Before the COVID-19 pandemic, tuberculosis (TB) was the deadliest infectious disease in the world. Now, just three years later, TB is unfortunately back on top. While the urgency of the pandemic necessitated the rapid development of diagnostic tests for SARS-CoV-2, TB diagnostics are not as simple as a nose or throat swab. In the low-resource settings most affected by TB, clinicians use smear tests that were developed more than 100 years ago to diagnose the infection. These tests vary in sensitivity from 32 to 94 percent depending on the administrator’s skill (1). To confirm the diagnosis, technicians culture the sample, which can take weeks for patients to get their results back, delaying treatment.
University of California, Los Angeles (UCLA) bioengineer Mireille Kamariza plans to change that. As a child growing up in the small central African country of Burundi, Kamariza wondered why infectious diseases like TB affected so many people in her country but not in wealthier nations. When she moved to the United States at age 17, she knew that she wanted to use her interest in chemistry to address the burden of infectious diseases facing her community back home.
During her PhD studies, Kamariza developed a fluorescent probe that selectively binds live TB bacteria, diagnosing patient samples within minutes. Soon after, she founded the biotechnology company OliLux Biosciences to further improve her TB diagnostic dye for people living in areas where TB is common such as sub-Saharan Africa, Asia, and Eastern Europe. With a fast and clear diagnosis of TB, Kamariza hopes that her new probe will lead to faster treatment and save more lives (2).
What made you decide to pursue science as a career?
When I was a kid, I had a natural curiosity about the world around me. I wanted to understand how a plant can thrive when there's virtually nobody watering it or people are stepping on top of it every day. I wanted to know things like why some stars shine brighter than others. Marveling at the world around me pushed my education towards science. I really enjoyed chemistry and learning how it explains how molecules and bonds and atoms interact with each other. Because of those initial interactions, there is now this polymer or this drug or this cell that behaves in a particular way.
A serendipitous string of events led me to study TB. I was born in Burundi, which is affected by a lot of infectious diseases, not just TB but also malaria and many others. I grew up understanding how impactful those are. When I moved to the United States, I was afforded the opportunity to do research, and I really wanted to work on a project that would help places like where I came from.
My training was in chemistry, and back in the day, there really wasn't a large overlap between chemistry and infectious disease. People often use chemistry to do drug discovery for cancer or brain research but almost never for infectious disease. When I went to the University of California, Berkeley for graduate school, there was only one person working on chemistry and infectious disease, Carolyn Bertozzi. When I talked to her about how I wanted to cure infectious diseases, she said, “Yes, you go girl!” I loved that energy, and I ended up joining her laboratory.
When I joined Carolyn's lab, she had a few graduate students and postdoctoral researchers doing a genome wide CRISPR screen to look for novel therapeutic targets in TB. They learned very quickly that to do this, they needed to grow these cells out, and TB has a doubling time of 24 hours. It would take months to be able to study them, and graduate students don't have that kind of time. This was also a chemistry lab, so people expected a faster turnaround.
Because Carolyn was studying sugar chemistry, the team decided to leverage the sugars that decorate the TB cell surface to deliver a probe for the genome screening. They wouldn’t have to grow the cells for months. They used click chemistry and bioorthogonal reactions, which Carolyn developed in her lab and won the Nobel Prize for last year, and they found that the sugar trehalose worked great for this purpose (3).
I joined the lab when that paper got published, and my original goal was to use this probe to understand the important metabolic pathways in TB and to find some therapeutic targets. But I wondered what else we could do with the probe. It worked so easily and quickly, so I wanted to test it in diagnostic spaces. Carolyn really latched onto that idea, but the original click probe required a laborious preparation process. That pushed us to design a probe that would work outside of the lab.
How did you develop that probe?
I had some of the best chemistry cohort mates. We did journal club together, and Peyton Shieh, a fellow graduate student in the laboratory, found this paper about a probe that inherently changes fluorescence if put in environments with different polarities (1,4). The particular one that we landed on was 4-N,N-dimethylamino-1,8-naphthalimide (DMN). In highly hydrophilic environments like water-based solvents, the vibrations of the photons in the probe are stable, so no fluorescence occurs. But if the dye is in a hydrophobic space, vibrations get induced in the molecule, and that kicks off photon emission and we see florescence. The beauty here is that the TB mycomembrane (the outer membrane layer) has this uniquely rich layer of fatty acids, making it hydrophobic. We hypothesized that if we put this probe outside of the bacteria, we wouldn’t be able to see it, but once it got inside the cell membrane, we should see it. So, we loaded the probe onto the sugar trehalose.
Carolyn didn't think that it would work because of the chemistry of the probe itself. It requires such a particular condition to turn on that it might not function properly in the messy and chaotic intracellular space, but we went ahead and tried it. Peyton and I were in the microscope room, and I thought I saw something. We zoomed in on the picture, and then he said, “Oh my God, it worked!” He took the picture, and we ran into Carolyn’s office. It was quite exciting, but of course because we’re scientists, she immediately asked, “Did you have controls?" We had to do a lot of characterization to make sure that what we thought was happening was actually happening, but it was a good day.
How is the development of your TB probe coming along now?
When I began my postdoctoral fellowship at Harvard University in the fall of 2019, I started OliLux Biosciences. We're a public benefit company because our target market is low-income countries, so we're not going to be the Eli Lilly or Pfizer of diagnostics. I did that on purpose. I want to work on projects that will help my community. There are also low-income areas here in the United States that could benefit from this technology, so the company is primarily funded by foundations that understand our mission.
When we founded the company, I had an ongoing Gates Foundation collaboration in South Africa where we were planning to start a large clinical trial testing the DMN probe for TB patients. Then in March of 2020, everything stopped. We were essentially shut down, but we started coming back in 2021. We split our clinical trial into two smaller studies: one in Uganda and one in Vietnam. We started recruitment this year in January, so that's finally on track.
You recently developed a brighter TB probe. Will that one also undergo clinical trials?
Right now, we’re using DMN in the clinical trials because that's the one that has the most data. DMN has been tested on patient samples in South Africa, and those results are in our earlier publication (1). But, when I was doing those experiments, I realized that DMN was actually quite dim. We went back to the drawing board and did a bit more chemistry. We developed a new probe based on the 3-hydroxychromone (3HC) dye that has a higher fluorescence intensity than DMN (2). Because it’s brighter, it could work better with some of the low-cost microscopes that are available in clinics in South Africa and elsewhere.
My goal right now is to validate that this probe works in patient samples and that it works better than DMN because we must see a net benefit. We have those two probes, and now we're developing a third one. We hope to create a class of molecules that can be selected depending on the conditions or available resources.
This interview has been edited for length and clarity.
- Kamariza, M. et al. Rapid detection of Mycobacterium tuberculosis in sputum with a solvatochromic trehalose probe. Sci Transl Med 10, eaam6310 (2018).
- Kamariza, M. et al. Toward Point-of-Care Detection of Mycobacterium tuberculosis: A Brighter Solvatochromic Probe Detects Mycobacteria within Minutes. JACS Au 1, 1368-1379 (2021).
- Swarts, B.M. et al. Probing the Mycobacterial Trehalome with Bioorthogonal Chemistry. J Am Chem Soc 134, 16123-16126 (2012).
- Loving, G. and Imperiali, B. A Versatile Amino Acid Analogue of the Solvatochromic Fluorophore 4-N,N-Dimethylamino-1,8-naphthalimide: A Powerful Tool for the Study of Dynamic Protein Interactions. J Am Chem Soc 130, 13630–13638 (2008).