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Biomedical subjects

Douglas Powell

Publications and source records attributed to Douglas Powell.

At least 19 recordsLinked to original sources
PubMed2006

Proteasome inhibition to maximize the apoptotic potential of cytokine therapy for murine neuroblastoma tumors.

Human neuroblastomas possess several mechanisms of self-defense that may confer an ability to resist apoptosis and contribute to the observed difficulty in treating these tumors in the clinical setting. These molecular alterations may include defects in proapoptotic genes as well as the overexpression of prosurvival factors, such as Akt among others. As a key regulator of the turnover of proteins that modulate the cell cycle and mechanisms of apoptosis, the proteasome could serve as an important target for the treatment of neuroblastoma. The present studies provide the first evidence that bortezomib, a newly approved inhibitor of proteasome function, inhibits phosphorylation of Akt, induces the translocation of proapoptotic Bid, and potently enhances the apoptosis of murine neuroblastoma tumor cells in vitro. Furthermore, in that inhibitors of the Akt pathway can sensitize otherwise resistant TBJ/Neuro-2a cells to apoptosis induced by IFN-gamma plus TNF-alpha, we hypothesized that bortezomib also could sensitize these cells to IFN-gamma plus TNF-alpha. We demonstrate for the first time that bortezomib not only up-regulates the expression of receptors for IFN-gamma and TNF-alpha on both TBJ neuroblastoma and EOMA endothelial cell lines, but also markedly enhances the sensitivity of these cells to apoptosis induced by IFN-gamma plus TNF-alpha in vitro. Furthermore, bortezomib enhances the in vivo antitumor efficacy of IFN-gamma/TNF-alpha-inducing cytokines, including both IL-2 and IL-12 in mice bearing well-established primary and/or metastatic TBJ neuroblastoma tumors. Collectively, these studies suggest that bortezomib could be used therapeutically to enhance the proapoptotic and overall antitumor activity of systemic cytokine therapy in children with advanced neuroblastoma.

Animals↗
PubMed2006

Thioamide pincer ligands with charge versatility.

This paper reports the synthesis and characterization of three complexes, two palladium and one platinum, with 2,6-bis-thioamido-phenyl and 2,6-bis-thioamido-pyridine ligands. The ligands show internal charge versatility by losing protons from a phenyl CH (I) or from amide NH's (II and III). The complexes were also examined as Heck catalysts, and the palladacycle, I, was found to be more effective compared to the others. The crystal structures of the complexes are also reported.

Crystallization↗
PubMed2006

Therapeutic modulation of Akt activity and antitumor efficacy of interleukin-12 against orthotopic murine neuroblastoma.

BACKGROUND: Patients with advanced neuroblastoma have a poor prognosis. The antiapoptotic protein Akt has been implicated as a possible mediator of the resistance of human neuroblastoma cells to apoptosis; the proapoptotic protein Bid, is inhibited by activated Akt. Neuroblastoma has demonstrated responsiveness to immunotherapeutic approaches in preclinical studies, prompting investigation of new therapeutic strategies based on potentiation of the host immune response, including the use of systemic cytokines. METHODS: We examined the antitumor efficacy and mechanisms of action of the central immunoregulatory cytokine interleukin-12 (IL-12) in mice bearing established orthotopic neuroblastoma tumors derived from murine TBJ and Neuro-2a cells. Cohorts of mice (10 mice/group) bearing established orthotopic neuroblastoma tumors were injected intraperitoneally with IL-12 or vehicle and monitored for survival. IL-12-induced apoptosis within the tumor microenvironment was investigated using ribonuclease protection assays, nuclear staining, and electron microscopy. Protein expression was determined via Western blot analysis and enzyme-linked immunosorbent assays. Confocal microscopy was used to examine the distribution of overexpressed Bid-enhanced green fluorescent protein fusion protein (Bid-EGFP) in TBJ cells. All statistical tests were two-sided. RESULTS: IL-12 induced complete tumor regression and long-term survival of 8 (80%) of 10 mice bearing established neuroblastoma tumors compared with 1 (10%) of 10 control mice (P = .0055) and profound tumor cell apoptosis in vivo despite the fact that TBJ and Neuro-2a cells were resistant to receptor-mediated apoptosis in vitro. These cells expressed high levels of phosphorylated Akt, a key prosurvival molecule, and Akt inhibitors sensitized neuroblastoma cells to apoptosis mediated by IL-12-inducible cytokines including tumor necrosis factor-alpha and interferon-gamma in vitro. IL-12 increased the expression of proapoptotic genes and decreased Akt phosphorylation within established TBJ tumors in conjunction with activation and subcellular translocation of Bid. CONCLUSIONS: Our results suggest that IL-12 overcomes a potentially critical mechanism of tumor self-defense in vivo by inhibiting Akt activity and imply that IL-12 may possess unique therapeutic activity against tumors that express high levels of activated Akt.

