Biomedical subjects

Duane D Miller

Publications and source records attributed to Duane D Miller.

43 records · Page 3Linked to original sources

PubMed2003

Key structural features of nonsteroidal ligands for binding and activation of the androgen receptor.

The purposes of the present studies were to examine the androgen receptor (AR) binding ability and in vitro functional activity of multiple series of nonsteroidal compounds derived from known antiandrogen pharmacophores and to investigate the structure-activity relationships (SARs) of these nonsteroidal compounds. The AR binding properties of sixty-five nonsteroidal compounds were assessed by a radioligand competitive binding assay with the use of cytosolic AR prepared from rat prostates. The AR agonist and antagonist activities of high-affinity ligands were determined by the ability of the ligand to regulate AR-mediated transcriptional activation in cultured CV-1 cells, using a cotransfection assay. Nonsteroidal compounds with diverse structural features demonstrated a wide range of binding affinity for the AR. Ten compounds, mainly from the bicalutamide-related series, showed a binding affinity superior to the structural pharmacophore from which they were derived. Several SARs regarding nonsteroidal AR binding were revealed from the binding data, including stereoisomeric conformation, steric effect, and electronic effect. The functional activity of high-affinity ligands ranged from antagonist to full agonist for the AR. Several structural features were found to be determinative of agonist and antagonist activities. The nonsteroidal AR agonists identified from the present studies provided a pool of candidates for further development of selective androgen receptor modulators (SARMs) for androgen therapy. Also, these studies uncovered or confirmed numerous important SARs governing AR binding and functional properties by nonsteroidal molecules, which would be valuable in the future structural optimization of SARMs.

Anilides↗

PubMed2003

Fatty alcohol phosphates are subtype-selective agonists and antagonists of lysophosphatidic acid receptors.

A more complete understanding of the physiological and pathological role of lysophosphatidic acid (LPA) requires receptor subtype-specific agonists and antagonists. Here, we report the synthesis and pharmacological characterization of fatty alcohol phosphates (FAP) containing saturated hydrocarbon chains from 4 to 22 carbons in length. Selection of FAP as the lead structure was based on computational modeling as a minimal structure that satisfies the two-point pharmacophore developed earlier for the interaction of LPA with its receptors. Decyl and dodecyl FAPs (FAP-10 and FAP-12) were specific agonists of LPA(2) (EC(50) = 3.7 +/- 0.2 microM and 700 +/- 22 nM, respectively), yet selective antagonists of LPA(3) (K(i) = 90 nM for FAP-12) and FAP-12 was a weak antagonist of LPA(1). Neither LPA(1) nor LPA(3) receptors were activated by FAPs; in contrast, LPA(2) was activated by FAPs with carbon chains between 10 and 14. Computational modeling was used to evaluate the interaction between individual FAPs (8 to 18) with LPA(2) by docking each compound in the LPA binding site. FAP-12 displayed the lowest docked energy, consistent with its lower observed EC(50). The inhibitory effect of FAP showed a strong hydrocarbon chain length dependence with C12 being optimum in the Xenopus laevis oocytes and in LPA(3)-expressing RH7777 cells. FAP-12 did not activate or interfere with several other G-protein-coupled receptors, including S1P-induced responses through S1P(1,2,3,5) receptors. These data suggest that FAPs are ligands of LPA receptors and that FAP-10 and FAP-12 are the first receptor subtype-specific agonists for LPA(2).

Animals↗

PubMed2003

Emerging trends in oral delivery of peptide and protein drugs.

