A new direction in HIV therapy

HIV is a worldwide problem – 35.3 million people are living with HIV or AIDS and 36 million people have died from AIDS related illnesses1. As the HIV/AIDS epidemic is such a big problem, there are many agencies working hard to come up with effective and efficient treatments. In a recent seminar at Macquarie University’s Australian School of Advanced Medicine, Professor Anthony Kelleher discussed the cutting edge HIV research his team has been conducting. His research at The Kirby Institute, University of NSW, focuses on how HIV reservoirs are established and maintained and how this information can help to develop a cure.

Currently, the only option for HIV treatment is the use of antiviral therapy. This treatment suppresses the replication of HIV virus particles by targeting different stages of the virus lifecycle, such as the entry of the virus particles into the cell, creation of DNA from virus RNA, insertion of the virus DNA into the cell’s nuclear DNA or preventing mature virus particles from leaving the cell (Figure 1)2.

Figure 1. Anti-viral drugs target different stages of the HIV lifecycle.

Figure 1. Anti-viral drugs target different stages of the HIV lifecycle.

Antiviral therapies work well, but they don’t eliminate the virus from the body3. As soon as patients stop anti-viral drug treatment, HIV levels can rapidly rise due to the stockpile of virus particles lurking in cells (the viral reservoir)4. This means patients with HIV need anti-viral drugs for the rest of their lives, otherwise the virus will return with vigor and the patient will relapse (Figure 2).

Figure 2. HIV+ patients currently take a cocktail of drugs to suppress the virus.

Figure 2. HIV+ patients currently take a cocktail of drugs to suppress the virus.

Staying on anti-viral drugs for a lifetime can have a lot of unwanted side effects, such as metabolic diseases and osteoporosis5. There are research teams developing a vaccine for HIV, but this is still in phase 1 of clinical trials (preliminary safety tests). A vaccine needs multiple rounds of trials and will take approximately 15 years before it can be used in the general population6.

Only one person has been cured of HIV – a patient with HIV developed an additional disease which required a bone marrow transplant. The bone marrow donor had a deletion of his CCR5 gene, a receptor for HIV. As the HIV receptor was no longer present the HIV could no longer bind and the patient was cured4. Unfortunately, a bone marrow transplant from a donor with a CCR5 gene deletion is not an option for everyone with HIV, so other avenues are being explored.

Professor Kelleher and his team are interested in developing alternatives to long-term medications and are looking at the genes of HIV and the immune system for a possible solution. The team has just recently received ethics clearance to start human clinical trials using siRNAs (small interfering RNAs) to degrade the CCR5 gene receptor for HIV. This method aims to stop expression of the HIV genes (silencing) which stops the virus particles from replicating7 (Figure 3). This will remove the viral reservoir, but the genes will still be present in the patient’s genome. This method has been effective for other viruses, such as the Human Papilloma Virus, Polio, Hepatitis B and Hepatitis C, and oncogenes have also been silenced using this method8,9.

The way retroviruses, such as HIV, replicate is by inserting their genetic material into the host cell’s genome and taking over the DNA replication machinery to produce messenger RNA and then proteins. The siRNAs can bind to the HIV messenger RNAs (Figure 3) and degrade them, stopping the creation of the proteins that create HIV particles10. Alternatively the siRNAs can bind to the DNA and change the chemical structure so the HIV genes can’t be transcribed4. The siRNAs are so specifically targeted to HIV strains that there is no chance of detrimental impacts on other genes10.

Figure 3. siRNAs degrade messenger RNA so it can’t be translated into protein (McManus & Sharp 2002).

Figure 3. siRNAs degrade messenger RNA so it can’t be translated into protein (McManus & Sharp 2002).

Another research group has shown that treatment with siRNAs works to silence an HIV-type virus in mice11 so the next step is to try this method with human HIV. Professor Kelleher’s team will start the treatment with a small group of HIV patients, following these steps:

  1. Identify HIV+ patients currently taking anti-viral drugs to suppress the virus;
  2. Treat with siRNA therapy;
  3. Stop anti-viral treatment; and
  4. Monitor the patients to see if the HIV stays suppressed.

If the HIV symptoms don’t return then the siRNAs have eliminated the viral reservoir and silenced the HIV. The siRNA treatment has been shown to be effective in human cells for up to 30 days in a laboratory setting. If the same is shown in human clinical trials, this could lead to a significant improvement for the quality of life for millions of HIV+ patients in the future.

Want to know more?

