Archive for March, 2011

InCROIable quatre!

3 March, 2011

This morning, I’m afraid I experienced rather more secondary effects from the previous night’s entertainment. Thanks to my friend Sylvie, I got invited to the Walker lab party, where I found myself hopelessly outclassed, both scientifically and alcoholically*. Over the course of the evening, I’m sure we worked out exactly how to both cure HIV infection, and produce an effective vaccine, but by the time I awoke (somewhat disorientedly) this morning, it had all disappeared in a mist of Sam Adams.

XMRV – the incredible vanishing virus

As you may recall, in 2009 a new retrovirus called XMRV was reported to be associated with chronic fatigue syndrome (CFS – Lombardi and colleagues 2009). It had previously been reported to be associated with prostate cancer. These results have been the subject of much controversy, and today there was a one-hour discussion session on XMRV. Speakers gave two-minute presentations of their recent results, and this was followed by comments from the floor. The highlights were as follows:

Four different labs, using different techniques reported that they basically did not find XMRV in humans.

William Switzer (CDC, USA) – Tested 45 CFS patients and 42 controls using the same technique as that reported in the Lombardi paper, and looked for serology by Western blot. ZERO POSITIVES.

Timothy Henrich (Brigham and Women’s Hospital, USA) – Tested 293 diverse and varied patients, and 3 CFS patients reported to be XMRV positive in a previous study by nested PCR. ZERO POSITIVES.

Mary Kearney (NCI Frederick, USA) – developed a quantitative PCR assay with single-copy sensitivity to detect XMRV. Reported experimental infection in two macaques. In those two animals, XMRV proviral DNA persisted in blood cells, and was consistently detected. Using this technique, they tested 134 prostate cancer patients, and 4 patients previously reported as XMRV positive in the Lombardi study. ZERO POSITIVES.

Finally, Oya Cingoz (Tufts, USA) and Vinay Pathak (NCI Frederick, USA) reported on the origins of XMRV. This virus was first described in a protstate cancer cell line called 22Rv1, which secretes XMRV. This cell line definitely carries the virus, but how did it get there?

Like many immortalized cell lines, 22Rv1 started out as a human tumor transplanted into immunodeficient “nude” mice in what is known as a xenograft. It was passaged in this way many times in different types of mouse – suggesting that 22Rv1 may have acquired XMRV from its mouse hosts. This is plausible because mice carry many types of endogenous retroviruses in their genomes. Cingoz and Pathak showed that althoug XMRV is not identical to any known mouse retroviruses, the left-hand (5′) half of XMRV is identical to one particular mouse retrovirus, while the right-hand (3′) half is identical to a different mouse retrovirus. XMRV is therefore a new virus produced by recombination between two distinct mouse viruses. This all happened since 1992, when the prostate cancer that gave rise to 22Rv1 was first transplanted into nude mice. It is not a virus that has been circulating in human beings.

One would have liked to have heard the other side of the story from the authors of the Lombardi paper, but they didn’t show up to face the data. I guess that tells its own story.

So just to wind up, XMRV is NOT associated with CFS, and does not appear to be present in the human population (although one might wonder whether researchers working with the 22Rv1 line might in fact be at risk of infection).

If you have CFS, do not buy a test for XMRV (they are entirely BOGUS, as Simon Singh might have said), and do not ask your doctor for antiretroviral medication (unless you are HIV positive, of course). It will be a waste of money, and you will just get the side effects of the medication, without any benefit.

And that was it for the 18th CROI!


* OK, maybe only scientifically

…and my thanks, Dorian, for a job really well done! – Ed

InCROIable trois…

2 March, 2011

Going into day 3, and the only ill effect carried over from last night’s French AIDS research party is a mild ringing in the ears from ANRS director François Delfraissy’s experiments with audio feedback, while he was thanking us all for our efforts. Next time, please don’t stand so close to the speakers when you’re talking into the microphone, Professor Delfraissy!

