Pyrimethamine upregulates BNIP3 to interfere SNARE-mediated autophagosome-lysosomal fusion in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of the most common tumor types and remains a major clinical challenge. Increasing evidence has revealed that mitophagy inhibitors can enhance the effect of chemotherapy on HCC. However, few mitophagy inhibitors have been approved for clinical use in humans. Pyrimethamine (Pyr) is used to treat infections caused by protozoan parasites. Recent studies have reported that Pyr may be beneficial in the treatment of various tumors. However, its mechanism of action is still not clearly defined. Here, we found that blocking mitophagy sensitized cells to Pyr-induced apoptosis. Mechanistically, Pyr potently induced the accumulation of autophagosomes by inhibiting autophagosome-lysosome fusion in human HCC cells. In vitro and in vivo studies revealed that Pyr blocked autophagosome-lysosome fusion by upregulating BNIP3 to inhibit synaptosomal-associated protein 29 (SNAP29)-vesicle-associated membrane protein 8 (VAMP8) interaction. Moreover, Pyr acted synergistically with sorafenib (Sora) to induce apoptosis and inhibit HCC proliferation in vitro and in vivo. Pyr enhances the sensitivity of HCC cells to Sora, a common chemotherapeutic, by inhibiting mitophagy. Thus, these results provide new insights into the mechanism of action of Pyr and imply that Pyr could potentially be further developed as a novel mitophagy inhibitor. Notably, Pyr and Sora combination therapy could be a promising treatment for malignant HCC.

Comparison of protein patterns between Plasmodium falciparum mutant clone T9/94-M1-1(b3) induced by pyrimethamine and the original parent clone T9/94

Objective:To 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).Methods:Proteins 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.Results:Differentially 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.Conclusions:These 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.

Effectiveness of non-pyrimethamine-based regimens for toxoplasma encephalitis:A systematic and meta-synthesis study

Objective:To examine the differences in effectiveness and side effects between pyrimethamine-based and non-pyrimethamine-based regimens for toxoplasma encephalitis since the availability of pyrimethamine in Indonesia is currently limited due to its withdrawal from the market.Methods:A systematic review and meta-synthesis study that was carried out by following a protocol guided by the Preffered Reporting Items for Systematic Review and Meta-analysis(PRISMA).Effectiveness measures included clinical improvement,mortality,and radiological improvement.We evaluated selected articles narratively because of the limitations of homogeneity.The risk of bias in RCTs was assessed using the Cochrane Risk of Bias tool for RCT(ROB 2.0)and cohort studies were assessed using the Risk of Bias In Non-Randomized Studies of Interventions(ROBINS-1)tool.Research quality was assessed using the GradePro software.Results:We included two retrospective cohort studies and one RCT.Narrative outcome assessment in these three studies did not show significant difference in effectiveness between pyrimethamine-based and non-pyrimethamine-based regimens for toxoplasma encephalitis treatment.However,drug side effects were consistently higher in the pyrimethamine-based regimen.Conclusions:This study has a high risk of bias.The quality of the research also has a low recommendation value.However,the results may be considered for application if a standard regimen is not available.

Vibrational Spectroscopic Investigations, Electronic Properties, Molecular Structure and Quantum Mechanical Study of an Antifolate Drug: Pyrimethamine

The computational modelling supported by experimental results can explain the molecular structure, vibrational assignments, reactive sites and several structural properties. In this context, the spectroscopic (FT-IR, FT-Raman and NMR) analysis, electronic properties (HOMO and LUMO energies) and molecular structure of pyrimethamine (Pyr) were investigated by density functional theory (DFT) method associated with three levels of theory viz., B3LYP, MN15 and wB97XD with 6-311++G(d,p) and def2TZVPP as basis sets, respectively in the Gaussian 16 programs. The 1H and 13C NMR chemical shifts were calculated with a gauge-independent atomic orbital (GIAO) approach by also applying the same levels of theory and basis sets. All experimental results were compared with theoretical data. Although the results revealed high degrees of correlation between the theoretical and experimental values for spectroscopic properties using the three methods. Furthermore, the atomic and natural charges, energy band gap and chemical reactivity were determined, while the frontier molecular orbital (FMO) and molecular electrostatic potential (MEP) surfaces were plotted to explain the reactive nature of the title molecule.

