An Investigation of In Vitro Anti-Cancer Efficacy of Dihydroartemisinin-Loaded Bovine Milk Exosomes Against Triple-Negative Breast Cancer.

Repurposing drugs offers several advantages, including reduced time and cost compared to developing new drugs from scratch. It leverages existing knowledge about drug safety, dosage, and pharmacokinetics, expediting the process of clinical trials and regulatory approval. Dihydroartemisinin (DHA) is a semi-synthetic and active metabolite of all artemisinin molecules and is FDA-approved for the treatment of malaria. Apart from having anti-malarial properties, DHA also possesses anticancer properties. However, its pharmacological actions are limited by toxicity and solubility problems. To overcome these challenges and enhance its anticancer effectiveness, we designed an exosomal formulation of DHA. We isolated exosomes from bovine milk using differential ultracentrifugation and loaded DHA using sonication. Scanning and transition electron microscopy revealed a size of roughly 100 nm, with a spherical shape. Furthermore, in pH 7.4 and 5.5, the exosomes exhibited burst release followed by sustained release. Multiple in vitro cell culture tests demonstrated that Exo-DHA exhibited enhanced anticancer activity, including cytotoxicity, cellular uptake, generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential, and inhibition of colony formation. Additional evidence supporting Exo-DHA's anti-migration ability came from transwell migration and scratch assays. Based on these results, it was concluded that the anticancer efficacy of DHA was improved when loaded into bovine milk-derived exosomes. While the in vitro results are encouraging, more in vivo testing in suitable animal models and biochemical marker analysis are warranted.

Artemisinin pre-treatment fore cisplatin dosage enhances high grade urothelial carcinoma treatment in male albino mice via reverse gene expression modulation of FGFR3, HRAS, P53 and KDM6A.

BACKGROUND: Urinary bladder cancer, is the 10th most common global cancer, diagnosed in over 600,000 people causing 200,000 deaths annually. Artemisinin and its derivatives are safe compounds that have recently been proven to possess potent anti-tumor effects in vivo, through inhibition of cancer cell growth. The aim of this study is to assess the efficiency of artemisinin as a cancer treatment alone and as a pre-treatment fore cisplatin therapy for high grade urothelial carcinoma. METHODS: Sixty male albino mice were divided into six groups, and BBN was used to induce urinary bladder cancer. Blood samples were tested for renal functions and complete blood counts, kidney and urinary bladder tissues were harvested for histopathological examination. Total RNAs from urinary bladder tissues was collected, and gene expression of FGFR3, HRAS, P53, and KDM6A was quantified using qRT-PCR. RESULTS: Compared to the induced cancer group, the results revealed that FGFR3 expression levels were down-regulated in the induced cancer group treated by artemisinin only and the induced cancer group pre-treated with artemisinin prior to cisplatin by ~ 0.86-fold and 0.4-folds, respectively, aligning with HRAS down-regulation by ~ 9.54-fold and 9.05-fold, respectively. Whereas, P53 expression levels were up-regulated by ~ 0.68-fold and 0.84-fold, respectively, in parallel with KDM6A expression, which is up-regulated by ~ 0.95-folds and 5.27-folds, respectively. Also, serum creatinine and urea levels decreased significantly in the induced cancer group treated by artemisinin alone and the induced cancer group pre-treated with artemisinin prior to cisplatin, whereas the induced cancer group treated by cisplatin their levels increased significantly. Moreover, Hb, PLT, RBC, and WBC counts improved in both cancer groups treated by artemisinin alone and pre-treated with artemisinin prior to cisplatin. Histologically, in kidney tissues, artemisinin pre-treatment significantly reduced renal injury caused by cisplatin. While Artemisinin treatment for cancer in bladder tissues reverted invasive urothelial carcinoma to moderate urothelial dysplasia. CONCLUSIONS: This study indicates that artemisinin demonstrated a significant effect in reversal of the multi-step carcinogenesis process of high grade urothelial carcinoma and could enhance the effect of cisplatin therapy using artemisinin pre-treatment.

Effects of artemisinin and cisplatin on the malignant progression of oral leukoplakia. In vitro and in vivo study.

