TWiV 326: Giving HIV a flat tyr

On episode #326 of the science show This Week in Virology, the sternutating TWiVers discuss preventing infection of cells and animals by a soluble CD4-CCR5 molecule that binds to HIV-1 virus particles.

You can find TWiV #326 at

The Berlin patient

HIV binding CD4 and ccrSince the beginning of the AIDS epidemic, an estimated 75 million people have been infected with HIV. Only one person, Timothy Ray Brown, has ever been cured of infection.

Brown was diagnosed with HIV while living in Berlin in 1995, and was treated with anti-retroviral drugs for more than ten years. In 2007 he was diagnosed with acute myeloid leukemia. When the disease did not respond to chemotherapy, Brown underwent stem cell transplantation, which involves treatment with cytotoxic drugs and whole-body irradiation to destroy leukemic and immune cells, followed by administration of donor stem cells to restore the immune system. When his leukemia relapsed, Brown was subjected to a second stem cell transplant.

The entry of HIV-1 into lymphocytes requires two cellular proteins, the receptor CD4, and a co-receptor, either CXCR4 or CCR5. Individuals who carry a mutation in the gene encoding CCR5, called delta 32, are resistant to HIV-1 infection. This information prompted Brown’s Berlin physician to screen 62 individuals to identify a stem cell donor who carried a homozygous CCR5∆32 mutation. Peripheral blood stem cells from the same donor were used for both transplants. 

Despite enduring complications and undergoing two transplants, Brown’s treatment was a success: he was cured both of his leukemia and HIV infection. Even though he had stopped taking antiviral drugs, there was no evidence of the virus in his blood following his treatment, and his immune system gradually recovered. Follow-up studies in 2011, including biopsies from his brain, intestine, and other organs, showed no signs of HIV RNA or DNA, and also provided evidence for the replacement of long-lived host tissue cells with donor-derived cells. Today Brown remains HIV-1 free.

Although Brown’s cure is somewhat of a medical miracle, and by no means a practical road map for treating AIDS, the example of the Berlin patient has galvanized research efforts and continues to inspire hope that a simpler and more general cure for infection may someday be achieved. Clinical trials have been conducted to test a variety of strategies in which CD4+ T or stem cells are obtained from a patient, the CCR5 gene is either mutated or its translation blocked by RNA interference, and then the resulting virus-resistant cells are returned to the patient. In one case zinc finger nucleases were used to delete the CCR5 gene in a patient’s cells, a procedure that we discussed in TWiV #278.

TWiV 293: Virology Down Under

On episode #293 of the science show This Week in VirologyVincent visits Melbourne, Australia and speaks with Melissa, Alex, Gilda, and Paul about their work on HIV infection of the central nervous system, West Nile virus, microbicides for HIV, and the Koala retrovirus.

You can find TWiV #293 at

HIV gets the zinc finger

zinc finger nucleaseBecause all animal viruses initiate infection by binding to a receptor on the cell surface, this step has long been considered a prime target for antiviral therapy. Unfortunately, drugs that block virus attachment to cells have never shown much promise. Another approach, which is to ablate the receptor from the cell surface, is also problematic because these molecules have essential cellular functions. Removing one of the receptors for human immunodeficiency virus type 1 might be an exception.

HIV-1 must interact with two cell surface proteins to initiate infection: a T lymphocyte protein called CD4, and a second receptor, which can be one of two molecules called CCR5 or CXCR4. For many years it has been known that humans can survive without the CCR5 protein: from 4-16% of people of European descent carry the ccr5-delta32 mutation, that prevents the protein from reaching the cell surface. Individuals who are homozygous for ccr5-delta32 (the mutation is present in both copies of the gene) are resistant to HIV infection. Because the vast majority of HIV viruses that are transmitted are those that require CCR5 for cell entry, absence of the protein on the cell surface confers resistance to infection.

The key role of CCR5 in HIV infection in humans was further confirmed when an AIDS patient was given a bone marrow transplant from a donor with the ccr5-delta32 mutation. The patient has been free of HIV for years despite not taking anti-retroviral drugs.

