• Toxicological Research
• /
• 제12권2호
• /
• pp.223-230
• /
• 1996

The increased embryotoxicity of the antifolate drug pyrimethamine (PYM) with concomitant dietary dosing of folic acid (FA) was examined in mice. The preventive effects of folinic acid (FNA) on PYM embryotoxicity were also examined. Six groups were constructed: PYM I (pyrimethamine 80 ppm), PYM II (pyrimethamine 150 ppm), PYM II+FNA (pyrimethamine 150 ppm and folinic acid 12 mg/kg/day), PYM II+FA (pyrimethamine 150 ppm and folic acid 350 ppm), FA (folic acid 350 ppm) and a control group. The agents were administered for 7 days from day 6 throughout 12 of gestation. PYM and FA were administered with mashed feed and FNA was intraperitoneally injected. The high incidence of fetal realformations was observed in the PYM II group; these included kinky tail, open eyelids, club foot, cleft palate, absence of the pulmonary lobe, diaphragmatic hernia, fused sternebrae, fused cervical or thoracic vertebral arch, among others. All embryos of the PYM II+FA group were resorbed. No realformed fetuses were observed in the PYM II+FNA group. These results show that the concomitant dosing of FA augments PYM embryotoxicity. The preventive effects of FNA were also observed.

• Bulletin of the Korean Chemical Society
• /
• 제33권8호
• /
• pp.2731-2736
• /
• 2012

The use of pyrimethamine as antibacterial drug is limited by the poor solubility. To enhance its solubility, we prepared complexes of pyrimethamine with low-molecular-weight succinoglycan isolated from Sinorhizobium meliloti. Low-molecular-weight succinoglycans are monomers, dimers, and trimers of the succinoglycan repeating unit. The monomers and dimers were separated into their three species (M1, M2, and M3) and four fractions (D1 to D4) using chromatographic techniques, which were shown to be nontoxic. The solubility of pyrimethamine was markedly increased up to 42 fold by succinoglycan D3, where the level of its solubility enhancement was even 8-20 fold higher comparing with cyclodextrin or its derivatives. The complex formation of succinoglycan D3 with pyrimethamine was confirmed by $^1H$ nuclear magnetic resonance spectroscopy, Fourier-transform infrared spectroscopy, differential scanning calorimetry, scanning electron microscopy, and molecular modeling studies. Herein, we suggest that the low-molecular-weight succinoglycans may be utilized as highly effective solubilizers of pyrimethamine for pharmaceutical purposes.

• Parasites, Hosts and Diseases
• /
• 제32권1호
• /
• pp.19-26
• /
• 1994

Toxoplasma 약독주(ME49 주)의 배양계 확립을 위하여 뇌내 cyst를 마우스 복강에 주입하고 1, 3, 5 그리고 7일후 다시 얻어 배양기 부착성 세포를 배양하는 방법으로 ME49주를 배양하였으며, ME49주의 성장에 미치는 CAMP 및 DHFR억제제의 영향을 관찰하였다. ME49주로 감염된 대식 세포의 형태학적 관찰은 Giemsa 염색방법을 이용하였고 성장정도는 $[^3H]-uracil$ 표지량을 대조군에 대한 비로 나타내었다. 감염 3일 및 5일이 경과된 후에 채취한 복수의 대식세포에서 ME49 주의 bradyzoite가 주로 판찰되었으며, 배양기에서 3일 이상 경과된 후에는 pseudocyst를 형성하기 시작하였고, 5일 10일이 경과되면서 pseudocyst의 크기가 증가하였다. CAMP를 농도별로 처리하였을 때 3일째와 5일째의 복수를 5일간 및 10일간 배양했을 때 농도의존적으로 성장을 촉진하였다. DHFR 억제제중 pyrimethamine의 경우 농도의존적인 성장억제효과를 나타냈고 methotrexate의 경우엔 ME49 주 bradyzoite의 성장에 영향을 미치지 않는 것으로 나타났다. 이상의 결과로 마우스 대식세포내에서 bradyzoite의 배양이 가능하고 그 배양조건은 3일째와 5일째 복수를 5일 이상 10일 정도 배양하는 것이 적당하며 CAMP 및 pyrimethamine이 bradyBoite의 성장을 각각 촉진 및 억제하는 것으로 나타났다.

