Tuesday, May 07, 2013

ZFN, VIP and bnAb are latest members of battle against HIV

Fig 1: HIV prevalence. Source
     Going back at my blog post archives, what is the most common thing I have talked about? HIV. Though I have just posted 4 posts that was much dedicated to HIV, I have posted about HIV in brief as a news or matter of interest in many blog posts (As an insert). Look for a topic on HIV to talk about and you would never run out of matter or material to write about. Why? HIV is a global problem and focus.As anyone could argue, when there is no vaccine available against a particular disease it is a global problem.
      HIV is a unique virus. Apart from the fact that it integrates its genetic material stably into the host genome (All retroviruses do that), the strike point is HIV infects CD4+ T cells that are itself crucial for immunity (There is a lot more detail in that, but for the time, I will skip complicated reasoning).In addition, the latency of HIV virus provides the benifit of immune silence to the virus. In simple terms Immunological memory is long lasting, so is the HIV that lasts in the cell involved with such memories. Drugs such as 5-aza-2'deoxycytidine and HDAC inhibitors (such as Vorinostat) have been tried to combat latency. Vaccine is a great answer. The International AIDS Vaccine Initiative, IAVI estimated that a vaccine with just 50% efficacy administered to 30% of the population of developing countries between 2015 and 2030 would prevent approximately a quarter of the infections that would otherwise occur.
    Anti-retroviral drugs have been of great use. HAART therapy uses a combination of drugs that can clear 99% of viral load. But you still got the latent virus left. The most obvious approach is to silence the viral genome or deplete latent cells. But if you ask me how good is HIV cure to date, it is probably better to cite an example of the first report of successfull treatment of a baby (Link). The paper presented in March 2013, was the First report of a case of functional HIV cure in a 26-month-old infected child who initiated ART at 30 hours of age. Mind you this is the first ever reported case.
    So we still have a lot to do. When the right target and methodology to make a vaccine is defined, it takes almost a decade atleast, to setup and bring out an industrial standard product. Currently we are not even close to knowing what that target must be. More recently, scientists have ventured into areas such as Zinc-Finger technology, Specific neutralising antibodies etc, and provided us some hope.
      Let me make a small digression here for the mere importance of the point. The easiest solution to look into a very complicated problem is to see how the lucky people have escaped from the problem. Same is the case with HIV. There are a group of people referred as "Elite Carriers" or commonly as Long-term nonprogressors (LTNPs), who seem to be exposed to infections but have no functional consequence. Studies indicated that this could be because of inherited mutations of the CCR5 receptor. In addition many Elites have elevated HIV-1 2-LTR circles, an episomal HIV-1 DNA form that results from aborted integration of HIV-1 DNA into host chromosomes. That indicates something antagnozies the HIV DNA integration (Reference). Even the binding of HIV to CD4 cells is not as simple as we had imagined. More recently Purinergic receptors (action via pannexin-1) were found to be involved in the process (Reference).

Fig 2: Zinc finger Nulcease. Source
       From apropos, it is clear that if we can block or eliminate CCR5  receptors from the cells, entry would be effected. The first question that is then asked, would be what about functions conferred by the receptor. Since at least a few elite's have been shown to lack these receptors and are perfectly normal (Appears to be so), it is reasonable to argue that such strategy will not effect the subject in question. The possible success can also be inferred from the maraviroc drug which has been used as a entry inhibitor. There have been several set of studies that tried to inhibit the CCR5 binding, approaches such as ribozymes, single-chain intracellular antibodies, trans-dominant coreceptor mutants, and RNAi, but stability was a question. But the best approach was use of Zinc finger nuclease (such as Fok1) to cut specific sequence of CXCR4 or CCR5, rendering the cells resistant to virus entry. The idea sounds very much pragmatic. The idea of applicability of such an approach comes from the case of famous "German patient".

