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Tag: malaria

ANTI-MALARIAL ACTIVITY OF AMENTOFLAVONE ISOLATED FROM LEAF OF CALOPHYLLUM TOMENTOSUM WEIGHT

Calophyllum tomentosum belonging to Clusiaceae family is an Indian medicinal plant used as folklore medicine to cure various kinds of diseases reported in Ayurveda, and the leaves of the plant are also used as an active ingredient for the preparation of a botanical medicine known as ‘Punnaga’, ‘Surapunnaga’ and ‘Tamoil’ among other common names. Chemical profiling of the methanol extract of the defatted leaf revealed the presence of amentoflavone as one of the constituents along with coumarins, terpenoids, steroids, and apetalic acids. Structural determination of these amentoflavone has been conducted by chemical, spectral, and spectrometric methods in comparison with spectral values available in the literature and confirmed by a single crystal X-ray diffraction study. Amentoflavone (1) and its derivative (2-5) tested to check the efficacy of anti-malarial activity against Plasmodium falciparum. Amongst them, only tetra methoxy amentoflavone, (2) exhibited moderate anti-malarial activity with IC50 value 1.99 ± 0.42 µM against Plasmodium falciparum in comparison with artemisinin as control, whereas the other products possessed almost negligible activity although their structural skeletons are identical with little variation of number and nature of substituents. The structure activity relationship (SAR) of the active constituent and its derivatives is reported herein.

Ajoy Kumar Bauri, Joshua H Butler, Maria B Cassera, Sabine Foro. Chem Biodivers. 2024 Oct 14:e202401576. doi: 10.1002/cbdv.202401576.

Optimization of diastereomeric dihydropyridines as antimalarials

graphical abstract

The increase in research funding for the development of antimalarials since 2000 has led to a surge of new chemotypes with potent antimalarial activity. High-throughput screens have delivered several thousand new active compounds in several hundred series, including the 4,7-diphenyl-1,4,5,6,7,8-hexahydroquinolines, hereafter termed dihydropyridines (DHPs). We optimized the DHPs for antimalarial activity. Structure-activity relationship studies focusing on the 2-, 3-, 4-, 6-, and 7-positions of the DHP core led to the identification of compounds potent (EC50 < 10 nM) against all strains of P. falciparum tested, including the drug-resistant parasite strains K1, W2, and TM90-C2B. Evaluation of efficacy of several compounds in vivo identified two compounds that reduced parasitemia by >75 % in mice 6 days post-exposure following a single 50 mg/kg oral dose. Resistance acquisition experiments with a selected dihydropyridine led to the identification of a single mutation conveying resistance in the gene encoding for Plasmodium falciparum multi-drug resistance protein 1 (PfMDR1). The same dihydropyridine possessed transmission blocking activity. The DHPs have the potential for the development of novel antimalarial drug candidates.

Kurt S Van Horn, Yingzhao Zhao, Prakash T Parvatkar, Julie Maier, Tina Mutka, Alexis Lacrue, Fabian Brockmeier, Daniel Ebert, Wesley Wu, Debora R Casandra, Niranjan Namelikonda, Jeanine Yacoub, Martina Sigal, Spencer Knapp, David Floyd, David Waterson, Jeremy N Burrows, James Duffy, Joseph L DeRisi, Dennis E Kyle, R Kiplin Guy, Roman Manetsch. Eur J Med Chem. 2024 Jun 18:275:116599. doi: 10.1016/j.ejmech.2024.116599.

The influence of oviposition status on measures of transmission potential in malaria-infected mosquitoes depends on sugar availability

graphical abstract

Background: Like other oviparous organisms, the gonotrophic cycle of mosquitoes is not complete until they have selected a suitable habitat to oviposit. In addition to the evolutionary constraints associated with selective oviposition behavior, the physiological demands relative to an organism’s oviposition status also influence their nutrient requirement from the environment. Yet, studies that measure transmission potential (vectorial capacity or competence) of mosquito-borne parasites rarely consider whether the rates of parasite replication and development could be influenced by these constraints resulting from whether mosquitoes have completed their gonotrophic cycle.

Methods: Anopheles stephensi mosquitoes were infected with Plasmodium berghei, the rodent analog of human malaria, and maintained on 1% or 10% dextrose and either provided oviposition sites (‘oviposited’ herein) to complete their gonotrophic cycle or forced to retain eggs (‘non-oviposited’). Transmission potential in the four groups was measured up to 27 days post-infection as the rates of (i) sporozoite appearance in the salivary glands (‘extrinsic incubation period’ or EIP), (ii) vector survival and (iii) sporozoite densities.

