Overview

Despite the great strides modern medicine has made in the management of diseases, a large population of the world today is still afflicted with entirely preventable and treatable diseases such as malaria, tuberculosis and diarrheal diseases. Many of these diseases have roots that stem from poverty, which often results in poor nutrition and lack of access to proper sanitation, healthcare and health education. In fact, the WHO estimates that diseases associated with poverty account for 45 per cent of the disease burden in the poorest countries (WHO, World Health Report, 2002).

Our project was thus motivated by the need to find a solution to this pressing problem which takes an immense toll on millions of vulnerable people around the world. We felt a strong social and moral responsibility to play our part in solving this crisis, using the particularly relevant tools and our exposure to and knowledge of synthetic biology, which would allow us to make fruitful contributions to this cause.

As we come from the tropical country of India, we looked around to find a tropical disease that we could work on as we felt we could make the most impact here. Given our own struggles with battling mosquitoes during the rainy season, a contender came to mind: malaria. In the beginning, we weren’t particularly aware of the actual extent and prevalence of the disease, but with every research article and study we stumbled across, our resolve to work on the disease grew stronger. Zeroing in on what exactly we would do to help eradicate the disease became particularly simple when we learnt about the increasing resistance of malarial parasites to existing treatment methods, and we set off to create a Library of drugs that would be cheap and accessible and overcome the problem of resistance. We also sought to create a diagnostic tool, since the highest incidences of malaria occur in rural areas, where the inaccurate diagnosis of malaria is a major issue due to a lack of appropriate infrastructure, causing both preventable deaths and underreporting of Malaria cases.^[Source] Our diagnostic kit’s innovation has been the development of a Deep Learning software that can diagnose infected samples based on images of blood smears. For more information about this, visit the Software wiki page.

At the end of the day, the main drivers of our project were the vulnerable, poor people afflicted with a preventable disease with no solution in sight.

The major stakeholders in our project are everyday people at risk for contracting Malaria, doctors, scientists, and officials in the healthcare industry. We set out to interact with people representing all these stakeholders.

A major player in malaria eradication is public health education as malaria is easily preventable by preventing stagnant water collection, using mosquito nets and insecticide sprays and so on, which are very simple and cheap strategies to prevent the disease. In order to understand where the gap was, we conducted surveys to gauge the attitudes of people toward the disease, their knowledge about the disease and ways to prevent it and accordingly designed a public education programme to fill in the pre-existing gaps in knowledge and to try and change the lax attitudes we commonly encountered, since people thought of malaria as equivalent to something like the seasonal flu, and to convey the seriousness of the disease, and the absolute importance of preventing it.

We also spoke to doctors like Dr. Priyanka Devgun, who helped us understand the need for a project like this and how to better it.

Dr. Velavan and Dr. Gamboa who are leading Malarial scientists, were some of the scientists we spoke to to gain insight into current research being undertaken to combat this deadly disease. We also spoke to executives and senior management from a healthcare allied company in India (Star Health) who enlightened us on some industrial perspectives.

Expert Interactions

Dr Dionicia Gamboa

Associate Professor, Department of Cellular and Molecular Sciences, Faculty of Sciences and Philosophy, Universidad Peruana Cayetano Heredia (UPCH)

"Working with Malaria is interesting but very difficult. The same tools and strategies cannot be applied everywhere."

Key Insights

Accessibility of rural/ tribal populations to drugs and diagnostic methods is an issue (in Latin America, but the same issue has been seen in India)
Asymptomaticity of the disease is a very pressing problem
New and improved diagnostic methods will be vital to eradicate Malaria

Incorporation into the Project

We started looking into the development of a diagnostic tool that could detect Malaria in remote areas.
We considered the development of a oral drug more strongly because of accessibility issues.

Summary

Dr. Gamboa and her fellow researcher, Elizabeth, said that targeting membrane interactions between Plasmodium falciparum and erythrocyte proteins should be enough to treat Malaria in the human body. The plasmodium proteins for the interactions we decide to target must be specifically chosen depending on the regional strains we decide to develop a drug against. Each strain of the parasite will have different surface proteins and thus, different interactions with erythrocyte proteins. A good idea would be to target one protein from each family of invasion proteins, thus ensuring we would be able to effectively treat the disease.

