Converting natural product leads into structure- and ligand-based drug discovery campaigns against leishmaniasis and Chagas' disease

Lead Research Organisation: University of Dundee
Department Name: School of Life Sciences

Abstract

The neglected parasitic diseases, visceral leishmaniasis (VL), and Chagas' disease (CD), together cause an estimated 50,000 deaths annually world-wide. Most of the people affected by these terrible diseases are amongst the poorest members of society and often live in remote rural areas. Despite this death toll, the current treatments available are inadequate for many reasons. These problems include side effects, compounds not working very effectively, resistance to current drugs and the requirement for intravenous administration or repeated injections which are not suitable for the rural environments where these diseases are typically found. Consequently, the development of new, effective and safe oral drugs provides an opportunity to save many lives and improve the quality of life for whole regions of the developing world, through the reduction of the morbidity, mortality and the economic consequences of these diseases. Unfortunately there are very few compounds being developed for VL and CD.

In this proposal we aim to identify potential new targets for drug discovery programmes. These are typically enzymes, which are vital to the life of the parasites that cause the diseases. We aim to identify these start points by testing natural products against the parasites. For those natural products that kill the parasites, we will then find out how the compounds are killing the parasites. Natural products are chemical compounds that are found from natural sources, such as plants and bacteria. They often have unique chemical structures, that have not been made before by humans and are difficult to make synthetically. Therefore they provide good tools to identify potential drug targets that often would not be found by other means. Once we have identified the molecular target (enzymes) we can then carry out a drug discovery programme, either by using these natural products as starting points or using conventional drug discovery approaches to find inhibitors of the enzyme and then optimise them into potential drugs.

An established, fully integrated team comprised of the University of Dundee and University of São Paulo, São Carlos brings together the broad range of expertise required to discover these much needed drugs. The team combines natural product expertise, world renowned parasitology, expertise in identifying the modes of action of compounds in these parasites, extensive experience developing molecules that can kill the parasites, and the substantial expertise and infrastructure required to develop drug candidates suitable for progress toward clinical trials.

Technical Summary

The development of new drugs for the treatment of kinetoplastid diseases is hampered by a lack of validated drug targets, and insufficient understanding of basic parasite biology. Here, we intend to leverage the more diverse chemistry available in natural product (NP) compounds active against Leishmania donovani and/or Trypanosoma cruzi to elucidate the MoA and enable initiation of novel structure based small molecule drug discovery programmes. Our approach to defining MoA utilizes a combination of established and novel techniques. These include:
1. High-throughput genetics: We will utilise our genome-wide RNAi library (RITseq) in T. brucei alongside our genome-scale overexpression libraries in T. brucei, L. donovani and T. cruzi. These global knock-down and overexpression methodologies enable us to rapidly assess virtually every gene in the parasite genome for their potential role(s) in resistance to the compounds we are screening. Specifically, we will identify genes, that when their levels are modulated (up or down), can confer compound resistance. We will also generate cell-lines that are resistant to our NPs in vitro. The resulting drug-resistant clones will be assessed by whole-genome sequencing to identify genomic changes responsible for resistance.
2. Cellular: We have developed a number of genetically modified parasite cell lines that enable us to report on specific cell and organellar functions (proteasome, glycosome) affected by our NPs. We also use a number of established reagents, e.g. MitoTracker, redox-active reporters. In a broad sense, these cell-based approaches provide an early and broad indication of the potential MoA our NPs.
3. Chemical proteomics: Our sophisticated mass spectrometry-based quantitative proteomics techniques allow us to directly identify proteins that selectively bind to bioactive compounds. Specific approaches that will be utilized include thermal shift proteomics (CETSA) and SILAC-facilitated pulldowns.

Planned Impact

Societal Impact:
The World Health Organization (WHO) has produced overwhelming evidence to show that the burden caused by many of the 17 neglected tropical diseases that affect more than 1 billion people worldwide can be effectively controlled and, in many cases, eliminated or even eradicated. This project aims to lift one of the barriers to effective drug discovery for visceral leishmaniasis (VL) and Chagas' disease (CD) through identification of well validated targets and thus help find new and more effective therapies to treat 2 of these 17 diseases; VL and CD that affect millions of people worldwide and cause an estimated 50,000 deaths annually. New treatments for either of these diseases would have a major impact on the health of millions of people.

Economic Impact:
The impact of neglected diseases, such as VL and CD, is also an economic one, impacting on the economic development of countries. This is due to loss of earnings and productivity due to ill-health. Unfortunately, the people affected by such diseases often cannot afford medicines to treat these diseases. Therefore it is important that the cost of goods for new drugs for these diseases is as low as possible. There is little or no commercial motivation or opportunity for profitability associated with the research and development for improved drug therapies for these diseases. Therefore, new affordable drugs to treat these diseases will have a huge impact on the economic prosperity of disease endemic areas, positively benefiting many millions of people.