OBJECTIVES: Chemoprevention can be a treatment for potentially malignant lesions (PMLs). We aimed to evaluate whether artemisinin (ART) and cisplatin (CSP) are associated with apoptosis and immunogenic cell death (ICD) in vitro, using oral leukoplakia (OL) and oral squamous cell carcinoma (OSCC) cell lines, and whether these compounds prevent OL progression in vivo. METHODS: Normal keratinocytes (HaCat), Dysplastic oral cells (DOK), and oral squamous cell carcinoma (SCC-180) cell lines were treated with ART, CSP, and ART + CSP to analyze cytotoxicity, genotoxicity, cell migration, and increased expression of proteins related to apoptosis and ICD. Additionally, 41 mice were induced with OL using 4NQO, treated with ART and CSP, and their tongues were histologically analyzed. RESULTS: In vitro, CSP and CSP + ART showed dose-dependent cytotoxicity and reduced SCC-180 migration. No treatment was genotoxic, and none induced expression of proteins related to apoptosis and ICD; CSP considerably reduced High-mobility group box-1 (HMGB-1) protein expression in SCC-180. In vivo, there was a delay in OL progression with ART and CSP treatment; however, by the 16th week, only CSP prevented progression to OSCC. CONCLUSION: Expression of proteins related to ICD and apoptosis did not increase with treatments, and CSP was shown to reduce immunogenic pathways in SCC-180, while reducing cell migration. ART did not prevent the malignant progression of OL in vivo; CSP did despite significant adverse effects.

A transcription factor of SHI family AaSHI1 activates artemisinin biosynthesis genes in Artemisia annua.

BACKGROUND: Transcription factors (TFs) of plant-specific SHORT INTERNODES (SHI) family play a significant role in regulating development and metabolism in plants. In Artemisia annua, various TFs from different families have been discovered to regulate the accumulation of artemisinin. However, specific members of the SHI family in A. annua (AaSHIs) have not been identified to regulate the biosynthesis of artemisinin. RESULTS: We found five AaSHI genes (AaSHI1 to AaSHI5) in the A. annua genome. The expression levels of AaSHI1, AaSHI2, AaSHI3 and AaSHI4 genes were higher in trichomes and young leaves, also induced by light and decreased when the plants were subjected to dark treatment. The expression pattern of these four AaSHI genes was consistent with the expression pattern of four structural genes of artemisinin biosynthesis and their specific regulatory factors. Dual-luciferase reporter assays, yeast one-hybrid assays, and transient transformation in A. annua provided the evidence that AaSHI1 could directly bind to the promoters of structural genes AaADS and AaCYP71AV1, and positively regulate their expressions. This study has presented candidate genes, with AaSHI1 in particular, that can be considered for the metabolic engineering of artemisinin biosynthesis in A. annua. CONCLUSIONS: Overall, a genome-wide analysis of the AaSHI TF family of A. annua was conducted. Five AaSHIs were identified in A. annua genome. Among the identified AaSHIs, AaSHI1 was found to be localized to the nucleus and activate the expression of structural genes of artemisinin biosynthesis including AaADS and AaCYP71AV1. These results indicated that AaSHI1 had positive roles in modulating artemisinin biosynthesis, providing candidate genes for obtaining high-quality new A. annua germplasms.

Evaluation of the binding interactions between Plasmodium falciparum Kelch-13 mutant recombinant proteins with artemisinin.

Malaria, an ancient mosquito-borne illness caused by Plasmodium parasites, is mostly treated with Artemisinin Combination Therapy (ACT). However, Single Nucleotide Polymorphisms (SNPs) mutations in the P. falciparum Kelch 13 (PfK13) protein have been associated with artemisinin resistance (ART-R). Therefore, this study aims to generate PfK13 recombinant proteins incorporating of two specific SNPs mutations, PfK13-V494I and PfK13-N537I, and subsequently analyze their binding interactions with artemisinin (ART). The recombinant proteins of PfK13 mutations and the Wild Type (WT) variant were expressed utilizing a standard protein expression protocol with modifications and subsequently purified via IMAC and confirmed with SDS-PAGE analysis and Orbitrap tandem mass spectrometry. The binding interactions between PfK13-V494I and PfK13-N537I propeller domain proteins ART were assessed through Isothermal Titration Calorimetry (ITC) and subsequently validated using fluorescence spectrometry. The protein concentrations obtained were 0.3 mg/ml for PfK13-WT, 0.18 mg/ml for PfK13-V494I, and 0.28 mg/ml for PfK13-N537I. Results obtained for binding interaction revealed an increased fluorescence intensity in the mutants PfK13-N537I (83 a.u.) and PfK13-V494I (143 a.u.) compared to PfK13-WT (33 a.u.), indicating increased exposure of surface proteins because of the looser binding between PfK13 protein mutants with ART. This shows that the PfK13 mutations may induce alterations in the binding interaction with ART, potentially leading to reduced effectiveness of ART and ultimately contributing to ART-R. However, this study only elucidated one facet of the contributing factors that could serve as potential indicators for ART-R and further investigation should be pursued in the future to comprehensively explore this complex mechanism of ART-R.

