Artemisinin resistance in the malaria parasite, Plasmodium falciparum, originates from its initial transcriptional response.
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ABSTRACT:
SUBMITTER: Zhu L
PROVIDER: S-EPMC8960834 | biostudies-literature | 2022 Mar
REPOSITORIES: biostudies-literature
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Artemisinin resistance in the malaria parasite, Plasmodium falciparum, originates from its initial transcriptional response.
Zhu Lei L van der Pluijm Rob W RW Kucharski Michal M Nayak Sourav S Tripathi Jaishree J White Nicholas J NJ Day Nicholas P J NPJ Faiz Abul A Phyo Aung Pyae AP Amaratunga Chanaki C Lek Dysoley D Ashley Elizabeth A EA Nosten François F Smithuis Frank F Ginsburg Hagai H von Seidlein Lorenz L Lin Khin K Imwong Mallika M Chotivanich Kesinee K Mayxay Mayfong M Dhorda Mehul M Nguyen Hoang Chau HC Nguyen Thuy Nhien Thanh TNT Miotto Olivo O Newton Paul N PN Jittamala Podjanee P Tripura Rupam R Pukrittayakamee Sasithon S Peto Thomas J TJ Hien Tran Tinh TT Dondorp Arjen M AM Bozdech Zbynek Z
Communications biology 20220328 1
The emergence and spread of artemisinin-resistant Plasmodium falciparum, first in the Greater Mekong Subregion (GMS), and now in East Africa, is a major threat to global malaria elimination ambitions. To investigate the artemisinin resistance mechanism, transcriptome analysis was conducted of 577 P. falciparum isolates collected in the GMS between 2016-2018. A specific artemisinin resistance-associated transcriptional profile was identified that involves a broad but discrete set of biological fu ...[more]
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Project description:BackgroundArtemisinin-based combination therapies are the recommended first-line treatments of falciparum malaria in all countries with endemic disease. There are recent concerns that the efficacy of such therapies has declined on the Thai-Cambodian border, historically a site of emerging antimalarial-drug resistance.MethodsIn two open-label, randomized trials, we compared the efficacies of two treatments for uncomplicated falciparum malaria in Pailin, western Cambodia, and Wang Pha, northwestern Thailand: oral artesunate given at a dose of 2 mg per kilogram of body weight per day, for 7 days, and artesunate given at a dose of 4 mg per kilogram per day, for 3 days, followed by mefloquine at two doses totaling 25 mg per kilogram. We assessed in vitro and in vivo Plasmodium falciparum susceptibility, artesunate pharmacokinetics, and molecular markers of resistance.ResultsWe studied 40 patients in each of the two locations. The overall median parasite clearance times were 84 hours (interquartile range, 60 to 96) in Pailin and 48 hours (interquartile range, 36 to 66) in Wang Pha (P<0.001). Recrudescence confirmed by means of polymerase-chain-reaction assay occurred in 6 of 20 patients (30%) receiving artesunate monotherapy and 1 of 20 (5%) receiving artesunate-mefloquine therapy in Pailin, as compared with 2 of 20 (10%) and 1 of 20 (5%), respectively, in Wang Pha (P=0.31). These markedly different parasitologic responses were not explained by differences in age, artesunate or dihydroartemisinin pharmacokinetics, results of isotopic in vitro sensitivity tests, or putative molecular correlates of P. falciparum drug resistance (mutations or amplifications of the gene encoding a multidrug resistance protein [PfMDR1] or mutations in the gene encoding sarco-endoplasmic reticulum calcium ATPase6 [PfSERCA]). Adverse events were mild and did not differ significantly between the two treatment groups.ConclusionsP. falciparum has reduced in vivo susceptibility to artesunate in western Cambodia as compared with northwestern Thailand. Resistance is characterized by slow parasite clearance in vivo without corresponding reductions on conventional in vitro susceptibility testing. Containment measures are urgently needed. (ClinicalTrials.gov number, NCT00493363, and Current Controlled Trials number, ISRCTN64835265.)
