The effects of tyrosine kinase inhibitors (TKIs) were evaluated on growth inhibition of intracellular Toxoplasma gondii in host ARPE-19 cells. The number of tachyzoites per parasitophorous vacuolar membrane (PVM) was counted after treatment with TKIs. T. gondii protein expression was assessed by western blot. Immunofluorescence assay was performed using Programmed Cell Death 4 (PDCD4) and T. gondii GRA3 antibodies. The TKIs were divided into 3 groups; non-epidermal growth factor receptor (non-EGFR), anti-human EGFR 2 (anti-HER2), and anti-HER2/4 TKIs, respectively. Group I TKIs (nintedanib, AZD9291, and sunitinib) were unable to inhibit proliferation without destroying host cells. Group II TKIs (lapatinib, gefitinib, erlotinib, and AG1478) inhibited proliferation up to 98% equivalent to control pyrimethamine (5 μM) at 20 μM and higher, without affecting host cells. Group III TKIs (neratinib, dacomitinib, afatinib, and pelitinib) inhibited proliferation up to 98% equivalent to pyrimethamine at 1–5 μM, but host cells were destroyed at 10–20 μM. In Group I, TgHSP90 and SAG1 inhibitions were weak, and GRA3 expression was moderately inhibited. In Group II, TgHSP90 and SAG1 expressions seemed to be slightly enhanced, while GRA3 showed none to mild inhibition; however, AG1478 inhibited all proteins moderately. Protein expression was blocked in Group III, comparable to pyrimethamine. PDCD4 and GRA3 were well localized inside the nuclei in Group I, mildly disrupted in Group II, and were completely disrupted in Group III. This study suggests the possibility of a vital T. gondii TK having potential HER2/4 properties, thus anti-HER2/4 TKIs may inhibit intracellular parasite proliferation with minimal adverse effects on host cells.
Antibodies ; Blotting, Western ; Cell Death ; Epidermal Growth Factor* ; Erlotinib Hydrochloride ; Fluorescent Antibody Technique ; Humans* ; Membranes ; Parasites ; Protein-Tyrosine Kinases ; Pyrimethamine ; Receptor, Epidermal Growth Factor* ; Toxoplasma

Toxoplasma gondii is an important opportunistic pathogen that causes toxoplasmosis, which has very few therapeutic treatment options. The most effective therapy is a combination of pyrimethamine and sulfadiazine; however, their utility is limited because of drug toxicity and serious side effects. For these reasons, new drugs with lower toxicity are urgently needed. In this study, the compound, (Z)-1-[(5-nitrofuran-2-yl)methyleneamino]-imidazolidine-2,4-dione (nitrofurantoin), showed anti-T. gondii effects in vitro and in vivo. In HeLa cells, the selectivity of nitrofurantoin was 2.3, which was greater than that of pyrimethamine (0.9). In T. gondii-infected female ICR mice, the inhibition rate of T. gondii growth in the peritoneal cavity was 44.7% compared to the negative control group after 4-day treatment with 100 mg/kg of nitrofurantoin. In addition, hematology indicators showed that T. gondii infection-induced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, biochemical parameters involved in liver injury, were reduced by nitrofurantoin significantly. Moreover, nitrofurantoin exerted significant effects on the index of antioxidant status, i.e., malondialdehyde (MDA) and glutathione (GSH). The nitrofurantoin-treated group inhibited the T. gondii-induced MDA levels while alleviating the decrease in GSH levels. Thus, nitrofurantoin is a potential anti-T. gondii candidate for clinical application.
Alanine Transaminase ; Animals ; Aspartate Aminotransferases ; Drug-Related Side Effects and Adverse Reactions ; Female ; Glutathione ; HeLa Cells ; Hematology ; Humans ; Liver ; Malondialdehyde ; Mice ; Mice, Inbred ICR ; Nitrofurantoin* ; Peritoneal Cavity ; Pyrimethamine ; Sulfadiazine ; Toxoplasma ; Toxoplasmosis*

OBJECTIVETo compare the protein patterns from the extracts of the mutant clone T9/94-M1-1(b3) induced by pyrimethamine, and the original parent clone T9/94 following separation of parasite extracts by two-dimensional electrophoresis (2-DE).