Artemisitene shows superiority over artemisinin in preventing Schistosoma japonica-induced liver disease.

BACKGROUND: Artemisinin (ART) analogs, such as dihydroartemisinin, arteether, artemether, and artesunate, all featuring an endoperoxide bridge, have demonstrated efficacy against schistosomiasis. Artemisitene (ATT), which contains an additional α, ß-unsaturated carbonyl structure, has shown enhanced biological activities. This study aims to evaluate the anti-schistosomaiasis japonica activity of ATT and compare it with ART. METHODS: We assessed liver inflammation and fibrosis in mice using hematoxylin and eosin staining and Sirius red staining, respectively. RNA sequencing analyzed transcriptomics in female and male Schistosoma japonicum (S. japonicum) adult worms and mice livers, with cytokine profiling and flow cytometry to study immune responses under ART or ATT treatment. RESULTS: ATT exhibits a marked reduction in female S. japonicum adult worms and egg numbers, damaging the adult worms' surface. It also influences the transcription of genes related to cellular anatomical structures. Notably, ATT treatment resulted in significant reductions in liver granuloma size and collagen area, alongside lowering serum levels of glutamic pyruvic and glutamic oxaloacetic transaminase more effectively than ART. Both ART and ATT markedly decreased neutrophil frequency in the liver and elevated eosinophil counts. However, only ATT treatment significantly reduced the M1/M2 and Th1/Th2 indices, indicating a pronounced shift in immune response profiles. ATT-affected host immunity correlated with the extent of liver fibrosis and the count of single males more strongly than ART. CONCLUSION: ATT, as a novel preventive strategy for schistosomiasis japonica in mice, significantly outperforms ART.

Adapting malaria indicator surveys to investigate treatment adherence: a pilot study on Bioko Island, Equatorial Guinea.

BACKGROUND: Adherence to anti-malarial treatment regimens is an important aspect of understanding and improving the impact of malaria case management. However, both adherence to artemisinin-based combination therapy (ACT) and the factors driving it vary widely. While many other evaluation activities have been conducted on Bioko Island, until now adherence to anti-malarial treatments, and in particular ACT has not been evaluated. METHODS: The implementation of a malaria indicator survey (MIS) conducted on Bioko in 2023 was leveraged to evaluate adherence to ACT provided to individuals testing positive following the survey. A follow-up team visited the targeted households, physically observed treatment blisters where possible, and provided messaging to household members on the importance of adhering to the treatment guidelines to household members. The team used survey data from the targeted households to make messaging as relevant to the household's particular context as possible. RESULTS: Overall ACT adherence on Bioko Island was low, around 50%, and this varied demographically and geographically. Some of the highest transmission areas had exceptionally low adherence, but no systematic relationship between proper adherence and Plasmodium falciparum prevalence was detected. Estimates of adherence from follow-up visits were much lower than survey-based estimates in the same households (52.5% versus 87.1%), suggesting that lack of proper adherence may be a much larger issue on Bioko Island than previously thought. CONCLUSION: Representative surveys can be easily adapted to provide empirical estimates of adherence to anti-malarial treatments, complementary to survey-based and health facility-based estimates. The large discrepancy between adherence as measured in this study and survey-based estimates on Bioko Island suggests a health facility-based study to quantify adherence among the population receiving treatment for symptomatic malaria may be necessary.

Molecular Engineering of a Doubly Quenched Fluorescent Probe Enables Ultrasensitive Detection of Biothiols in Highly Diluted Plasma and High-Fidelity Imaging of Dihydroartemisinin-Induced Ferroptosis.