| S-EPMC3495232 | biostudies-literature
Project description:BackgroundArtemisinin resistance in Plasmodium falciparum has emerged in Southeast Asia and now poses a threat to the control and elimination of malaria. Mapping the geographic extent of resistance is essential for planning containment and elimination strategies.MethodsBetween May 2011 and April 2013, we enrolled 1241 adults and children with acute, uncomplicated falciparum malaria in an open-label trial at 15 sites in 10 countries (7 in Asia and 3 in Africa). Patients received artesunate, administered orally at a daily dose of either 2 mg per kilogram of body weight per day or 4 mg per kilogram, for 3 days, followed by a standard 3-day course of artemisinin-based combination therapy. Parasite counts in peripheral-blood samples were measured every 6 hours, and the parasite clearance half-lives were determined.ResultsThe median parasite clearance half-lives ranged from 1.9 hours in the Democratic Republic of Congo to 7.0 hours at the Thailand-Cambodia border. Slowly clearing infections (parasite clearance half-life >5 hours), strongly associated with single point mutations in the "propeller" region of the P. falciparum kelch protein gene on chromosome 13 (kelch13), were detected throughout mainland Southeast Asia from southern Vietnam to central Myanmar. The incidence of pretreatment and post-treatment gametocytemia was higher among patients with slow parasite clearance, suggesting greater potential for transmission. In western Cambodia, where artemisinin-based combination therapies are failing, the 6-day course of antimalarial therapy was associated with a cure rate of 97.7% (95% confidence interval, 90.9 to 99.4) at 42 days.ConclusionsArtemisinin resistance to P. falciparum, which is now prevalent across mainland Southeast Asia, is associated with mutations in kelch13. Prolonged courses of artemisinin-based combination therapies are currently efficacious in areas where standard 3-day treatments are failing. (Funded by the U.K. Department of International Development and others; ClinicalTrials.gov number, NCT01350856.).
| S-EPMC4143591 | biostudies-literature
Project description:Malaria genetic variation has been extensively characterized, but the level of epigenetic plasticity remains largely unexplored. Here we provide a comprehensive characterization of transcriptional variation in the most lethal malaria parasite, Plasmodium falciparum, based on highly accurate transcriptional analysis of isogenic parasite lines grown under homogeneous conditions. This analysis revealed extensive transcriptional heterogeneity within genetically homogeneous clonal parasite populations. We show that clonally variant expression controlled at the epigenetic level is an intrinsic property of specific genes and gene families, the majority of which participate in host-parasite interactions. Intrinsic transcriptional variability is not restricted to genes involved in immune evasion, but also affects genes linked to lipid metabolism, protein folding, erythrocyte remodeling, or transcriptional regulation, among others, indicating that epigenetic variation results in both antigenic and functional variation. We observed a general association between heterochromatin marks and clonally variant expression, extending previous observations for specific genes to essentially all variantly expressed gene families. These results suggest that phenotypic variation of functionally unrelated P. falciparum gene families is mediated by a common mechanism based on reversible formation of H3K9me3-based heterochromatin. In changing environments, diversity confers fitness to a population. Our results support the idea that P. falciparum uses a bet-hedging strategy, as an alternative to directed transcriptional responses, to adapt to common fluctuations in its environment. Consistent with this idea, we found that transcriptionally different isogenic parasite lines markedly differed in their survival to heat-shock mimicking febrile episodes and adapted to periodic heat-shock with a pattern consistent with natural selection of pre-existing parasites.