Adrenal Gland Neoplasms↗
PubMed2006

Ground reaction force and 3D biomechanical characteristics of walking in short-leg walkers.

Short-leg walking boots offer several advantages over traditional casts. However, their effects on ground reaction forces (GRF) and three-dimensional (3D) biomechanics are not fully understood. The purpose of the study was to examine 3D lower extremity kinematics and joint dynamics during walking in two different short-leg walking boots. Eleven (five females and six males) healthy subjects performed five level walking trials in each of three conditions: two testing boot conditions, Gait Walker (DeRoyal Industries, Inc.) and Equalizer (Royce Medical Co.), and one pair of laboratory shoes (Noveto, Adidas). A force platform and a 6-camera Vicon motion analysis system were used to collect GRFs and 3D kinematic data during the testing session. A one-way repeated measures analysis of variance (ANOVA) was used to evaluate selected kinematic, GRF, and joint kinetic variables (p<0.05). The results revealed that both short-leg walking boots were effective in minimizing ankle eversion and hip adduction. Neither walker increased the bimodal vertical GRF peaks typically observed in normal walking. However, they did impose a small initial peak (<1BW) earlier in the stance phase. The Gait Walker also exhibited a slightly increased vertical GRF during midstance. These characteristics may be related to the sole materials/design, the restriction of ankle movements, and/or the elevated heel heights of the tested walkers. Both walkers appeared to increase the demand on the knee extensors while they decreased the demand of the knee and hip abductors based on the joint kinetic results.

Adult↗
PubMed2006

Ii-Key/MHC class II epitope peptides as helper T cell vaccines for cancer and infectious disease.

Potent MHC class II antigenic peptide vaccines are created by covalently linking the N-terminus of a MHC class II epitope through a polymethylene bridge to the C-terminus of the Ii-Key segment of the Ii protein. Such hybrids enhance potency of presentation in vitro of the MHC class II epitope about 200 times relative to the epitope-only peptide. In vivo, as measured by IFN-gamma ELISPOT assays, the helper T cell response to vaccination is enhanced up to 8 times. The design of such hybrid vaccine peptides comes from insight into the mechanism of action of the Ii-Key motif within the Ii protein, in regulating antigenic peptide binding into the antigenic peptide binding groove of MHC class II molecules. Here we present the logic and experimental history of the development of these vaccine peptides, with particular attention to the hypothesized mechanism of action. Methods for the design and testing of these peptides are presented. Experience in developing peptide vaccines for immunotherapy of cancer is reviewed, focusing on the clinical potential of Ii-Key/MHC class II epitope hybrids.

Algorithms↗
PubMed2006

High frequency of genetic recombination is a common feature of primate lentivirus replication.

Recent studies indicate that human immunodeficiency virus type 1 (HIV-1) recombines at exceedingly high rates, approximately 1 order of magnitude more frequently than simple gammaretroviruses such as murine leukemia virus and spleen necrosis virus. We hypothesize that this high frequency of genetic recombination is a common feature of primate lentiviruses. Alternatively, it is possible that HIV-1 is unique among primate lentiviruses in possessing high recombination rates. Among other primate lentiviruses, only the molecular mechanisms of HIV-2 replication have been extensively studied. There are reported differences between the replication mechanisms of HIV-1 and those of HIV-2, such as preferences for RNA packaging in cis and properties of reverse transcriptase and RNase H activities. These biological disparities could lead to differences in recombination rates between the two viruses. Currently, HIV-1 is the only primate lentivirus in which recombination rates have been measured. To test our hypothesis, we established recombination systems to measure the recombination rates of two other primate lentiviruses, HIV-2 and simian immunodeficiency virus from African green monkeys (SIVagm), in one round of viral replication. We determined that, for markers separated by 588, 288, and 90 bp, HIV-2 recombined at rates of 7.4%, 5.5%, and 2.4%, respectively, whereas SIVagm recombined at rates of 7.8%, 5.6%, and 2.7%, respectively. These high recombination rates are within the same range as the previously measured HIV-1 recombination rates. Taken together, our results indicate that HIV-1, HIV-2, and SIVagm all possess high recombination frequencies; hence, the high recombination potential is most likely a common feature of primate lentivirus replication.