Results: In the two groups of oviposited mosquitoes, rates of sporozoite appearance and densities in the salivary glands were clearly dependent on sugar availability, with shorter EIP and higher sporozoite densities in mosquitoes fed 10% dextrose. In contrast, rates of appearance and densities in the salivary glands were independent of sugar concentrations in non-oviposited mosquitoes, although both measures were slightly lower than in oviposited mosquitoes fed 10% dextrose. Vector survival was higher in non-oviposited mosquitoes.

Conclusions: Costs to parasite fitness and vector survival were buffered against changes in nutritional availability from the environment in non-oviposited but not oviposited mosquitoes. Taken together, these results suggest vectorial capacity for malaria parasites may be dependent on nutrient availability and oviposition/gonotrophic status and, as such, argue for more careful consideration of this interaction when estimating transmission potential. More broadly, the complex patterns resulting from physiological (nutrition) and evolutionary (egg-retention) trade-offs described here, combined with the ubiquity of selective oviposition behavior, implies the fitness of vector-borne pathogens could be shaped by selection for these traits, with implications for disease transmission and management. For instance, while reducing availability of oviposition sites and environmental sources of nutrition are key components of integrated vector management strategies, their abundance and distribution are under strong selection pressure from the patterns associated with climate change.

Justine C Shiau, Nathan Garcia-Diaz, Dennis E Kyle, Ashutosh K Pathak. Parasit Vectors. 2024 May 23;17(1):236. doi: 10.1186/s13071-024-06317-2.

Trainee Spotlight: Grace Vick

Ph.D. student Grace Woods

My name is Grace Vick and I am a 4th year infectious diseases PhD candidate in Vasant Muralidharan’s lab. I’m originally from North Carolina and received my Bachelor’s of Science in Biology from Western Carolina University. After graduating undergraduate, I completed an internship at the Defense Forensic Science Center doing forensic biology research. After that, I spent 2 years as an ORISE Fellow at the Centers for Disease Control and Prevention, studying and identifying genetic markers of multi-drug resistant strains of Neisseria gonorrhoeae. I came straight to UGA through the ILS program after my fellowship at CDC.

What made you want to study science?

Ever since I was little, I’ve always spent a lot of time being outside in nature and enjoyed figuring out the intricacies of how things work. During my undergraduate, I was able to explore the different areas of science and found the molecular biology of genetics to be an interesting field that is highly translatable and still vastly unknown. After I spent a few years gaining lab experience and an appreciation for the public health concerns of infectious diseases at the CDC, I knew I wanted to pursue a PhD in that field which brought me to UGA.

Why did you choose UGA?

My experience at the CDC offered the opportunity to learn about diseases and public health issues across all sectors and countries, which led me to learn more about parasitic diseases. Previously, I knew nothing about these diseases but as I learned more about their complex and fascinating life cycles and how these diseases of poverty impact people around the world, I was captivated by this research. Because I was really interested in spending my PhD studying infectious and parasitic diseases, I found out about the CTEGD at UGA and that is what brought me here. The CTEGD is a really wonderful environment for trainees to be exposed to exciting and diverse parasitology research, and I’ve really enjoyed my experience here.

What is your research focus and why did you choose it?

Our lab works on the deadliest form of malaria, Plasmodium falciparum. P. falciparum kills over half a million people each year, with the majority of those deaths being children under the age of 5. Our lab is interested in understanding the molecular mechanisms that are essential to asexual blood stage of this parasite. My work specifically focuses on determining the role of previously unknown proteins that we have discovered are essential for asexual stage invasion of merozoites into host red blood cells. Using a combination of genetic engineering, molecular, and cellular biology techniques, I aim to determine the molecular function of these proteins in the human asexual stage invasion of red blood cells.

Have you received any awards or honors?

In addition to receiving the NIH T32 Predoctoral Fellowship, I have been invited to present at multiple national and international conferences such as Molecular Parasitology Meeting in Massachusetts and Molecular Approaches in Malaria in Lorne, Australia where I won a poster award.

What are your career goals?

When I graduate with my doctoral degree, I hope to either join governmental research or the industry sector. If I decided to head into governmental work, I would choose a career at the CDC where I could continue working in the parasitology research field and apply current public health policies to the international parasitology field. If I decide to join the biomedical industry sector, I would want to work in Research and Design at a company that designs therapeutics and diagnostics for disease prevention and treatment.

What do you hope to do for your capstone experience?

I would really love to experience fieldwork in a malaria-endemic region. I think having the experience of meeting people and learning firsthand how this disease affects millions of people every day would be very eye-opening for me since I have only seen the lab side of malaria. The ability to experience fieldwork would give me a broader experience with how malaria is researched and treated outside of the lab environment and in rural lab environments. I would love to visit Africa or South East Asia to conduct fieldwork in a malaria-endemic environment.

What is your favorite thing about Athens?