| S-EPMC3337437 | biostudies-literature
Project description:For more than five decades, Southeast Asia (SEA) has been fertile ground for the emergence of drug-resistant Plasmodium falciparum malaria. After generating parasites resistant to chloroquine, sulfadoxine, pyrimethamine, quinine, and mefloquine, this region has now spawned parasites resistant to artemisinins, the world's most potent antimalarial drugs. In areas where artemisinin resistance is prevalent, artemisinin combination therapies (ACTs)-the first-line treatments for malaria-are failing fast. This worrisome development threatens to make malaria practically untreatable in SEA, and threatens to compromise global endeavors to eliminate this disease. A recent series of clinical, in vitro, genomics, and transcriptomics studies in SEA have defined in vivo and in vitro phenotypes of artemisinin resistance, identified its causal genetic determinant, explored its molecular mechanism, and assessed its clinical impact. Specifically, these studies have established that artemisinin resistance manifests as slow parasite clearance in patients and increased survival of early-ring-stage parasites in vitro; is caused by single nucleotide polymorphisms in the parasite's K13 gene, is associated with an upregulated "unfolded protein response" pathway that may antagonize the pro-oxidant activity of artemisinins, and selects for partner drug resistance that rapidly leads to ACT failures. In SEA, clinical studies are urgently needed to monitor ACT efficacy where K13 mutations are prevalent, test whether new combinations of currently available drugs cure ACT failures, and advance new antimalarial compounds through preclinical pipelines and into clinical trials. Intensifying these efforts should help to forestall the spread of artemisinin and partner drug resistance from SEA to sub-Saharan Africa, where the world's malaria transmission, morbidity, and mortality rates are highest.
| S-EPMC4992992 | biostudies-literature
Project description:Artemisinin combination therapies (ACTs) have led to a significant decrease in Plasmodium falciparum malaria mortality. This progress is now threatened by emerging artemisinin resistance (ART-R) linked originally in SE Asia to polymorphisms in the Kelch propeller protein (K13) and more recently to several other seemingly unrelated genetic mutations. To better understand the parasite response to ART, we are characterizing a P. falciparum mutant with altered sensitivity to ART that was created via piggyBac transposon mutagenesis. The transposon inserted near the putative transcription start site of a gene defined as a "Plasmodium-conserved gene of unknown function," now functionally linked to K13 as the Kelch13 Interacting Candidate 5 protein (KIC5). Phenotype analysis of the KIC5 mutant during intraerythrocytic asexual development identified transcriptional changes associated with DNA stress response and altered mitochondrial metabolism, linking dysregulation of the KIC5 gene to the parasite's ability to respond to ART exposure. Through characterization of the KIC5 transcriptome, we hypothesize that this gene may be essential under ART exposure to manage gene expression of the wild-type stress response at early ring stage, thereby providing a better understanding of the parasite's processes that can alter ART sensitivity.
| S-EPMC9829687 | biostudies-literature
Project description:RNAseq data profiling the artemisinin-sensitive P. falciparum piggyBac mutant of the KIC5 gene (PF3D7_1138700, Kelch13 Interacting Candidate 5).
2022-11-29 | GSE205012 | GEO
Project description:Artemisinins are the cornerstone of anti-malarial drugs. Emergence and spread of resistance to them raises risk of wiping out recent gains achieved in reducing worldwide malaria burden and threatens future malaria control and elimination on a global level. Genome-wide association studies (GWAS) have revealed parasite genetic loci associated with artemisinin resistance. However, there is no consensus on biochemical targets of artemisinin. Whether and how these targets interact with genes identified by GWAS, remains unknown. Here we provide biochemical and cellular evidence that artemisinins are potent inhibitors of Plasmodium falciparum phosphatidylinositol-3-kinase (PfPI3K), revealing an unexpected mechanism of action. In resistant clinical strains, increased PfPI3K was associated with the C580Y mutation in P. falciparum Kelch13 (PfKelch13), a primary marker of artemisinin resistance. Polyubiquitination of PfPI3K and its binding to PfKelch13 were reduced by the PfKelch13 mutation, which limited proteolysis of PfPI3K and thus increased levels of the kinase, as well as its lipid product phosphatidylinositol-3-phosphate (PI3P). We find PI3P levels to be predictive of artemisinin resistance in both clinical and engineered laboratory parasites as well as across non-isogenic strains. Elevated PI3P induced artemisinin resistance in absence of PfKelch13 mutations, but remained responsive to regulation by PfKelch13. Evidence is presented for PI3P-dependent signalling in which transgenic expression of an additional kinase confers resistance. Together these data present PI3P as the key mediator of artemisinin resistance and the sole PfPI3K as an important target for malaria elimination.