Cell Line↗
PubMed2006

Contact urticaria from curcumin.

Turmeric, a spice derived from the rhizome of the plant Curcuma longa, contains the chemical curcumin, which is responsible for turmeric's taste, color, and biologic properties. Curcumin is used as a spice in foods, as a treatment in traditional medicine, as a dye for fur, and as a component in nutritional supplements. A few cases of allergic contact dermatitis from curcumin have been reported. We report two cases of contact urticaria from curcumin. These cases are mediated by two different mechanisms of contact urticaria: nonimmunologic and immunologic (immunoglobulin-E mediated).

Adult↗
PubMed2005

Anion binding motifs: topicity and charge in amidocryptands.

An expanded amidocryptand with propyl linkages provides multitopic sites for binding anions and water molecules. Upon quaternization of the two bridgehead amines, the molecular shape changes from an inverted Y to that of a bowl, which is filled with water and topped by the anion.

Journal Article↗
PubMed2005

Tritopic (cascade) and ditopic complexes of halides with an azacryptand.

Structural aspects of the binding of halides in the octaaza cryptand (1,4,11,14,17,24,29,36-octa-azapentacyclo[12.12.12.2(6,9).2(19,22).2(31,34)]-tetratetraconta-6(43),7,9(44),19(41),20,22(42),31(39),32,34(40)-nonaene, N(CH2CH2NHCH2-p-xylyl-CH2NHCH2CH2)3N), L1, were examined for fluoride, chloride, and bromide. Crystallographic results for two different fluoride complexes indicated cascade-like coordination, with two fluoride ions inside the tren-based cavity, bridged by a water molecule. In the two different chloride structures, a single chloride and a water molecule occupied the cavity. The bromide structure contained two crystallographically independent cationic cryptands. Unit A consisted of a bromide on one side of the cavity and three disordered water molecules situated between the cryptand arms on the other side. Unit B also had a bromide inside the cavity at one side, but a single molecule of water was centered at the other side of the cavity. Association constants for the three ions, determined by NMR titrations in aqueous solution at pH 5, revealed log Ka=3.15(5), 3.37(3), and 3.34(4) for fluoride, chloride, and bromide, respectively.

Journal Article↗
PubMed2005

Scientific conviction amidst scientific controversy in the transatlantic livestock and meat trade.

A century before the outbreak of bovine spongiform encephalopathy (BSE), Great Britain and North America grappled with pleuro-pneumonia - a disease in cattle that had equally maddening consequences. Towards the end of the 19th century, this condition was at the heart of a transatlantic trade dispute that lasted for decades and attracted the attention of livestock farmers, diplomats, shipping moguls, veterinarians, public health regulators and journalists the world over. Scientific controversy aggravated the situation when there were doubts about the scientific judgment of Privy Council veterinary officials, who were simultaneously conducting disease-diagnosis activities and pushing for tighter regulations at British ports. At this point, William Williams, principal of the New Veterinary College in Edinburgh, waded into this troubled arena. His strong convictions spawned a long-running disagreement with the British Government over the diagnoses of pleuro-pneumonia in cattle imported from the USA and Canada.

Animal Husbandry↗
PubMed2005

Genetic recombination of human immunodeficiency virus type 1 in one round of viral replication: effects of genetic distance, target cells, accessory genes, and lack of high negative interference in crossover events.

Recombination is a major mechanism that generates variation in populations of human immunodeficiency virus type 1 (HIV-1). Mutations that confer replication advantages, such as drug resistance, often cluster within regions of the HIV-1 genome. To explore how efficiently HIV-1 can assort markers separated by short distances, we developed a flow cytometry-based system to study recombination. Two HIV-1-based vectors were generated, one encoding the mouse heat-stable antigen gene and green fluorescent protein gene (GFP), and the other encoding the mouse Thy-1 gene and GFP. We generated derivatives of both vectors that contained nonfunctional GFP inactivated by different mutations. Recombination in the region between the two inactivating mutations during reverse transcription could yield a functional GFP. With this system, we determined that the recombination rates of markers separated by 588, 300, 288, and 103 bp in one round of viral replication are 56, 38, 31, and 12%, respectively, of the theoretical maximum measurable recombination rate. Statistical analyses revealed that at these intervals, recombination rates and marker distances have a near-linear relationship that is part of an overall quadratic fit. Additionally, we examined the segregation of three markers within 600 bp and concluded that HIV-1 crossover events do not exhibit high negative interference. We also examined the effects of target cells and viral accessory proteins on recombination rate. Similar recombination rates were observed when human primary CD4(+) T cells and a human T-cell line were used as target cells. We also found equivalent recombination rates in the presence and absence of accessory genes vif, vpr, vpu, and nef. These results illustrate the power of recombination in generating viral population variation and predict the rapid assortment of mutations in the HIV-1 genome in infected individuals.

