Probe Detects HIV Protease and Toxicity of Drugs

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This image illustrates HIV-1 protease activity detected by Sha Jin’s molecular probe.

FAYETTEVILLE, Ark. – A biomedical engineer at the University of Arkansas has developed a molecular probe that can simultaneously detect the presence of HIV-1 protease and toxicity levels of chemical compounds used to combat the deadly virus that causes AIDS. The probe can be used to investigate the efficacy and efficiency of HIV drugs, some of which are so toxic that many patients elect to stop treatment.

“Because our screening system detects biochemical activity related to HIV and the toxicity of drug compounds at the same time, it can be used to determine whether a given combination of drugs is effective,” said Sha Jin, assistant professor in the College of Engineering. “The ultimate goal is to identify more effective and affordable drugs to treat HIV.”

Human immunodeficiency virus was discovered in the early 1980s and has been responsible for an estimated 25 million deaths. Today, about 33 million people worldwide live with HIV. About 1 million people in the United States have HIV. If untreated, HIV causes acquired immunodeficiency syndrome, or AIDS.

Although it is currently incurable, HIV can be controlled by anti-HIV drugs, which suppress or inhibit virus replication in patients. Currently, treatment of HIV infection depends heavily upon a strategy referred to as highly active antiretroviral therapy, or HAART, in which drugs that inhibit HIV-1 protease – a family of enzymes that break down polyproteins and are essential for the life cycle of HIV – are combined with drugs that suppress virus replication. Although HAART has reduced mortality, it has significant side effects. A quarter of patients with HIV stop therapy within the first year due to symptoms related to its high toxicity.

None of the anti-HIV drugs, even when combined, represent the ideal therapy. In addition to side effects, the high cost of treatment and the emergence of drug-resistant viruses are further obstacles to successful therapy. Because the drugs are expensive, patients in low-income countries start treatment at the AIDS stage of the disease, which leads to higher morbidity and mortality rates in low-income countries. Most importantly, existing treatment for HIV cannot eradicate the virus from patients.

To address these issues, Jin created a screening system by applying fluorescent proteins, one green and one red, to generate Förster resonance energy transfer signals responsive to HIV-1 protease inhibition and activity. Förster resonance energy transfer is a mechanism of energy transfer between a donor and receptor chromophore, which is the part of a molecule that is responsible for its color. The energy transfer prompts reactions from cells in the form of color. In the absence of protease inhibitors from various compounds, the researchers observed green and yellow cells. When protease inhibitors were added to cells, they showed a red color.

“We confirmed that compounds nelfinavir and lopinavir were toxic to cells, since many healthy cells died after two days incubation,” Jin said. “Indinavir, sequinavir, and ritonavir showed less toxicity.”

Jin is establishing a cell line that will consistently express the detection system. To do this, she will use HeLa cells, the first immortal line of cancer cells kept alive in cell culture. HeLa cells, which were originally taken from a tissue sample removed from the cervical-cancer tumor of Henrietta Lacks, an African American woman at Johns Hopkins Hospital in Baltimore in 1951, were chosen because:

• they grow rapidly and can be handled straightforwardly,

• they grow in large quantity and can be grown in cell culture at much less expense than many other mammalian tissue-culture cells, and

• targeted genes can be easily delivered to HeLa cells.

Jin’s research has been submitted for publication to Biotechnology Progress.