Raymond E. Gibson

[125I] 3-quinuclidinyl 4-iodobenzilate: A high affinity, high specific activity radioligand for the M1 and M2-acetylcholine receptors

We have prepared a radioiodinated ligand which binds with high affinity to the muscarinic acetylcholine receptor (m-AChR). A derivative of 3-quinuclidinyl benzilate, [125I] labeled (R) 1-aza-bicyclo(2.2.2)oct-3-yl (R,S)-alpha-hydroxy-alpha-(4-[125I]iodophenyl)phenyl acetate (4- IQNB ) exhibits an affinity for the m-AChR from corpus striatum higher than that of (R) [3H] QNB. Additionally, [125I] 4- IQNB exhibits receptor selectivity for the M1 receptor since the affinity for the receptor from dog and rat heart is lower than that using dog or rat corpus striatum.

Differences in affinities of muscarinic acetylcholine receptor antagonists for brain and heart receptors.

The affinities of atropine, scopolamine, 3-quinuclidinyl benzilate and twelve analogues of 3-quinuclidinyl benzilate were determined for the muscarinic acetylcholine receptor (m-AChR) using membrane preparations from caudate/putamen. The affinity constants thus obtained were compared with affinities previously reported for the m-AChR obtained from ventricular muscle. The affinities differed significantly for six of the compounds, the largest difference being 16-fold. Neither solubilization nor variation of physiologically significant salts led to a significant change in the affinity of that compound. These results are interpreted as supporting the subclassification of the muscarinic acetylcholine receptor.

An autoradiographic study of muscarinic cholinoceptors in blood vessels: no localization on vascular endothelium

In vitro labelling and autoradiographic techniques were used to examine the localization of [3H]quinuclidinyl benzilate ([3H]QNB) and [125I]4-iodo-QNB ([125I]4IQNB) to slide-mounted sections of rabbit aorta and pulmonary artery, cat aorta, pulmonary and superior mesenteric arteries. These vessels all respond to acetylcholine (ACh) with endothelium-dependent relaxation, yet there was no evidence for endothelium-related binding of either [3H]QNB or [125I]4IQNB. Muscarinic receptors were localized over the medial smooth muscle and, in the rabbit pulmonary artery, the density of binding increased towards the adventitia. Binding of either radioligand to sections of rabbit pulmonary artery was not affected by the muscarinic M1 receptor antagonist pirenzepine (20 nM) but was markedly reduced by the muscarinic M2 antagonist 4DAMP (4-diphenylacetoxy-N-methyl-piperidine methobromide) (1 nM). This study provides evidence for muscarinic receptors located directly on smooth muscle cells, indicating that endothelium-dependent relaxation to ACh results from an indirect mechanism involving smooth muscle muscarinic receptors.

[77Br]-17-α-Bromoethynylestradiol: In vivo and In vitro characterization of an estrogen receptor radiotracer

Abstract [ 77 Br]17-α-Bromoethynylestradiol has been prepared by direct, no-carrier-added bromination of 17-α-iodoethynylestradiol. The product thus obtained has an effective specific activity up to 185 Ci/mmol and an association constant( K A ) of 7.1 × 10 9 M −1 . In vivo distribution studies indicate accumulation in rat uterus to the same levels as [ 3 H]E 2 (10% dose/g wet weight) and a target to blood ratio of 5–6.

Calculation of binding isotherms when ligand and receptor are in different volumes of distribution

Abstract An equation for the determination of the concentration of ligand bound in a system where the receptor and ligand occupy different volumes of distribution (nonhomogeneous) is derived. The validity of the equation is demonstrated using equilibrium dialysis as the model experimental system. The nonhomogeneous equation and the equation describing the homogeneous system are contrasted and the results discussed in terms of application to in vivo steady-state systems.

