Biomedical subjects
Publications and source records attributed to V M Kolb.
Doisynolic acids are alkaline degradation products of steroidal estrogens. While these doisynolic acids are potent estrogens, some being more estrogenic than estradiol itself, they bind to cytoplasmic estrogen receptors only feebly compared with estradiol.
Approximate ab initio molecular orbital methods are used to examine the structural and electronic properties of oxymorphone. The most stable conformation of the molecule is found to include an intramolecular hydrogen bond between the C-14 hydroxyl group and the nitrogen atom in agreement with available experimental data. The total molecular electron density is transformed to a set of localized molecular orbitals, one of which corresponds to the lone electron pair on nitrogen. The hydrogen bond is shown to produce substantial bending and stretching of the lone pair when compared to its shape when such hydrogen bonding is precluded.
Anomalous variations in the pKa values of variously substituted morphine-type agonists and antagonists are interpreted as a reflection of long-range substituent effects operating in these molecules. Based on the operation of long-range effects, a mechanism is proposed by which substitution into the N-normorphine portion of morphine-type agonists and antagonists changes the activity of the parent molecule. Thus, a remote substituent would distort the whole molecule via a conformational transmission effect and thereby (a) change the fit between the opiate and its receptor; (b) change the electron density distribution throughout the molecule and, therefore, at the nitrogen; (c) modify the directionality of the lone electron pair on the nitrogen; and (d) affect the pKa of the drug. The operation of long-range effects as proposed here could account for some of the anomalous changes in opiate activity effected by substitution into the parent molecule.
A new opiate-receptor model is proposed in which only one conformation of the receptor is needed for binding of both agonists and antagonists. There are two different spacially fixed amine-binding sites in this model: one agonist and one antagonist. The opiates undergo binding to their amine-binding sites via the lone electron pair on nitrogen. The role of the N-allyl or other such group in imparting antagonist properties is explained in terms of the steric requirements of this group. For this group to be accommodated without imposing severe steric interactions in the rest of the opiate molecule, the piperidine ring must assume a flexible (skew boat) conformation; in this conformation, the N-lone-pair electron lobe assumes the characteristic directionality of an antagonist toward its amine-binding site. If the N-lone-pair lobe is not rigorously maintained in this direction, the opiate molecule assumes both antagonist and agonist conformations and mixed antagonist-agonist activity is observed. The observed differences in the effect of sodium on the degree of binding of an agonist versus an antagonist can be explained in this model by the different effects of sodium on the two amine-binding sites. The antagonist activity of an N-methyl antagonist can be rationalized on the basis of the proposed model.
Fluorescent-labeled hormones can be used to study hormone-receptor interactions by means of fluorescence polarization, visualization by fluorescence microscopy, or separation methods, e.g., dextran-coated charcoal. Subcellular fragments, single cells, and tissue preparations are amenable to study; in this work rat uterine cytosol was used unless otherwise noted. Estrone labeled with fluorescein at position 17 gives 50% inhibition in the radiometric dextran-coated charcoal assay at 8.3 X 10(-7) M as compared to 3.4 and 3.5 X 10(-8) M for diethylstilbestrol and estradiol, respectively. Scatchard plots from fluorescence polarization are hyperbolic and consistent with two classes of binding sites having association constants 5.6 X 10(10) and 6.4 X 10(7) M-1. Binding by high-affinity sites, which were present at about 3 times the concentraion of "specific" sites (radiometric dextran-coated charcoal assay), was abrogated by estradiol or diethylstilbestrol. Kinetic measurements showed that binding sites that can be blocked by excess estradiol or diethylstilbestrol are those that are both slowly associating and slowly dissociating. Staining of tissues by estrone labeled with fluorescein at position 17 as seen in the fluorescence microscope showed specificity. In normal rat uterus only epithelial cells were stained. In one human infiltrating ductal carcinoma only the malignant ductoid elements stained, while in another there was essentially no staining.