Drug Cytotoxicity Assay for African Trypanosomes and Leishmania Species

Bodley, Annette L.; McGarry, Michael W.; Shapiro, Theresa A.

doi:10.1093/infdis/172.4.1157

Cited by 92 publications

(70 citation statements)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Bloodstream form trypanosomes were grown in microtiter plates for 20 or 40 h in the presence of serial dilutions of the inhibitors (Fig. 6), and cell proliferation was determined by an acid phosphatase-based assay (41). All four compounds inhibited trypanosome growth with IC 50 values that were about 10-fold higher than those determined with the soluble enzyme.…”

Section: Resultsmentioning

confidence: 96%

“…Cytotoxicity Determination-Cytotoxicity of PDE inhibitors was determined for bloodstream forms in culture by determining acid phosphatase activity as described (41). Exponentially growing monomorphic bloodstream forms MiTat 1.2 were transferred into colorless medium (42) (cell density 3 ϫ 10 5 cells/ml culture) and were seeded into microtiter wells (199 l/well) containing 1 l of inhibitor or solvent control.…”

Section: Tacgccggtattccaatgtagg-3ј; Openmentioning

confidence: 99%

See 1 more Smart Citation

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Zoraghi¹,

Kunz

Gong³

et al. 2001

Journal of Biological Chemistry

View full text Add to dashboard Cite

This study reports the identification and characterization of a cAMP-specific phosphodiesterase from the parasitic hemoflagellate Trypanosoma brucei. TbPDE2A is a class I phosphodiesterase. Its catalytic domain exhibits 30 -40% sequence identity with those of all 11 mammalian phosphodiesterase (PDE) families, as well as with PDE2 from Saccharomyces cerevisiae, dunce from Drosophila melanogaster, and regA from Dictyostelium discoideum. The overall structure of TbPDE2A resembles that of human PDE11A in that its N-terminal region contains a single GAF domain. This domain is very similar to those of the mammalian PDE2, -5, -6, -10, and -11, where it constitutes a potential cGMP binding site. TbPDE2A can be expressed in S. cerevisiae, and it complements an S. cerevisiae PDE deletion strain. Recombinant TbPDE2A is specific for cAMP, with a K m of ϳ2 M. It is entirely resistant to the nonselective PDE inhibitor 3-isobutyl-1-methylxanthine, but it is sensitive to trequinsin, dipyridamole, sildenafil, and ethaverine with IC 50 values of 5.4, 5.9, 9.4, and 14.2 M, respectively. All four compounds inhibit proliferation of bloodstream form trypanosomes in culture, indicating that TbPDE2A is an essential enzyme.

show abstract

Section: Resultsmentioning

confidence: 96%

Section: Tacgccggtattccaatgtagg-3ј; Openmentioning

confidence: 99%

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Zoraghi¹,

Kunz

Gong³

et al. 2001

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…The absorbance was determined in an EMS Reader MF Version 2.4-0, at a wavelength of 405 nm. The 50% inhibitory concentration (IC 50 ) was obtained from doseresponse curves fit to data by means of the equation for the sigmoidal E max model (Bodley et al, 1995).…”

Section: Antipromastigote Activitymentioning

confidence: 99%

Antileishmanial activity of leaf extract from Calophyllum rivulare against Leishmania amazonensis

Oubada

García

Bello

et al. 2014

Emir. J. Food Agric

View full text Add to dashboard Cite

Calophyllum rivulare is an endemic tree from Cuba that belongs to Clusiaceae family. This plant had revealed it to be a rich source of pyranochromanone acids including compounds with a variety of biological potentialities such as antioxidant, antiulcer, anticancer and anti-Helicobacter pylori activities. Herein, antileishmanial and cytotoxic activities of extracts obtained from C. rivulare leaves (rich in pyranochromanone acids) and amentoflavone were compared. Extracts were prepared by maceration employing ethyl acetate (CR-1) and methanol (CR-2) as solvents and amentoflavone was isolated from both extracts. The CR-2 extract showed the better activity against the parasite, with an IC 50 value of 95.1 ± 5.4 µg/mL and 20.6 ± 1.4 µg/mL against promastigote and amastigote forms, respectively. This extract showed lower cytotoxicity (IC 50 =182.4 ± 2.3 µg/mL) against peritoneal macrophage from BALB/c mice and a selectivity index of 9. Amentoflavone showed an IC50 of 5.9 ± 0.3 µg/mL against amastigote form and a selectivity index of 9. Results suggest that CR-2 extract and amentoflavone could be explored as new antileishmanial alternatives.

