Considerable Cytotoxic Activity Research Articles

Synthesis, spectral and pharmacological studies on lanthanide(III) complexes of 3,5-pyrazoledicarboxylic acid

Complexes of cerium(III), lanthanum(III) and neodymium(III) with 3,5-pyrazoledicarboxylic acid (H3pdc) were synthesized and their compositions determined by elemental analysis. To identify the binding of Ce(III), La(III) and Nd(III) with H3pdc, detailed vibrational analysis was performed comparing experimental vibrational spectra of the ligand and its Ln(III) complexes with theoretically predicted and with literature data from related compounds. Significant differences in the IR and Raman spectra of the complexes were observed as compared to spectra of the ligand. The ligand and the complexes were tested for cytotoxic activities on the chronic myeloid leukemia derived K-562, overexpressing the BCR-ABL fusion protein and the non-Hodgkin lymphoma derived DOHH-2, characterized by an overexpression of the antiapoptotic protein bcl-2 cell lines. The results indicate that the tested compounds exerted considerable cytotoxic activity upon the evaluated cell lines in a concentration dependent manner; we constructed dose-response curves and calculated corresponding IC50 values. The lanthanide complexes exhibited potent cytotoxic activity, even more than cisplatin towards K-562 and DOHH-2 cell lines. In order to elucidate some of the mechanistic aspects of the observed cytotoxic effects, we evaluated whether the established cytotoxicity of the most active complex La(H2pdc) is related to its capacity to induce cell death through apoptosis.

Transcriptome analysis approaches for the isolation of trichome-specific genes from the medicinal plant Cistus creticus subsp. creticus

Cistus creticus subsp. creticus is a plant of intrinsic scientific interest due to the distinctive pharmaceutical properties of its resin. Labdane-type diterpenes, the main constituents of the resin, exhibit considerable antibacterial and cytotoxic activities. In this study chemical analysis of isolated trichomes from different developmental stages revealed that young leaves of 1-2 cm length displayed the highest content of labdane-type diterpenes (80 mg/g fresh weight) whereas trichomes from older leaves (2-3 or 3-4 cm) exhibited gradual decreased concentrations. A cDNA library was constructed enriched in transcripts from trichomes isolated from young leaves, which are characterized by high levels of labdane-type diterpenes. Functional annotation of 2,022 expressed sequence tags (ESTs) from the trichome cDNA library based on homology to A. thaliana genes suggested that 8% of the putative identified sequences were secondary metabolism-related and involved primarily in flavonoid and terpenoid biosynthesis. A significant proportion of the ESTs (38%) displayed no significant similarity to any other DNA deposited in databases, indicating a yet unknown function. Custom DNA microarrays constructed with 1,248 individual clones from the cDNA library facilitated transcriptome comparisons between trichomes and trichome-free tissues. In addition, gene expression studies in various Cistus tissues and organs for one of the genes highlighted as the most differentially expressed by the microarray experiments revealed a putative sesquiterpene synthase with a trichome-specific expression pattern. Full length cDNA isolation and heterologous expression in E. coli followed by biochemical analysis, led to the characterization of the produced protein as germacrene B synthase.

Synthesis and cytotoxic activities of 2-alkyl-2,3-dihydro-1H-2,6,9-triazacyclopenta[b]anthracene-5,10-diones

A series of 2-alkyl-2,3-dihydro-1H-2,6,9-triazacyclopenta[b]anthracene-5,10-diones (4a-f) were synthesized and their in vitro cytotoxic activities were evaluated against six human cancer cell lines (HCT15, SK-OV-3, A549, SNB19, MCF7 and MCF7/ADR). A number of compounds including 4c and 4d showed 2-180 times more potent cytotoxic activity than doxorubicin against all human cancer cell lines tested. Furthermore, these compounds retained considerable cytotoxic activity against the doxorubicin-resistant cell line MCF7/ADR, implying their therapeutic potential to treat doxorubicin-resistant tumors.

