Ebola outbreak poses major drug research and development challenges

Published: 7-Nov-2014

The severity of the current Ebola crisis in West Africa has resulted in a rush to speed up development of potential treatments. But despite its simplicity, the virus presents significant challenges. Dr Sarah Houlton reviews drugs and vaccines in the pipeline

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Outbreaks of deadly infectious diseases make for terrifying headlines. However distant the outbreak is, and however difficult it is to catch, human nature makes us assume the worst. The Ebola virus disease outbreak in West Africa is a prime example. It is a horrible disease and frequently fatal, yet it is only transmitted via bodily fluids from people with active infection. It cannot be caught from an infected person during the incubation phase, which can last anywhere between two and 21 days.

Once infection sets in, the patient experiences a rapid onset of fever, along with weakness, muscle pain, headache and a sore throat. Next come diarrhoea, vomiting, impaired liver and kidney functions and a rash, plus, in some cases, internal and external bleeding. Currently, intense supportive care, including rehydration, is the only real therapy, and the sooner this starts the more likely the patient is to survive. The death rate can be as high as 90%. Health workers are at particular risk because of their close contact with patients, and strict infection control is essential. Despite this, several health workers have been infected, most likely through failures in gowning procedures.

Health workers are at particular risk because of their close contact with patients, and strict infection control is essential

The virus was first identified after two outbreaks in 1976, one in Sudan and one in Zaire (now the Democratic Republic of Congo), near the Ebola river, from which the virus takes its name. The current outbreak in West Africa was recognised in March 2014, first in Guinea, and spreading to Sierra Leone and Liberia. The epidemic was declared an international public health emergency by the World Health Organization in August, and several thousand people have died.

Five species of Ebola virus have thus far been identified, including the Zaire strain, responsible for the current outbreak. The Bundibugyo and Sudan strains have also caused big outbreaks in Africa; the other known two species are Reston and Taï Forest.

Ebola is a filovirus, the only other known members of this class being the closely related Marburg virus and the more distantly related cuevavirus. Ebola is an RNA virus that encodes just seven structural proteins and one non-structural one. The polymerase L protein is a particularly interesting therapeutic target, as if it is blocked it should prevent almost all RNA synthesis, and there is no similar protein in mammalian cells. Two other genes, virion proteins 24 and 35, also have potential as targets, as they inhibit interferon responses and thus if they are blocked virulence could be reduced.

As such a simple virus, Ebola presents a huge challenge to drug and vaccine developers. But various products are in the – largely – early stage of development, with research driven by the potential threat of Ebola as a bioterrorism agent. Because of the huge public health emergency, there has been a rush to give experimental drugs to patients, particularly infected health workers, in spite of the fact that minimal – or even no – human clinical trials have been carried out.

As such a simple virus, Ebola presents a huge challenge to drug and vaccine developers

The one that has gained most media attention, the antibody cocktail ZMapp, is being developed by Mapp Pharmaceuticals’ commercialisation arm, LeafBio in San Diego, CA, US. ZMapp combines the best three antibodies from two other antibody cocktails – one from MB-003, a three-antibody cocktail developed by Mapp with funding from the US government, and two from another three-antibody combination, ZMab, licensed from Toronto, Canada-based Defyrus, a biodefence company, and created with Public Health Canada funding.

MB-003 combines three different human and human-murine chimeric antibodies, first produced in Chinese hamster ovary and subsequently in whole-plant tobacco cells in a process developed by Kentucky BioProcessing to speed up manufacturing. Early studies in rhesus macaques showed that it protected the monkeys from infection when dosed an hour after infection; four out of six survived – and neither of two controls – when dosed 24 or 48 hours after infection.1 In a further trial, infected macaques were treated with MB-003 after the onset of disease symptoms. Three out of seven survived, compared with none of the controls, either in this or any historical trials.2

Defyrus’s ZMab is another three-antibody cocktail, with the antibodies directed against the virus’s envelope glycoproteins. In an initial trial in cynomolgus macaques, the monkeys were given three doses three days apart, starting 24 hours after infection with Ebola. All four survived. When started 48 hours after the lethal challenge with virus, two out of four fully recovered.3 When ZMab was combined with an adenovirus-vectored interferon-alpha, three out of four cynomolgus macaques survived, and all four rhesus macaques, when treatment was initiated after the detection of viraemia.4

The ZMapp cocktail has now been tested in rhesus macaques, and was able to rescue all of them when treatment was started up to five days after challenge with Ebola virus. Advanced disease could be reversed, including mucosal haemorrhages, elevated liver enzymes, and generalised petechia.5

A molecule already approved in Japan for influenza is being investigated in Ebola by Boston-based MediVector. Favipiravir is a broad-sprectrum nucleoside analogue that inhibits RNA-dependent RNA polymerase. Now in Phase III in North America for flu, it has shown promise in mice in a study by a team at the Bernhard-Nocht-Institute for Tropical Medicine in Hamburg, Germany. The mice were dosed with the drug six days after infection, at which point the disease had become symptomatic. All untreated mice died within 10 days; all of those treated with the drug survived.6 With Japan having already stockpiled the drug for use in case of an influenza pandemic, it offered to supply it for experimental treatment of Ebola. In early October, it was reported that a French nurse who contracted Ebola while working for Médicins sans Frontières in Liberia recovered after treatment with favipiravir.