The occurrence and development of diseases are accompanied by abnormal activity or concentration of biomarkers in cells, tissues, and blood. However, the insufficient sensitivity and accuracy of the available fluorescence probes hinder the precise monitoring of associated indexes in biological systems, which is generally due to the high probe intrinsic fluorescence and false-negative signal caused by the reactive oxygen species (ROS)-induced probe decomposition. To resolve these problems, we have engineered a ROS-stable, meso-carboxylate boron dipyrromethene (BODIPY)-based fluorescent probe, which displays quite a low background fluorescence due to the doubly quenched intrinsic fluorescence by a combined strategy of the photoinduced electron transfer (PET) effect and "ester-to-carboxylate" conversion. The probe achieved a high S/N ratio with ultrasensitivity and good selectivity toward biothiols, endowing its fast detection capability toward the biothiol level in 200×-diluted plasma samples. Using this probe, we achieved remarkable distinguishing of liver injury plasma from normal plasma even at 80× dilution. Moreover, owing to its good stability toward ROS, the probe was successfully employed for high-fidelity imaging of the negative fluctuation of the biothiol level in nonsmall-cell lung cancer (NSCLC) during dihydroartemisinin-induced ferroptosis. This delicate design of suppressing intrinsic fluorescence reveals insights into enhancing the sensitivity and accuracy of fluorescent probes toward the detection and imaging of biomarkers in the occurrence and development of diseases.

Receipt of antimalarials among children aged 6-59 months in Nigeria from 2010 to 2021.

BACKGROUND: Nigeria has the highest malaria burden globally, and anti-malarials have been commonly used to treat malaria without parasitological confirmation. In 2012, Nigeria implemented rapid diagnostic tests (RDTs) to reduce the use of anti-malarials for those without malaria and to increase the use of artemisinin-based combination therapy (ACT) for malaria treatment. This study examined changes in anti-malarial receipt among children aged 6-59 months during a 12-year period of increasing RDT availability. METHODS: A cross-sectional analysis was conducted using the Nigeria Malaria Indicator Survey (NMIS) data from 2010 (before RDT implementation in 2012), 2015, and 2021. The analysis assessed trends in prevalence of malaria by survey RDT result, and fever and anti-malarial/ACT receipt in the 2 weeks prior to the survey. A multivariable logistic regression was used to account for the complex survey design and to examine factors associated with anti-malarial receipt, stratified by survey RDT result, a proxy for recent/current malaria infection. RESULTS: In a nationally-representative, weighted sample of 22,802 children aged 6-59 months, fever prevalence remained stable over time, while confirmed malaria prevalence decreased from 51.2% in 2010 to 44.3% in 2015 and 38.5% in 2021 (trend test p < 0.0001). Anti-malarial use among these children decreased from 19% in 2010 to 10% in 2021 (trend test p < 0.0001), accompanied by an increase in ACT use (2% in 2010 to 8% in 2021; trend test p < 0.0001). Overall, among children who had experienced fever, 30.6% of survey RDT-positive and 36.1% of survey RDT-negative children had received anti-malarials. The proportion of anti-malarials obtained from the private sector increased from 61.8% in 2010 to 80.1% in 2021 for RDT-positive children; most of the anti-malarials received in 2021 were artemisinin-based combinations. Factors associated with anti-malarial receipt for both RDT-positive and RDT-negative children included geographic region, greater household wealth, higher maternal education, and older children. CONCLUSION: From 2010 to 2021 in Nigeria, both malaria prevalence and anti-malarial treatments among children aged 6-59 months decreased, as RDT availability increased. Among children who had fever in the prior 2 weeks, anti-malarial receipt was similar between children with either positive or negative survey RDT results, indicative of persistent challenges in reducing inappropriate anti-malarials uptake.

Based on NF-κB and Notch1/Hes1 Signaling Pathways, the Mechanism of Artesunate on Inflammation in Osteoporosis in Ovariectomized Rats was Investigated.