In Peru, most Malaria cases are in the Amazonian region, and around 80% of cases are caused by P. vivax. Accessibility to some of these regions is very difficult, and this causes differences in the epidemiological context of how the disease spreads. Microscopy is the gold standard to detect and diagnose Malaria, but other diagnostic techniques are also used. These other methods show that asymptomaticity is a huge problem. Many people have such a low parasite load that microscopy can’t detect it. Sometimes, LAM based techniques and PCR can’t detect the cases. Multiple test trials are required to show that the person is uninfected. According to Dr. Gamboa’s observations, asymptomatic cases are 6x and 4x more likely than symptomatic infections of P. falciparum and P. vivax respectively.

Dr Velavan

Dr Thirumalaisamy P Velavan

Professor and Group Leader, Institute of Tropical Medicine, Universitätsklinikum Tübingen, Tübingen, Germany

"Ambition combined with a systematic approach is the need of the hour."

Key Insights

Plasmodium falciparum is responsible for most deaths due to Malaria; it causes a virulent strain as it sequesters inside the human body
Different strains and mutations in Plasmodium falciparum has led to the rise of ‘delayed parasite clearances’/ resistance to currently used drugs.
Drug development process has multiple phases and the process can takes 12 - 20 years and around 30 - 60 million USD

Incorporation into the Project

Decided to continue working primarily on Plasmodium falciparum, and not shift focus to Plasmodium vivax.

Summary

Plasmodium falciparum is responsible for the more virulent kind of Malaria as sequestration happens with falciparum malaria. This means it would make more sense for our team to work on developing a drug to combat Malaria caused by Plasmodium falciparum, as compared to Plasmodium vivax.

Artemisinin Combination Therapies have been developed and involve the drug Artemisinin in combination with other antimalarial drugs. Each country or region of the world has its own combination therapy consisting of Artemisinin and other secondary drugs. These combination therapies work based on the fact that Artemisinin is an effective drug to combat Malaria, but it has a short half-life, allowing it to quickly wipe out most of the parasite load. The other drugs in the therapies are less effective but have longer half-lives, allowing them to clear out the remaining load over time.

Different countries also have different Plasmodium parasites responsible for Malaria; strains such as Plasmodium falciparum 3d7 and Plasmodium falciparum 7g8 are strains that have entirely different geographical distributions. The different alleles are also different for different MSP-I and MSP-II markers once these parasites circulate in the human body.

Mutations that confer resistances to many of these drugs (such as the mutation A76T in the Chloroquine Resistance Transporter - PfCRT gene that confers resistance to Chloroquine) are on the rise, that this means the drugs used in these therapies get swapped out for other drugs. This has happened multiple times, and these drugs have different targets. Instead of calling it resistances, it is better to term it ‘delayed parasite clearances’, as although the parasite gets cleared out of the human body, it takes much longer for the drug to do this. These strains that show delayed parasite clearances to drugs are especially common in Southeast Asia. The bioavailability of drugs also differs between populations.

He said that our team should also cross-reference and look at available databases, such as the one from Glaxo Smith-Kline, that has candidate drug targets listed. We can check if our candidate is in that database and look at whether testing, either in-vivo or in-vitro has already occurred for this candidate.

The process of drug development is very complex. As students, he urged us to atleast go on with in-vitro experiments that would prove the efficacy of the candidate drug before proceeding to in-vivo experiments. Testing would have to be done carefully, as mouse models prove to be useless when studying Plasmodium.

Provided our experiments are successful, Professor Velavan also suggested that there were multiple international organizations we could team up with organizations such as the Medicines for Malaria Venture (MMV). The pharmacokinetic profile of our candidate drugs would need to be studied. We would have to find its ADME profile (absorption, distribution, metabolism, and excretion) to see how it would react in the human body. We would also need to study its bioavailability to understand toxicity.

