On Friday at the International Workshop on HIV & Hepatitis Virus Drug Resistance and Curative Strategies in Sitges, Spain, Steve Yukl from UCSF presented new data on the case of Timothy Brown, the “Berlin Patient." Yukl described multiple experiments performed by several independent laboratories with the aim of searching intensively for any signs of residual HIV infection in plasma, peripheral blood mononuclear cells (PBMC) and biopsies from the gut and cerebrospinal fluid (CSF). The nature of these analyses is a testament to Brown’s extremely laudable willingness to undergo an array of unappealing procedures in order to advance research into curing HIV.
No infectious HIV was detectable in any sample (including samples containing huge numbers of cells). In most cases, no HIV RNA or DNA could be found either, but there were some exceptions: a minority of samples, analyzed by some labs, intermittently tested positive for extremely low levels of HIV RNA. A very small proportion of the rectal samples also tested positive for very low levels of HIV DNA. Genetic sequencing results were not available but the abstract indicates that the RNA positive samples did not show any relationship with each other or the original infecting HIV (a finding perhaps suggestive of PCR contamination). Levels of antibodies against HIV have continued to decline over 18 months of follow up, while CD4 and CD8 T cell counts have reached near normal levels. The researchers make it very clear that because the assays being used are at the limits of their sensitivity and specificity, it cannot and should not be concluded from these data that Brown is still infected. Although it is possible that there is some residual virus present and that Brown is a case of a “functional cure” rather than complete HIV eradication (or “sterilizing cure”), further work will be needed to explore that possibility. But it is far more likely that—as the study authors state—these new results are just evidence of the technological challenges associated with looking for miniscule amounts of viral genetic material.
Unfortunately, it is all too easy to envision the mainstream media picking up news of this presentation and wildly misinterpreting it (e.g. “Man Said Cured of HIV Still Infected!”). Alain Lafeuillade, who runs the biannual HIV Persistence Workshop and the HIV Reservoir Portal website, has not helped matters by writing a bizarrely misleading post on the study which suggests that the authors interpretation of the data is wrong and that Brown is either not cured, or—in an even stranger piece of speculation—that he may have been reinfected. The evidence supports neither claim.
In a related development, on June 7th the scientist Lawrence Petz held a press conference with Timothy Brown at a symposium in San Francisco on the use of cord blood to facilitate stem cell transplants. Petz revealed that around 100 cord blood donors homozygous for the CCR5 delta-32 deletion have been identified (out of 17,000 tested), and one HIV-positive individual in the Netherlands has recently received such a transplant as part of a course of treatment for another disease. Another similar transplant is to be performed soon for an HIV-positive individual in Madrid. These cases will be carefully followed to see if the beneficial outcome experienced by Timothy Brown can be duplicated.
International Workshop on HIV & Hepatitis Virus Drug Resistance and Curative Strategies, Sitges, Spain, June 5-9 2012.
Challenges inherent in detecting HIV persistence during potentially curative interventions.
SA Yukl1, T Chun2, MC Strain3, J Siliciano4, E Eisele4, R Buckheit4, YC Ho4, JK Wong1, MP Busch5, G Hütter6, DD Richman3, RF Siliciano4, SG Deeks1
1University of California, San Francisco, San Francisco, CA 2National Institute of Health, Bethesda, MD 3Unversity of California, San Diego, San Diego, CA 4Johns Hopkins University, Baltimore, MD 5Blood Systems Research Institute, San Francisco, CA 6Institute of Transfusion Medicine and Immunology, University Heidelberg, Mannheim, Germany
Background. The size of the HIV reservoir during long-term effective antiretroviral therapy and in "elite" controllers is close to the limit of detecting using standard assays. This imposes challenges for the design and assessment of potentially curative interventions. We applied a series of measurements of HIV persistence to the study of the "Berlin Patient", who underwent a hematopoietic stem cell transplant from a donor who was homozygous for the CCR5 delta-32 deletion and who had exhibited clinical evidence of being cured. Our objectives were to (1) determine if HIV had been fully eradicated as a consequence of the transplant and (2) define the potential role of various reservoir measurements in cure research.
Methods. The subject underwent a series of intensive virologic and immunologic studies beginning approximately five years after this transplantation. Replication-competent virus was measured in two laboratories, and HIV DNA and RNA levels (from blood and rectal mucosa) were measured in several laboratories using different approaches.
Results. A large volume apheresis was performed and 9 billion PBMCs evaluated for the presence of replication-competent virus. All wells were negative for HIV p24, indicating that the frequency of replication-competent HIV was therefore estimated to less than one infected cell per 1.4 billion CD4+ T cells. A repeat experiment in a second laboratory confirmed these findings. Using a variety of assays and approaches, very low levels of HIV RNA were intermittently detected in plasma, although sequence analysis of these variants were different from each other and different from those present before the transplant. Digital PCR for HIV DNA was negative for 1 copy per 2 million cells with a 95% confidence limit of less than 1.9 copies per million cells. Collagenase-digested rectal biopsy-derived cells were positive for very low levels of HIV DNA but not RNA; no sequence for confirmatory studies could be obtained. HIV antibodies levels were low and declined over the course of approximately 18 months.
Conclusion. Although the subject has had intermittent evidence for HIV persistence in some assays in some laboratories, the extremely low levels of virus which were detected, while pushing the limits of sensitivity and specificity, and the inability to match sequence with the subject's pre-therapy virus make it impossible to conclude that the subject remains HIV infected.