BACKGROUND: Artesunate (ART) has the potential to modulate the nuclear factor kappa B (NF-κB) and Notch1/Hes1 signaling pathways, which play crucial roles in the pathogenesis of osteoporosis. This study aims to explore whether ART participates in the progression of osteoporosis by regulating these signaling pathways. METHODS: In the in vitro experiments, we treated bone marrow mesenchymal stem cells (BMSCs) with different concentrations of ART (0, 3, 6, 12 µM) and evaluated osteogenic differentiation using alkaline phosphatase staining (ALP) and alizarin red S staining (ARS) staining. The expression levels of osteocalcin (OCN), RUNT-related transcription factor 2 (RUNX2), osteoprotegerin (OPG), and receptor activator of the nuclear factor kappa ligand (RANKL) were detected by real-time quantitative PCR (RT-qPCR). The effects of ART on NF-κB p65 and Notch1 protein expression were analyzed by Western blot (WB) and immunofluorescence (IF). In the in vivo experiments, a postmenopausal osteoporosis rat model was established via ovariectomy. Bone tissue pathological injury was evaluated using hematoxylin eosin (HE) staining. Serum ALP levels were measured using a kit, bone density was determined by dual-energy X-ray absorptiometry, and serum levels of bone gla protein (BGP), OPG, RANKL, tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and IL-1ß were measured by enzyme-linked immunosorbent assay (ELISA). Additionally, the expression of NF-κB p65 and Notch1 in tissues was assessed by immunohistochemistry. RESULTS: In vitro experiments revealed that compared to the control group, ART dose-dependently promoted BMSCs proliferation and enhanced their osteogenic differentiation capability. The expression of OCN, RUNX2, and OPG significantly increased in the ART-treated group, while RANKL expression decreased significantly (p < 0.05). ART significantly inhibited the expression of NF-κB p65 and Notch1/Hes1 signaling pathway proteins (p < 0.05). Compared to ART treatment alone, combined treatment with ART and phorbol myristate acetate (PMA) or valproic acid (VPA) resulted in increased expression of NF-κB p65 and Notch1 proteins and decreased osteogenic differentiation capability (p < 0.05). In vivo experiments showed that in rats treated with ART, bone damage was significantly reduced, bone density and mineral content were restored considerably, and the expression of inflammatory factors (TNF-α, IL-6, IL-1ß) decreased significantly (p < 0.05). Additionally, ART treatment significantly reduced the expression of NF-κB p65 and Notch1 proteins, increased OPG expression, and decreased BGP and RANKL levels (p < 0.05). CONCLUSION: In summary, ART facilitates the osteogenic differentiation of BMSCs by inhibiting the NF-κB and Notch1/Hes1 signaling pathways, thereby exerting significant protective effects against osteoporosis.

Modulation of the Antimelanoma Activity Imparted to Artemisinin Hybrids by the Monoterpene Counterpart.

Molecular hybridization is a widely used strategy in drug discovery and development processes that consists of the combination of two bioactive compounds toward a novel entity. In the current study, two libraries of hybrid derivatives coming from the linkage of sesquiterpene counterparts dihydroartemisinin and artesunic acid, with a series of monoterpenes, were synthesized and evaluated by cell viability assay on primary and metastatic melanoma cell lines. Almost all the obtained compounds showed micromolar antimelanoma activity and selectivity toward the metastatic form of this cancer. Four hybrid derivatives containing perillyl alcohol, citronellol, and nerol as monoterpene counterpart emerged as the best compounds of the series, with nerol being active in combination with both sesquiterpenes, dihydroartemisinin and artesunic acid. Preliminary studies on the mechanism of action have shown the dependence of the pharmacological activity of newly synthesized hybrids on the formation of carbon- and oxygen-centered radical species. This study demonstrated the positive modulation of the pharmacodynamic effect of artemisinin semisynthetic derivatives dihydroartemisinin and artesunic acid due to the hybridization with monoterpene counterparts.

Indigenous emergence and spread of kelch13 C469Y artemisinin-resistant Plasmodium falciparum in Uganda.

Artemisinin-based combination therapies (ACTs) were introduced as the standard of care for uncomplicated malaria in Africa almost two decades ago. Recent studies in East Africa have reported a gradual increase in kelch13 (k13) mutant parasites associated with reduced artesunate efficacy. As part of the Community Access to Rectal Artesunate for Malaria project, we collected blood samples from 697 children with signs of severe malaria in northern Uganda between 2018 and 2020, before and after the introduction of rectal artesunate (RAS) in 2019. K13 polymorphisms were assessed, and parasite editing and phenotyping were performed to assess the impact of mutations on parasite resistance. Whole-genome sequencing was performed, and haplotype networks were constructed to determine the geographic origin of k13 mutations. Of the 697 children, 540 were positive for Plasmodium falciparum malaria by PCR and were treated with either RAS or injectable artesunate monotherapy followed in most cases by ACT. The most common k13 mutation was C469Y (6.7%), which was detected more frequently in samples collected after RAS introduction. Genome editing confirmed reduced in vitro susceptibility to artemisinin in C469Y-harboring parasites compared to wild-type controls (P < 0.001). The haplotypic network showed that flanking regions of the C469Y mutation shared the same African genetic background, suggesting a single and indigenous origin of the mutation. Our data provide evidence of selection for the artemisinin-resistant C469Y mutation. The realistic threat of multiresistant parasites emerging in Africa should encourage careful monitoring of the efficacy of artemisinin derivatives and strict adherence to ACT treatment regimens.