Animals↗
PubMed2005

Mechanisms of nonrandom human immunodeficiency virus type 1 infection and double infection: preference in virus entry is important but is not the sole factor.

We previously demonstrated that human immunodeficiency virus type 1 (HIV-1) infection is nonrandom and that double infection occurs more frequently than predicted from random events. To probe the possible mechanisms for nonrandom infection, we examined the role of HIV-1 entry pathways by using viruses pseudotyped with either CCR5-tropic HIV-1 Env or vesicular stomatitis virus G protein (VSV G). These two proteins use different receptors and entry pathways. We found that regardless of the protein used, double infection occurred more frequently than random events, indicating nonrandom HIV-1 infection in both entry pathways. However, the frequency of double infection differed significantly, depending on the envelope protein. In primary CD4(+) T cells, double infection occurred most frequently when both viruses had CCR5-tropic HIV-1 Env and least frequently when the two viruses had different envelopes. These results indicated that the preference in virus entry was a significant but not the only factor contributing to nonrandom double infection. Furthermore, we demonstrated that the CD4 expression level in primary T cells affects their susceptibility to CCR5-tropic HIV-1 infection but not VSV G-pseudotyped HIV-1 infection. We have also examined infection with two viruses pseudotyped with CCR5- or CXCR4-tropic HIV-1 Env and have found that double infection occurred more frequently than random events. These results indicate that coreceptor usage is not a barrier to recombination between the two virus populations. In our previous study, we also demonstrated nonrandom double infection via dendritic cell (DC)-mediated HIV-1 transmission. To test our hypothesis that multiple HIV-1 virions are transmitted during DC-T-cell contact, we used two populations of DCs, each capturing one vector virus, and added both DC populations to T cells. We observed a decreased frequency of double infection compared with experiments in which DCs captured both viruses simultaneously. Therefore, these results support our hypothesis that multiple virions are transmitted from DCs to T cells during cell-mediated HIV-1 transmission.

CD4 Antigens↗
PubMed2004

Encapsulated sulfates: insight to binding propensities.

Two crystal structures of sulfate inclusion complexes in an aza- and amido-cryptand represent the first examples of encapsulated sulfate in synthetic cryptand receptors and indicate penta- and octa-coordination, respectively.

Amides↗
PubMed2004

Ditopic double pincer palladacycle catalyst for C-C coupling.

A ditopic palladacycle with SCS pincer coordination, L(PdCl)(2) (1), was isolated and structurally characterized and represents the first example of a transition metal complex with a polythioamide-based macrocycle. Preliminary studies of 1 in the catalytic coupling of 4-iodotoluene and styrene indicated it to be robust in the presence of oxygen and high temperatures, with high turnover numbers in relatively short times.

Journal Article↗
PubMed2004

Fluoride-facilitated deuterium exchange from DMSO-d6 to polyamide-based cryptands.

Multiple deuterium exchange between DMSO-d6 and amide hydrogens in two hexaamido cryptand fluoride receptors has been verified by 19F and 2H NMR and FAB mass spectral studies. Structural results for one of the complexes indicate a tricapped trigonal prism hydrogen bond coordination geometry around an encapsulated fluoride, with hydrogen bonds from fluoride to six amide and three phenyl hydrogens.

Journal Article↗
PubMed2004

Anion binding with a tripodal amine.

Binding studies of the tren-based amine, L (N,N',N' '-tris(2-benzylaminoethyl)amine), with inorganic anions and two crystal structures, [H(3)L][H(2)PO(4)](3).H(3)PO(4) and [H(3)L][Br](3), are reported. NMR titration results indicate that the ligand binds H(2)PO(4)(-) and HSO(4)(-) more strongly than NO(3)(-) and halides. In the crystal structure of the phosphate complex, the ligand is triprotonated with the three arms pointing outward in a trigonal-planar-like arrangement. Four phosphate species are associated with the receptor, and have been assigned as three H(2)PO(4)(-) counterions located between each of the tren arms, and an additional H(3)PO(4) molecule above the quasi-planar tren. The structure of the bromide complex is slightly different, although again the tren receptor is triprotonated and quasi-planar, but in this case C(2v)-like symmetry is seen with two of the arms pointed in the same direction with a bromide ion in between. The other two bromides lie outside of the tren arms.

Journal Article↗