Synthesis, receptor binding and tissue distribution of 17α-E[125I]iodovinyl-11β-ethyl-estradiol

Abstract In this study we prepared and evaluated a derivative of estradiol with an ethyl group at the 11β-position and an E-iodovinyl group at the 17α-position. This new ligand binds to the estrogen receptor with an affinity slightly less than estradiol (RBA = 43%) at 0 °C but much greater (RBA = 890%) at 25 °C. The 125 I-labeled derivative was obtained by radioiododestannylation of the tri- n -butylstannyl precursor in good radiochemical yield with a specific activity exceeding 1500Ci/mmol. The tissue distribution in immature female rats was evaluated over a 48 h period to determine uterine uptake and selectivity. Peak uterine uptake at 2 h was 6% ID/g and was significantly greater than that of [ 3 H]estradiol, 2.4% ID/g. Substantial uptake in the uterus was still present at 48 h (2.4% ID/g). Co-administration of estradiol reduced the uptake at 2 and 24 h by 85%. Uterus-to-plasma ratios increased with time, from about 25:1 at 2 h to nearly 90:1 at 48 h. The affinity, ease of radiosynthesis and tissue distribution of the 17α-E-[ 125 I]iodovinyl-11β-ethyl-estradiol suggest that further evaluation of this agent as an imaging agent for estrogen-receptor-positive breast cancer is warranted.

In vivo dissociation kinetics of [3H]quinuclidinyl benzilate: relationship to muscarinic receptor concentration and in vitro kinetics

The in vivo washout kinetics of [3H]quinuclidinyl benzilate ([3H]QNB) varies significantly in various structures in the rat brain. The slowest washout rates are from the hippocampus, corpus striatum, and cortex, intermediate rates are exhibited from the thalamus and colliculi, while the fastest washout rate is from the cerebellum. We have also demonstrated a difference in the in vitro dissociation rates (k-1) of [3H]QNB from various structures. The k-1 for the hippocampus, corpus striatum and cortex, is two-fold slower than that observed in the thalamus, colliculi, and cerebellum. The differences in the in vitro dissociation kinetics are not, however, sufficient to explain the differences in the in vivo washout kinetics. We have developed a theoretical formulation which describes conditions under which the washout kinetics are a function of the concentration of receptor in a structure. Furthermore, we present a graphical method in which a plot of the reciprocal of the observed washout rate constant, 1/k(obs), vs receptor concentration is linear. Analysis of the washout kinetics of [3H]QNB from various structures of the CNS of rat were well described by this theory when the differences in in vitro k-1 are included.

The use of tritium labeled compounds to develop gamma-emitting receptor-binding radiotracers.

In an effort to evaluate receptor binding drugs for their potential as gamma labeled radiopharmaceuticals suitable for clinical heart scanning, in vivo data were compared with the results obtained from a theoretical model. The distribution of selected tritium-labeled, receptor-binding radiotracers was studied in animals to determine if the heart to blood ratios agree with those obtained using a theoretical model of receptor binding. In general, the in vivo studies agree with the theoretical model when the concentration of the radiotracer in the heart is due to specific receptor binding. The use of the theoretical model for a first approximation followed by in vivo biodistribution studies is an efficient strategy to select those few from among the large number of receptor binding compounds that will ultimately yield an efficacious radiopharmaceutical to study receptor changes in the intact human heart.

In vivo and in vitro characteristics of the N-Methyl derivative of [125I]3-Quinuclidinyl 4-Iodobenzilate

Abstract Radiolabeled analogues of 3-quinuclidinyl benzilate (QNB) localize in the myocardium by virtue of ligand-receptor interactions. The radioiodinated derivative exhibits less favorable non-receptor interactions than tritiated QNB in the myocardium, probably due to increased lipophilicity. In order to prevent intracellular localization and partitioning into lipid membranes, we prepared the N-methyl quaternary analogue of the iodinated derivative [ 125 I]3-quinuclidinyl 4-iodobenzilate. In vitro and in vivo results indicate that the quaternization reduces the affinity of the analogue to a value too low for advantageous receptor-mediated localization.

Receptor-selective localization in pancreas

We examined the distribution of three tritiated ligands and two radioiodinated ligands for their ability to localize in the pancreas of rat and rabbit. The ligands examined are selective for the alpha- and beta-adrenoceptors and the muscarinic acetylcholine receptor. Of the ligands examined, the results indicate that only (R) 3H-3-quinuclidinyl benzilate (QNB) localized in the pancreas by the receptor-mediated mechanism. The % dose/g tissue, the pancreas-to-blood and pancreas-to-liver ratios are such that a 18F-labeled derivative of QNB should provide images of the pancreas.