After this is when we would move on to clinical trials in multiple phases to verify safety and tolerability. Here is where we would study interactions between multiple drugs as well to study any effects. Genetic heterogeneity might play a role in the efficacy of the drug, so that would have to be studied. After data from phase 3 trials, certification from regulatory drug agencies, such as the FDA, would be possible. Phase 4 trials occur after this; the entire process can take 12 -20 years and costs 30 - 60 million USD. He advised to look at the PlasmoDB and other databases for available literature, both for candidate molecules for drugs and candidate drugs.

Finally, Dr. Velavan also briefed us on new diagnostics methods to detect Malaria that were being tested out in the field.

Dr Priyanka Devgun

Professor & Head of the Department of Community Medicine, Sri Guru Ramdas Institute of Medical Sciences and Research, Amritsar, India

"Nothing will work without community participation."

Key Insights

Malaria still persists today because it is not a major priority in society.
To effectively fight Malaria, community participation is of utmost importance.
How to design effective public awareness programs.
To ensure maximal accessibility our drug should be orally administered.

Incorporation into the Project

Re-designed and refined our public health awareness strategy.
Decided on creating an orally administrable drug instead of an injectable and modified drug design accordingly.

Summary

We met with Dr Priyanka Devgun to gain insights into the attitudes of the general public toward Malaria and their perception of the disease. We realised that the high prevalence of Malaria even today is a result of a complex interaction of many factors and that we could not hope to eradicate the disease without addressing all the individual factors.

One of those factors happens to be community participation or lack thereof. According to Dr Devgun, one of the main reasons that Malaria still persists in India today is that it isn’t a priority, either for the government or the general populace. Malaria is not a “felt need” in society and thus efforts to prevent and eradicate it are dismal.

If we wanted to make a meaningful impact on the pervasiveness of Malaria in society, we would need to not only make people aware of the mechanisms of transmission and the ways to prevent contracting the disease but also make people understand the severity of the impact of Malaria and its implications in society. We would need to make people care enough so that they felt a need to participate in efforts to curtail Malaria.

She gave us a lot of tips on how to build an effective awareness campaign, through the use of social media, webinars, infographics, informational videos etc, after which we were able to come up with a comprehensive public health awareness program.

We also talked to her about drug design since we wanted to ensure our drug was accessible to as many people as possible, cheap, and wouldn’t require much technical knowledge to administer. She suggested we design an oral drug as opposed to an injectable one since that would be the most effective way to reach even remote populations.

Dr David Norman

Reader in Structural Biology, College of Life Sciences, University of Dundee, Dundee

"If you are going to design a cyclotide with a therapeutic peptide insertion, there are some good places to put it but remember to consider the innate functionality of cyclotides which is often somewhat cytotoxic."

Key Insights

Helped us by providing a tutorial for doing homology modelling of cyclic peptides
Gave insights on future aspects of dry lab works

Incorporation into the Project

With his help, we were able to graft our designed inhibitors into the cyclotide backbone and perform MD simulations on it. Hence we could study the stability of the grafted cyclotide in silico.

Summary

Dr David Norman is a structural biologist who has expertise in Magnetic Resonance, FRET, EPR, Spin-labeling and PELDOR. He uses a variety of biophysical steps to investigate the structure and function of Proteins and Nucleic acids.

We first came to know about Dr Norman from the Svangård et al. 2003 article while we were searching for ways to do homology modelling for circular peptides. Dr Norman had provided the authors of this paper with a special patch for MODELLER for making a homology model for a cyclic peptide. He was very excited that we contacted him for help regarding the work he had done long back and tried to locate the patch but with no luck. Dr Norman then provided us with a tutorial which was on the MODELLER wiki describing ways to model a cyclic peptide, which we had never come across. The tutorial had errors which he pointed to us and gave solutions.

He told us that since the innate functionality of the cyclotide is somewhat cytotoxic we should insert the designed peptide at a good place to overcome this. He also suggested trying to shorten and lengthen the inhibitor to see how restrained things were. He also generously provided insights into the results we got from the MD simulations.

Poornima Raveendran

Senior Teaching Associate

"Your project is a very good attempt in applying available knowledge and using pocket-friendly resources for problem solving. An ergonomic approach!"

Key Insights

Software needs to be further developed to better aid the diagnostic kit.
Accessibility to the kit might still be an issue. There is a need for instructional videos to be made available in local languages.