show abstract

“…Extensive phosphorylation is predicted in the N-terminal non- conserved region, phosphorylation would account for slower than expected migration in SDS͞PAGE, and a difference in phosphorylation was evident in peptide i from the two proteins. Trypanosomes have vigorous phosphatase activity (30) and phosphatase inhibitors were not used during purification, hence phosphorylation differences may be artifactual. Alternatively, they may reflect a mechanism for intracellular regulation, in keeping with topoisomerases in other systems (5).…”

Section: Discussionmentioning

confidence: 99%

An unusual type IB topoisomerase from African trypanosomes

Bodley

Chakraborty

Xie

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

African trypanosomes are ancient eukaryotes that cause lethal disease in humans and cattle. Available drugs are inadequate and the need for new therapeutic targets is great. Trypanosoma brucei and related pathogens differ strikingly from higher eukaryotes in many aspects of nucleic acid structure and metabolism. We find yet another example of this in their unusual DNA topoisomerase IB. Type IB topoisomerases relieve the supercoils that accumulate during DNA and RNA synthesis, and are of considerable importance as the target for antitumor camptothecins. Dozens of type IB topoisomerases sequenced from eukaryotes, bacteria, and pox viruses are all encoded by a single gene that predictably contains a highly conserved DNA binding domain and C-terminal catalytic domain, linked by a nonconserved hydrophilic region. We find that topoisomerase IB in T. brucei is encoded by two genes: one for the DNA-binding domain and a second for the C-terminal catalytic domain. In keeping with this, highly purified fractions of native T. brucei topoisomerase IB catalytic activity contain two proteins, of 90 and 36 kDa. The native enzyme is conventional in its Mg 2؉ -independence, ability to relax positive and negative supercoils, and inhibition by camptothecin. Camptothecin promotes the formation of a covalent complex between 32 P-labeled substrate DNA and the small subunit. This unusual structural organization may provide a missing link in the evolution of type IB enzymes, which are thought to have arisen over time from the fusion of two independent domains. It also provides another basis for the design of selectively toxic drug candidates.T he African trypanosome, Trypanosoma brucei, is a flagellated protozoan that causes sleeping sickness in humans. This tsetse fly-transmitted meningoencephalitis is of increasing incidence and is fatal if not treated (1). Closely related organisms cause Chagas' disease and leishmaniasis. Available therapies for sleeping sickness are widely acknowledged to be inadequate: they require multiple parenteral doses for cure, are expensive, toxic, and are losing efficacy against drug-resistant parasites. Trypanosomes and leishmania are ancient eukaryotes whose distinctive features include structurally and metabolically unusual DNAs. In African trypanosomes, the nuclear genome is comprised of 11 chromosome pairs, several additional chromosomes of unknown ploidy, and Ϸ100 minichromosomes, each containing a single gene that encodes a variable surface protein (2). Surface protein genes can be activated by recombinationbased transfer from minichromosomes to transcriptionally active sites in larger chromosomes. Large polycistronic transcripts are trans-spliced to form mRNAs with a 5Ј-leader sequence and 3Ј poly(A) tail. Even more unusual is their mitochondrial DNA, termed kinetoplast or kDNA, which is in the form of a single gigantic network of interlocked relaxed DNA circles (3, 4). Mature mitochondrial messages are created by an editing process that involves systematic insertion or removal of U residues.DNA top...

show abstract

Drug Cytotoxicity Assay for African Trypanosomes and Leishmania Species

Cited by 92 publications

References 11 publications

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Antileishmanial activity of leaf extract from Calophyllum rivulare against Leishmania amazonensis

An unusual type IB topoisomerase from African trypanosomes

Contact Info

Product

Resources

About