  1. World Health Organisation (2014.) HIV/AIDS, http://www.who.int/gho/hiv/en/, accessed 22 May 2014.
  2. Mehellou Y & De Clercq E (2010). Twenty-Six Years of Anti-HIV Drug Discovery: Where Do We Stand and Where Do We Go? Journal of Medicinal Chemistry, 53(2), 521-538. doi: 10.1021/jm900492g.
  3. Suzuki K, Marks K, Symonds G, Cooper DA, Kelleher AD, et al. (2013). Promoter targeting shRNA suppresses HIV-1 infection in vivo through transcriptional gene silencing. Molecular Therapy – Nucleic Acids, 2, e137; doi: 10.1038/mtna.2013.64.
  4. Kent SJ, Reece JC, Petravic J, Martyushev A, Kramski M, et al. (2013). The search for an HIV cure: tackling latent infection. The Lancet, 13(7), 614-621. doi: 10.1016/S1473-3099(13)70043-4.
  5. Grund BA, Peng GA, Gibert CLB, Hoy JFC, Isaksson RLA, et al. (2009). Continuous antiretroviral therapy decreases bone mineral density. AIDS, 23(12), 1519-1529. doi: 10.1097/QAD.0b013e32832c1792.
  6. National Health and Medical Research Council, Commonwealth of Australia (2014). Australian Clinical Trials. http://www.australianclinicaltrials.gov.au/node/5, accessed 22 May 2014.
  7. McManus MT and Sharp PA (2002). Gene silencing in mammals by small interfering RNAs. Nature Reviews Genetics, 3, 737-747. doi: 10.1038/nrg908.
  8. Saleh MC, Van Rij RP, Andino R (2004). RNA silencing in viral infections: insights from poliovirus. Virus Research, 102, 11–17. doi: 10.1016/j.virusres.2004.01.010.
  9. Li S-D, Chono S & Huang L (2008). Efficient Oncogene Silencing and Metastasis Inhibition via Systemic Delivery of siRNA. Molecular Therapy, 16(5), 942-946. doi: 10.1038/mt.2008.51.
  10. Suzuki, K, Ishida T, Yamagishi M, Ahlenstiel C, Swaminathan S, et al. (2011). Transcriptional gene silencing of HIV-1 through promoter targeted RNA is highly specific. RNA Biology, 8(6), 1035-1046. doi: 10.4161/rna.8.6.16264.
  11. Mitsuyasu RT, Merigan TC, Carr A, Zack JA, Winters MA, et al. (2009). Phase 2 gene therapy trial of an anti-HIV ribozyme in autologous CD34+ cells. Nature Medicine, 15, 285-292. doi: 10.1038/nm.1932.

Going a bit batty – How do bats withstand so many viruses?

Dr Michelle Baker from the CSIRO Australian Animal Health Laboratory spoke at Macquarie University last week about her work on bat immune systems. Her lab recently contributed to the high impact Hendra virus vaccine for horses. Dr Baker’s work has implications for disease control and prevention and management of virus spillovers into human communities worldwide.

Bats make up 20% of mammalian diversity, are long lived for their body size and are the only mammals with powered flight. Bats are vital to ecosystem functions such as pollination, fertilisation and insect control (Calisher et al. 2006). Despite these unique characteristics they are not intensively studied like most other mammal groups. Before Dr Baker’s research group focused on bats not much was known about their immune systems (Baker et al. 2013).

Figure 1. Possible virus transmission routes from bats to humans. (adapted from image presented by Dr Michelle Baker)

Figure 1. Possible virus transmission routes
(adapted from image presented by Dr Michelle Baker).

Bats act as viral reservoirs, meaning they carry a range of viruses (He et al. 2010; Ng & Baker 2013). Fatal human viruses that can be traced back to bats include Rabies, Hendra, Ebola, Marburg and the SARS coronavirus (Ng & Baker 2013). These viruses occasionally spill over from bats into other animals and that’s normally how humans become infected (Figure 1). But even with this load of up to 100 different viruses, bats are hardly ever sick (Baker et al. 2013).

The bat’s lack of symptoms from viral infections was puzzling, so Dr Baker’s team looked more closely at bat immune systems. When a viral infection occurs in other mammals the immune system quickly delivers a generic response (innate response) and then a specific response occurs more slowly (adaptive response; Katze et al. 2002). The researchers observed that bats don’t develop many antibodies in response to infections, so they thought the bat immune system might be knocking down the viruses before the immune system could mount an adaptive response (Baker et al. 2013).

Figure 2. The Black Flying Fox, Pteropus alecto.

Figure 2. The Black Flying Fox, Pteropus alecto.

To test this idea the researchers sequenced the genomes and studied the immune responses of two species of bat: Myotis davidii, a micro bat, and Pteropus alecto, a megabat (Figure 2). They found the collection of immune genes in bats is different to other mammals. For example, bats have fewer genes for interferon production (Papenfuss et al. 2012).