Incorporation into the Project

As part of the future implementation of our project, we plan to develop the software further so that it can detect Malaria without the currently required preprocessing of the image.
We aim to translate our instructional videos into vernacular languages so maximum people can benefit from it.

Summary

One problem Poornima mentioned that could arise is the stain that is used for the blood smear and the concentration of the stain. Giemsa is commonly used but there could arise several problems with respect to its preparation in remote areas. Often this stain has to be made fresh and used immediately. Sometimes another stain is used like Leishman’s but she was unable to inform us of how that would work as she hadn’t worked with it.

Currently the software is able to recognise only RBC images that have been cropped and processed as given in the dataset. But what would really make this tool complete is if the software also identified the RBCs from a given Field of View, and then processed them accordingly to make them suitable for analysis.

Another suggestion was based on something known as Percentage Parasitemia. This is something that is calculated based on the number of infected RBCs in a given blood sample. It is usually used in the diagnosis of P. falciparum malaria. We can add another function to the software of calculating the percentage parasitemia (since it would already be able to identify RBCs and whether they are infected or not).

Poornima also recommended that the explanations in all videos need to be made available in the local languages. She pointed out that the instructional paper centrifuge video does not explain what sample holders are provided/can be used, how much sample needed, where and how to place the samples.

Feedback from Doctors

Our team developed a diagnostic kit that would be easy to use in rural areas. Oftentimes, there is a lack of a technical expertise of healthcare workers to effectively diagnose Malaria using blood smears - currently the gold standard of testing. The software our team developed would automatically be able to detect and diagnose infected samples using images of blood smears that had been made using the easily assembled, and cheap foldscope and paper centrifuge.

To use such a kit, we made an Instruction Manual in English with links to some videos that would help the users understand how our kit could be used.

To understand the feasibility of this kit and to get feedback on the instruction manual we’d made, we reached out to doctors who had previously worked in/ currently work in rural Primary Health Centres (PHCs). Primary Health Centres are the basic centres of public health that are set up by the Government to provide healthcare services to people.

This was their feedback:

Dr. Nilofar Bijali, previously worked in a public health organization in a tribal area (Gadchiroli, Maharashtra, India): She said tests would need to be conducted to establish the advantage of such a system of testing over Rapid Diagnostic Tests (RDTs) currently used in rural areas. She appreciated the innovation we had made.

Dr. Rituja Sardesai, previously worked in a rural PHC (Parinche, Pune district, Maharashtra, India): She said that the videos in the instruction manual were extremely self explanatory. The paper centrifuge video was quite pictorial and easy to understand by just watching. The foldscope video might require translations into regional languages.

Dr Akshay Wagh, currently works at a rural PHC (Taluka Bhamragad, Gadchiroli, Maharashtra, India): He said that it would be important to perform field studies in regions of India severely affected by Malaria before continuing the development of such a kit. He praised the use of the foldscope in a kit like this but mentioned that efficiency studies would need to be performed on the efficiency of a foldscope versus a conventional microscope.

Dr. Roshani Rameshwar Sagane, currently works at a rural PHC (Jogawadi, Pune district, Maharashtra, India): She recommended that we add maximum number of pictures and flowcharts to our instruction manual that we have developed. She also recommended that a minimum of one or two foldscopes must be made available to each PHC.

Based on their feedback, we have plans to translate the foldscope video into regional languages. We also plan to make our manual easier to use and more pictorial. Check out our Future Implementation page here to know how we plan to take this forward!

Sriram Raghavendran

Joint Executive Director, Star Health and Allied Insurance

"The simpler the solution, lower is the cost and greater are the chances to achieve higher scale."

Key Insights

In order to succeed with an endeavour like this, it must be simplified in an efficient way.
Splitting the kit into two components, Reusable (foldscope, paper centrifuge, etc.) and Consumables (disposables needed for every patient), will ensure that maximum patients can be targeted with this technique.
Pre-assembly of foldscopes is recommended as assembling them might be difficult for rural healthcare workers.

Incorporation into the Project

Decided to proceed with the splitting of the kit into two components, Reusable and Consumables..
Our team is now looking into whether pre-assembling the foldscope before sending it out could be possible.