Dihydroartemisinin inhibits follicular helper T and B cells: implications for systemic lupus erythematosus treatment.

Systemic lupus erythematosus (SLE) is a common autoimmune disease, and its pathogenesis mainly involves the aberrant activation of B cells through follicular helper T (Tfh) cells to produce pathogenic antibodies, which requires more effective and safe treatment methods. Dihydroartemisinin (DHA) is the main active ingredient of artemisinin and has immunosuppressive effects. In this study, in vitro experiments confirmed that DHA inhibited Tfh cell induction and weakened its auxiliary function in B cell differentiation; furthermore, DHA directly inhibited B cell activation, differentiation, and antibody production. Furthermore, a mouse model of SLE was established, and we confirmed that DHA significantly reduced the symptoms of SLE and lupus nephritis, and decreased serum immunoglobulin (Ig)G, IgM, IgA, and anti-dsDNA levels. Moreover, DHA reduced the frequencies of total Tfh cells, activated Tfh cells, and B cell lymphoma 6, and interleukin (IL)-21 levels in Tfh cells from the spleen and lymph nodes, as well as the levels of B cells, germinal center B cells, and plasma cells in the spleen, lymph nodes, and kidneys. Additionally, DHA inhibited Tfh cells by blocking IL-2-inducible T cell kinase (ITK) signaling and its downstream nuclear factor (NF)-κB, nuclear factor of activated T cell, and activating protein-1 pathways, and directly inhibited B cells by blocking Bruton's tyrosine kinase (BTK) signaling and the downstream NF-κB and Myc pathways. Overall, our results demonstrated that DHA inhibited Tfh cells by blocking ITK signaling and also directly inhibited B cells by blocking BTK signaling. Therefore, reducing the production of pathogenic antibodies might effectively treat SLE.

The effect of single low-dose primaquine treatment for uncomplicated Plasmodium falciparum malaria on haemoglobin levels in Ethiopia: a longitudinal cohort study.

BACKGROUND: To interrupt residual malaria transmission and achieve successful elimination of Plasmodium falciparum in low-transmission settings, the World Health Organization (WHO) recommends the administration of a single dose of 0.25 mg/kg (or 15 mg/kg for adults) primaquine (PQ) combined with artemisinin-based combination therapy (ACT), without glucose-6-phosphate dehydrogenase (G6PD) testing. However, due to the risk of haemolysis in patients with G6PD deficiency (G6PDd), PQ use is uncommon. Thus, this study aimed to assess the safety of a single low dose of PQ administered to patients with G6PD deficiency. METHODS: An observational cohort study was conducted with patients treated for uncomplicated P. falciparum malaria with either single-dose PQ (0.25 mg/kg) (SLD PQ) + ACT or ACT alone. Microscopy-confirmed uncomplicated P. falciparum malaria patients visiting public health facilities in Arjo Didessa, Southwest Ethiopia, were enrolled in the study from September 2019 to November 2022. Patients with uncomplicated P. falciparum malaria were followed up for 28 days through clinical and laboratory diagnosis, such as measurements of G6PD levels and haemoglobin (Hb) concentrations. G6PD levels were measured by a quantiative CareSTART™ POCT S1 biosensor machine. Patient interviews were also conducted, and the type and frequency of clinical complaints were recorded. Hb data were taken on days (D) 7, 14, 21, and 28 following treatment with SLD-PQ + ACT or ACT alone. RESULTS: A total of 249 patients with uncomplicated P. falciparum malaria were enrolled in this study. Of these, 83 (33.3%) patients received ACT alone, and 166 (66.7%) received ACT combined with SLD-PQ treatment. The median age of the patients was 20 (IQR 28-15) years. G6PD deficiency was found in 17 (6.8%) patients, 14 males and 3 females. There were 6 (7.2%) and 11 (6.6%) phenotypic G6PD-deficient patients in the ACT alone and ACT + SLD-PQ arms, respectively. The mean Hb levels in patients treated with ACT + SLD-PQ were reduced by an average of 0.45 g/dl (95% CI = 0.39 to 0.52) in the posttreatment phase (D7) compared to a reduction of 0.30 g/dl (95% CI = 0.14 to - 0.47) in patients treated with ACT alone (P = 0.157). A greater mean Hb reduction was observed on day 7 in the G6PDd ACT + SLD-PQ group (- 0.60 g/dL) than in the G6PDd ACT alone group (- 0.48 g/dL); however, there was no statistically significant difference (P = 0.465). Overall, D14 losses were 0.10 g/dl (95% CI = - 0.00 to 0.20) and 0.05 g/dl (95% CI = - 0.123 to 0.22) in patients with and without SLD-PQ, respectively (P = 0.412). CONCLUSIONS: This study's findings indicate that using SLD-PQ in combination with ACT is safe for uncomplicated P. falciparum malaria regardless of the patient's G6PD status in Ethiopian settings. Caution should be taken in extrapolating this finding in other settings with diverse G6DP phenotypes.

