Host and Viral Determinants of Interferon Resistance During HIV-1 Transmission

Context. To sustain themselves, all viruses must be transmitted to a new host. Accordingly, strategies interfering with this stage of the viral lifecycle can have an enormous impact on human health, wellbeing and productivity. Interventions such as vaccination, basic hygiene, sterile medical devices and the screening of blood products have been hugely beneficial because they can prevent infection altogether (rather than treating an already infected patient). Thus, there is an urgent need to improve our understanding of virus transmission as this knowledge will underpin future strategies of viral infection prevention.

Although effective HIV-1 treatments are available for those who can afford them, more than 2 million people become newly infected with HIV-1 every year worldwide. Infected patients can have more than a million HIV-1 virus particles in a single ml of blood. But even as viral loads can be high, the majority of new HIV-1 infections originate from infection by a single virus particle. There is great interest in understanding exactly why these particular particles are successfully transmitted, as interventions that specifically block transmission of these particles could prevent lifelong infection.

Interferons are an early defence against invading pathogens that can signal infection and alert cells to increase the levels of their antiviral defences. Because these defences are increased in response to interferons, they are known as interferon-stimulated genes (ISGs). As a result of the hostile environment created by ISGs, most viruses (including HIV-1) are less able to replicate in the presence of interferons. However, unlike the average HIV-1 variant, the transmitted HIV-1 particles are far more resistant to interferon-mediated inhibition. This observation underpins the idea that host interferon responses are a substantial barrier to HIV-1 transmission, such that an HIV-1 variant that is interferon resistant is more likely to be successfully transmitted.

Aims. Very little is known about which ISGs constrain HIV-1 transmission and how transmitted HIV-1 avoids this inhibition. Our overall aim is to illuminate the molecular details of this important and vulnerable stage of the HIV-1 lifecycle. We will do this by identifying: (i) the host factors that transmitted HIV-1 overcomes, and (ii) the mechanistic details that enable transmitted HIV-1 to resist interferon inhibition.

To map the determinants of interferon resistance in transmitted HIV-1 particles, we will make hybrid viruses comprised of interferon sensitive and interferon resistant HIV-1 viruses, in order to locate the specific region(s) of HIV-1 that confer interferon resistance. We will also grow interferon sensitive HIV-1 in the presence of interferons, in order to learn how the virus adapts to this inhibition. We can then examine any identified resistance motifs in HIV-1 sequence data (from infected patients) to determine whether these motifs might influence HIV-1 transmission.

To identify the host factors resisted by transmitted HIV-1 we will carry out screens to systematically measure the ability of hundreds of individual ISGs to inhibit HIV-1. Crucially, we will compare transmitted interferon resistant HIV-1 virus variants to interferon inhibited HIV-1 variants to identify ISGs that are specifically resisted by transmitted HIV-1.

Applications and benefits. During the timeframe of this award we plan to illuminate an exciting area of HIV-1 biology and stimulate further research. In the longer term, our research defining the molecular details of HIV-1 transmission should aid the design of novel prevention/intervention strategies (such as vaccines) designed to target transmitted HIV-1. Furthermore, because sustainable transmission is an essential facet of pandemic viruses, the insight our work will provide on transmission of HIV-1 should improve our ability to assess the pandemic potential of other emerging viral pathogens.

The hypothesis underlying this proposal is that transmitted/founder (TF) HIV-1 is more interferon resistant than other HIV-1 genetic variants, and that specific interferon (IFN)-stimulated genes (ISGs) underlie this phenomenon. We will identify and characterise ISGs that limit HIV-1 transmission and define how TF viruses avoid this inhibition.

The most important restriction factors targeting HIV-1, such as APOBEC3G, TRIM5, tetherin and SAMHD1, were identified using cell lines. However, TF HIV-1 interferon resistance is typically only revealed in primary cell systems. In our preliminary experiments, we unexpectedly observed that TF viruses can exhibit striking IFN resistance in cell lines. Thus, this important phenotype is now amenable to investigation using more tractable experimental systems.