Summary

We spoke to Mr. Sriram Raghavendran, an executive at Star Health and Allied Insurance, as we felt that feedback from someone in the healthcare industry with managerial experience would be important for us to better our project. According to him, based on the YouTube video of the foldscope, assembling the foldscope from scratch might seem like a fun exercise but it seems intellectually demanding and totally unnecessary. He suggested that it would be much better to distribute the pre-assembled foldscope to rural areas rather than expecting them to follow instructions and assemble the same. There is no apparent advantage to the healthcare workers self-assembling the foldscopes, as there is no difference in shipment size between the unassembled and assembled foldscope.

He also mentioned that a malaria testing kit would have two distinct parts: Reusable Equipment (such as an assembled Foldscope, paper centrifuge, etc) and consumables per test (such as syringe, paper strip for smear, etc). The consumables will be for each patient. Mr. Sriram recommended that creating a single diagnostic kit with both the reusable and consumables was a waste of space and was inefficient, as when packed together, we would practically only be offering a few tests per kit. When you create separate Equipment and Consumables kits, the same Foldscope can be reused for hundreds of tests, and one consumable kit can contain consumables that would be enough to test a given number of people (say 50). These consumables could then be resupplied whenever necessary. These couple of simple design modifications will simplify the solution and reduce the cost per test further.

Survey

Our survey aimed to collect inputs from people on their understanding of Malaria and its severity. In order to reach the most people we could, we translated our survey into 11 vernacular languages. Through a collaboration with UPCH Peru, we translated our survey into Spanish as well!

The findings from the survey served as a reinforcement of why this project is so important and how people severely lack awareness about the extent and seriousness of Malaria.

In terms of who actually responded to our survey, around 50.3% were male and 45.6% were female. 0.1% of our respondents selected the option ‘Other’, while 4% of our respondents chose not to answer.

44% of our respondents said that they never used a mosquito net while sleeping during the monsoon seasons, while the remaining 56% of respondents used them in varying degrees. This figure shocked us, as mosquito nets are a cheap and effective way to significantly reduce the risk of contracting mosquito borne diseases.

According to the WHO, globally, around 405,000 people die due to Malaria every year. This translates to an average of 1,100 deaths every single day. A little over half of our respondents thought less than 1,000 people died every day.

Around 30% of our respondents were not aware of any cure or treatment existing against Malaria! This was shocking given the fact that Antimalarial drugs such as Hydroxychloroquine have been in the news with regards to CoVID-19. This reinforced the fact that there is low awareness amongst people of treatments against Malaria.

There has only been one vaccine that has been approved for use against Malaria, the RTS, S vaccine. This requires four doses and has a low efficacy. This vaccine has not been publicly released in India and trials are still ongoing.

36% of our respondents said they would strongly prefer an oral method of medication while only 14% said they would prefer an injectable form. Around 50% of our respondents said either method would work. This showed us that there was a preference for an oral drug.

Accessibility to hospitals is a huge issue in India, especially in rural areas. Due to the pandemic situation, our surveys were online, which might have caused some bias in the sort of people who were able to access and respond to our surveys. Our translated surveys were an attempt to correct some of this. Surprisingly, 8% of our respondents lived over 10 km away from the nearest hospital that could treat Malaria, and 1% of the respondents lived over 50 km away from the nearest hospital that could treat malaria!

Malaria has multiple symptoms such as fever, sweating, headache, nausea and vomiting, muscle pains and diarrhoea. We asked our respondents to pick the symptoms of Malaria they were aware of. Most of our respondents (94%) were aware of fever and chills being a symptom. Fewer people were aware of the other symptoms of Malaria - sweating (46%), headache (55%), nausea and vomiting (55%).

We have compiled our survey data into a Table.

The results of the survey gave us some very useful insights. We learnt about the general public's preference for an oral versus an injectable drug and modified our drug design approach in alignment to engineer an orally administrable drug. We also got insights about the exact knowledge gaps about malaria that existed in the minds of the general public and geared our Human Practices accordingly to bridge this gap through activities like a webinar, radio show and a song to disseminate information about how to tackle Malaria.

Team:IISER-Pune-India/Human Practices

Overview