Brincidofovir is a second antiviral being repurposed as a potential Ebola treatment. North Carolina-based Chimerix has been investigating the orally available nucleotide analogue in cytomegalovirus and adenovirus infections, and it is now in Phase III trials for those indications. It has also shown in vitro activity against Ebola, and thus the company has made stocks of the drug available for use as an experimental rescue therapy in patients with confirmed Ebola infection; the company says that the FDA has approved it for emergency use, and some patients in the US have received treatment. Formal Phase II trials are being planned.

Brincidofovir

Brincidofovir

Another molecule that inhibits RNA-dependent RNA polymerase is being investigated by BioCryst Pharmaceuticals. The broad-spectrum antiviral adenosine analogue BCX4430 was shown to have a favourable preclinical safety programme. In rodent models, it protected against filovirus infection, and completely protected cynomolgus macaques from Marburg virus infection up to 40 hours after infection.7 The US National Institute of Allergy and Infectious Diseases (NIAID) has provided several tranches of funding in recent months for its development in haemorrhagic fever virus diseases. This includes finance for safety and efficacy studies in non-human primates, and for the molecule’s manufacture.

BCX4430

BCX4430

Inhibikase Therapeutics, based in Atlanta, GA, US, is working on a potential Ebola treatment with a different mechanism of action. IkT-001 Pro, an extended release form of the leukaemia treatment imatinib (marketed by Novartis as Glivec) is being developed to treat rare diseases caused by polyomaviruses, but it has the potential for activity against Ebola, and also smallpox. The drug inhibits c-Abl1 tyrosine kinase, interfering with the release of viruses from cells.8 The company says that, although the drug hits the same targets in all these diseases, in polyomavirus infection it interferes with viral entry into cells, but in Ebola it blocks the spread of infection around the body.

Antisense compounds could have potential, too. Tekmira’s TKM-Ebola viral RNAi therapeutic was granted fast track designation by the FDA in March. The small interfering RNAs target Ebola’s RNA polymerase L protein, and are formulated in stable nucleic acid–lipid particles. Early studies showed that it could protect guinea pigs when they were dosed shortly after a challenge with Ebola.9 In a subsequent study in non-human primates, three rhesus macaques were given the siRNA product 30min after viral challenge, and on days 1, 3 and 5. A second group of four macaques was given the first dose plus six subsequent daily doses. All those given seven doses survived; two of those that received four doses were protected. A Phase I clinical trial in humans commenced in January of this year.

A further antisense oligonucleotide has been developed by Sarepta Therapeutics. AVI-7537 is an antisense phosphorodiamidate morpholino oligomer that targets the VP24 gene of the Ebola virus.10 The company’s funding from the US Department of Defense was cancelled in 2012 thanks to a tightening of the federal budget in the light of the impending fiscal cliff. The company says it had a cure rate of 60–80 % in rhesus macaques, and while it has been evaluated for safety in healthy humans, it has not been given to those infected with Ebola. The current outbreak could well see its return.

The potential of vaccines

As with so many viral diseases, prevention will most probably be more effective than cure, and multiple vaccines are under development – both DNA vaccines and viral vaccines. A viral vaccine from GSK, cAd3-ZEBOV, has been fast-tracked into development, and the first volunteers were dosed with the product in the UK and the US in September. The first safety indicators were good, with no serious adverse reactions in those initial trial subjects. The Phase I trials will look at whether the vaccine raises antibodies to Ebola virus as well as evaluating its safety.

Ebola outbreak poses major drug research and development challenges

These studies should be complete by the end of the year, by which time it should be clearer whether it is likely to confer any degree of protection against Ebola infection. Only once the trials in the UK and the US are done will the trial be expanded, with the plan to run further trials in the Gambia and Mali. If it proves both safe and immunogenic, there is the potential to fast-track it into the wider at-risk population.

The product is a recombinant chimpanzee adenovirus engineered to deliver Ebola genes, whose identity has not been disclosed, from the Zaire species of the virus responsible for the current outbreak.11 Preclinical studies of the vaccine in monkeys looked promising. It was developed by Okairos, acquired by GSK last year, in conjunction with the US National Institute of Allergy and Infectious Diseases.