Obviously, I love the food in Athens! I love going downtown to grab food and drinks on the weekend. Otherwise, I enjoy getting out and exploring the green spaces and parks that Athens has to offer such as Sandy Creek and the North Oconee Greenway with my husband and dog.

Any advice for a student interested in this field?

I would say the best advice is to read and soak up as much as you can about parasitology both before you get into the field and after. A lot of research has overlap between different parasites and it’s helpful to know about other parasitic diseases that might not be your main focus. Plus, parasites are fun! 🙂 My other advice in general for starting graduate school is to always reach out to students in labs you’re interested in joining. Students are pretty much always willing to help give clear insight into lab dynamics, mentorship of the PI, and generally how life working in that lab is. That information is all really helpful to know when choosing which lab to join!

 

Support trainees like Grace by giving today to the Center for Tropical & Emerging Global Diseases.

Hepatocytes and the art of killing Plasmodium softly

Figure 1. The gap in our understanding of how hepatocytes eliminate Plasmodium.
Figure 1. The gap in our understanding of how hepatocytes eliminate Plasmodium.

 

The Plasmodium parasites that cause malaria undergo asymptomatic development in the parenchymal cells of the liver, the hepatocytes, prior to infecting erythrocytes and causing clinical disease. Traditionally, hepatocytes have been perceived as passive bystanders that allow hepatotropic pathogens such as Plasmodium to develop relatively unchallenged. However, now there is emerging evidence suggesting that hepatocytes can mount robust cell-autonomous immune responses that target Plasmodium, limiting its progression to the blood and reducing the incidence and severity of clinical malaria. Here we discuss our current understanding of hepatocyte cell-intrinsic immune responses that target Plasmodium and how these pathways impact malaria.

Camila Marques-da-Silva, Clyde Schmidt-Silva, Samarchith P Kurup. Trends Parasitol. 2024 May 6:S1471-4922(24)00086-2. doi: 10.1016/j.pt.2024.04.004.

UGA researchers received prestigious grant to develop malaria drug

by Amy Horton

Chet Joyner and Steven Maher
Principal Investigators Chet Joyner (left) and Steven Maher (right). Photo credit: Donna Huber

 

New compound targets P. vivax, source of recent U.S. infections

Two University of Georgia researchers have been awarded approximately $770,000 from the Global Health Initiative Technology (GHIT) Fund to develop a new drug to kill the dormant liver stages of Plasmodium vivax, the most widespread of the malaria parasites. This amount is part of a total of JPY 334,238,778 awarded by the GHIT Fund to a partnership consisting of UGA, Medicine for Malaria Venture and Mitsubishi Tanabe Pharma Corporation.

P. vivax often persists in the liver of patients, causing a relapse infection following treatment of the symptomatic blood infection,” said Steven Maher, associate research scientist in the Office of Research’s Center for Tropical and Emerging Global Diseases (CTEGD). “In many parts of the world, relapses account for the majority of total P. vivax cases.”

The announcement comes on the heels of reports of the first locally acquired cases of malaria in the United States in 20 years. In the summer of 2023, seven cases of locally acquired P. vivax malaria were reported in Sarasota, Fla., and one in Cameron County, Texas. These are in addition to a case of P. falciparum diagnosed in a Maryland resident living in the National Capital Region.

Most malaria cases diagnosed in the United States occur in people who have traveled to countries in South America, Africa, and southeast Asia where malaria is endemic. While locally acquired mosquito-transmitted malaria cases can occur, as Anopheles mosquito vectors exist throughout the United States, they are rare. The last reported outbreak was in 2003 when eight cases of locally acquired P. vivax malaria were identified in Palm Beach County, Fla.

The GHIT award will allow Maher and Chet Joyner to develop a compound series drug-screening program. Joyner is an assistant professor in the College of Veterinary Medicine’s Department of Infectious Diseases and Center for Vaccines and Immunology and jointly appointed to CTEGD.

Microscopy image of Plasmodium vivax
Microscopy image of a P. vivax dormant (left, green) and growing (right, green) liver parasites inside of human liver cells (nuclei in purple). Image taken using 100x magnification. The dormant form survives most antimalarial treatments, but the new series of antimalarials kills both forms of the parasite. (Image credit: Wayne Cheng)

The compound series identified by Maher, the result of testing more than 100,000 samples using infected liver cells, is the first new chemical class discovered in more than 70 years with efficacy against the persisting liver stage. Over the next two years, Maher and Joyner will be collaborating with Medicine for Malaria Venture and Mitsubishi Tanabe Pharma Corporation to alter the chemistry of the compound to improve drug-like properties, including half-life and potency, necessary to achieve single dose criteria.

“Discovering a drug to kill dormant, non-proliferating cells is extremely difficult, yet with the novel assay the team developed we now have the first new target and drug class with potential to accelerate global malaria elimination efforts,” said Dennis Kyle, director of the CTEGD.