• Parasites, Hosts and Diseases
• /
• 제49권2호
• /
• pp.125-131
• /
• 2011

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.

• Parasites, Hosts and Diseases
• /
• 제60권2호
• /
• pp.109-116
• /
• 2022

Drug resistance is an important problem hindering malaria elimination in tropical areas. Point mutations in Plasmodium falciparum dihydrofolate reductase (Pfdhfr) and dihydropteroate synthase (Pfdhps) genes confer resistance to antifolate drug, sulfadoxine-pyrimethamine (SP) while P. falciparum chloroquine-resistant transporter (Pfcrt) genes caused resistance to chloroquine (CQ). Decline in Pfdhfr/Pfdhps and Pfcrt mutations after withdrawal of SP and CQ has been reported. The aim of present study was to investigate the prevalence of Pfdhfr, Pfdhps, and Pfcrt mutation from 2 endemic areas of Thailand. All of 200 blood samples collected from western area (Thai-Myanmar) and southern area (Thai-Malaysian) contained multiple mutations in Pfdhfr and Pfdhps genes. The most prevalent haplotypes for Pfdhfr and Pfdhps were quadruple and double mutations, respectively. The quadruple and triple mutations of Pfdhfr and Pfdhps were common in western samples, whereas low frequency of triple and double mutations was found in southern samples, respectively. The Pfcrt 76T mutation was present in all samples examined. Malaria isolated from 2 different endemic regions of Thailand had high mutation rates in the Pfdhfr, Pfdhps, and Pfcrt genes. These findings highlighted the fixation of mutant alleles causing resistance of SP and CQ in this area. It is necessary to monitor the re-emergence of SP and CQ sensitive parasites in this area.

• Parasites, Hosts and Diseases
• /
• 제55권5호
• /
• pp.491-503
• /
• 2017

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 ($5{\mu}M$) at $20{\mu}M$ and higher, without affecting host cells. Group III TKIs (neratinib, dacomitinib, afatinib, and pelitinib) inhibited proliferation up to 98% equivalent to pyrimethamine at $1-5{\mu}M$, but host cells were destroyed at $10-20{\mu}M$. 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.

• Parasites, Hosts and Diseases
• /
• 제36권3호
• /
• pp.191-198
• /
• 1998

톡소포자충의 pyrimethamine (PBM)과 methotrexate (MYX)에 대한 효소활성 및 유전자 발현 의 변화를 살펴본 결과 두 약제를 처리한 경우 모두 약제 농도의 증가에 따라 생존력이 감소하였으며, 계대가 진행에 따라 DHFR 효소활성은 증가하였다. PBM의 경우, 0.01 mM을 사용한 1차 계대시 효소 활성이 33.9%를 나타낸 반면, 10배로 농도를 증가시킨 4차 계대에서는 63.5%로 약 2배 증가하였다. WIBC의 경우에는 1차 계대시 77.4% ($1{\;}{\mu\textrm{m}}$), 4차 계대시 82.2% ($10{\;}{\mu\textrm{m}}$), 7 차 계대시에는 141.3% ($100{\;}{\mu\textrm{m}}$)로 1차 계대와 비교하여 볼 때 효소활성이 약 183% 증가한 것으로 나타났다. 특히 DHFRWIRNA의 양은 4차 계대시에 1차 계대시와 비교하였을때, PBM 농도 증가에 따라 각각 1. 5배 이상씩 증가하였으며, WRC의 경우 역시 rnRNA양의 변화는 관찰되었으나, 그 변화에 따른 유의성은 없었다. 이상의 결과로 PlM에 대한 톡소포자충의 반응은 DHFR 효소환성 증가 및 mRNA 수준 즉, transcriptional love떼서 조절되는 것임을 알 수 있었으며, Mn(에 대한 톡소포자충의 반응은 DHFR 효소활성 증가에 기인하나, PW에 의해 유발되는 반응과는 다른 약제 극복 기전이 존재하는 것으로 추정되었다.