Community education training to optimize the use of artemisinin-based combination therapy in Kamuli District, Uganda.

BACKGROUND: Community health education improves members health-seeking and utilization behaviours. To enhance the community knowledge and optimize the use of Artemisinin-based combination therapy (ACT), we carried out a community training in Kamuli District, Uganda. METHODS: The Analysis, Design, Development, Implementation and Evaluation (ADDIE) model was adopted. A total of 3420 community members were trained, 384 sampled to participate in pre-post-test assessment, with 76 healthcare workers (HCW). Community members were sampled by simple random sampling while the HCW were purposively selected. Community trainings occurred for two days at each of 42 public health facilities and one day at 27 parishes. A paired sample t-test and effect size was computed to establish effect with statistical significance tested at p < 0.05. RESULTS: Overall, a total of 3496 participants, majority 2705 (77.4%) females were trained. A total of 3420 community members, majority 2659 (77.7%) females trained, and 76 HCW, majority 46 (60.5%) females trained. The median age of community participants was 32 years, and interquartile range (IQR) = 17 years. The median age of HCW was 32 years, and IQR = 8 years. The training had a positive and significant effect on the community members knowledge: malaria transmission (T-test = 9.359; p < 0.0001) causes of malaria (T-test = 6.738; p < 0.0001), malaria symptoms (T-test = 5.403; p < 0.0001), dangerous malaria species (T-test = 12.088; p < 0.0001), Plasmodium vivax malaria cycle and occurrence every 48 h (T-test = 7.470; p < 0.0001), assessing whether a patient with malaria may suffer from jaundice (T-test = 7.228; p < 0.0001), organs affected by Plasmodium falciparum (T-test = 12.214; p < 0.0001), malaria diagnosis (T-test = 9.765; p < 0.0001), Plasmodium associated with malaria relapse (T-test = 10.250; p < 0.0001), and malaria prevention and control (T-test = 9.278; p < 0.0001). The intervention also had a significant and positive effect on HCW knowledge on all domains except on malaria transmission (T-test = 1.217; p = 0.228) where it didn't have any statistically significant increase on their knowledge. CONCLUSION: The education intervention improved the knowledge of participants significantly. There is need to adopt and scale-up the current intervention at all levels of care to enhance proper use of medicines.

An overview of artemisinin-resistant malaria and associated Pfk13 gene mutations in Central Africa.

Malaria caused by Plasmodium falciparum is one of the deadliest and most common tropical infectious diseases. However, the emergence of artemisinin drug resistance associated with the parasite's Pfk13 gene, threatens the public health of individual countries as well as current efforts to reduce malaria burdens globally. It is of concern that artemisinin-resistant parasites may be selected or have already emerged in Africa. This narrative review aims to evaluate the published evidence concerning validated, candidate, and novel Pfk13 polymorphisms in ten Central African countries. Results show that four validated non-synonymous polymorphisms (M476I, R539T, P553L, and P574L), directly associated with a delayed therapy response, have been reported in the region. Also, two Pfk13 polymorphisms associated to artemisinin resistance but not validated (C469F and P527H) have been reported. Furthermore, several non-validated mutations have been observed in Central Africa, and one allele A578S, is commonly found in different countries, although additional molecular and biochemical studies are needed to investigate whether those mutations alter artemisinin effects. This information is discussed in the context of biochemical and genetic aspects of Pfk13, and related to the regional malaria epidemiology of Central African countries.