One of this morning’s plenary talks was from Stephen Cherepanov, on the structure of the HIV integrase complex, but since this has already been covered in Viroblogy, I don’t need to say any more about it. Rather fortunately, because describing the 3-D structures of the integrase-DNA complex would have been far, far beyond my literary prowess. “Yes, well, try to imagine a couple of short tube-shaped sections, close together, but held at an angle – not parallel to one another. Those are the ends of the proviral DNA, just before strand-transfer. They’re being held in place by what looks something like a Henry Moore sculpture, and the wiggly orange bit, close to one end of short tube-shaped DNA ends – that’s the active site of the enzyme.” You see what I mean?

Anyway, later on there was a session on HIV-host cell interactions, one of which harked back to those pesky microRNAs from the other day. Carlos de Noronha (Albany Med Coll, USA) told a story that led from Vpr – one of the HIV’s small proteins – to micro RNA. Vpr has several effects on infected host cells, including cell cycle arrest (infected cells stop dividing) and inducing expression of molecules on the surface of the infected cell that prevent infected cells being killed by “Natural Killer” cells of the immune system. The way Vpr does this, apparently, is by interacting with a ubiquitin ligase complex (DECAF1-CRL4). Ubiquitin ligases stick a protein called ubiquitin onto other proteins, and this ubiquitin tag marks its victim for destruction. De Noronha’s group set out to identify what other cellular proteins are ubiquitinylated by DECAF1-CRL4, and could therefore be influenced by Vpr. Their hunt turned up Dicer, which is involved in producing miRNAs. They showed that Vpr does indeed induce DECAF1-CRL4 to tag Dicer for degradation, and that viruses deficient in Vpr replicate efficiently  only when Dicer is artificially depleted. Now it’s not at all clear why destroying Dicer is useful for the virus, but in answer to a question, de Noronha suggested that infected cells may use miRNA to shut down expression of host factors necessary for HIV replication. In that case, it would be useful for HIV to block production of cellular miRNA.

Micro RNA came back again in the afternoon when Mary Carrington (NCI Frederick) presented data in press in Nature dissecting an association between a genetic polymorphism in HLA-C, and control of HIV infection. The HLA region of the genome controls, to a large extent, the immune response against infectious diseases, including viruses. It is also extremely polymorphic (that is, variable between individuals) and this polymorphism is what ensures that the human race would not be entirely wiped out if an extremely nasty, new infection were to appear. Because of the variability in the immune response between individuals, no virus can be perfectly adapted to every individual in the whole population. Variations in HLA-B genes modify the HLA-B proteins, and this alters their ability to present HIV epitopes, which in the end results in people with certain HLA-B variants (or alleles) such as HLA-B57 and B27 controlling HIV infection better.

The HLA-C polymorphism associated with control of HIV infection, however, does not alter the HLA-C protein, so until this afternoon, it has been rather mysterious how it might work. Well, to cut a long story illustrated by several slides short, it turns out that the protective HLA-C alleles have modifications in the 3′ non-coding region of the gene, and these changes occur in a microRNA (miR-148) binding site. In variants which can be targeted by miR-148, the level of HLA-C expression on the surface of cell are lower. Variants associated with better control of HIV infection “escape” from miRNA148 control, and result in higher HLA-C expression. Moral of the story – even “silent” gene polymorphisms can in fact be functional, and rather strangely, it appears that avoiding control by microRNA can be a mechanism of host defence as well as a means of virus attack.

Also, a very interesting talk from David Evans (Harvard, USA) about how different primate lentiviruses avoid being retained on the surface of the infected cell by Tetherin. One interesting point that he illustrated was that HIV-1 type M viruses are much better at escaping from Tetherin’s grip than HIV-1 type O and type N viruses. This could be one reason why HIV-1 type M viruses are more infectious, and why they, rather than the other two types of HIV-1, caused the current HIV pandemic.


InCROIable Deux

1 March, 2011

In which the redoubtable Dorian reports further on the doings at CROI 2011.