Bioactive metabolites from the endophytic fungus Ampelomyces sp. isolated from the medicinal plant Urospermum picroides

Extracts of cultures grown in liquid or on solid rice media of the fungal endophyte Ampelomyces sp. isolated from the medicinal plant Urospermum picroides exhibited considerable cytotoxic activity when tested in vitro against L5178Y cells. Chromatographic separation yielded 14 natural products that were unequivocally identified based on their 1H and 13C NMR as well as mass spectra and comparison with previously published data. Six compounds ( 2, 4, 5, 7, 9 and 11) were natural products. Both fungal extracts differed considerably in their secondary metabolites. The extract obtained from liquid cultures afforded a pyrone ( 2) and sulfated anthraquinones ( 7 and 9) along with the known compounds 1, 3, 6 and 8. When grown on solid rice medium the fungus yielded three compounds 4, 5 and 11 in addition to several known metabolites including 6, 8, 10, 12, 13 and 14. Compounds 4, 8 and 10 showed the strongest cytotoxic activity against L5178Y cells with EC 50 values ranging from 0.2–7.3 μg/ml. Furthermore, 8 and 10 displayed antimicrobial activity against the Gram-positive pathogens, Staphylococcus aureus, S. epidermidis and Enterococcus faecalis at minimal inhibitory concentrations (MIC) of 12.5 μg/ml and 12.5–25 μg/ml, respectively. Interestingly, 6 and 8 were also identified as constituents of an extract derived from a healthy plant sample of the host plant U. picroides thereby indicating that the production of bioactive natural products by the endophyte proceeds also under in situ conditions within the host plant.

Use of self-organizing maps and molecular descriptors to predict the cytotoxic activity of sesquiterpene lactones

Some sesquiterpene lactones (SLs) are the active compounds of a great number of traditionally medicinal plants from the Asteraceae family and possess considerable cytotoxic activity. Several studies in vitro have shown the inhibitory activity against cells derived from human carcinoma of the nasopharynx (KB). Chemical studies showed that the cytotoxic activity is due to the reaction of α,β-unsaturated carbonyl structures of the SLs with thiols, such as cysteine. These studies support the view that SLs inhibit tumour growth by selective alkylation of growth-regulatory biological macromolecules, such as key enzymes, which control cell division, thereby inhibiting a variety of cellular functions, which directs the cells into apoptosis. In this study we investigated a set of 55 different sesquiterpene lactones, represented by 5 skeletons (22 germacranolides, 6 elemanolides, 2 eudesmanolides, 16 guaianolides and nor-derivatives and 9 pseudoguaianolides), in respect to their cytotoxic properties. The experimental results and 3D molecular descriptors were submitted to Kohonen self-organizing map (SOM) to classify (training set) and predict (test set) the cytotoxic activity. From the obtained results, it was concluded that only the geometrical descriptors showed satisfactory values. The Kohonen map obtained after training set using 25 geometrical descriptors shows a very significant match, mainly among the inactive compounds (∼84%). Analyzing both groups, the percentage seen is high (83%). The test set shows the highest match, where 89% of the substances had their cytotoxic activity correctly predicted. From these results, important properties for the inhibition potency are discussed for the whole dataset and for subsets of the different structural skeletons.

Anti-cancer volatile constituents from the flowers of lonicera japonica thunb. Cultivated in Egypt

The volatile constituents of the flowers of Lonicera japonica Thunb. (Japanese honeysuckle), obtained by two methods [viz., floral absolute volatiles (FAV) and hydrodistilled/hexane extracted volatiles (HHV)], were comparatively investigated as regards composition and cytotoxicity. Hydrodistillation/hexane extraction was performed in Likens-Nickerson apparatus. The volatile constituents were identified and quantitated using GC/MS. Sixty compounds, 44 of which have not been previously reported in L. japonica, were identified among the volatiles prepared by the two methods. Linalool was the major constituent in both cases. However, its percentage is higher in the floral absolute volatiles (47.92%). On the other hand, 4-terpineol, nerolidol, cis-jasmone, cis-3-hexenyl tiglate, methyl palmitate and trans-linalool oxide, were detected in lesser but appreciable quantities in both samples. The volatiles prepared by the two methods were tested for their cytotoxic activities against four human cancer cell lines. The floral absolute showed a notable cytotoxic activity on the brain cancer cell line U251 (IC50 value of 7.92 μg/ml (and was found to be more potent than cisplatin reference drug (IC50 =8.66 μg/ml). Meanwhile, the volatiles prepared by hydrodistillation/hexane extraction showed a considerable cytotoxic activity on the liver cancer cell line Hepg2 with IC50 value of 7.58 μg/ml.Egyptian Journal of Biomedical Sciences Vol. 23 (1) 2007: pp. 135-145

