TWiV 278: Flushing HIV down the zinc

On episode #278 of the science show This Week in Virology, Vincent, Dickson, Alan, and Kathy discuss disruption of the ccr5 gene in lymphocytes of patients infected with HIV-1.

You can find TWiV #278 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 144: HIV gets the (zinc) finger

zinc finger nucleaseHosts: Vincent Racaniello, Rich Condit, and Alan Dove

Vincent, Rich, and Alan discuss live blogging of scientific meetings, the current outbreak of Hendra virus is Australia, and using zinc finger nucleases to make HIV-resistant CD4 cells.

Click the arrow above to play, or right-click to download TWiV 144 (75 MB .mp3, 104 minutes).

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Links for this episode:

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