Interferon is a protein produced by the immune system in response to the detection of viral invaders. It starts a signaling cascade that creates an anti-viral state in cells (He et al. 2010; Figure 3). High interferon levels can be toxic for cells, so normally the interferon level is very low. When a viral infection is detected, the interferon level is dramatically increased which signals cells to start fighting the infection (Katze et al. 2002). There are multiple types of interferon, but the type Dr Baker spoke most about is interferon alpha (IFNA).

Figure 3. Interferon signaling cascade - causes expression of immune system genes and creates an antiviral state in cells (Katze et al. 2002).

Figure 3. Interferon signaling cascade – causes expression of immune system genes and creates an antiviral state in cells (Katze et al. 2002).

In contrast to expectations, bat cells were found to have their IFNA genes constantly switched on and there is no increase when cells are infected with viruses (Figure 4). Even with this high IFNA level the toxicity effect observed in other mammals isn’t seen in bats. This IFNA level in bats may be part of the reason they can carry so many viruses, but don’t often get sick from them. Other research groups have found that bat IFNA genes have been positively selected which means they must have been beneficial to bats as they lived with viruses in the past (Calisher et al. 2006; He et al. 2010). Recent work has hypothesised that there is a link between the evolution of flight and the ability of bats to harbor viruses without becoming sick (Zhang et al. 2013; O’Shea et al. 2014).

These findings were very new and unexpected so there were lots of questions from the audience after the seminar. It seems like everywhere Dr Baker turned there were more questions! These are the ‘top 5’ questions asked:

  1. Why aren’t the bats harmed by high levels of interferon in uninfected cells like other mammals?
  2. What triggers the spillover of viruses into other animals that cause outbreaks?
  3. Could there be a link between the interferon level and their long lifespan relative to their body size?
  4. What is the bat immune response to bacterial infection?
  5. Why doesn’t the bat immune response completely wipe out the viruses? How come the viruses can persist and then spill over into other animals?
Figure 4. Interferon alpha levels in infected and uninfected cells in bats and other mammals  (adapted from image presented by Dr Michelle Baker).

Figure 4. Interferon alpha levels in infected and
uninfected cells in bats and other mammals
(adapted from image by Dr Michelle Baker).

So much is currently unknown about how bats can carry so many viruses without being sick. Dr Baker and her team are working to find answers to these questions and more. As human populations increasingly overlap with bat habitats there is more chance of spillover events affecting human and animal populations. Dr Baker’s research could be used to understand human responses to viruses and develop anti-viral treatments in the future.

Learn more:

Baker ML, Schountz T & Wang L-F (2013). Antiviral Immune Responses of Bats: A Review. Zoonoses and Public Health, 60, 104-116. doi: 10.1111/j.1863-2378.2012.01528.x

Calisher CH, Childs JE, Field HE, Holmes KV & Schountz T (2006). Bats: Important Reservoir Hosts of Emerging Viruses. Clinical Microbiology Reviews, 19(3), 531-545. doi:  10.1128/CMR.00017-06

He G, He B, Racey PA & Cui J (2010). Positive Selection of the Bat Interferon Alpha Gene Family. Biochemical Genetics, 48(9-10), 840-846. doi: 10.1007/s10528-010-9365-9

Katze M, He Y & Gale M (2002). Viruses and interferon: A fight for supremacy. Nature Reviews Immunology, 2(9), 675-687. doi: 10.1038/nri888

Ng J & Baker ML (2013). Bats and bat-borne diseases: a perspective on Australian megabats. Australian Journal of Zoology, 61, 48-57. doi: 10.1071/ZO12126

O’Shea T, Cryan P, Cunningham A, Fooks A, Hayman D, Luis A, Peel A, Plowright R, & Wood J (2014). Bat Flight and Zoonotic Viruses. Emerging Infectious Diseases, 20(5), 741-745. doi: 10.3201/eid2005.130539

Papenfuss AT, Baker ML, Feng Z-P, Tachedjian M, Crameri G, Cowled C, Ng J, Janardhana V, Field HE, Wang L-F (2012). The immune gene repertoire of an important viral reservoir, the Australian black flying fox. BMC Genomics, 13:261, doi: 10.1186/1471-2164-13-261.

Zhang G, Cowled C, Shi Z, Huang Z, Bishop-Lilly KA, Fang X, Wynne JW, Xiong Z, Baker ML, Zhao W, Tachedjian M, Zhu Y, Zhou P, Jiang X, Ng J, Yang L, Wu L, Xiao J, Feng Y, Chen Y, Sun X, Zhang Y, Marsh GA, Crameri G, Broder CC, Frey KG, Wang L-F & Wang J (2013). Comparative Analysis of Bat Genomes Provides Insight into the Evolution of Flight and Immunity. Science, 339, 456-460. doi: 10.1126/science.1230835

Silhouette images in Figure 1 sourced from: horse, bat, pig, humans.