Synthesis and biologic evaluation of a new ligand for potential SPECT imaging of opioid receptors

Objectives To synthesize a novel iodinated analog of diprenorphine (DPN) as a potential radioligand for SPECT imaging of cerebral opioid receptor and evaluate its biological characterization.

Methods E-3-(Tri-butylstannyl)prop-2-en-1-ol, esters of p-Toluenesulfonate as prosthetic group for radioiodination, 3-acetyldiprenorphine and 7α-O-[[(3-tri-butylstannyl)prop-2- enyl]oxy]-diprenorphine were synthesized, respectively. This compound was purified after AC-DPN reaction with Radioiododestannylation with no-carrier iodine-125 using iodogen as oxidant in one step to give the corresponding 125I-7α-O-Iodoallyl DPN analogue. The binding studied with membranes from rat brain without cerebellum. Kd values were calculated according to the Scachard equation. All assays were performed in triplicate. The 25 mice were distributed randomly into 5 groups, each was administrated 125I-7α-O-Iodoallyl DPN (20 μCi, 10 pmol) through the tail vein. At various time after injection, the animals were euthanized by cervical dislocation. Organs were excised, rinsed with saline and then blotted dry, weighed and determined their radioactivity in γ counter. Another 25 mice were co-inject DPN (10 μmol) with 125I-7α-O-Iodoallyl DPN (20 μCi, 10pmol) through the tail vein. Then the rats repeated the same procedure as above.

Results Prosthetic groups which imparted less steric bulk and lipophilicity were especially desirable for radioiodination of small molecules. 3-acetylation DPN was to protect the 3-OH on phenolic ring of diprenorphine. Armed with appropriate standards, radioiodination of the vinylstannanes at the no-carrier-added level was explored. 125I-7α-O-Iodoallyl DPN was obtained in good radiolabelled yields (81%). After puriefied by HPLC, we got a promising radioligand with high specific activity ( 2250 Ci/mmol). In vitro opioid receptor binding assay showed that Iodoallyl diprenorphine had a very high affinity as a ligand to opioid receptor (Kd= 0.23 nM, Bmax=38 pmol/g ) in mouse brain. Biodistribution in mice showed that the greatest amount (0.67 ID%/g) of the cerebral uptake occurred 20 min after i.v. The uptake rate could be competition-inhibited by co-injection of DPN.

Conclusions 7α-O-IA-DPN can be successfully obtained from an opioid antagonist DPN by radioiododestannylation. The characterization of this compound, high specific activity, good yield, simple rapid radiolabel procedure and very high-affinity in vitro provides a promising potential radioligand for SPECT imaging of opioid receptor.