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Browsing Health and Biomedical Sciences by Author "Campbell, Christopher"
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Item A Method and Composition Using a Dual-specificity Pseudophosphatase as an Antimalarial Drug Target(2012-02-24) Campbell, ChristopherItem A Method and Composition Using a Dual-specificity Pseudophosphatase as an Antimalarial Drug Target(2012-02-17) Campbell, ChristopherItem A Method and Composition Using a Dual-specificity Pseudophosphatase as an Antimalarial Drug Target(2012-04-05) Campbell, ChristopherItem Atypical Mitogen-activated Protein Kinase Phosphatase Implicated in Regulating Transition from Pre-S-Phase Asexual Intraerythrocytic Development of Plasmodium falciparum(Eukaryotic Cell, 2013) Campbell, ChristopherIntraerythrocytic development of the human malaria parasite Plasmodium falciparum appears as a continuous flow through growth and proliferation. To develop a greater understanding of the critical regulatory events, we utilized piggyBac insertional mutagenesis to randomly disrupt genes. Screening a collection of piggyBac mutants for slow growth, we isolated the attenuated parasite C9, which carried a single insertion disrupting the open reading frame (ORF) of PF3D7_1305500. This gene encodes a protein structurally similar to a mitogen-activated protein kinase (MAPK) phosphatase, except for two notable characteristics that alter the signature motif of the dual-specificity phosphatase domain, suggesting that it may be a low-activity phosphatase or pseudophosphatase. C9 parasites demonstrated a significantly lower growth rate with delayed entry into the S/M phase of the cell cycle, which follows the stage of maximum PF3D7_1305500 expression in intact parasites. Genetic complementation with the full-length PF3D7_1305500 rescued the wild-type phenotype of C9, validating the importance of the putative protein phosphatase PF3D7_1305500 as a regulator of pre-S-phase cell cycle progression in P. falciparum.Item Evaluation of a Putative Plasmodium Pseudophosphatase for Antimalarial Drug Discovery(2013-03-02) Campbell, ChristopherItem Evaluation of Putative Plasmodium MAPK-like Phosphatase for Antimalarial Drug Discovery(2013-02-22) Campbell, ChristopherItem Evaluation of Putative Pseudophosphatase Function for Antimalarial Drug Discovery(2012-04-01) Campbell, ChristopherItem In Silico Characterization of an Atypical MAPK Phosphatase of Plasmodium Falciparum as a Suitable Target for Drug Discovery(Chemical Biology and Drug Design, 2014) Campbell, ChristopherPlasmodium falciparum , the causative agent of malaria, contributes to significant morbidity and mortality worldwide. Forward genetic analysis of the blood-stage asexual cycle identified the putative phosphatase from PF3D7_1305500 as an important element of intraerythrocytic development expressed throughout the life cycle. Our preliminary evaluation identified it as an atypical MAPK phosphatase. Additional bioinformatics analysis delineated a conserved signature motif and three residues with potential importance to functional activity of the atypical dual-specificity phosphatase (DUSP) domain. A homology model of the DUSP domain was developed for use in high-throughput in silico screening of the available library of antimalarial compounds from ChEMBL-NTD. Seven compounds from this set with predicted affinity to the active site were tested against in vitro cultures and three had reduced activity against a ΔPF3D7_1305500 parasite, suggesting PF3D7_1305500 is a potential target of the selected compounds. Identification of these compounds provides a novel starting point for a structure-based drug discovery strategy that moves us closer towards the discovery of new classes of clinical antimalarial drugs. These data suggest that MAPK phosphatases represent a potentially new class of P. falciparum drug target.Item Mapping Epitopes of the Plasmodium vivax Duffy Binding Protein with Naturally Acquired Inhibitory Antibodies(Infection and immunity, 2010) Campbell, ChristopherPlasmodium vivax Duffy binding protein (DBP) is a merozoite microneme ligand vital for blood-stage infection, which makes it an important candidate vaccine for antibody-mediated immunity against vivax malaria. A differential screen with a linear peptide array compared the reactivities of noninhibitory and inhibitory high-titer human immune