• Parasites, Hosts and Diseases
• /
• 제54권2호
• /
• pp.155-161
• /
• 2016

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.

• Parasites, Hosts and Diseases
• /
• 제28권2호
• /
• pp.79-84
• /
• 1990

Pyrimidine salvage과정 및 de novo 합성과정에 작용하는 억제제들이 Toxeplasma gondii의 특이한 uracil 도입 과정에 미치는 영향을 방사능을 표지한 uracil과 thymidine을 사용하여 검토하였다. Dihydrofolate reductase (DHFR)에 작용하는 억제제인 methotreBate, pyrimethamine 그리고 thymidylate synthase(75)의 경쟁적 억제제인 fluoro-uridine, fluoro-dUMP, fluoro-uracil을 각각 처리한 경우, 그 농도에 비례적으로 uracil 표지가 감소 하였다. OMP decarboxylase에 작용하는 억제제인 azauridine에 대해서는 100$\mu$M 농도 이상에서도 uracil 표지량에 변화가 거의 없었다. 본 결과로부터 Toxoplasma가 uracil을 DNA로 도입할 때 dihydrofolate reductase와 thymidylate synthase가 관여하는 TMP biosynthesis 과정이 중요함을 알 수 있었고, uracil salvage 과정이 있는 다른 기생성 원충의 경우와 비교했을 때 Toxoplasmn에 있어서도 효율적인 다기능의 DHFR-75 system의 존재를 예상할 수 있었다. Thymidine 도입의 양상은 모든 경우에 있어 HL-60세포만의 경우와 Toxoplasma가 함께 배양된 HL-60세포에서 차이가 없었다. 이로부터 thymidine은 Toxoplasmn 성장을 반영하지 않음을 알 수 있었다. Toxoplasma의 특이한 uracil 표지와 Toxoplasma에 배타적인 thymidine 표지는 기생원충과 숙주세포의 성장, DNA합성 정도 및 이들 과정에 미치는 제제들의 영향을 쉽고 간단하게 연구할 수 있는 방법이 된다.

• 대한약침학회지
• /
• 제20권3호
• /
• pp.220-226
• /
• 2017

Objectives: The currently available agents for use against toxoplasmosis have serious limitations. Thus, the aim of the present study was to investigate the anti-Toxoplasma gondii (T. gondii) activities of methanol extracts of Feijoa sellowiana (F. sellowiana) (leaves and fruits), Quercus castaneifolia (Q. castaneifolia) (fruits), and Allium paradoxum (A. paradoxum) (leaves) in vitro and in vivo. Methods: Vero cells were treated with different concentrations (from 0 to $400{\mu}g/mL$) of the above extracts or with pyrimethamine at a dose of 50 mg/mL (positive control). Then, the viabilities of the T. gondii-infected cells were measured by using colorimetric MTT (3-(4, 5-dimethylthiazol-2-yl) 2, 5-diphenyltetrazolium bromide) assays. In addition, the survival rates of mice acutely infected with $2{\times}10^4$ RH strain tachyzoites of T. gondii were examined in vivo after intraperitoneal injection of the extracts at doses of 100 and 200 mg/kg/day for 5 days. Results: In the in vitro anti- T. gondii assay, the $IC_{50}$ values were 12.77, 180.2, 74.73, 213.2 and $163.8{\mu}g/mL$, and the selectivity indices were 6.05, 1.31, 0.35, 0.69 and 1.30 for the F. sellowiana (leaves and fruits), Q. castaneifolia, and A. paradoxum extracts and pyrimethamine, respectively. Moreover, the mice treated with F. sellowiana (leaves and fruits) achieved better results in terms of survival than the others (P < 0.05). Conclusion: The results of the current study indicate that methanol extract of F. sellowiana has significant anti-Toxoplasma activity. Further study should be conducted to investigate the potential bioactivity of this extract through bioactivity-guided fractionation.