Scientific Impact:
This UK-Brazil partnership will expand and enhance the scientific knowledge on the effective drug development opportunities for these neglected parasitic diseases. It aims to deliver new drug targets and novel compounds effective in the treatment of VL and/ or CD. Treatment of these diseases is dependent on very few, poorly tolerated drugs. Resistance is a real concern as is poor efficacy with current treatment options, particularly for chronic CD.

The data and knowledge that are generated during this project will also be important to un-ravel the basic biology of these parasites. New understanding of pathways and identification of potential new drug targets will arise from investigative mode of action work in this project. This knowledge will be made available to the wider community through publication and presentation at key meetings.

This project forms a powerful platform to drive translational research between the UK and Brazil, fueling the drug discovery pipeline against an important public health threat in Brazil. The partners will benefit from important transfer of technology and expertise in various aspects of kinetoplastid drug discovery. Building a strong working relationship between the two centres in the UK and Brazil through this project will, going forward, further benefit the two centres through future exchange of students and additional project development.

Publications

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Ortega HE (2021) Antileishmanial macrolides from ant-associated Streptomyces sp. ISID311. in Bioorganic & medicinal chemistry

 
Title Emily Fong WCAIR digital residence 
Description The works are primarily hand-drawn illustrations, painted with watercolours. They depict the researchers and Emily's interpretation of their conversations around their work and situations. They are then shown online, on both a website and on social media. 
Type Of Art Artwork 
Year Produced 2020 
Impact The work has been widely liked and commented on, particularly on Instagram. The posts themselves have received between 80 and 160 likes, with comments from both Wellcome centre of ant-infective research scientists and artists who had previously been entirely unconnected from the Centre. Comments have included: "Language can have magical qualities if it's used the right way" - Sha Sarwari, artist 
URL https://wcair.dundee.ac.uk/public-engagement/science-art/emily-fong-intro/keys-to-curiosity-you-give...
 
Description There have been substantial delays (technical challenges and COVID related) to being able to provide the exciting novel natural product isolates active against the parasites of interest (but selective against mammalian cells) to the quantity and quality required to start assessment of mode of action in Dundee. As previously reported, initial isolate supply in Brazil had decomposed over an extended period of time. Attempts to re-purify would not leave sufficient compound amount from the current isolate supply for the mechanism of action studies to proceed. New fermentations of the producing bacterial strain were therefore initiated in order to obtain more extracts for purification of the target compounds. However, there has been difficulty in reproducing the culturing conditions to get the same yields of compounds. The original condition and a further 6-7 additional conditions have been tried but the yields are very low. It is possible the original strain is losing the biosynthetic capability, a factor not so uncommon in microbial natural products research. To potentially overcome this issue, the first culturing of the bacterial strain that led to the isolation of cyphomycin and analogues was carried out by a former PhD student of the Brazilian team (Humberto Ortega) during a research internship at UW-Madison, with him then reproducing the conditions in the Brazil lab to finish structural elucidation. A supply of the crude extract remains in the US, so the Brazilian team have asked their US collaborator (Tim Bugni) to send it to them and they will try this as an alternative way to obtain more compounds as soon as the extract arrives.
As previously reported, a week of joint face to face meetings in Brazil in autumn 2019 significantly strengthened the relationship of the two research teams in Sao Paulo and Dundee, building the collaboration. Bringing together an industry experienced unit with world class infrastructure and capabilities in kinetoplastid drug discovery and mode of action deconvolution in Dundee with natural product expertise and disease biology expertise in Brazil offers significant opportunity for transfer of technology, novel approaches and drug discovery expertise in kinetoplastid research between the two teams and their respective units. It was hoped that movement of staff for specific training needs would commence in 2020. A specific one week drug discovery training programme was also scheduled in Brazil in April 2020 and a visit of the Brazilian team to Dundee in Summer 2020 planned. Unfortunately, however, this all had to be cancelled due to COVID. Discussions are now ongoing with regard to potential online training and development as travel between the two research groups remains highly unlikely in 2021 due to the continuing COVID pandemic.
In summary, the best part of a year has now been lost to the COVID pandemic on delivering progress in this research award. Brazil went into lockdown in late February 2020, only partially returning to lab activities at the end of October 2020, and then with only 30% of people allowed to work at the same time. Staff in Brazil are therefore currently doing lab rotations and all experiments are substantially delayed. Furthermore, since early January 2021 a new intense wave of COVID is underway in Brazil, worse than previous due to more transmissable virus variants and this has had a further substantial impact on research activities. As example, every time a COVID case is reported, the lab closes for 10 days.
Exploitation Route The aim of the proposed project is to identify novel pharmacologically validated drug targets and early leads for visceral leishmaniasis and Chagas disease with the ultimate goal to deliver much needed further potential new medicines for these diseases. This work will address multiple of the UN Strategic Development Goals (SDG): SDG1-No Poverty. These diseases keep people in poverty; SDG3- Good Health and Wellbeing. There is an urgent need for new drugs to treat these diseases; SDG4- Quality Education. This funding will provide training in drug discovery and mode of action studies in Brazil; SDG9- Industry, Innovation and Infrastructure. This will help develop this within Brazil.
Sectors Pharmaceuticals and Medical Biotechnology