As with so many viral diseases, prevention will most probably be more effective than cure

A second potential Ebola vaccine has just entered Phase I human trials, following successful studies in animals. rVSV-EBOV-GP was developed by the Public Health Agency of Canada, and licensed to BioProtection Systems, a subsidiary of Iowa-based biotech NewLink Genetics. The vaccine targets the virus envelope protein, inducing antibodies that neutralise the virus. It was created using the rVSV platform, based on attenuated strains of vesicular stomatitis virus that has been modified to express the Ebola envelope protein.

Both mice and guinea pigs were immunised with the vaccine, and then challenged with a lethal dose of Ebola virus several months after vaccination. Almost all survived.12 Numerous studies in non-human primates have also been carried out, including in immuno-compromised rhesus macaques. Four out of six survived infection with Ebola.13

Other experimental vaccines are set to enter the early stages of clinical evaluation. For example, Inovio Pharmaceuticals’ experimental product is a DNA vaccine that is being advanced into Phase I in collaboration with GeneOne Life Sciences, a specialist in the manufacture of this type of vaccine. They expect the study will start in early 2015.

The product is based on Inovio’s SynCon technology, which the company claims can elicit immune responses against multiple disease-specific antigens, giving protection against diverse strains of pathogens in humans. The vaccines incorporate a DNA fragment that causes cells to produce just the targeted antigen; it cannot replicate and thus is unable to cause the disease. Importantly, they rely on the genetic code for a specific antigen from multiple strains of the target pathogen, conferring protection against more than one strain of a disease – a strategy that early clinical results in influenza support. The vaccines can be made through standard fermentation techniques, speeding up production in an emergency, and are more stable at room temperature than a traditional vaccine.

Despite recent efforts there are still no licensed products, and human testing remains minimal for all potential treatments

This Ebola vaccine was designed to give broad protective antibody and T-cell responses against multiple strains of the virus. It contains three consensus plasmids that target strains of three different families of Ebola and Marburg viruses, and is delivered via the company’s proprietary electroporation technique. Initial studies on animals showed its promise. All of a group of twice-vaccinated guinea pigs survived after a virus challenge, with significant increases in neutralising antibody titres, and strong, broad levels of vaccine-induced T-cells. A subsequent study in single-vaccinated mice also prevented death.14

Bavarian Nordic and Johnson & Johnson are teaming up to create a product that combines their anti-Ebola vectors. Bavarian Nordic’s vaccine uses the company’s MVA-BN technology, whereby modified vaccinia Ankara (already employed in its marketed smallpox vaccine) is used to deliver Ebola genes which, again, have not been disclosed. The product from J&J’s Crucell unit, meanwhile, is made using its AdVac recombinant adenovirus technology, used to deliver Ebola GP1. In collaboration with NIAID, the combination has already shown promise as a prime-boost regimen against Ebola in macaques, and human trials are scheduled to begin in 2015.

Despite recent efforts there are still no licensed products, and human testing remains minimal for all potential treatments bar those already investigated in other indications. Funding issues put some of the potential anti-Ebola products on the back-burner, slowing development down further. But a public health crisis like the current Ebola virus disease outbreak concentrates the collective mind, and with the infection having such catastrophic consequences, there is little wonder that those products that are under development are being fast-tracked, and given to patients as a last resort. We are a long way from being able to treat the disease – and probably still further from being able to prevent it. But without the current efforts, and the willingness of regulators to take risks, these solutions would be even more of a distant dream.

References

1. G.G. Olinger et al. PNAS, 2012, 109, 18030

2. J. Pettitt et al. Sci. Transl. Med. 2013, 5, 199ra113

3. X. Qiu et al, Sci. Trans. Med. 2012, 4, 138ra81

4. X. Qiu et al, Sci. Transl. Med. 2013, 5, 207ra143

5. X. Qiu et al. Nature 2014, 514, 47

6. L. Oestereich et al. Antivir. Res. 2014, 105, 17

7. T.K. Warren et al. Nature 2014, 508, 402

8. M. García et al. Sci. Trans. Med. 2012, 4, 123ra24

9. T.W. Geisbert et al. J. Infect. Dis. 2006, 193, 1650

10. P.L. Iversen et al. Viruses 2012, 4, 2806

11. S. Colloca et al. Sci. Trans. Med. 2012, 4, 115ra2

12. G. Wong et al. Vaccine 2014, 32, 5722

13. T.W. Geisbert et al. PLoS Pathog. 2008. 4, e1000225

14. D.J. Shedlock et al. Molecule. Ther. 2013, 21, 1432

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