These findings suggest that one possible therapy for AIDS would be to disrupt the ccr5 gene in patient lymphocytes. The development of gene-targeting technologies has brought this approach closer to reality. One approach uses zinc finger nucleases, which are artificial proteins made by joining a protein that can specifically bind DNA with an enzyme that can cleave DNA. A zinc finger nuclease can be designed, for example, to specifically cut within the ccr5 gene. When the cell tries to repair the cut, the gene may be damaged so that the CCR5 protein is no longer made (illustrated).

This approach works: when CD4 T lymphocytes are removed from humans, cultured, and treated with a ccr5 zinc finger nuclease, they become resistant to HIV infection. We discussed this experiment on episode #144 of This Week in Virology.

The next step has now been done: to remove CD4 T lymphocytes from HIV positive donors, treat the cells with the ccr5 zinc finger nuclease (delivered using an adenovirus vector), and infuse the cells back into the patients (each person receives his or her own modified cells). Half of the donors were removed from anti-retroviral therapy, and then the levels of HIV, and CD4 lymphocytes, were measured over the next 250 days.

The result were encouraging: not only were the infusions safe, but the overall levels of CD4 lymphocytes increased, and a good fraction of these had modified ccr5 genes. The initial rise of HIV viremia after interruption of treatment was followed by a decline in virus load. These results show that the CD4 T lymphocytes with modified ccr5 were able to expand in the recipients, and survived better than the unaltered lymphocytes, probably because they were at least partially resistant to HIV infection.

This important clinical trial is only the beginning of a new approach to HIV therapy, and several substantial problems still remain to be solved. Both copies of the ccr5 gene were modified in only 33% of the CD4 T lymphocytes; the remaining cells can still be infected by HIV, albeit less efficiently. New approaches are needed to disrupt both copies of the ccr5 gene in most of the T lymphocytes.

Another issue is that the modified T cells can proliferate for a long time, but not indefinitely. As these cells divide from a limited number of infused cells, they will not have the broad repertoire needed to fight pathogens. T cells are also known to become “exhausted”: they eventually lose their protective functions. Patients given modified lymphocytes still harbor a pool of long-lived T cells which contain HIV DNA. These cells will likely always be present and could give rise to viremia. CD4 T lymphocytes with normal levels of ccr5 protein will always be produced, serving as potential hosts for HIV replication. Modifying stem cells so that they do not produce CCR5 is one long-term solution, but more difficult and dangerous for the patient.

Despite these drawbacks, it is amazing that we can now remove cells from patients, modify their genes, and place them back in patients with little harm and some clear benefit. This is a complicated set of procedures, made even more difficult because humans are involved. It’s truly a landmark clinical trial.

TWIV 266: A pathogenic vicious cycle

On episode #266 of the science show This Week in Virology, Vincent, Alan, and Kathy discuss finding viruses in outer space, varying results obtained from personal genetic testing, and depletion of CD4 cells during HIV infection by pyroptosis.

You can find TWiV #266 at

TWiV 260: Badgers go viral

On episode #260 of the science show This Week in Virology, Vincent visits the University of Wisconsin, Madison and speaks with Tom, Tony, and David about their work on virus discovery at the AIDS Vaccine Research Laboratory.

You can find TWiV #260 at

TWiV 254: Ninety-nine macaques on the wall

On episode #254 of the science show This Week in Virology, Vincent, Dickson, Alan, and Kathy review clearance of simian immunodeficiency virus infection from macaques by immunization with SIV proteins encoded in a rhesus cytomegalovirus vector.

You can find TWiV #254 at

TWiV 244: Back in the CVVR

Episode #244 of the science show This Week in Virology was recorded before an audience at the Beth Israel Deaconess Medical Center, where Vincent and Alan spoke with Dan and Jeff about AIDS vaccines.

You can find TWiV #244 at

TWiV 240: Virology in Vermont

On episode #240 of the science show This Week in Virology,  Vincent travels to the University of Vermont to talk with Markus and Jason about their work on HIV, influenza virus, arenaviruses and hantaviruses.

You can find TWiV #240 at

TWiV 237: Paleovirology with Michael Emerman

Episode #237 of the science show This Week in Virology was recorded at the Fred Hutchinson Cancer Research Center in Seattle, WA, where Vincent and Rich met up with Michael to talk about his work on the molecular and evolutionary basis of HIV replication and pathogenesis.

You can find TWiV #237 at