The current drug class used to treat P. vivax malaria, 8-aminoquinolines, often results in serious side effects and cannot be administered to pregnant women, who are one of the patient groups most in need of treatment.

“We have the first validated compound that kills vivax while it lies dormant in the liver,” Joyner said. “We hope in the next two years to help advance the new compounds to clinical testing.”

Lisa K. Nolan, dean of the College of Veterinary Medicine, said the work Maher and Joyner are doing could deliver a better quality of life to millions of people around the world.

“This great research is a shining example of our commitment to translational research, which will take this drug from the lab to preclinical testing to the patient rapidly,” Nolan said.

Time-resolved proximity biotinylation implicates a porin protein in export of transmembrane malaria parasite effectors

Figure 1 Generation of SBP1TbID mutants.
Generation of SBP1TbID mutants.

The malaria-causing parasite, Plasmodium falciparum completely remodels its host red blood cell (RBC) through the export of several hundred parasite proteins, including transmembrane proteins, across multiple membranes to the RBC. However, the process by which these exported membrane proteins are extracted from the parasite plasma membrane for export remains unknown. To address this question, we fused the exported membrane protein, skeleton binding protein 1 (SBP1), with TurboID, a rapid, efficient, and promiscuous biotin ligase (SBP1TbID). Using time-resolved, proximity biotinylation, and label-free quantitative proteomics, we identified two groups of SBP1TbID interactors: early interactors (pre-export) and late interactors (post-export). Notably, two promising membrane-associated proteins were identified as pre-export interactors, one of which possesses a predicted translocon domain, that could facilitate the export of membrane proteins. Further investigation using conditional mutants of these candidate proteins showed that these proteins were essential for asexual growth and localize to the host-parasite interface during early stages of the intraerythrocytic cycle. These data suggest that they may play a role in ushering membrane proteins from the PPM for export to the host RBC.

David Anaguano, Watcharatip Dedkhad, Carrie F Brooks, David W Cobb, Vasant Muralidharan. J Cell Sci. 2023 Sep 29;jcs.260506. doi: 10.1242/jcs.260506

Sheptide A: an antimalarial cyclic pentapeptide from a fungal strain in the Herpotrichiellaceae

Structure and amino acid sequence of the cyclic pentapeptide, sheptide A (1)

As part of ongoing efforts to isolate biologically active fungal metabolites, a cyclic pentapeptide, sheptide A (1), was discovered from strain MSX53339 (Herpotrichiellaceae). The structure and sequence of 1 were determined primarily by analysis of 2D NMR and HRMS/MS data, while the absolute configuration was assigned using a modified version of Marfey’s method. In an in vitro assay for antimalarial potency, 1 displayed a pEC50 value of 5.75 ± 0.49 against malaria-causing Plasmodium falciparum. Compound 1 was also tested in a counter screen for general cytotoxicity against human hepatocellular carcinoma (HepG2), yielding a pCC50 value of 5.01 ± 0.45 and indicating a selectivity factor of ~6. This makes 1 the third known cyclic pentapeptide biosynthesized by fungi with antimalarial activity.

Robert A Shepherd, Cody E Earp, Kristof B Cank, Huzefa A Raja, Joanna Burdette, Steven P Maher, Adriana A Marin, Anthony A Ruberto, Sarah Lee Mai, Blaise A Darveaux, Dennis E Kyle, Cedric J Pearce, Nicholas H Oberlies. J Antibiot (Tokyo). 2023 Sep 20. doi: 10.1038/s41429-023-00655-6.

Generating Genetically Modified Plasmodium berghei Sporozoites

Malaria is a deadly disease caused by the parasite Plasmodium and is transmitted through the bite of female Anopheles mosquitoes. The sporozoite stage of Plasmodium deposited by mosquitoes in the skin of vertebrate hosts undergoes a phase of mandatory development in the liver before initiating clinical malaria. We know little about the biology of Plasmodium development in the liver; access to the sporozoite stage and the ability to genetically modify such sporozoites are critical tools for studying the nature of Plasmodium infection and the resulting immune response in the liver. Here, we present a comprehensive protocol for the generation of transgenic Plasmodium berghei sporozoites. We genetically modify blood-stage P. berghei and use this form to infect Anopheles mosquitoes when they take a blood meal. After the transgenic parasites undergo development in the mosquitoes, we isolate the sporozoite stage of the parasite from the mosquito salivary glands for in vivo and in vitro experimentation. We demonstrate the validity of the protocol by generating sporozoites of a novel strain of P. berghei expressing the green fluorescent protein (GFP) subunit 11 (GFP11), and show how it could be used to investigate the biology of liver-stage malaria.

Carson Bowers, Samarchith P Kurup. J Vis Exp. 2023 May 5;(195). doi: 10.3791/64992.