Production of gallium-67 is undertaken by the proton bombardment of an isotopically enriched zinc-68 target at 21 MeV on medium energy (30 MeV) cyclotrons, via the 68Zn(p,2n)67Ga nuclear reaction. A number of other production routes are available (8), including natZn(p,xn)67Ga, 67Zn(p,n)67Ga and natZn(d,xn)67Ga (9), but these are not routinely utilised. Chemical purification of gallium-67 from the target material and radioisotopic impurities can be achieved by ion exchange chromatography or solvent extraction, both of which allow recycling of the enriched target (10).
Despite a decline in the number of sites worldwide producing gallium-67 commercially it remains readily available as both the chloride salt, [67Ga]GaCl3 in dilute hydrochloric acid and as the citrate complex, [67Ga][Ga(C6H5O5)2]3- (11) in sterile formulation for human administration. Radiolabelling directly with the formulated citrate complex can be problematic due to the low radioactive concentration (74 MBq/mL (2 mCi/mL) at calibration) and presence of excess citrate. However, the formulated citrate complex can be easily converted to the chloride salt in higher activity concentration using a simple silica based SPE method (12).
Gallium (III) is classified as a hard metal with a small ionic radius of 0.62 Å and in aqueous solution it exists solely as redox stable 3+ state. Ga(III) typically adopts octahedral geometries, preferring hard donor atoms such as amine nitrogens and carboxylate oxygens. It is chemically very similar to iron (III), hence its tendency to be taken up by iron proteins, such as transferrin, in vivo. It readily hydrolyses above pH 4 to form ill-defined chemical species and colloids. However, the presence of weakly coordinating ligands such as acetate, citrate or glycinate reduce the propensity to undergo hydrolysis, providing access to a wider useful pH range (13).
The value of radiolabelling biomolecules was recognised in the mid 1970s with the development of bifunctional chelators based on EDTA (14, 15). This was quickly extended to include DTPA (16). Some of the first examples of gallium-67 and indium-111 labelled antibodies were presented in the early 1980s (17-20) with indium-111 labelled antibodies quickly coming to dominate the field. This is highlighted by the development and FDA approval (in 1996) of [111In]capromab-pendetide (ProstaScint®). Since the early developments, a wide range of chelators have been developed and utilised to prepare radiogallium labelled biomolecules, particularly in the last two decades due to the explosive growth in both the research and clinical use of gallium-68. This topic has been covered in numerous reviews (21-25). Aspects of chelator design and radiolabelling conditions have been optimised to meet the demands of gallium-68, namely rapid labelling to match the short 68 minute half-life. Galllium-67 does not have this kinetic constraint due to the longer half-life so efforts can focus on developing and using thermodynamically stable complexes and mild radiolabelling conditions suitable for sensitive biomolecules. Additionally, thermodynamic stability is likely to be a key factor in the stability of the complex in vivo and resistance towards trans-chelation with transferrin. NOTA (1,4,7-triazacyclononane-N,N’,N’’-triacetic acid) and its derivatives remain the benchmark for 67Ga/68Ga labelling with high thermodynamic stability, mild reaction conditions (25C, <1 h) and excellent in-vivo stability (26). While the thermodynamic stability constant for Ga(DOTA) (log K = 21.3) is significantly lower than that of Ga(NOTA) (log K = 31.0) it is worthy of consideration as DOTA-conjugates have direct applicability to therapeutic application with radiolanthanides (153Sm, 161Tb, 177Lu) and other group 3 radiometals such as 47Sc and 90Y. The gallium complex of desferrioxamine (DFO) also has a high stability constant (log K = 29.7) (27) and was extensively used in the early developments of radiolabelled antibodies (see above). DFO is also the ligand of choice for most [89Zr]immunoPET (28) applications, these agents could easily be adapted to use with gallium-67. Recent studies (29, 30) have demonstrated the utility of the THP (1,6-dimethyl-3-hydroxypyridin-4-one) chelator for theranostic use of gallium-67 as and imaging and an Auger therapy agent.
The method and chemistry of conjugation also plays a pivotal role in the performance of the antibody-drug conjugates (31, 32). Early studies (33) of [67Ga]DFO-mAbs with three different linkers demonstrated the relationship between linker stability and pharmacokinetic behaviour. Arguably one of the most common methods utilised has been the straightforward acylation of biomolecules, including antibodies, with the anhydride of DTPA (16). However, conjugation strategies have improved and been embellished since this time, particularly recently by the boom in antibody-drug conjugates (34-36) and immunoPET (28, 37) within our own field. The triumvirate of new site-specific conjugation techniques, novel ligands and improved biological vectors signals an opportunity to reinvigorate the development of gallium-67 based radiopharmaceuticals.