| S-EPMC4417027 | biostudies-literature
Project description:Despite a significant decline in morbidity and mortality over the last two decades, in 2018 there were 228 million reported cases of malaria and 405000 malaria-related deaths. Artemisinin, the cornerstone of artemisinin-based combination therapies, is the most potent drug in the antimalarial armamentarium against falciparum malaria. Heme-mediated activation of artemisinin and its derivatives results in widespread parasite protein alkylation, which is thought to lead to parasite death. Alarmingly, cases of decreased artemisinin efficacy have been widely detected across Cambodia and in neighboring countries, and a few cases have been reported in the Guiana Shield, India, and Africa. The grim prospect of widespread artemisinin resistance propelled a concerted effort to understand the mechanisms of artemisinin action and resistance. The identification of genetic markers and the knowledge of molecular mechanisms underpinning artemisinin resistance allow prospective surveillance and inform future drug development strategies, respectively. Here, we highlight recent advances in our understanding of how parasite vesicle trafficking, hemoglobin digestion, and cell stress responses contribute to artemisinin resistance.
| S-EPMC8528195 | biostudies-literature
Project description:BackgroundGenetic variation is an essential means of evolution and adaptation in many organisms in response to environmental change. Certain DNA alterations can be carried out by site-specific recombinases (SSRs) that fall into two families: the serine and the tyrosine recombinases. SSRs are seldom found in eukaryotes. A gene homologous to a tyrosine site-specific recombinase has been identified in the genome of Plasmodium falciparum. The sequence is highly conserved among five other members of Plasmodia.Methodology/principal findingsThe predicted open reading frame encodes for a ∼57 kDa protein containing a C-terminal domain including the putative tyrosine recombinase conserved active site residues R-H-R-(H/W)-Y. The N-terminus has the typical alpha-helical bundle and potentially a mixed alpha-beta domain resembling that of λ-Int. Pf-Int mRNA is expressed differentially during the P. falciparum erythrocytic life stages, peaking in the schizont stage. Recombinant Pf-Int and affinity chromatography of DNA from genomic or synthetic origin were used to identify potential DNA targets after sequencing or micro-array hybridization. Interestingly, the sequences captured also included highly variable subtelomeric genes such as var, rif, and stevor sequences. Electrophoretic mobility shift assays with DNA were carried out to verify Pf-Int/DNA binding. Finally, Pf-Int knock-out parasites were created in order to investigate the biological role of Pf-Int.Conclusions/significanceOur data identify for the first time a malaria parasite gene with structural and functional features of recombinases. Pf-Int may bind to and alter DNA, either in a sequence specific or in a non-specific fashion, and may contribute to programmed or random DNA rearrangements. Pf-Int is the first molecular player identified with a potential role in genome plasticity in this pathogen. Finally, Pf-Int knock-out parasite is viable showing no detectable impact on blood stage development, which is compatible with such function.
| S-EPMC3466309 | biostudies-literature
Project description:Human to vector transmission of malaria requires that some blood-stage parasites abandon asexual growth and convert into non-replicating sexual forms called gametocytes. The initial steps of gametocytogenesis remain largely uncharacterized. Here, we study this part of the malaria life cycle in Plasmodium falciparum using PfAP2-G, the master regulator of sexual conversion, as a marker of commitment. We demonstrate the existence of PfAP2-G-positive sexually committed parasite stages that precede the previously known committed schizont stage. We also found that sexual conversion can occur by two different routes: the previously described route in which PfAP2-G-expressing parasites complete a replicative cycle as committed forms before converting into gametocytes upon re-invasion, or a direct route with conversion within the same cycle as initial PfAP2-G expression. The latter route is linked to early PfAP2-G expression in ring stages. Reanalysis of published single-cell RNA-sequencing (RNA-seq) data confirmed the presence of both routes. Consistent with these results, using plaque assays we observed that, in contrast to the prevailing model, many schizonts produced mixed plaques containing both asexual parasites and gametocytes. Altogether, our results reveal unexpected features of the initial steps of sexual development and extend the current view of this part of the malaria life cycle.
| S-EPMC6294672 | biostudies-literature