Cytotoxic Activity of Gd(III)‐ and Dy(III)‐Complexes

The complexes of gadolinium(III) and dysprosium(III) were synthesized by reaction of the respective inorganic salts with 3,5-pyrazoledicarboxylic acid in amounts equal to metal to ligand molar ratio of 1 : 2. The structures of the final complexes were determined by means of spectral and elemental analyses. To help further the binding mode elucidation in the new Gd(III)- and Dy(III)-complexes of H(3)pdc, detailed vibrational analysis was performed on the basis of comparison of experimental vibrational spectra of the ligand and its Ln(III)-complexes using data theoretically predicted by us earlier, as well as data from the literature about related compounds. Significant differences in the IR and Raman spectra of the complexes were observed as compared to the spectra of the ligand. The ligand and the complexes were tested for their cytotoxic activities on the chronic myeloid leukemia-derived K-562, overexpressing the BCR-ABL fusion protein and the non-Hodgkin lymphoma-derived DOHH-2, characterized by a re-expression of the anti-apoptotic protein bcl-2 cell lines. The results obtained indicate that the tested compounds exerted a considerable cytotoxic activity upon the evaluated cell lines in a concentration-dependent manner, which enabled the construction of dose-response curves and the calculation of the corresponding IC(50) values. The inorganic salts exerted a very weak cytotoxic effect on these cells. This is in contrast to the lanthanide complexes, which exhibited potent cytotoxic activity, even more than the activity of cisplatin towards K-562 and DOHH-2 cell lines.

Structure elucidation of compounds extracted from the Chinese medicinal plant Patrinia heterophylla

For several hundred years, Patrinia heterophylla has been used in traditional Chinese medicine as a treatment for abscesses, hepatitis, tonsillitis, ulcers, etc. Recent research suggests that it may also have some anti-cancer activity. We have extracted five pure compounds from this plant; two known flavonols without bio-activity, one known isocoumarin glucoside that exhibits some cytotoxic activity toward HeLa cervical cancer cells, and two novel compounds that show considerable cytotoxic activity toward HeLa cells.

Quantitative structure–activity relationship of sesquiterpene lactones with cytotoxic activity

Some sesquiterpene lactones (SLs) are the active compounds of a great number of traditionally medicinal plants from the Asteraceae family and possess considerable cytotoxic activity. Several studies in vitro have shown the inhibitory activity against cells derived from human carcinoma of the nasopharynx (KB). In this study, we investigated a set of 37 different sesquiterpene lactones, represented by 4 skeletons (14 germacranolides, 6 elemanolides, 9 guaianolides and nor-derivatives, and 8 pseudoguaianolides), in what it says respect of their cytotoxic properties. The experimental results were submitted to a QSAR study. A single model for the entire data set was described using 3D molecular descriptors and genetic algorithms establishing structure–activity relationships among the compounds. Important properties for the inhibition potency are discussed for the whole data set and for subsets of the different structural skeletons.

New Lanthanum (III) Complex – Synthesis, Characterization, and Cytotoxic Activity

The complex of lanthanum (III) was synthesized reacting the respective inorganic salt with 5-aminoorotic acid in amounts equal to the metal:ligand molar ratio of 1:3. The complex was prepared by adding an aqueous solution of lanthanum (III) nitrate to an aqueous solution of the ligand, subsequently raising the pH of the mixture gradually to approx. 5.0 through addition of a dilute solution of sodium hydroxide. The structure of the final complex was determined by means of spectral data (IR, Raman,( 1)H-NMR) and elemental analysis. Significant differences in the IR spectrum of the complex were observed as compared to the spectrum of the ligand. A comparative analysis of the Raman spectrum of the complex with that of the free 5-aminoorotic acid allowed a straightforward assignment of the vibrations of the ligand groups involved in coordination. The ligand and the complex were tested for the cytotoxic activities on the chronic myeloid leukemia derived K-562, overexpressing the BCR-ABL fusion protein and the non-Hodgkin lymphoma derived DOHH-2, characterized by a re-expression of the anti-apoptotic protein bcl-2 cell lines. The results obtained indicate that the tested compounds exerted a considerable cytotoxic activity upon the evaluated cell lines in a concentration-dependent matter, which enabled the construction of dose-response curves and the calculation of the corresponding IC(50 )values. The inorganic salt exerted a very weak cytotoxic effect on these cells, which is in contrast to the lanthanum (III) complex.

