Synthesis of the Novel AT₁ Receptor Tracer [¹⁸F]Fluoropyridine–Candesartan via Click Chemistry

Abstract

A novel 7-((4-(3-((2-[ 18 F]fluoropyridin-3-yl)oxy)propyl)-1 H -1,2,3-triazol-1-yl)methyl)-1 H -benzo[ d ]imidazole derivative of the angiotensin II type-1 receptor (AT 1 R) blocker candesartan, [ 18 F]fluoropyridine–candesartan, was synthesized via the copper-catalyzed azide–alkyne cycloaddition click reaction between 2-[ 18 F]fluoro-3-(pent-4-yn-1-yloxy)pyridine ([ 18 F]FPyKYNE) and the tetrazole-protected azido-candesartan derivative, followed by acid deprotection. This three-step, two-pot, and two-step purification synthesis was done within 2 h. The use of tris[(1-hydroxypropyl-1 H -1,2,3-triazol-4-yl)methyl]amine (THPTA) as a Cu(I) stabilizing agent increased the overall radiochemical yield by 4-fold (10 ± 2%, n = 13) compared to the reaction without THPTA (2.4 ± 0.2%, n = 3; decay-corrected from 18 F produced at the end-of-beam). Complete separation of [ 18 F]FPyKYNE from its nitro precursor and [ 18 F]fluoropyridine–candesartan from the deprotected azido-candesartan allowed for high molar activities (>380 GBq/μmol) of the tracer. The use of 0.1% trifluoroacetic acid in water for reformulation and the addition of sodium ascorbate to the final formulation (1.6 ± 0.2 GBq/mL, n = 3) prevented tracer radiolysis with >97% radiochemical purity for a period of up to 10 h after the end-of-synthesis. A significant reduction in the uptake (86 ± 3%, n = 8) of the tracer was observed ex vivo in rats (at 20 min postinjection) in the AT 1 R-rich kidney cortex following pretreatment with saturating doses of the AT 1 R antagonist candesartan or losartan. This specific binding to AT 1 R was confirmed in vitro in the rat renal cortex (autoradiography) by a reduction of 26 ± 5% ( n = 12) with losartan coincubation (10 μM). These favorable binding properties support further studies to assess the potential of [ 18 F]fluoropyridine–candesartan as a tracer for the positron emission tomography imaging of renal AT 1 R.

[18F]Fluoropyridine‐losartan: A new approach toward human Positron Emission Tomography imaging of Angiotensin II Type 1 receptors

“…The addition of Cu(I)‐stabilizing ligands was demonstrated to accelerate the click reaction kinetics under mild conditions 20,26–29 . Indeed, we found that TBTA, BPDS, or THPTA enhanced the conversion of [ 18 F]FPyKYNE to the desired product (Table 1, entries 4–6), resulting in RCYs ranging from 7% to 17%.…”

“…In order to minimize the volatilization of the prosthetic group, we reduced the reaction temperature (150°C) and time (5 min). Furthermore, the purification was improved by using semipreparative silica‐gel HPLC 20 . It is important to note that we were not able to find a proper solvent to completely separate [ 18 F]FPyKYNE from NO 2 PyKYNE by C18 HPLC (Table S1).…”

“…Indeed, we found that TBTA, BPDS, or THPTA enhanced the conversion of [ 18 F]FPyKYNE to the desired product (Table 1, entries 4–6), resulting in RCYs ranging from 7% to 17%. We also tested the reaction conditions employed in the synthesis of [ 18 F]fluoropyridine‐candesartan, 20 which gave a similar conversion (Table 1, entry 7). The best results were attained with BPDS (Table 1, entry 5), that diminished the formation of 18 F‐fluorinated by‐products (Figure 1C).…”

“…[ 18 F]Fluoride was then eluted with a solution of K 2 CO 3 (5.5 mg, 0.04 mmol) and 4,7,13,16,21,24‐hexaoxa‐1,10‐diazabicyclo[8.8.8]hexacosane (Kryptofix® 2.2.2, 30 mg, 0.08 mmol) in 95% CH 3 CN/H 2 O (1.5 mL) into the first reactor and azeotropically dried after addition of acetonitrile (0.4 mL). [ 18 F]FPyKYNE was produced by radiofluorination of NO 2 PyKYNE (Scheme 1), completely separated from the nitro‐precursor by silica‐gel HPLC (84.5:15:0.5 hexane/dichloromethane [CH 2 Cl 2 ]/isopropanol, 9 mL/min, switched to 70:30 CH 2 Cl 2 /isopropanol after peak collection) and transferred to the second reactor as previously reported 20 . The HPLC conditions were tested earlier to optimize the purification of FPyKYNE from the nitro‐precursor (Tables S1–S3).…”

Development of a novel [18F]fluorobenzyl derivative of the AT1 receptor antagonist Candesartan

“…Incorporation of 18 F‐labeled prosthetic groups onto biological active compound has been accomplished via acylation, amidation, imidation, alkylation, and photochemical conjugation 11 . Recently, our group developed the novel tracer [ 18 F]fluoropyridine‐Candesartan labeled with the [ 18 F]FPyKYNE prosthetic via the copper‐catalyzed azide‐alkyne cycloaddition at the imidazole 7‐position in 10% yield, high molar activity, and radiochemical purity (≥97%) 12 . Biological studies revealed specific binding of [ 18 F]fluoropyridine‐Candesartan in AT 1 R‐rich tissues such as the kidney cortex.…”