Most peptide and protein drugs are currently used as parenteral formulations because of their poor oral bioavailability. Development of an effective oral delivery system for these macromolecular drugs requires a thorough understanding of their physicochemical properties, such as molecular weight, hydrophobicity, ionization constants, and pH stability, as well as biological barriers that restrict protein and peptide absorption from the gastrointestinal (GI) tract, including pH variability, enzymatic degradation, and membrane efflux. Various strategies currently under investigation include amino acid backbone modifications, formulation approaches, chemical conjugation of hydrophobic or targeting ligand, and use of enzyme inhibitors, mucoadhesive polymers, and absorption enhancers. However, there is only limited success because of the hostile environment of the GI tract--e.g., strong pH extremes and abundant presence of potent luminal enzymes. This review focuses on the challenges posed by the GI system and how different pharmaceutical approaches can be used to make oral delivery of protein and peptide drugs more feasible. The roles of P-glycoprotein and CYP3A4 in controlling the extent of intestinal absorption and metabolism will also be discussed.

Administration, Oral↗

PubMed2002

Novel nonsteroidal ligands with high binding affinity and potent functional activity for the androgen receptor.

While nonsteroidal androgen receptor (AR) antagonists have been known for many years, and used in the clinic for the treatment of hormone dependent prostate cancer, very little is known about nonsteroidal AR agonists. We designed and synthesized a series of chiral bicalutamide analogs, which bear electron-withdrawing groups (either a cyano or a nitro group at the 4-position and a trifluoromethyl group at the 3-position) in the aromatic A ring, and different substituents at the para position in the aromatic B ring of the parent molecule. We also synthesized a series of racemic bicalutamide analogs, which have a trifluoromethyl group instead of a methyl group at the R(2) position. We examined AR binding affinities of our compounds in a competitive binding assay with a radiolabeled high affinity AR ligand, 3H-mibolerone, and also measured their abilities to stimulate AR-mediated transcriptional activation in a cotransfection assay. These studies demonstrated that (1) nonsteroidal ligands can be structurally modified from known nonsteroidal antiandrogens to generate ligands capable of activating AR-mediated transcriptional activation. (2) R-isomer analogs exhibit higher AR binding affinity and more potent functional activity than their corresponding S-isomers in all cases. (3) All sulphide analogs show higher AR binding affinity and more potent functional activity than their corresponding sulphone analogs, with the exception of ligand R-8. Those ligands which exhibit high AR binding affinity and potent functional activity for human AR may provide effective clinical uses for male fertility, male contraception, and hormone replacement therapy.

Androgens↗

PubMed2002

Yohimbine dimers exhibiting selectivity for the human alpha 2C-adrenoceptor subtype.

Yohimbine is a potent and selective alpha2- versus alpha1-adrenoceptor antagonist. To date, drugs with high specificity for the alpha2-adrenoceptor show marginal selectivity among the three alpha2-adrenoceptor subtypes. Initial studies showed that yohimbine was about 4- and 15-fold more selective for the human alpha2C-adrenoceptor in comparison with the alpha2A- and alpha2B-adrenoceptors, respectively. To improve on this alpha2-adrenoceptor subtype selectivity, a series of yohimbine dimers (varying from n = 2 to 24 spacer atoms) were prepared and evaluated for receptor binding on human alpha2-adrenoceptor subtypes expressed in Chinese hamster ovary cells. Each dimeric analog showed higher affinities for alpha2A- and alpha2C-adrenoceptor versus the alpha2B-adrenoceptor; and yohimbine dimers with spacers of n = 2, 3, 4, 18, and 24 exhibited selectivity for the alpha2C-adrenoceptor. The yohimbine dimers n = 3 and n = 24 showed the highest potency and selectivity (32- and 82-fold. respectively) for the alpha2C-adrenoceptor in receptor binding and in functional studies (42- and 29-fold, respectively) measuring cAMP changes using a cell-based luciferase reporter gene assay. The dimers (n = 3 and n = 24) had high selectivity (>1000-fold) for the alpha2C-adrenoceptor compared with the three alpha1-adrenoceptor subtypes. These findings demonstrate that the addition of spacer linkages to bivalent yohimbine molecules provides a successful approach to the development of ligands that are potent and highly selective for the alpha2C-adrenoceptor.