Cytotoxic Abietane Diterpenes from Hyptis martiusii Benth.

From roots of Hyptis martiusii Benth. two tanshinone diterpenes were isolated, the new 7beta-hydroxy-11,14-dioxoabieta-8,12-diene (1) in addition to the known 7alpha-acetoxy-12-hydroxy-11,14-dioxoabieta-8,12-diene (7alpha-acetoxyroyleanone) (2). Structures of 1 and 2 were established by spectroscopic means. The cytotoxic activity against five cancer cell lines was evaluated. Compounds 1 and 2 displayed considerable cytotoxic activity against several cancer cell lines with IC50 values in the range of 3.1 to 11.5 microg/ml and 0.9 to 7.6 microg/ml, respectively. The cytotoxic activity seemed to be related to inhibition of DNA synthesis, as revealed by the reduction of 5-bromo-2'-deoxyuridine incorporation and induction of apoptosis, as indicated by the acridine orange/ethidium bromide assay and morphological changes after 24 h of incubation in leukemic cells.

New cerium(III) and neodymium(III) complexes as cytotoxic agents

The cerium(III) and neodymium(III) complexes of 5-aminoorotic acid were synthesized and characterized by means of spectral data (IR, Raman, 1H NMR and 13C NMR) and elemental analysis. Significant differences in the IR spectra of the complexes were observed as compared with the spectrum of the ligand. A comparative analysis of the Raman spectra of the complexes with that of the free 5-aminoorotic acid allowed a straightforward assignment of the vibrations of the ligand groups involved in coordination. 1H NMR and 13C NMR spectra confirmed the formation of the complexes. The ligand and the complexes were tested for the cytotoxic activities on the chronic myeloid leukemia-derived K-562, overexpressing the BCR-ABL fusion protein, and the non-Hodgkin lymphoma-derived DOHH-2, characterized by a rexpression of the antiapoptotic protein bcl-2 cell lines. The results obtained indicate that the tested compounds exerted a considerable cytotoxic activity upon the evaluated cell lines in a concentration-dependent manner, which enabled the construction of dose–response curves and the calculation of the corresponding IC50 values. Cytotoxicity towards tumor cells was determined for a broad concentration range. The inorganic salts exerted a very weak cytotoxic effect on these cells that is in contrast to the lanthanide complexes, which exhibited potent cytotoxic activity towards K-562 and DOHH-2 cell lines. Copyright © 2006 John Wiley & Sons, Ltd.

Bioconversion of methyl protodioscin by Penicillium melinii cells

Bioconversion of methyl protodioscin (1) by Penicillium melinii was investigated. Seven bioconversion products were isolated and identified. Three of them were glycosidation products and were new compounds. Their structures were identified to be 3- O-[α- l-rhamnopyranosyl-(1 → 2)-{α- l-rhamnopyranosyl-(1 → 4)}-β- d-glucopyranosyl]-26- O-[β- d-glucopyranosyl-(1 → 6)-β- d-glucopyranosyl]-25( R)-furost-5,20(22)-diene-3β,26-triol (2), 3- O-[α- l-rhamnopyranosyl-(1 → 2)-{α- l-rhamnopyranosyl-(1 → 4)}-β- d-glucopyranosyl]-26- O-[β- d-glucopyranosyl-(1 → 6)-β- d-glucopyranosyl]-25( R)-furost-5-ene-3β,26-triol (3), 16β-[4′-methyl-5′- O-(β- d-glucopyranosyl-(1 → 6)-β- d-glucopyranosyl)-pentanoxyl]-pregn-5-en-3β-ol-20-one- O-α- l-rhamnopyranosyl-(1 → 2)-[α- l-rhamnopyranosyl-(1 → 4)]-β- d-glucopyranoside (4), respectively. To the best knowledge we known, this was the first time to find the type of furostanol saponins that had disugar at C-26. The proposed biosynthetic pathways of methyl protodioscin were drawn. Most bioconversion products showed considerable cytotoxic activities against HepG2, NCI-H460, MCF-7 and HeLa cell lines compared to methyl protodioscin.

