60) | Global Medical Solutions Pty Ltd (Australia) | 1559-84 | Used to calibrate the Gamma Spectometer |
| High performance liquid chromatography (HPLC) system | Shimadzu Scientific Instruments (NSW, Australia) | LC-20 | HPLC equipment |
| Hydrophobic air vent needle | Baldwin Medical (Victoria, australia) | 1088 | Used with final product vial |
| IGG100 | Eckert & Ziegler Isotope Products (Berlin, Germany) | IGG100-65M-NT | 68Ge/68Ga generator |
| 5 mL syringe (Injekt luer lock solo syringe) | B Braun (Melsungen, Germany) | 4606710V | Polypropylene (PP)/polyethylene (PE) syringes, free of latex, PVC, and silicone oil free syringe used for reagents |
| iTLC-SG paper | Agilent technologies (Vic, Australia) | SGI0001 | Used to for iTLC analysis |
| LabLogic software (LAURA) | LabLogic Limited (VA, USA) | LAURA software version 6.1 | Used to for radio-TLC analysis |
| L-Ascorbic acid Trace select | Fluka Sigma | 05878-100G | Used as a radical scavenger in the reaction mixture |
| Lichrosolv Acetonitrile (ACN) | Sigma Aldrich Pty Ltd (NSW, Australia) | 1.00030.2500 | Used to make HPLC organic mobile phase |
| Lichrosolv Water | Sigma Aldrich Pty Ltd (NSW, Australia) | 1.15333.2500 | Used to make HPLC aqueous mobile phase |
| Methanol (MeOH) | Sigma Aldrich Pty Ltd (NSW, Australia) | 1060182500 | Used to make iTLC mobile phase |
| Na+I- detector | LabLogic Limited (VA, USA) | 1"NaI / PMT | Radiodetector used for radio-HPLC |
| Oasis HLB (30 mg) | Waters (Milford, MA, USA) | WAT094225 | Postpurification copolymer SPE |
| Oasis HLB Plus short (225 mg) | Waters (Milford, MA, USA) | 186000132 | Postpurification copolymer SPE |
| pH strips | Thermo Fisher Scientific Australia Tty Ltd | 90424 | Used to measure product pH |
| PS detector | LabLogic Limited (VA, USA) | PS plastic/PMT | Radiodetector used for radio-TLC |
| Safe Lock tube (1.5 mL) | Eppendorf | 0030 120.086 | Used for quality control samples |
| (+)-Sodium L-ascorbate | Merck Pty Ltd (Victoria (Australia) | 11140-250G | Stabilizer of the final product |
| Sodium chloride (NaCl) solution (saline) | Pfizer | PS111 | 0.9%, for injection, USP grade |
| Sterican 100 Needles | B Braun (Melsungen, Germany) | 4667093 | Used for final product |
| Sterile syringe filter (0.22 µm) | Millipore Sigma (Burlington, MA, USA) | SLGSV255F | Millex-GV |
| Strata SCX (in Hardware cassette kit) | Phenomenex inside hardware kit from iPHASE Technologies (Melbourne, Australia) | MSH-120 | Prepurification silica SPE inside Hardware Cassette |
| Strata X (in Hardware cassette kit) | Phenomenex inside hardware kit from iPHASE Technologies (Melbourne, Australia) | MSH-120 | Postpurification silica SPE inside Hardware Cassette |
| Trace Select Water for trace analysis | Honeywell Riedel-de-Haen | 95305-2.5L | Used for reaction mixture and to precondition the prepurification SPE cartridge |
| Trifluoracetic acid (TFA) | Sigma Aldrich Pty Ltd (NSW, Australia) | 302031-10X1mL | Used to make HPLC aqueous mobile phase |
| Ultra Fine insulin syringe (0.5 mL) | BD | 326769 | Used for dispensing quality control samples |
| Vented filter Cathivex-GV 0.22 µm, low protein binding Durapore PVDF membrane | Merk Millipore (Cork, Ireland) | SLGV02505 | Used to filter the final product |
References
- Imlimthan, S., et al. New frontiers in cancer imaging and therapy based on radiolabeled fibroblast activation protein inhibitors: a rational review and current progress. Pharmaceuticals. 14 (10), 1023 (2021).
- Siveke, J. T.
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