Antiplasmodial and antiulcer activities of Melanthera scadens

Objective:To evaluate the antimalarial and antiulcerogenic activities ofleaf exlracl and fractions of Melanthera scandens(M.scandens).Methods:The crude leaf extract(37-111 mg/kg)and fractions(chloroform,ethylacetale and methanol;78 mg/kg)of M.scadens were investigated for antiplasmodial activity against chloroquine-sensitive Plasmodium berghei infections in mice and for antiulcer activity against experimentally-induced ulcers.The antimalarial activity during early and established infections as well as prophylactic was investigated.Artesunate(5 mg/kg)and pyrimethamine(1.2 mg/kg)were used as positive controls.Thin films made from tail blood of each mouse were used to assess the level of parasitaemia of the mice.Antiulcer activity of the crude extract was also evaluated against indomethacin,ethanol and histamine induced ulcers.Results:The extract and its fractions dose-dependently reduced parasitaemia induced by chloroquine-sensitive Plasmodium berghei infection in prophylactic,suppressive and curative models in mice.These reductions were statistically significant(P<0.00l),They also improved the mean survival time(MST)from 9.28 to 17.73 days as compared with the control(P<0.0l-0.001).The activities of extract/fractions were incomparable to that of the standard drugs ie.artesunate and pyrimethamine.On experimentally-induced ulcers,the extract inhibited indomethacin,ethanol and histamine induced ulcers.These inhibitions were statistically significant(P<0.001)and in a dose-dependent fashion.Conclusions:The antiplasmodial and antiulcerogenic effects of this plant may in part be mediated through the chemical constituents of the plant.

The antiplasmodial effect of the extracts and formulated capsules of Phyllanthus amarus on Plasmodium yoelii infection in mice

Objective:To investigate the antiplasmodial activity of the extracts of Phyllanthus amarus(P. amarus) on Plasmodium yoelii(P.yoelii)(a resistant malaria parasite strain used in animal stuthes) infection in mice.Methods:The aqueous and ethanol extracts of the whole plant of Phyllanthus amarus was administered to Swiss albino mice at doses of 200 mg/kg/day,400 mg/ kg/day,800 mg/kg/day and 1 600 mg/kg/day and the prophylactic and chemotherapeutic effect of the extracts against P.yoelii infection in mice was investigated and compared with those of standard antimalaria drugs used in the treatment of malaria parasite infection.Acute toxicity test was carried out in mice to determine the safety of the plant extract when administered orally. Results:The results showed that the extracts demonstrated a dose-dependent prophylactic and chemotherapeutic activity with the aqueous extracts showing slightly higher effect than the ethanol extract.The antiplasmodial effects of the extracts were comparable to the standard prophylactic and chemotherapeutic drugs used in chloroquine resistant Plasmodium infection although the activity depended on the dose of the extract administered.The extracts showed prophylactic effect by significantly delaying the onset of infection with the suppression of 79% at a dose of 1 600 mg/kg/day.Conclusions:The results obtained indicate that the extracts of the whole plant of P.amarus possess repository and chemotherapeutic effects against resistant strains of P.yoelii in Swiss albino mice.The findings justify the use of the extract of P.amarus in traditional medicine practice,for the treatment of malaria infections.

Knowledge and Perceived Barriers towards Intermittent Prevention of Malaria in Pregnancy: A Cross-Sectional Study

Background: Intermittent prevention of malaria is a key strategy adopted globally to ensure the wellbeing of pregnant women. By extension, these strategies enhance the healthy development of the fetus. This study assesses the knowledge and practices of pregnant women towards intermittent malaria prevention strategies in the Juaben Government Hospital of Ghana. Methods: A cross-sectional, quantitative, descriptive survey was conducted among 120 pregnant women at the antenatal clinic of the Juaben Government hospital in the Ashanti region of Ghana using a simple random sampling technique in selecting the study participants. The participant response rate was 94.5%. Data were analyzed using Statistical Package for the Social Sciences (SPSS) version 22.0. Results: Majority (54.2%) of respondents were between 20 - 29 years of age. Majority of study respondents attended Antenatal visits in their first trimester of pregnancy. Majority (69.2%) have knowledge of malaria preventive strategies in pregnancy, with about 70% of respondents stating these strategies were designed only for pregnant women. Only 37.5% of study respondents could rightly identify the chemoprophylaxis indicated for malaria prevention in pregnancy. Whereas majority (93.3%) of respondents owned insecticide-treated mosquito nets, only 63.3% utilized these nets for their purpose. Barriers to utilization of insecticide-treated nets include difficulty in setting up the nets (26.7%), using mosquito coils (10.8%), and feeling uncomfortable sleeping in the nets (36.7%). Barriers to taking malaria prophylaxis include distance to health facilities (28.3%), thoughts of not being sick with malaria (55.8%), using herbs (10.0%), and not being aware of the essence of prophylaxis (7.5%). Conclusion: Knowledge of malaria preventive strategies in pregnancy is high. However, utilization of these strategies is low due to perceived human and institutional barriers. A comprehensive community-wide approach is required to improve the utilization of these services among pregnant women.