sera to identify target epitopes associated with protective immunity. Naturally acquired anti-DBP-specific serologic responses observed in the residents of a region of Papua New Guinea where P. vivax is highly endemic exhibited significant changes in DBP-specific titers over time. The anti-DBP functional inhibition for each serum ranged from complete inhibition to no inhibition even for high-titer responders to the DBP, indicating that epitope specificity is important. Inhibitory immune human antibodies identified specific B-cell linear epitopes on the DBP (SalI) ligand domain that showed significant correlations with inhibitory responses. Affinity-purified naturally acquired antibodies on these epitopes inhibited the DBP erythrocyte binding function greatly, confirming the protective value of specific epitopes. These results represent an important advance in our understanding of part of blood-stage immunity to P. vivax and some of the specific targets for vaccine-elicited antibody protection.Item Partial Characterization of PF13_0027: A Putative Phosphatase of Plasmodium Falciparum(2013) Campbell, ChristopherSignal transduction and stage-specific gene expression are essential components of Plasmodium falciparum development. In this study, the putative phosphatase PF13_0027 is investigated as a critical component of intraerythrocytic development contributing to maturation of the late trophozoite. This putative phosphatase was identified during the course of a large-scale insertional mutagenesis project by insertion of the piggyBac (pB) element, containing a human dihydrofolate reductase (hDHFR) drug selection cassette into the open reading frame (ORF) preventing expression and attenuating parasite development. PF13_0027 codes for a protein with a rhodanese (RHD) and dual specificity phosphatase (DUSP) in a tandem arrangement typically identified with mitogen-activated protein kinase (MAPK) phosphatases (MKP). Despite numerous INDELs, the tertiary structure is conserved when compared to the solved structures of MKP homologs. The expression profile reveals transcripts at all stages of the blood cycle with a highest relative abundance in the late trophozoite. Restoration of the phenotype was achieved through genetic complementation using the complete PF13_0027 open reading frame (ORF) under the control of its endogenous promoter. A homology model of PF13_0027 was developed for structural analysis and evaluated using in silico high-throughput screening (HTS) to identify antimalarial compounds with predicted affinity to the active site and used to challenge parasites in vitro. This study reveals that PF13_0027 is a vital component of asexual development and a potential target for a new class of antimalarial compounds targeting phosphorylation pathways in P. falciparum. Discovery of the functional role of this unknown ORF provides additional insight into the importance of MAPK signaling in P. falciparum.Item Site Specific/Targeted Delivery of Gemcitabine through Anisamide Anchored Chitosan/Poly Ethylene Glycol Nanoparticles: An Improved Understanding of Lung Cancer Therapeutic Intervention(European Journal of Pharmaceutical Sciences, 2012) Campbell, ChristopherGemcitabine (2', 2'-difluorodeoxycytidine) is a deoxycytidine analog with significant antitumor activity against variety of cancers including non-small cell lung cancer. However, rapid metabolism and shorter half-life of drug mandate higher dose and frequent dosing schedule which subsequently results into higher toxicity. Therefore, there is a need to design a vector which can reduce the burden of frequent dosing and higher toxicity associated with the use of gemcitabine. In this study, we investigated the possibility of improving the targeting potential by employing the surface modification on Chitosan/poly(ethylene glycol) (CTS/PEG) Nanoparticles. We demonstrate formulation and characterization of chitosan/poly(ethylene glycol)-anisamide (CTS/PEG-AA) and compared its efficiency with CTS/PEG and free gemcitabine. Our results reveal its sizeable compatibility, comparatively less organ toxicity and higher antitumor activity in vitro as well as in vivo. This wealth of information surfaces the potential of CTS/PEG-AA nanoparticles as a potent carrier for drug delivery. In brief, this novel carrier opens new avenues for drug delivery which better meets the needs of anticancer research.