Animals↗

PubMedDate not supplied

Cationic lipids with increased DNA binding affinity for nonviral gene transfer in dividing and nondividing cells.

Effect of headgroup structure on catonic lipid-mediated transfection was investigated with either a (i) tertiary amine, (ii) quaternary amine with a hydroxyl, or (iii) quaternary amine with mesylate as headgroups. Liposomes were formulated using cholesterol or dioleoyl phosphatidyl ethanolamine (DOPE) as colipids, and transfection efficiencies were determined in rapidly dividing colon carcinoma (CT 26) and rat aortic smooth muscle (RASM) cells as well as in nondividing human pancreatic islets using luciferase and green fluorescent protein expression plasmids, pcDNA3-Luc and pCMS-EGFP, respectively. Liposome/pDNA complexes were evaluated for DNA conformational state by circular dichroism (CD), DNA condensation by electrophoretic mobility shift assay (EMSA), particle size and zeta potential by laser diffraction technique, and surface morphology by transmission electron microscopy (TEM). Encouraging transfection results were obtained with the mesylate headgroup based lipid in liposome formulations with DOPE as a colipid, which were higher than the commercially available Lipofectamine formulation. We hypothesize that the additional hydrogen bonding or covalent interactions of the headgroup with the plasmid DNA, leading to higher binding affinity of the cationic lipids to pDNA, results in higher transfection. This hypothesis is supported by TEM observations where elongated complexes were observed and more lipid was seen associated with the DNA.

Animals↗

PubMedDate not supplied

Novel branched poly(ethylenimine)-cholesterol water-soluble lipopolymers for gene delivery.

A novel water-soluble lipopolymer was synthesized by linking cholesteryl chloroformate to the secondary amino groups of branched poly(ethylenimine) (PEI) of 1,800 and 10,000 Da. Conjugation through PEI secondary amines gives this newly synthesized lipopolymer (abbreviated as PEI-Chol) special advantage over our previously synthesized lipopolymers, which utilized the primary amino groups for conjugation, as the primary amino groups have a significant role in DNA condensation. Also, significantly, only one cholesterol molecule was grafted onto each PEI molecule (confirmed by (1)H NMR and MALDI-TOF mass spectrometry), leaving enough space for the steric interactions of the PEI's primary amines with the DNA. The PEI-Chol lipopolymer was characterized for the critical micellar concentration (cmc), buffer capacity, DNA condensation (by band retardation and circular dichroism), in vitro transfection efficiency, and cell viability. The cmcs of PEI-Chol 1,800 and PEI-Chol 10,000 were 496.6 and 1,330.5 microg/mL, respectively. The acid-base titration indicated high buffering capacity of the polymers around the pH range of 5-7, which indicated their potential for buffering in the acidic pH environment of the endosomes. The band retardation studies indicated that efficient condensation of the plasmid DNA could be achieved using these lipopolymers. The circular dichroism spectra indicated a change in DNA conformation and adoption of lower energy state upon condensation with these lipopolymers when an N/P ratio of 2.5/1 or above was formulated. The mean particle size of these complexes was in the range 110-205 nm, except for the complexes prepared using PEI of 1,800 Da, which had a mean particle size of 384 +/- 300 nm. The zeta potential of DNA complexes prepared using PEI-Chol 1,800, PEI-Chol 10,000 and PEI of 1,800, 10,000, and 25,000 Da at an N/P ratio of 15/1 was in the range 23-30 mV and was dependent on the N/P ratios. The in vitro transfection of PEI-Chol/pCMS-EGFP complexes in Jurkat cells showed high levels of expressed Green Fluorescent Protein (GFP) with little toxicity as determined by flow cytometry. These novel water-soluble lipopolymers provided good transfection efficiency with other desirable characteristics such as water solubility, free primary amino groups for efficient DNA condensation and high buffer capacity that indicated the possibility of efficient endosomal release.

Cell Survival↗