Characterization by NMR Spectroscopy, X‐ray Analysis and Cytotoxic Activity of the Ruthenium(II) Compounds [RuL3](PF6)2(L = 2‐Phenylazopyridine or o‐Tolylazopyridine) and [RuL'2L''](PF6)2(L', L'' = 2‐Phenylazopyridine, 2,2'‐Bipyridine)

AbstractTris(ligand) complexes [RuL3](PF6)2 (L = 2‐phenylazopyridine or o‐tolylazopyridine) and mixed ligand [RuL'2L''](PF6)2 (L' and L'' are 2‐phenylazopyridine or 2,2'‐bipyridine) have been synthesized, structurally characterized and investigated for cytotoxic activity. These complexes are important to study the hypothesis that the compound α‐[Ru(azpy)2Cl2] (azpy = 2‐phenylazopyridine) exhibits a high cytotoxicity due to its two cis chloride ligands, which might be exchanged for biological targets as DNA. Molecular structures of mer‐[Ru(azpy)3](PF6)2 (1) and mer‐[Ru(tazpy)3](PF6)2 (5) (tazpy = o‐tolylazopyridine) have been determined by X‐ray diffraction. Series of complexes [RuL3](PF6)2 and [RuL'2L''](PF6)2 show interesting NMR spectroscopic data; e.g. the spectrum of mer‐[Ru(azpy)3](PF6)2 (1) shows extremely broadened resonances at room temp. but sharpened resonances at low temperature. In the 1H NMR spectra of compounds [Ru(azpy)2(bpy)]2+ and [Ru(bpy)2(azpy)]2+ (bpy = 2,2‐bipyridine), respectively, less broadened (room temp.) or completely sharp resonances (room temp.) occur in comparison to 1 (under same conditions). By selecting the right temperature and/or concentration, NMR spectra of these series of compounds have been resolved using 2D COSY and NOESY NMR spectroscopy. Remarkably, the cytotoxicity data against a series of human tumor cell lines (A498, EVSA‐T, H226, IGROV, M19, MCF‐7 and WIDR) show a moderate cytotoxicity for these series of tris(ligand) complexes. So, even though no chloride ligands are present in these tris(ligand) complexes, a considerable cytotoxic activity is observed. This would imply that the 2‐phenylazopyridine ruthenium(II) complexes act by a completely different mechanism than the well‐known cisplatin. This finding is important, because an anticancer compound acting via a different mechanism is a prerequisite in designing new anticancer drugs. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2005)

Microbial transformation of methyl protodioscin by Cunninghamella elegans

Biotransformation of methyl protodioscin ( 1) by Cunninghamella elegans (AS 3.1207) was investigated. Nine bioconversion products were isolated and identified. Eight of the bioconversion products were pregnane glycoside or steroidal saponins. It was found that steroidal saponin skeleton could be converted to pregnenolone skeleton only using microbial methods, which must have chemical procedures in the reported literatures. The found enriched the types of bioconversion reaction and provided a new way for the production of androstenedione. Most bioconversion products showed considerable cytotoxic activities against HepG2, NCI-H460, MCF-7 and HeLa cell lines compared to methyl protodioscin.

A Cytotoxic Meroterpenoid Benzoquinone from Roots ofCordia globosa

(1a S*,1b S*,7a S*,8a S*)-4,5-Dimethoxy-1a,7a-dimethyl-1,1a,1b,2,7, 7a,8,8a-octahydrocyclopropa cyclopenta[1,2-b]naphthalene-3,6-dione (1), a new meroterpenoid benzoquinone, and microphyllaquinone (2), a known naphthoquinone, have been isolated from roots of Cordia globosa. Both structure determinations were performed by conventional spectroscopic methods, including inverse detection NMR techniques, and by comparison with data from the literature for related compounds. Compound 1 displayed considerable cytotoxic activity against several cancer cell lines with IC50 values in the range of 1.2 to 5.0 microg/mL. The cytotoxic activity seemed to be related to DNA synthesis inhibition, as revealed by the reduction of 5-bromo-2'-deoxyuridine incorporation, and apoptosis induction, as indicated by the acridine orange/ethidium bromide assay and morphological changes after 24 h of incubation on leukemic cells.