 
Description The award is currently at a very early stage. However, the outputs of this project could ultimately lead to better understanding of the basic biology of the parasites causing visceral leishmaniasis and Chagas disease. New understanding of pathways and identification of potential new drug targets may arise and this will be made available to the wider kinetoplastid biology and drug discovery community. This will ultimately lead to potential new drug treatments for these diseases. Our track record in neglected disease drug discovery and world class infrastructure will allow rapid transition through early drug discovery. This work will address multiple of the UN Strategic Development Goals (SDG): SDG1-No Poverty. These diseases keep people in poverty; SDG3- Good Health and Wellbeing. There is an urgent need for new drugs to treat these diseases; SDG4- Quality Education. This will provide training in drug discovery and mode of action studies in Brazil; SDG9- Industry, Innovation and Infrastructure. This will help develop this within Brazil. Chagas disease and visceral leishmaniasis affect both genders and any new medicine will be used equally for both male and female patients. Indeed, one of the current standards of care against visceral leishmaniasis is teratogenic, and cannot be prescribed to women of child bearing age. A new treatment will therefore offer significant benefit to this group of patients.
First Year Of Impact 2019
Sector Pharmaceuticals and Medical Biotechnology
Impact Types Economic

 
Description Sao Paulo, Brazil 
Organisation Universidade de São Paulo
Country Brazil 
Sector Academic/University 
PI Contribution We are an industry experienced drug discovery unit with world class infrastructure and capabilities in kinetoplastid drug discovery and mode of action deconvolution of phenotypic actives against the parasites of interest. Our expertise, infrastructure and capabilities will be applied to this project to identify natural product isolate mode of action. However, our aim is not merely to identify the targets of natural products but also to develop tools that will allow us to leverage these targets for future drug discovery programmes. Training in specific techniques as well as broader drug discovery training will be provided to the Brazilina team by us as part of this partnership.
Collaborator Contribution The team in USP, Brazil bring disease biology expertise and significant natural product expertise. The team have already identified a number of novel natural product isolates active against the parasites responsible for visceral leishmaniaisis and Chagas disease, and demonstrating selectivity over host cells. They will use their natural product expertise to isolate and purify further quantities of these interesting actives for mode of action work at Dundee and drug discovery projects in partnership.
Impact Decomposition over time of some of the initial identified natural product isolates has occurred. Scale up and purification of further material is therefore currently ongoing in Brazil in order to provide sufficient pure material for mode of action studies to be triggered in Dundee. This collaboration is multidisciplinary involving parasitologists, medicinal chemists and pharmacologists. The significant opportunity that exists in this award for transfer of technology, novel approaches and drug discovery expertise in kinetoplastid research between the two partners and their respective units, has started. A synthetic series of chemistry in a drug discovery campaign against Chagas Disease at USP, Brazil has been profiled and closed down, unsuitable for continued drug development. This project has started to transfer drug discovery expertise to Brazil.
Start Year 2019
 
Title Oxadiazole derivatives 
Description At USP, Campinas, Brazil, a drug discovery programme on a series of oxadiazole derivatives active against Trypanosoma cruzi and Leishmania infantum, the parasites responsible for Chagas disease and visceral leishmaniasis respectively, was shared under this award collaboration. Mode of action was investigated and key compounds profiled to assess suitability for further progress toward development in these disease areas. Unfortunately, a generally reactive structural feature likely explains the pan-activity and delivers the lack of selectivity seen for these compounds. This series is therefore of no further interest to develop and further work is now focused on removing the reactive feature whilst retaining activity against the parasites. 
Type Therapeutic Intervention - Drug
Current Stage Of Development Initial development
Year Development Stage Completed 2020
Development Status Closed
Impact Drug discovery knowledge exchange between the Dundee and Brazil chemists. Improved understanding of compound developability issues in Brazil. 
 
