The world around you is teeming with pathogens. Lucky, your immune system is well trained to protect and guard you from any unwanted invaders. Pathogens, such as viruses, are in a permanent arms race with our immune system, constantly evolving to evade our defences. Perhaps the most successful pathogens are the ones which have learned to disguise their weapons. And what better disguise can there be than becoming a physical part of the target- just the kind of tactics used by HIV.
HIV, or human immunodeficiency virus, is a pathogen, which infects humans and causes AIDS (acquired human immunodeficiency syndrome). Around 38M people worldwide are infected with HIV and, while with appropriate treatment patients can live for many years, there is still no cure or vaccine against the virus (World Health Organization 2016). The major treatment for the people with AIDS is an antiretroviral therapy (ART). ART is essentially a cocktail of various drugs, which stop the virus at different stages in its life cycle. However, preventing the virus from replicating is only a part of the challenge, as HIV is very good at playing hide-and-seek. HIV primarily targets our immune cells- the very same cells which are meant to seek and destroy the invaders. When HIV encounters an immune cell it enters and becomes part of the cell by integrating into the cell’s genome. Integration into a host genome is an essential part of HIV’s life cycle and it is also a perfect Trojan horse strategy. When HIV becomes part of the cell’s genome it is no longer recognised as a foreign entity and can remain dormant there until the environment is safe to reveal itself. The ART drugs are only able to kill an actively replicating virus but the dormant virus can remain integrated into the genome for many years. Consequently, HIV patients have to use ART drugs every single day for the rest of their lives to prevent virus from re-emerging, killing the immune cells and leading to AIDS.
The challenges for treating HIV infections still remain great, however, the incredible advancements in gene editing and therapy technologies are paving a way for a brighter future. Gene editing is basically an on-demand ability to change and modify any gene within a given genome. Because human genome has thousands of genes, the targeting of specific genes, without interfering with the functions of other genes, has always been a challenge. However, experiments using CRISPR (clustered regularly interspaced short palindromic repeats) technology (Figure 1) suggest that there is a way of eliminating the dormant form of HIV.
Figure 1. How CRISPR Works
The dormant HIV virus, also called a provirus, is essentially a gene in disguise. The cell’s surveillance system cannot distinguish the provirus from any other cellular gene. With the use of CRISPR, however, there is a possibility of cutting out this non-native gene from the genome (Figure 2). CRISPR can be specifically targeted to a single gene via a guide RNA molecule and the guide RNA can be made to recognise the HIV sequence in the genome. Once the CRISPR system has identified the site of the provirus, it can cut it out, which leaves the cell free of the HIV.
While in laboratory experiments this strategy has been very successful (Liao et al. 2015, Kaminski et al. 2016a, Kaminski et al. 2016b), the challenge has remained in being able to deliver the CRISPR system into a living organism. Recently, however, a combination of CRISPR and decades-long advancements in gene delivery methods showed the potential of this strategy in animals too. Adeno-associated virus vectors (AAVs) have for many years been used as treatment-delivery systems. AAVs can be made to carry almost any gene and target specific cell types. The genes carried by AAVs integrate into a genome and are expressed inside the cells like any normal gene. Last year the AAVs engineered to carry a provirus-targeting CRISPR have been used to remove an integrated HIV genome from mice (Ebina et al. 2013). As much as 90% of blood cells were successfully depleted of provirus in the mice, indicating the potential for the clinical use of this approach.
Figure 2. Using CRISPR against HIV
While gene therapy approaches to remove the provirus from a genome have still a long way to go, gene therapy methods that prevent the virus from entering a cell in the first place have already reached clinical trials. When HIV encounters an immune cell it has to bind a receptor on its surface. The receptor, called CCR5, acts like a lock into which HIV inserts its key and opens the door into the cell. Just like with any lock, a slight change in the CCR5 can prevent the key from fitting in. Interestingly, it has been observed that a small deletion in CCR5 gene can prevent or attenuate HIV infection. 1% of Caucasians naturally carry the CCR5 deletion, indicating that it is not deleterious for humans. This observation suggested that engineering patient cells to carry the CCR5 deletion could be a potential treatment against the HIV. Indeed, experiments in a lab using CRISPR to introduce the CCR5 deletion have successfully produced immune cells that are resistant to HIV infection (Ye et al. 2014). First clinical trial exploring the same principles has also showed some promise. The patients whose blood cells where isolated and modified to carry CCR5 deletion showed a much slower re-emergence of the virus in their blood in the absence of ART (Tebas et al. 2014).
Gene therapy has traditionally been used to treat genetic diseases, however, the advancements in gene editing technologies now enable the use of gene therapy to treat infectious diseases as well. Many challenges still remain for the use of CRISPR against HIV. HIV is renowned for its ability to change and evade many treatment strategies. Potentially, HIV could outsmart CRISPR by changing its sequence, so that CRISPR can no longer recognise it. Even more so, the use of CRISPR in humans has still many ethical and safety issues to be resolved. However, until effective vaccines against HIV are available, combination of ART and gene therapy approaches may be the only way of developing a cure against HIV.
Ebina, H., et al. (2013). “Harnessing the CRISPR/Cas9 system to disrupt latent HIV-1 provirus.” Sci Rep 3: 2510.
Kaminski, R., et al. (2016a). “Excision of HIV-1 DNA by gene editing: a proof-of-concept in vivo study.” Gene Ther 23(8-9): 690-695.
Kaminski, R., et al. (2016b). “Elimination of HIV-1 Genomes from Human T-lymphoid Cells by CRISPR/Cas9 Gene Editing.” Sci Rep 6: 22555.
Liao, H. K., et al. (2015). “Use of the CRISPR/Cas9 system as an intracellular defense against HIV-1 infection in human cells.” Nat Commun 6: 6413.
Tebas, P., et al. (2014). “Gene editing of CCR5 in autologous CD4 T cells of persons infected with HIV.” N Engl J Med 370(10): 901-910.
World Health Organization. (2016). “HIV/AIDS.” Retrieved Jan 16, 2017, from http://www.who.int/mediacentre/factsheets/fs360/en.
Ye, L., et al. (2014). “Seamless modification of wild-type induced pluripotent stem cells to the natural CCR5Delta32 mutation confers resistance to HIV infection.” Proc Natl Acad Sci U S A 111(26): 9591-9596.
By Bernadeta Dadonaite; PhD Student, University of Oxford.