Antitumor Activity of Pulsatilla koreana Saponins and Their Structure-Activity Relationship

Seventeen saponins isolated from the root of Pulsatilla koreana were examined for their in vitro cytotoxic activity against the human solid cancer cell lines, A-549, SK-OV-3, SK-MEL-2, and HCT15, using the SRB assay method, and their in vivo antitumor activity using BDF1 mice bearing Lewis lung carcinoma (LLC). The saponins 5-17, with a free acidic functional group at C-28 of aglycon, exhibited moderate to considerable cytotoxic activity, however, the saponins 1-4, esterified with a trisaccharide at C-28 of aglycon, did not exhibit cytotoxic activity (ED50; >300 microM). Among them, oleanolic acid 3-O-alpha-L-rhamnopyranosyl-(1-->2)-[beta-D-glucopyranosyl-(1-->4)]-alpha-L-arabinopyranoside (10) exhibited the most potent cytotoxic activity (ED50; 2.56, 2.31, 1.57, 8.36 microM, respectively). In vivo test, hederagenin 3-O-alpha-L-rhamnopyranosyl-(1-->2)-[beta-D-glucopyranosyl-(1-->4)]-alpha-L-arabinopyranoside (6, Inhibition Ratio, IR; 66.9%) exhibited more potent antitumor activity than taxol (IR; 35.8%) and doxorubicin (IR; 62.1%). Also, hedragenin 3-O-beta-D-glucopyranosyl-(1-->4)-O-beta-D-glucopyranosyl-(1-->3)-O-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside (17, IR; 50.3%) exhibited potent antitumor activity. These two saponins were identically comprised of a hederagenin aglycon moiety and a sugar sequence O-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranoside at C-3 of the hederagenin, suggesting that the two elements are essential factors for the antitumor activity.

Cytotoxic effect of a new 1,3,4-thiadiazolium mesoionic compound (MI-D) on cell lines of human melanoma.

The structural characteristics of mesoionic compounds, which contain distinct regions of positive and negative charges associated with a poly-heteroatomic system, enable them to cross cellular membranes and interact strongly with biomolecules. Potential biological applications have been described for mesoionic compounds. 1,3,4-Thiadiazolium mesoionic compound (MI-D), a new mesoionic compound, has been demonstrated to be extremely cytotoxic to B16-F10 murine melanoma cells when compared to fotemustine and dacarbazine, drugs of reference in melanoma treatment protocols, describing inhibition of tumours grown in vitro and in vivo. We now evaluate the effects of mesoionic compound MI-D on different human melanoma cell lines. The drug decreased the viability and proliferation of MEL-85, SK-MEL, A2058 and MEWO cell lines in vitro, showing a considerable cytotoxic activity on these human cells. Adhesion of MEL-85 cells was evaluated in the presence of the drug using different extracellular matrix (ECM) constituents. MI-D decreased MEL-85 adhesion to laminin, fibronectin and matrigel. The morphology and actin cytoskeleton organisation of MEL-85 cells were also modified on MI-D treatment. These results on human melanoma cell lines indicate that MI-D is a very encouraging drug against melanoma, a tumour that is extremely resistant to chemotherapy.

Synthesis and in vitro Evaluation of 1,8‐Diazaanthraquinones Bearing 3‐Dialkylaminomethyl or 3‐(N‐Alkyl‐ or N‐Aryl)carbamoyloxymethyl Substituent.

AbstractFor Abstract see ChemInform Abstract in Full Text.

CD8(+) T cells with parasite-specific cytotoxic activity and a Tc1 profile of cytokine and chemokine secretion develop in experimental visceral leishmaniasis.

Recently, a prominent role for CD8(+) T cells in immunity against pathogens has emerged. The mode of action of CD8(+) T cells in murine visceral leishmaniasis and their contribution to the clearance of the parasite has been addressed in the present study. We showed that during the course of experimental infection cytotoxic clones specific for Leishmania infantum antigens developed in the spleen of susceptible BALB/c mice, showed an activated phenotype and became susceptible to apoptotic cell death late in the course of the disease. CD8(+) T cells exhibited considerable cytotoxic activity against cells expressing Leishmania antigens. This activity was mediated by both the perforin and the Fas/FasL pathway, as judged from in vitro and in vivo assays. The CD8(+) T cells also up-regulated mRNAs for cytokines (IFN-gamma and TNF-alpha) and C-C chemokines (RANTES and MIP-1alpha), which have a major role in immunity against the pathogen. CD8(+) T-cells thus displayed a Tc1 pattern of differentiation. In conclusion, CD8(+) T cells appear to play multiple roles in an experimental model of visceral leishmaniasis comprising both cytotoxic activity and secretion of cytokines and chemokines.