Description Drug Discovery and Design 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact This was an event to engage with the general public around the science that we are doing. There were various hands-on activities for children
Year(s) Of Engagement Activity 2019
 
Description Emily Fong artist in digital residence project 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact This project is a digital equivalent of an artist-in-residence programme, as part of the Wellcome Centre of Anti-infectives Research (WCAIR) wider science art work. Emily is an artist who specialises in illustration, sculpture, and interdisciplinary collaborations. She interviews a number of WCAIR researchers about their work, with a focus on how the Covid lockdown of 2020 had affected it. She then created illustrations of them and wrote a blog post for each person. These were shown on the WCAIR website. She also shared work to social media, particularly Instagram, where posts often received over 100 likes. Blog posts on the WCAIR website have received over 500 hits.
We are planning for the work to be exhibited in person when the situation allows, and for Emily to speak about the work at various festivals.
Year(s) Of Engagement Activity 2020,2021
URL https://wcair.dundee.ac.uk/public-engagement/science-art/emily-fong-intro/
 
Description In conversation with Ian and Kev: Industry, Academia and Drug Discovery 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact This was a discussion around career paths for 2 members of the Senior Leadership Team of the DDU, Ian Gilbert and Kevin Read
Year(s) Of Engagement Activity 2021
 
Description Networking and Workshops in Brazil 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Study participants or study members
Results and Impact The drug discovery unit team on this award from Dundee visited the University of Sao Paolo, Brazil for one week, meeting the key Brazilian researchers on this award based at three different sites in Sao Paolo: Campinas, Sao Carlos and Ribeirao Preto. Networking and drug discovery workshops took place at each site with the lead researchers and their staff. A drug discovery case study presentation was also given to a broader audience at each site. This visit built and strengthened the collaboration as well as set goals and plans for further activities, including a focused medicinal chemistry and DMPK drug discovery training programme in Brazil to be held spring 2020.
Year(s) Of Engagement Activity 2019
 
Description Para-site-seeing Departure Lounge (Lifespace: Science Art Research Gallery Exhibition) 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact This was a science art exhibition, created as a collaboration between artists from the University of Lancaster and scientists from the Wellcome Centre for Anti-Infectives Reseach. Set up as a 'travel blog' written by one of the parasites the Centre fights (leishmania), it attracted over 500 visitors. Scientists, including myself, acted as a source of information and inspiration for the art, as well as providing technical support on some of the microscopy elements. The work has since toured to Dundee Science Centre and we have ambitions to take it further around the world, to disease-endemic countries. Feedback from the exhibition showed that audiences had learned more about the parasite and disease it causes, as very few had had much previous knowledge. It also showed that the public saw the scientists as real people carrying out a worthwhile mission, and an increased closeness to science.
Year(s) Of Engagement Activity 2019
URL http://lifespace.dundee.ac.uk/exhibition/para-site-seeing-departure-lounge
 
Description Parasites: Battle for Survival, National Museum of Scotland, Edinburgh 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact Susan Wyllie was involved in recording an extract for the exhibit describing her work in parasitology and also the impact of the work that her group does. This recording and several images form part of the museum exhibit. This exhibit attracted 25,000 visitors since Dec 2019 and ran until April 2020.
Year(s) Of Engagement Activity 2019,2020
URL https://www.nms.ac.uk/exhibitions-events/exhibitions/national-museum-of-scotland/parasites/
 
Description Parasites: Battle for survival exhibition 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact One of our team made a recording describing her work in parasitology that features in the exhibit at the National Museum of Scotland, Edinburgh. This exhibit is focused on drawing attention to the field of parasitology and specifically Scotland's contribution to this field. To date more than 25,000 visitors have viewed the exhibition. It is hoped that this exhibit will inspire the next generation of parasitologists.
Year(s) Of Engagement Activity 2019,2020
URL http://www.nms.ac.uk/exhibitions-events/exhibitions/national-museum-of-scotland/parasites
 
Description Rotary Club visit 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact I gave a talk to Dundee Rotary Club on neglected tropical diseases and the drug discovery efforts ongoing in the Drug Discovery Unit, Dundee to combat some of these diseases.
Year(s) Of Engagement Activity 2019
 
Description Street Food 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact 65 adults attended the School of Life Sciences for an evening event as part of both Dundee Science Festival and the University's Festival of the Future. Included in the attendees were secondary school teachers from local schools and members of local community groups. They were invited specifically to allow them to learn and ask questions inspired by our research and facilities which will allow us to continue to build relationships with these groups for future activity. The event combined food and drink makers, from Dundee, Angus, Fife and beyond, with our own researchers to explore the surprising, inspiring ways that research meets repast. Scientists and makers set up stalls together with activities and samples, explaining their work and the links. Examples included the links between malaria and quinine in tonic and artemisinins in absinthe, as well as bacterial biofilms and ice cream. Lab tours were also available. The audience reported that they had enjoyed the event, particularly the chance to meet scientists in such an informal, friendly setting. Their general feedback about Street Food ranged from "How approachable all the different research information tables were" to "loved how interactive it was" to "speaking to the researchers about their amazing work".
Year(s) Of Engagement Activity 2019
URL https://wcair.dundee.ac.uk/events/street-food-3/