Neutralizing HIV

Michel Nussenzweig (Rockefeller, USA) gave everyone an immunology lesson in order to explain what makes broadly neutralizing anti-HIV antibodies so special. So carrying on with the immunology lesson theme, I should just point out that neutralizing antibodies are those that not only stick to the surface of a virus, but actually prevent it from infecting a susceptible cell. So far, all effective antiviral vaccines work because they can induce these neutralizing antibodies. So that’s what neutralization is, now where does the “broadly” part come in? HIV is of course a highly variable virus, so “narrowly” neutralizing antibodies only neutralize a small number of HIV variants, while “broadly” neutralizing antibodies can block infection from a wide range of different HIV variants.

To date, none of the HIV vaccine candidates tested has been able to induce broadly neutralizing anti-HIV antibodies effectively, and most HIV-infected people do not make this type of antibody during natural infection. However some people with HIV infection do produce broadly neutralizing antibodies (It should be stressed however, that HIV+ individuals who make broadly neutralizing antibodies are not cured of their infection). The reason for studying antibodies from such people is that if we can understand how broadly neutralizing antibodies are formed during natural infection, then perhaps we might find a way to induce the same kind of antibodies with a HIV vaccine.

Using a variety of fantastically ingenious techniques, Nussenzweig showed us that the magical processes of hypermutation and affinity maturation are essential for the potency and the breadth of broadly neutralizing anti-HIV antibodies. These processes occur in the germinal centres of lymph nodes, and he presented some amazing imagery data to show that the maturation of antibodies is controlled by the CD4+ T-cells in the germinal centre that “help” B-cells produce antibodies. So the final message, I guess, is that CD4+ T-cell responses are going to be essential for a vaccine to be able to induce a good neutralizing antibody response.

However, that still doesn’t resolve the “broad” part of the problem – how to focus the antibody response onto the sensitive parts of the virus. Indeed, as a presentation in the afternoon from Laurent Verkoczy (Duke Univ. USA) showed, this may be extremely difficult to achieve. For one broadly neutralizing epitope on HIV (the so-called MPER epitope), the antibodies that bind to this site on the virus are also auto-reactive. In a mouse model, he showed that the cells that carry these antibodies are “strangled at birth” by the mechanisms that prevent our immune system from damaging ourselves. These antibodies have therefore probably been deleted from most people’s immune repertoire, and are therefore not available to be selected and amplified by vaccination.

So I’m afraid no-one has yet found the way to induce these broadly neutralizing antibodies.

A virus that slows down HIV

GBV-C is a virus infecting humans that is transmitted by sex, blood transfusion, and from mother to child – rather like HIV. It is a flavivirus (other family members include yellow fever virus, and hepatitis C virus), and because of its mode of transmission, GBV-C is often found in HIV seropositive people. It does not seem to cause disease in people who are infected either acutely, or chronically. Now, you might expect that being infected by two different viruses at the same time would be worse than just being infected by one. But remarkably, the 20-40% of HIV+ individuals who have chronic GBV-C infection have SLOWER disease progression than those who only have HIV infection (at least in European/North American patient cohorts).

There were two talks presenting results trying to explain this intriguing observation. Molly Perkins (NIAID, USA) presented data from a study of HIV-infected patients in the Gambia. She found that GBV-C coinfection did not change T-cell activation, but reduced expression of the HIV coreceptor CCR5 on T-cells. In direct contrast to these results, Jack Stapleton (U Iowa, USA) presented data showing the exact opposite. In his study, GBV-C lowered T-cell activation, but had no effect on CCR5 expression.

How can two groups looking at the same question get such discordant results? Jack Stapleton noted that the different studies on this topic have been conducted in different regions of the world. Both HIV and GBV-C show geographical variation – that is to say, the HIV that infects people in Iowa is not the same as the HIV that infects people in the Gambia, and the same goes for GBV-C. So one plausible explanation may be that different types of GBV-C have different biological effects.

Not wanting to send the room into an uproar, I didn’t ask the question that immediately sprung to my mind – when are we going to test GBV-C infection as a therapeutic intervention?

Lecturer in Microbiology, University of Nantes


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