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Curing HIV Mystery, The Berlin Patient
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http://m.theatlantic.com/health/archive/2014/09/the-berlin-patient-and-the-mysterious-cure-for-hiv/380895/
In a study published last week in the journal PLOS Pathogens, a team from Emory University winnowed down the potential factors by disproving the notion that radiation played a part. The researchers infected six rhesus monkeys with a hybrid of HIV and SIV (simian immunodeficiency virus) known as SHIV. All six monkeys received a course of antiretroviral drugs; mirroring the circumstances of Brown's illness, three monkeys underwent radiation, which destroyed the majority of their white blood cells, followed by transplants of stem cells from their own bone marrow, which had been stored before they were infected.
When all six monkeys were subsequently taken off of their antiretroviral drugs, five of them-the three that hadn't received transplants and two that had-saw their viral levels shoot back up almost immediately. (The sixth monkey, who had also received a transplant, was put down by the researchers after its kidneys failed.)
While not a definitive answer, the experiment does resolve one question about Brown's treatment: The radiation for his leukemia, even though it destroyed many of his HIV-infected cells in the short term, likely had little to do with his eventual permanent cure.
But other questions still linger; with one factor out of the way, researchers are now left with the two factors that the PLOS study didn't address. Because the monkeys received transplants from their own bodies, as opposed to donor stem cells, graft-versus-host disease is still in the running, as is the possibility that the donor's CCR5 mutation simply left Brown's immune system insulated against HIV. In their paper, the Emory researchers declared that bone-marrow transplant is still "a feasible intervention that can lead to a marked reduction of the virus reservoir," though recent results have offered more setbacks than promises-the "Boston patients," two men thought to be cured of HIV after bone-marrow transplants last year, suffered relapses a few months after going off their medications.
But still, by process of elimination, researchers are inching toward an understanding of the virus' Achilles' heel-and by extension, toward new treatments for an infection that currently affects around 36 million people worldwide.
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Persistence of Virus Reservoirs in ART-Treated SHIV-Infected Rhesus Macaques after Autologous Hematopoietic Stem Cell Transplant - (09/26/14) PLOS Pathogens
Discussion
The apparent cure of HIV infection in the "Berlin patient" [5]-[7] has energized efforts to understand the mechanisms of virus persistence despite ART-mediated suppression of virus replication. The factors thought to be involved in the favorable outcome of the Berlin patient following HSCT include (i) the myeloablative conditioning regimen; (ii) the donor's homozygosity for ~32ccr5; and (iii) the graft versus host effect. In this test-of-concept study of autologous HSCT in SHIV-infected RMs we interrogated the relative contribution of a myeloablative conditioning regimen in eliminating the persistent reservoir of latently infected cells. To the best of our knowledge this is the first time that a study of similar design has been conducted.
The key findings of this study are the following: (i) autologous HSCT using apheresis products collected prior to infection is feasible in SHIV-infected RMs; (ii) as expected, the myeloablative TBI used for conditioning induced a massive reset of the lympho-hematopoietic compartment, consequently resulting in the depletion of 94.2-99.2% of circulating CD4+ T-cells; (iii) animals receiving autologous HSCT under ART exhibited a prompt and pronounced decline in the peripheral blood viral reservoir (with undetectable SHIV-DNA in PBMCs in two out of three RMs) and maintained undetectable SHIV-RNA viremia with the exception of a few minor blips; (iv) two of the three transplanted RMs showed a very rapid rebound of viremia after ART interruption; and (v) the third transplanted RM, who was sacrificed for clinical reasons at day fourteen post ART interruption, had no detectable virus in plasma, PBMCs, tonsils, and GI tract, low but detectable levels of SHIV-DNA in sorted peripheral CD4+ T-cells and lymph nodes, and moderate levels of SHIV-DNA in the spleen.
