A target for an Ebola antiviral?

A CRISPR screen identified the cellular gene GNPTAB as being important for Ebola infection
A target for an Ebola antiviral?

At the time of writing, there have been 628 cases of Ebola virus disease in the Democratic Republic of Congo (DRC) since July 2018, making it the second largest Ebola outbreak ever recorded. The largest was the West African epidemic of 2013-2016, in which more than 11,000 people died. Today the world is better prepared for an Ebola epidemic than it was in late 2013: an experimental vaccine and therapies are being administered under compassionate use protocols in DRC. However, there is still much work to be done to ensure we have better countermeasures to prevent and treat Ebola virus disease.

Of the eleven authors on this manuscript, seven were directly involved in Ebola response activities in the West African outbreak, mostly performing diagnostic testing supporting the Médicins Sans Frontières unit in Bo, Sierra Leone. Andrew Tai, from the University of Michigan, first contacted us in September of 2014, just as this epidemic was escalating. Andrew and his graduate student, David Lin, were using a CRISPR screening library to identify host factors for dengue virus infection and they proposed performing a similar screen with some highly pathogenic viruses, such as Ebola. Through their dengue studies, Andrew and David had worked out a pipeline for screening and the subsequent next-generation sequencing and bioinformatic analysis.

We were extremely interested in their proposal as the CRISPR system was, and still is, an exciting system with huge possibilities for manipulating sequences on a genome-wide basis. In addition, while others had performed genome-wide screens using surrogate systems under BSL-2 containment, we had access to a BSL-4 laboratory and could use live Ebola virus in our work. Using the authentic virus, we thought that we might find something that others did not.

Unfortunately, the epidemic continued to escalate through late 2014 and into 2015. Media interest flared as infected healthcare workers were transported to Western hospitals for treatment. In this environment, basic research projects were pushed to the back burner as other, more pressing demands arose.

An aside: people often ask what working in BSL-4 containment is like. The over-riding sensation is that it’s loud! Wearing earplugs helps to block out the noise of the air being pumped through your positive-pressure suit. The work is often physically and mentally demanding, requiring a high level of concentration. Techniques that are simple at BSL-2 are considerably more difficult in a suit; if something takes an hour at BSL-2, allow for three hours at BSL-4. Working in BSL-4 can be strangely peaceful though, focusing completely on your work without outside distractions - a few hours’ escape from phone calls and emails.

After the West Africa Ebola outbreak began to subside, David in Michigan transduced cells with the CRISPR library and shipped them to us in Atlanta. We infected them with Ebola in our BSL-4 lab. Some cells survived and were expanded, and had their DNA sequenced. We found a number of hits in our screen, but initially decided to focus on the gene GNPTAB. This was partly because GNPTAB variants are associated with a known disease, the slowly progressing metabolic disorder mucolipidosis. A key aspect of our paper was being able to use fibroblasts from mucolipidosis patients, and show that these do not support Ebola virus infection.

As always happens, our work raises a number of follow-up questions. Does GNPTAB-knockout impair the course of Ebola infection in vivo? This could be answered using a GNPTAB-knockout mouse and mouse-adapted Ebola virus. Could GNPTAB be targeted by an antiviral? Such a drug would be likely to induce symptoms of mucolipidosis, but might be beneficial over a short period. Perhaps if given in combination with a direct-acting antiviral, such as a polymerase inhibitor like remdesivir, a relatively low dose could be given for a brief time to avoid serious side effects. Finally, a number of gene hits in our screen are yet to be investigated for their potential roles in Ebola infection. We hope to be able to explore these further in the coming months.

Pictures: Jessica Spengler, editing: Tanya Klimova