METHODSProteins were solubilized and separated according to their charges and sizes. The separated protein spots were then detected by silver staining and analyzed for protein density by the powerful image analysis software.

RESULTSDifferentially expressed protein patterns (up- or down-regulation) were separated from the extracts from the two clones. A total of 223 and 134 protein spots were detected from the extracts of T9/94 and T9/94-M1-1(b3) clones, respectively. Marked reduction in density of protein expression was observed with the extract from the mutant (resistant) clone compared with the parent (sensitive) clone. A total of 25 protein spots showed at least two-fold difference in density, some of which exhibited as high as ten-fold difference.

CONCLUSIONSThese proteins may be the molecular targets of resistance of Plasmodium falciparum to pyrimethamine. Further study to identify the chemical structures of these proteins by mass spectrometry is required.

Antimalarials ; metabolism ; Drug Resistance ; Electrophoresis, Gel, Two-Dimensional ; Humans ; Image Processing, Computer-Assisted ; Mutation ; Plasmodium falciparum ; chemistry ; drug effects ; genetics ; Proteome ; analysis ; Protozoan Proteins ; analysis ; Pyrimethamine ; metabolism ; Staining and Labeling

The use of sulfadoxine and pyrimethamine (SP) for treatment of vivax malaria is uncommon in most malarious areas, but Plasmodium vivax isolates are exposed to SP because of mixed infections with other Plasmodium species. As P. vivax is the most prevalent species of human malaria parasites in Iran, monitoring of resistance of the parasite against the drug is necessary. In the present study, 50 blood samples of symptomatic patients were collected from 4 separated geographical regions of south-east Iran. Point mutations at residues 57, 58, 61, and 117 were detected by the PCR-RFLP method. Polymorphism at positions 58R, 117N, and 117T of P. vivax dihydrofolate reductase (Pvdhfr) gene has been found in 12%, 34%, and 2% of isolates, respectively. Mutation at residues F57 and T61 was not detected. Five distinct haplotypes of the Pvdhfr gene were demonstrated. The 2 most prevalent haplotypes were F57S58T61S117 (62%) and F57S58T61N117 (24%). Haplotypes with 3 and 4 point mutations were not found. The present study suggested that P. vivax in Iran is under the pressure of SP and the sensitivity level of the parasite to SP is diminishing and this fact must be considered in development of malaria control programs.
Amino Acid Substitution/genetics ; Antimalarials/*pharmacology ; Drug Combinations ; *Drug Resistance ; Haplotypes ; Humans ; Iran ; Malaria, Vivax/*parasitology ; *Mutation, Missense ; Plasmodium vivax/*enzymology/genetics/isolation & purification ; Polymorphism, Genetic ; Pyrimethamine/*pharmacology ; Sulfadoxine/*pharmacology ; Tetrahydrofolate Dehydrogenase/*genetics

Toxoplasmosis is a rare but fatal complication in hematopoietic stem cell transplant recipients, usually associated with allogeneic hematopoietic stem cell transplantation (HSCT). We report a case of cerebral toxoplasmosis in a patient with multiple myeloma, following tandem autologous stem cell transplantation. A 55-year-old Korean male presented with weakness in both legs that had progressed to both arms. A magnetic resonance imaging scan of the brain revealed multiple, variable-sized ring-enhancing lesions with surrounding edema in the cerebral hemispheres and brain stem. Stereotactic biopsy revealed bradyzoites of Toxoplasma gondii in the brain tissue. The patient received trimethoprim-sulfamethoxazole, followed by pyrimethamine and sulfadiazine, accompanying treatment for progressive multiple myeloma. Cerebral toxoplasmosis should be considered as one of the differential diagnoses in patients with neurologic signs following autologous HSCT.
Arm ; Biopsy ; Brain ; Brain Stem ; Cerebrum ; Diagnosis, Differential ; Edema ; Hematopoietic Stem Cell Transplantation ; Hematopoietic Stem Cells ; Humans ; Leg ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Multiple Myeloma ; Neurologic Manifestations ; Pyrimethamine ; Stem Cell Transplantation ; Stem Cells ; Sulfadiazine ; Toxoplasma ; Toxoplasmosis ; Toxoplasmosis, Cerebral ; Trimethoprim, Sulfamethoxazole Drug Combination