Due to many logistical challenges of this experiment we chose to conduct the study in a temporally compressed fashion, with 37-53 days of ART before autologous HSCT, and interruption of ART after hematopoietic reconstitution, rather than prolonged continuation of therapy. This study was therefore designed to determine the impact of myeloablative irradiation on the viral reservoir, rather than the impact of prolonged viral suppression in conjunction with myeloablation. It is therefore possible that a similarly designed study, in which ART is maintained for a significantly longer period both before and after autologous HSCT, would have a different outcome, possibly demonstrating a more dramatic effect of autologous HSCT on the persistent reservoir of latently infected cells. Moreover, we cannot rule our the possibility that the level of virus suppression achieved by the short-term ART regimen in this experiment might not be as complete as what is observed in HIV-infected individuals on long-term ART. In this model of SHIV-infected RM, 5 to 7 weeks on ART pre-transplant may have been insufficient to fully suppress viral replication and the transient low-level viremia observed immediately post-transplant could be attributed to an insufficient period of ART pre-transplant. However, similar viral blips were observed in one patient who received allogeneic stem cell transplant after many years on combined ART [8]. Although the origin of these transient blips is unknown, it may represent release of the virus from latently infected cells in the setting of cell activation during conditioning and the peri-transplant period. In keeping with this hypothesis, it should be noted that in our study the post-transplant period was characterized by an expansion of CD4+ T-cells expressing CCR5 as well as proliferation and activation markers.
Together with the observed increased proportion of memory CD4+ T-cells post-transplant, these results suggest that the CD4+ T-cell compartment recovered primarily through homeostatic proliferation of memory CD4+ T-cells.
The myeloablative TBI used for conditioning resulted in the depletion of 94.2-99.2% of circulating CD4+ T-cells. Unfortunately, due to the clinical challenges of this innovative experiment, no tissue biopsies could be obtained immediately post-transplant to evaluate the TBI-induced CD4+ T-cell depletion in tissues. However, this study shows that myeloablative TBI and autologous HSCT did not prevent a rebound of viremia post-ART interruption in two out of three RMs despite relatively early ART initiation (day 28 post-infection). Moreover, while the SHIV-DNA level in PBMCs was undetectable or close to undetectable post autologous HSCT, it rapidly rebounded after ART interruption to levels that were similar or higher than those observed in the control animals at the same time-point. While in the third animal (T2) there was no sign of virus present in the plasma, PBMCs, and various tissues at the time of necropsy, this RM had to be sacrificed due to kidney failure at day fourteen after ART interruption making the interpretation of these data somewhat difficult. Of note, this study was not designed to identify the cellular and anatomic sources of the rapid plasma viral rebound observed in two transplanted RMs following ART interruption. Determining the relative contribution of tissue CD4+ T-cells, macrophages, and potentially other sources represents an important area for future investigation, amenable for interrogation with this model.
We acknowledge a number of limitations in our study including the small number of animals and the foreshortened time line involved. However, the demonstrated feasibility of this test-of-concept study in a non-human primate model of AIDS virus infection is per se an important result given the extreme complexity of the experimental protocol. The RMs included in this study underwent a series of procedures that have been only rarely, if ever, used in the same animal, including stem cell mobilization and harvesting by apheresis, RT-SHIV infection, daily four-drug ART administration, total body irradiation, re-infusion of HSCs, repeated platelet transfusions, and receipt of several antimicrobial prophylaxes. The feasibility of HSCT in SIV- or SHIV-infected RMs suggests, in our view, that further studies using this model in conjunction with longer term ART as well as additional interventions aimed at purging both the peripheral blood and lymphoid tissue-based viral reservoirs will provide critical information for the requirements to cure HIV infection in humans.
With respect to our understanding of the mechanisms responsible for "curing" HIV infection in the Berlin patient, our study supports the hypothesis that myeloablative TBI can cause a significant decrease in the viral reservoir in circulating PBMCs, even though it was not sufficient to eliminate all reservoirs. While the conditioning regimen in the Berlin patient also included antithymocyte globulin and chemotherapy, the use of a ~32ccr5 homozygous donor and/or the presence of graft versus host disease likely played a significant role in that clinical context. The importance of graft versus host disease that effectively results in a "graft versus reservoir" effect is also emphasized by the recent observation of two HIV-infected patients in which a prolonged (i.e., 3-8 months) period of undetectable viremia in absence of ART was observed after allogeneic HSCT from donors with wild-type ccr5 alleles [9], although these patients did eventually develop rebound of viremia [16]. Future studies of allogeneic HSCT in SIV- or SHIV-infected RMs in the presence or absence of gene therapy interventions to knock out ccr5 would be very informative in this regard, and may elucidate the mechanism of the sustained cure seen in the Berlin patient but not the above mentioned recipients of donor cells wild type for ccr5.
In conclusion, we have conducted the first test-of-concept study of myeloablative irradiation and autologous HSCT in ART-treated SHIV-infected RMs. This experiment demonstrated that autologous HSCT is a feasible intervention that can lead to a marked reduction of the virus reservoir in the peripheral blood, and can be used as an experimental in vivo platform to test innovative interventions aimed at curing HIV infection in humans.
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