Thiol-Amino Bifunctional Metal-Organic-Framework-Based Membrane Regulating Hydrophobic Sites for Selective Separation of Artesunate.

The selective separation and purification of artesunate (ARU) and artemisinin (ART) using zirconium-based metal-organic frameworks (MOF), especially UiO-66 MOF, are receiving increasing attention. In this study, tunable "hydrophobic" sites of thiol (-SH) were introduced to amino-functionalized MOFs (UiO-66-NH2) to fabricate a thiol-amino bifunctional UiO-66/polyvinylidene fluoride (PVDF)-blended membrane (S1-UiO/PVDF-DPIM) via the delayed-phase-inversion method for selective separation of ARU/ART. The adsorption results indicated that the modification of UiO-66-NH2 with thiol can indeed increase the ARU adsorption. The thiol-functional MOF (S1-UiO-66-NH2) was chosen as the optimal thiol-amino bifunctional MOF, as it possessed the maximum ARU adsorption capacity (111.14 mg g-1) and the highest selective-separation factor (α = 51.84). The ATR FT-IR dynamic spectrum disclosed the recognition mechanism, indicating that incorporating thiol groups into a hydrophilic MOF as hydrophobic sites can boost adsorption efficiency. Moreover, the static-selective permeation results showed that the S1-UiO/PVDF-DPIM preferentially transfers ARU when mixed with ART, even achieving complete ARU/ART separation. The most crucial aspect was the introduction of a hydrophobic core of -SH and new spontaneously formed disulfide bonds to S1-UiO/PVDF-DPIM, creating alternated hydrogen bonds and hydrophobic interactions. This work provides an alternative strategy to prepare hydrophobic-hydrophilic MOF-based membranes for the highly efficient and selective separation of complex analogue systems.

Pharmacokinetics of piperaquine and its association with intermittent malaria preventive therapy outcomes during pregnancy.

BACKGROUND: Dihydroartemisinin-piperaquine (DHP) recently showed superior effectiveness over sulfadoxine-pyrimethamine for malaria intermittent preventive treatment in pregnancy (IPTp). We investigated day 7 piperaquine pharmacokinetics and its therapeutic efficacy in preventing malaria during pregnancy. METHODS: Malaria-free (mRDT) pregnant women (n = 400) who received monthly IPTp-DHP were enrolled and followed till delivery. Day 7 Plasma piperaquine concentrations were determined after each IPTp dose using UPLC/MS/MS. IPTp outcomes (symptomatic malaria and parasitemia during pregnancy, placental malaria, and maternal malaria at delivery) were monitored. Linear mixed model and Cox regression were used to assess predictors of day 7 piperaquine concentration and treatment outcome, respectively. RESULTS: The incidences of symptomatic malaria and parasitemia during pregnancy per 100 person-year at risk were 2 and 33, respectively. The prevalence of histopathologically confirmed placental malaria and maternal malaria at delivery were 3% and 9.8%, respectively. Repeated monthly IPTp-DHP resulted in significantly increased day 7 plasma piperaquine concentration (p < 0.001). Following the 1st, 2nd, and 3rd monthly IPTp-DHP doses, the proportions of women with day 7 piperaquine concentration below the therapeutic threshold (< 30 ng/mL) were 6.1%, 4.1% and 3.6%, respectively. Factors such as maternal age, body weight and trimester were not significant predictors of day 7 piperaquine concentration. However, having a low day 7 piperaquine plasma concentration (< 30 ng/mL) was significantly associated with a higher risk of parasitemia during pregnancy (p = 0.004). CONCLUSION: Lower day 7 piperaquine plasma concentration is a risk factor for parasitemia during pregnancy. Single plasma sampling at day 7 can be used to monitor piperaquine effectiveness during IPTp-DHP. TRIAL REGISTRATION: Registered 09/12/2016, PACTR201612001901313.