In 1998-2002 year period, those studies were developed in LaoCai, LaiChau, QuangTri, Daklak, GiaLai and KonTum. Malarial patients with P. falciparum without complication were administered orally by a total dose of Chloroquine 25mg/kg in 3 day treating course or an unique dose of sulfadoxin-pyrimethamin.The use of medicines was observed and the parasitic density was monitored within 28 days after the treatment. Results showed the failure rate at almost studied locations
Malaria ; Therapeutics ; Pharmaceutical Preparations ; Malaria, Falciparum ; Drug Resistance ; Chloroquine ; Pyrimethamine

PCR-RFLP and DNA sequencing techniques were used for Pyrimethamine (PYR) and Cycloguanil (CYC) resistant mutations. The study results showed that 46 of 58 Plasmodium falciparum isolates were genotype resistant to pyrimethamine (79.31%). Thirty-one isolates were taken in vitro microscopic examination and revealed that 26 of 31 isolates were PYR resistance (83.87%) and 6 of 31 isolates of CYC resistance (19.35%). The resistant levels of these isolates were related to the mutations in DHFR genes
Plasmodium falciparum ; Pyrimethamine ; Chloroguanide ; malaria

The efficacy of chloroquine or sulfadoxine-pyrimethamine given combined with artesunate was assessed in Vietnamese patients with uncomplicated falciparum malaria from 2 Southern provinces, where there was in vitro evidence that the sensitivity of the parasite to conventional antimalarial therapies had returned in the absence of drug pressure, from October to December of 2000. In Dak Lak province, 57 patients (mean age 9.6 years) were randomized to artesunate-chloroquine (group 1) or artesunate-sulfadoxine/pyrimethamine (group 2). In Binh Phuoc province, 66 patients, who have just migrated in period of 1-7 years (mean age 24.2 years) were assessed with the 2 regimens. The results of 28 days in vivo response were over 96% and lower 52% of Dak Lak and Binh Phuoc respectively. PCR evidence of cure closely paralleled the in vivo results. The successful reintroduction of chloroquine and sulfadoxine-pyrimethamine as artemisinin partner drugs depends heavily on epidemiological and parasite factors
malaria ; Malaria, Falciparum ; Therapeutics ; Pharmaceutical Preparations ; artesunate ; Chloroquine ; Pyrimethamine

