To identify well tolerated, new molecules active against the dormant liver form of P. vivax (the hypnozoite) MMV has put in place a pragmatic cascade of tests. Compounds known to be active against blood-stage parasites are screened first against a rodent malaria liver assay to test them for activity against liver-stage schizonts. Active compounds are then progressed to a primate malaria in vitro assay followed by an in vivo model to test for activity against the hypnozoites.
The limitation of the current test cascade, however, is that we are not testing against parasites that infect humans (P. vivax) and so might miss some molecules. Additionally, the throughput of our current assays that look at the hypnozoite is limited. To overcome this, MMV and the Bill & Melinda Gates Foundation are working with different research groups to develop a cost-effective P. vivax cell assay.
In a major step towards that goal, a team led by Massachusetts Institute of Technology (MIT) researchers has now developed a system to grow liver tissue that can support the liver stage of both Plasmodium falciparum and P. vivax malaria.1 Prof Sangeeta Bhatia explains her interest in this area of research and how the system works.
1. As a professor of electrical engineering specializing in computer science, how did you come to be involved in malaria research?
A lot of our work is about leveraging engineering tools for medicine. Many tools for computer chip manufacturing have been very useful for manipulating cells. We had been using these chips to manipulate liver cells and grow implantable livers for patients. About 5 years ago, after the renewed call for malaria eradication, the Gates Foundation hosted a liver-stage meeting. They had seen our work and invited us. That’s where it all began. It was a combination of opportunism, experience and interest in making a global impact.
2. What progress has been made in the development of a P. vivax (hypnozoite) cell-based assay to identify new anti-relapse medicines?
We knew it would be a big challenge to grow the hypnozoite in the lab; everyone we spoke to in the malaria field was understandably sceptical. Sanaria, a biotechnology firm, was able to provide us with cyropreserved parasites, which means we were not dependant on fresh supplies. As the project progressed, led by Sandra March, we were able to get full liver-stage maturation of P. falciparum parasites.
We then started the P. vivax experiments with the help of many different groups, including Sanaria. Eventually we were able to see a sub-population of liver forms progress, while a sub-population persisted and became dormant. It is the latter population that we termed “persistent small forms”. Before we can really call them hypnozoites we need to see reactivation. The other thing we are looking at is differential drug sensitivity, which is work planned for next year. It has been really gratifying to get a glimpse at what we believe is the hypnozoite. Of course, there will be lots more to do if we can really reproduce it in this way. We’re really excited!
3. How do the assays work?
The assay we have working at the moment is really for P. falciparum; for P. vivax we have only achieved feasibility. For both assays, you start with a well plate of microcultured human livers, with hepatocytes in colonies of 250 cells, on islands of collagen surrounded by feeder cells that help to support their differentiation.We then add parasites directly onto these media and essentially allow them to progress through the liver stages. First, they glide and traverse several hepatocytes and then they will choose to set up shop in one. You are then left with a well plate of infected liver cells, to which you add the drug. You then stain and count the forms that remain to determine if the drug worked.
4. What has it been like to work in the malaria community?
Once we were introduced to the community and they embraced the potential of our assay, their input was transformative. We have had access to expertise, reagents, parasites and decades of research that wouldn’t have been available to us otherwise. Credit goes to the Gates Foundation and MMV for being visionary. They really took a chance on us, a group of complete outsiders to the malaria community, and we hope to live up to their expectations.
1. March S et al. “A microscale human liver platform that supports the hepatic stages of Plasmodium falciparum and vivax.” Cell Host Microbe. 14(1):104-15 (2013).