To probe the determinants of TF HIV-1 IFN resistance, we will perform fine mapping using chimeric viruses comprised of our already identified IFN sensitive and IFN resistant infectious molecular clones (IMCs). We will also expand this approach to other IFN-phenotypes using chimeric IMCs, and will use in vitro evolution to identify additional viral IFN sensitivity/resistance determinants. Importantly, we will validate our observations using primary cells and patient sequence data.

To identify the ISGs resisted by TF HIV-1, we will screen over 500 individual human ISGs to identify genes that inhibit HIV-1. Using matched TF and chronic control (CC) IMCs, we will uncover anti-HIV-1 ISGs that are resisted by TF HIV-1. We have substantial expertise with ISG screens and have previously studied restriction factors inhibiting multiple stages of the HIV-1 lifecycle, leaving us well placed to characterise how these ISGs inhibit HIV-1.

Our ultimate goal is to generate and disseminate important knowledge illuminating HIV-1 transmission. This will stimulate further research in this area, and in the long term, inform the design of intervention strategies targeting TF HIV-1.

Our research seeks to contribute to the MRC's strategic aims by addressing a global health pandemic in a fashion that will deliver basic information about a fundamental host-pathogen interface. By uncovering critical information about what happens during the earliest events of HIV-1 transmission, we will produce research of both short- and long-term value to diverse segments of society. For fellow academics, our work will contribute immediate benefit through the attainment of new knowledge that can inspire further research, perhaps for many years to come. For pharmaceutical companies and healthcare professionals, our research will be of longer-term benefit as it may aid the development of novel vaccines, antiviral treatments and biotechnologies. And perhaps most importantly, our research will be of great value to the wider public as it will not only aid our understanding of viral emergence and resilience, but it has the potential to contribute to interventions with real human health benefits, which, in turn, will have economic benefits.

The specific areas in which we believe our work can have significant impact include:

Viral emergence: HIV-1 has entered humans from non-human primates multiple, chronologically distinct times, yet only a single cross-species transmission resulted in the HIV-1 group M pandemic, highlighting the likelihood that specific conditions were required for HIV-1 emergence. As HIV and related SIV exist as extremely diverse populations within hosts, these conditions were likely dependent upon the specific genetic variant of the virus that was transmitted. Our work seeks to reveal more about how HIV is transmitted-namely to identify some of the phenotypic characteristics of the virus that are required for transmission between humans, as well as some of the early immune defenses that must be overcome for productive infection. By understanding these facets of transmission, not only do we grasp more about how this virus (and perhaps other viruses) emerged, but we also gain a better grip on how resilience might be engineered, and we learn better strategies for predicting or preventing further epidemics or pandemics. The economic and social benefits of preventing major virus outbreaks are enormous and cannot be underestimated.

Vaccine development: Despite the introduction of antiretroviral therapies, there were more than 2 million individuals newly infected with HIV-1 in 2014 alone (WHO/UNAIDS/Unicef), suggesting that development of a successful vaccine may offer the greatest hope of eliminating HIV-1 transmission. Several HIV-1 vaccine strategies have been proposed or tested, and these largely rely upon either priming immune cells to eliminate infected cells, or eliciting antibodies that either directly neutralize the virus or instigate killing of infected cells. In all of these strategies, the success of the vaccine relies upon the interaction of the host defenses with specific transmitted founder (TF) virus particles. As our work seeks to reveal more about TF viruses, namely how they resist host interferon defenses (and interferon-stimulated genes), we believe our research will contribute information that will critically inform future vaccine efforts, hopefully to global public health and economic benefit.

Novel therapies/prevention/biotechnologies: For reasons similar to those described above, we believe that a better understanding of TF viruses will reveal points of transmission vulnerability. These vulnerabilities can then be exploited for either prevention (alternative pre-exposure prophylaxis methods) or other novel therapies.

The history of HIV-1 research has shown that basic molecular virology, like the kind described herein, plays a vital role in pushing forward more translational approaches. While the immediate beneficiaries of our research will primarily be other academics, the longer-term benefits and beneficiaries over a period of many years may be immeasurable.