Breaking through the basic biology of malaria relapse

Prof. Dennis Kyle, Prof. Elizabeth Winzeler and Dr Jetsumon Sattabongkot Prachumsri

Three discovery teams led by Prof. Dennis Kyle (Distinguished University Health Professor, University of South Florida, USA), Prof. Elizabeth Winzeler (Professor of Pediatrics & Director of Translational Research, University of California, San Diego) and Dr Jetsumon Sattabongkot Prachumsri (Head of the Mahidol Vivax Research Unit, Thailand) jointly received the MMV Project of the Year 2016 award for their impressive progress in developing new assay platforms to test compounds for activity against the liver stages of malaria. These new assays are now making it possible to screen and identify novel compounds that could stop the relapse and protect against malaria.

They talk about their achievements, the unique features of the assays they have developed and their plans for the future.

1. What can your team’s assay tell us and what’s unique about it?

Elizabeth Winzeler: Our assay uses the P. berghei (rodent) malaria parasite− we measure its ability to infect human hepatocytes (liver cells) in the presence of test compounds. We've shown that activity in this assay can predict whether or not a compound has chemoprotective activity. As the assay screens for activity against the liver stage it provides a kind of ‘filter’ to help prioritize compounds to be tested for anti-relapse activity. 

Speed and efficiency are really the key features we have been able to contribute. We now have a very robust and reproducible assay that allows us to screen literally hundreds of thousands of compounds at low cost. So we're able to cast a very wide net.

Jetsumon Sattabongkot Prachumsri: We have developed an in vitro culture system that enables us to see the impact of compounds on the small (hypnozoite) and large (schizont) liver forms of P. vivax. The unique element is that it uses a homogenous cell line that can be cultured in the lab, which means we are not limited by the supply of cells from donors. This cell line has minimal metabolic activity and so doesn’t degrade compounds, which makes it easier to identify those that are active.

Dennis Kyle: We have developed a 384-well plate assay and can routinely do in vitro radical cure screens and assess the activity of compounds against the hypnozoite. One of the unique features of our assay is that we've enabled primary human hepatocytes to maintain their physiology for over 30 days, allowing us to do experiments over a longer period, when required, which is key when looking at mature hypnozoites. We are then able to infect these cells with the sporozoites from mosquitoes and get higher infection development rates than we've ever seen, meaning we have a robust and reliable assay for screening campaigns.

2. How many compounds have you been able to screen and what has the assay told us so far?

Jetsumon Sattabongkot Prachumsri: We have tested 32 compounds while optimizing the 384-well plate format using two MMV compounds as a positive control for prophylactic mode. As our assay was the first to test compounds against P. vivax hypnozoites, it has helped us better understand how the ‘surrogate’ animal malaria assays that were being used before are predictive of the situation in human relapsing malaria. Once fully optimized, the 384-well plate assay will allow us to test up to 5,000 compounds this year in three concentrations, for their prophylactic and anti-relapse activity.

Elizabeth Winzeler: We have screened over 750,000 compounds provided to us by MMV and an additional ~250,000 compounds from MMV partners. There are some promising prospects – I am personally interested in compounds that could be used to protect people from getting malaria. My vision is to try to find compounds that could be used as a ‘chemical vaccine’ or a ‘chemical bed net’.

Dennis Kyle: We have screened close to 1,500 compounds, mostly in the previous 4−5 months. The breakthrough came at the end of 2016 and since then we’ve already rescreened all of these compounds to reconfirm our findings. We've also conducted concentration–response assays to quantify the potency of the hits. The assay has allowed us for the first time to screen against true P. vivax mature hypnozoites and has produced lots of information on these forms of the parasite. We now plan to scale-up this assay to be able to screen up to 15,000 compounds in 2017.

3. How did you feel when you found out your project had been selected for MMV’s Project of the Year 2016?

Dennis Kyle: I was happy for our team because they put in a lot of work to get to this point. I feel quite strongly, with MMV, that it is critical for us to identify new anti-relapse drugs, and this assay is an important tool in that endeavour.”

Elizabeth Winzeler: We really appreciate the recognition. It makes everyone on the team feel like their work is that bit more worthwhile. It's good for our team morale.

Jetsumon Sattabongkot Prachumsri: Our team feels very honoured to be selected as MMV’s Project of the Year for 2016. It indicates that MMV recognizes the difficulty of developing the assay and the great progress we have achieved in the past year.