Since the national malaria eradication service (NMES) was implemented in 1959, extensive malariometric survey were conducted throughout the country. A total of 73,859 malaria cases of Plasmodium vivax were confirmed out of 268,259 blood smears through passive and active case detection (PCD and ACD) in 1960~1969 including chloroquine/ pyrimethamine combined tablets for the presumptive treatment of all of fever cases. The majority of the positive cases occurred in Gyeongsang buk-do, revealing 62,859 cases as of 85.2%, followed by Gyeonggi-do 5,844 cases (7.9%), Chungcheong buk-do 3,458 cases (4.7%), Gangwon-do 812 cases (1.1%), and 29 cases developed in Jeju-do. ACD was more sensitive and advantageous for detecting the malaria cases over PCD in the pilot study area. The high endemic area was located in the north Gyeongsang buk-do and north east Gyeonggi-do. The malaria incidence was high in the age group under 14 years in malaria foci, and that was low in non-malarious area. High parous rate (77.7%) and survival rate (91.4%) of the vector mosquitoes were shown in malaria foci, which were hilly or mountainous area. On the other hand, low parous rate (50.6%) and survival rate (42.7%) were found in non-malarious area, the rice field of south western part of the country even in abundance of vector mosquitoes. NMES had contributed the control of the malaria by carring out the residual DDT focal spray, mass drug administration, radical treatment of the positive cases, and mass blood survey in the heavy malarious area. Since then malaria disappeared rapidly, and no indigenous case was reported after 1984. In order to ascertain whether this country was malaria-free or not, mass blood survey was carried out for the juveniles, adolescents and adults in Yeongju-gun Gyeongsangbuk-do in 1986 where the highest incidence of malaria was recorded during the 1960s. The result showed no positive case at all. However vivax malaria reemerged, and made a dramatic resurgence since 1993 in the demilitarized zone (DMZ) bordering North Korea, and expanded toward eastern and southern part of the country year after year, totaling 14,089 cases including U.S. army by the end of 2000 in spite of the strong activities of antimalaria in the Republic of Korea. So vector control including the encouragement of the livestock farming in the malaria endemic area must be more sensitive and effective measure to reduce the risk of malaria transmission by the eliminating the dangerous age of mosquitoes in the points of characteristics of the Korean strain of parasites such as the mild symptoms, long-term incubation period and asymptomatic parasitaemia. Focal application of thermal fogging and mist spray of insecticides would be certainly effective in preventing the vector mosquitoes from infecting the gametocytes of malaria for the first peak of multi malaria cases in the early season of the year. All of the civilians who are in the high risk area for malaria should be forced to sleep under the permethrin impregnated mosquito nets. All of the soldiers near the DMZ are strongly recommended to take the combined tablets of chloroquine/ pyrimethamine instead of the single base of chloroquine at present to prevent the action of source of infection of the reservoirs and the completion of the sporogonic stages of the vector mosquitoes because the vector control activities are not allowed in the DMZ. The experts of entomologists and malariologist of the South and North Korea should discuss the antimalaria programme to eradicate the malaria from the Korean peninsular near future.
Adolescent ; Adult ; Chloroquine ; Culicidae ; DDT ; Democratic People's Republic of Korea ; Epidemiology ; Fever ; Gangwon-do ; Gyeonggi-do ; Gyeongsangbuk-do ; Hand ; Humans ; Incidence ; Insecticides ; Jeju-do ; Korea* ; Livestock ; Malaria* ; Malaria, Vivax ; Military Personnel ; Mosquito Nets ; Parasites ; Permethrin ; Pilot Projects ; Plasmodium vivax ; Pyrimethamine ; Republic of Korea ; Seasons ; Survival Rate ; Tablets ; Weather

Vivax malaria, which was once thought to be eradicable since more than 20 years ago, has made a dramatic resurgence since 1993 in the northern parts of Kyeonggi-do in the background of 0.13 of the annual parasite incidence in the population of 956,773 and 2 positives (0.07%) of asymptomatic parasitaemia among the 2,937 residents of mass blood survey in the area in 1969. The majority of the cases were soldiers infected in several counties located within 5 km from the Demilitarized Zone (DMZ) in 1994. Since then, malaria cases have increased and have tended to expand toward eastern and southern areas. Korean malaria is characterized by asymptomatic parasitaemia with long prepatent period, so malaria cases, including those manifesting as general fever should be treated with a combination of chloroquine and pyrimethamine to prevent the completion of sporogonic cycle in the mosquitoes. For the most effective control of vector mosquitoes, residual insecticides should be applied in the military houses and animal shelters with thermal fogging, including the control of the pre-stages of physiological age of the mosquitoes within 25 km from the DMZ. It is strongly suggested that military uniforms be treated with 0.5% permethrin complex (permethrin 39%+cyfluthrin 1%) or permethrin formulation. Further genetic studies are desired to determine the subspecies of Plasmodium vivax because Korean strain of malaria is likely to be a complex form of Plasmodium vivax vivax and Plasmodium vivax hibernans.
Animals ; Chloroquine ; Culicidae ; Fever ; Gyeonggi-do ; Humans ; Incidence ; Insecticides ; Korea* ; Malaria* ; Malaria, Vivax ; Military Personnel ; Parasites ; Permethrin ; Plasmodium vivax ; Pyrimethamine ; Weather