       Let me now try some Vaccinology here. As, every person who has studied some basic immunology knows how an antibody is produced. Antibodies are produced against the HIV in all cases (We use it as a diagnostic test). However, the problem is that the antibodies dont neutralise. The HIV gets around the problem of neutralisation by high somatic mutation. The antibodies mature into highly specific antibodies, against the newly evolving types, simply to see new variants arising. Point is we need a Broad specific antibodies. Immune system never makes that (Its always high affinity and ultra specific antibody).

Fig 3: Vectored Immunoprophylaxis
        So now, there is a possible solution. If you could get around the problem and constantly produce broad specific neutralising antibody, then we have a good candidate. It could act like a vaccine. This was shown to be achievable by a technique referred as "Vectored Immune-prophylaxis (VIP)". In the proof of concept experiment, self-complementary adeno-associated virus (scAAV or rAAV) which expressed SIV-neutralizing immunoadhesins consisting of small, artificially fused antibody fragments. The gene was incorporated into muscle cells when injected (Principle of vecotred gene therapy) and produced the antibody. The experiment showed good results in humanised mouse model with very promising result.

     Nothing can beat this. The study was done in mouse model using 4 epitopes- 2F5, 4E10, 2G12 and b12. Now the question is what would be the best monoclonal antibody (U cant expect to deliver infinite broad specific antibody produing genes).

      In a seperate set of experiments, a large population is screened for broad neutralising antibodies (bnAb). By repeated screening and improvisation a class of antibodies referred as VRC01 (Vaccine Research Center-01) was isolated by Dr. Xueling Wu. This was one of the best broadly neutralising antibody against HIV. VRC01 and VRC02 are a closely related pair of somatic variants that neutralize over 90% of diverse HIV-1 primary isolates. VRC03 has also been isolated. VRC01 and VRC01-like antibodies bind to gp120 and are able to bind to CD4's binding site on gp120, preventing the CD4 receptor binding, covering about 98% of important site. The representation of same is shown below in Fig 4.

Fig 4: The contact surface of VRC01 and CD4 are shown on gp120. The green represents VRC01's contact surface and yellow represents CD4's contact surface. Source

   In the past 2 years many more lineage of bnAb have been identified. This includes many classess such as PG9, PG16, NIH45-46 and the latest CH103 have been published. CH103 has been isolated by Hayne's and colleagues. The antibody series recognises less than that of VRC01 but is less complicated. “A message of our paper is that broad neutralizing antibodies don’t have to be as complicated as we thought, and therefore may be more easily induced,” says Haynes. Source

     Scientists are trying hard to get the germline B cells to produce a constant bnAb by variety of methods. However, I was wondering why none has engineered a VIP (Vectored Immune-prophylaxis) by using VRC01 or related class of antibodies. Or did i miss that paper?
ResearchBlogging.orgWilen CB, Wang J, Tilton JC, Miller JC, Kim KA, Rebar EJ, Sherrill-Mix SA, Patro SC, Secreto AJ, Jordan AP, Lee G, Kahn J, Aye PP, Bunnell BA, Lackner AA, Hoxie JA, Danet-Desnoyers GA, Bushman FD, Riley JL, Gregory PD, June CH, Holmes MC, & Doms RW (2011). Engineering HIV-resistant human CD4+ T cells with CXCR4-specific zinc-finger nucleases. PLoS pathogens, 7 (4) PMID: 21533216
Balazs AB, Chen J, Hong CM, Rao DS, Yang L, & Baltimore D (2011). Antibody-based protection against HIV infection by vectored immunoprophylaxis. Nature, 481 (7379), 81-4 PMID: 22139420
Li Y, O'Dell S, Walker LM, Wu X, Guenaga J, Feng Y, Schmidt SD, McKee K, Louder MK, Ledgerwood JE, Graham BS, Haynes BF, Burton DR, Wyatt RT, & Mascola JR (2011). Mechanism of neutralization by the broadly neutralizing HIV-1 monoclonal antibody VRC01. Journal of virology, 85 (17), 8954-67 PMID: 21715490