454 Sequencing Identifies Early Stage HIV Drug Resistance: Low-level Mutations may have Significant Impact on Clinical Outcomes

Retrospective research study shows that even the lowest levels of resistance mutations could lead to early treatment failure.

A research study published online this week in the Journal of Infectious Disease reports that HIV resistance mutations present in as little as 1% of the viral population have a significant impact on clinical outcomes. The paper outlines the results of a collaborative study between 454 Life Sciences, a Roche company, and a Yale School of Medicine researcher that used 454 Sequencing Systems to identify previously undetectable drug resistant HIV variants in research samples from an earlier performed clinical trial, called the FIRST Study (1).

The research study, lead by Michael Kozal, M.D. of the Yale University School of Medicine and the VA CT Health Care System, was a blinded-retrospectives analysis of 264 blood samples taken from HIV infected individuals before initiating drug treatment for research purposes. The FIRST Study samples were analyzed using Ultra-Deep Sequencing with the Genome Sequencer FLX System, a platform ideally suited for the sensitive detection of low-frequency mutations. Surprisingly, results showed that the fraction of patients harboring resistance variants was twice as high as previously thought. The second question answered by the study was whether or not the low-level mutations undetected with current methods may affect patient outcome. Remarkably, drug resistance levels as low as 1% were found which could lead to early antiretroviral treatment failure with statistical significance.

“Current technology available to clinicians is limited to detecting resistance mutations that are present at levels of approximately 20% or greater in the circulating viral population in a patient. Thus, the current test used in the clinic may miss many low-level resistant HIV strains which can grow rapidly under drug selection pressure and lead to therapy failure. “This retrospective research study shows that even resistance mutations present at the 1% level can lead to premature failure of therapy," explained Michael Kozal, M.D., the senior author of the study. "In the future, hopefully clinicians may use this knowledge to choose better antiretroviral drug combinations that have the ability to suppress these resistant HIV strains which will lead to better clinical responses in patients."

While HIV survival trends have increased tremendously over the past decade, a significant number of patients develop drug resistance shortly after treatment is initiated. This is particularly true in developed nations where antiretroviral drugs have been widely accessible for years, and is an increasing concern as more treatments reach developing nations with high prevalence of HIV infected populations. The availability of long-term clinical data from the FIRST Study, which lasted five years, enabled correlation of the sequence data with patient outcomes.

“We developed the Ultra Deep Sequencing method exactly to answer this type of question and are very pleased with the power demonstrated in this research study,” explained co-lead author Michael Egholm, Ph.D., Chief Technology Officer at 454 Life Sciences. “HIV drug-resistance is just one example of a real world problem that we may tackle with this powerful technology in future.”