Outline of Ebola Presentation

The Past: Summary of the West Africa Outbreak with contextual history of Ebola.

The Present:

  1. Ebola Survivor syndrome
  2. Immune privilege site where Ebola continues to live
  3. Eye, Testes, Brain
  4. Ebola Diagnostics
  5. Ebola Treatment
  6. Ebola Vaccines

The Future: What did we (could we) learn?

  1. More people died from other diseases in West Africa than from Ebola
  2. Coordinating global health response
  3. Politics of Fear

Time: 12:05 to 12:45 40 minutes

  1. Pre-Test questions: 5 minutes
  2. Presentation: 30 minutes
  3. Post-test questions: 5 minutes

Literature:

  1. Medley GF and Vassall A. 2017. When an emerging disease becomes endemic. Science 357, 156-158 (2017). 14 July 2017.

The “public health paradox”: If the epidemic is successfully controlled, then it is highly likely that the eventual impact of the epidemic disease will have been less than the opportunity cost of the resources allocated to it from other health areas.

  1. Venkatraman N, Silman D, Foleggati PM, Hill AVS. 2017. Vaccines against Ebola Virus. Vaccine.

12/26/2013: 18-month old boy in Meliandou, Guinea – fatal illness with fever, black stools, vomiting. The start of Zaire Ebola Virus Disease

03/24/2014: WHO public announcement of 49 cases ad 29 deaths

08/08/2024: WHO declares Public Health Emergency of International Concern (PHEIC) – 1711 reported cases and 932 deaths

03/26/2016: PHEIC lifted: 28,646 confirmed, probably and suspected cases in Guinea, Liberia and Sierra Leone with 11,323 deaths and estimated 17,306 survivors

881 healthcare workers were infected and 513 died

May 2017: DRC had 8 cases and 4 deaths

1976: simultaneous outbreaks in Sudan and Democratic Republic of Congo.

29 isolated outbreaks since then

Negative-strand RNA virus – 7 non-segmented genes encoding viral proteins and a single glycoprotein (GP

GP contains 2 subunits as trimeric spikes onviral surface. Pivotal role in cell attachment, fusion and cell entry – key antigenic target

History of Vaccine development:

1st attempts were inactivated whole virus. No clinical trials because potential safety concerns, failure to demonstrate efficacy in non-human primate (NHP) models.

1990s: pre-clinical studies expressing envelope GP or nucleocapsid (NP) genes demonstrated efficacy in ‘gold-standard’ model of cynomolgus macaques

Replication deficient viral vector vaccines:

2003: 1st ever human clinical trial: 3-plasmid DNA vaccine encoding transmembrane deleted GP from the Zaire and Sudan viruses and NP, showed that s 3-dose schedule was safe and immunogenic.

2010: replication-defective recombinant human adenovirus serotype 5 vaccine (rAd5) encoding GP genes with a point-mutation. Single dose – safety trial. Problem: pre-existing immunity again Ad5

2015: 2ndhuman clinical trial assessing safety and immunogenicity of constructs encoding wild type GP from Ebolavirus Zaire (EBOV), Sudan (SUDV and Marbrgvirus Angola strain. Result: vaccine safe but multiple doses required and immune response waned at 32 weeks

Work-around pre-existing immunity to Ad5 – use chimpanzee Ad3: 100% efficacious with zero viremia against both EBOV and SUDV. Booster with replication deficient modified vaccinia Ankara (MVA) 8 weeks later, enhanced durable protection.

Recombinant, replication competent vesicular stomatitis virus (rVSV)-based vaccine encoding EBOV GP showed encouraging results in NHP. Given to 1 patient on compassionate basis after needle-stick injury (2011)

Start of Ebola epidemic: 2 leading vaccine candidates entered Phase 1 clinical studies in centers in 3 contenents:

  1. Monovalent and bivalent ChAd3-vectored vaccine
  2. rVSV-vectored vaccine

Chimpanzee adenovirus 3 vectored vaccine (ChAd3):

  • given to humans in UK, Europe, US and in Mali (2016)
  • Large phase III trial in Liberia amended to a phase II because of waning cases
  • Being assessed in children ages 1 to 17 years in Nigeria, Mali and Senegal (ongoing)

Vesicular stomatitis virus vectored vaccine (rVSV):

  • Phase 1 clinical trials started in Europe and Africa shortly after ChAd3) trial
  • 10 – 20 % of healthy volunteers suffered severe adverse effects: arthralgia, arthritis with entry of VSV into CSF and 25 – 30% suffered fever.
  • Chosen by WHO for Phase III trial Ebola Ca Suffit!) in Guinea in a ring vaccination strategy – it worked 100% protection

Modified vaccinia Ankara vectored vaccine (MVA):

  • MVA-Bn-Filo – quadrivalent vaccine encoding GPs from EBOV and SUDV and GP from Marburg and NP from the Tai forest strain was tested with a 1-2 week booster dose (rather than 8 week)

Human adenovirus vectored vaccines:

  • rAd26 along with MVA-Bn-Filo was trialed as a prime and booster vaccination at intervals of 2, 4 and 8 weeks. Responses were noted up to 8 months.
  • Ad26 tested with MVA in a large phase 3 trial in Sierra Leone (in progress)
  • rAd5 vaccine containing the GP of 2014 strain of EBOV was safe and immunogenic in healthy adults in China and Sierra Leone.

Ring Vaccination strategy

  • developed in 1960s for smallpox eradication
  • vaccination of contacts and people at risk (contacts of contacts)
  • used for MMR outbreaks
  • Eboa ca Suffit! (Ebola, that’s enough!) trial (2017) showed 100% protection up to 10 days after vaccination (trial limitation)

Correlates of protection – a marker

  • None yet! Looking at GP antibody titers
  • What titer level determines protection?

ZMapp treated patient showed reversal of viral load, also got convalescent plasma, brincidofovir – a nurse treated at Royal Free Hospital in London; 9 months later, resurgence of virus in CNS and systemic circulation ((Jacobs et al, 2016)

Vaccines – Lessons learned

  • Survivors show persistent virus in semen, breast milk and immune privilege sites (eye, brain): this may lead to recurrence
  • We may see further outbreaks – spill-over events
  • Accelerated development of vaccine candidates during the epidemic was remarkable and demonstrated in the epidemic
  • Ring vaccination strategy was a innovative trial design implemented
  • Gained great insight into the role of open communication and research collaboration when doing clinical trials in resource-poor countries with limited infrastructure.
  • We need to find an immunological correlate of vaccine protection
  • There is still no licensed vaccine that can be manufactured, stockpiled and rapidly deployed
  1. Shantha, JG, Crozier I, Yeh, S. 2017. An update on ocular complications of Ebola Virus Disease.CurrOpinOphthalmol 28 (0). Sep 1. doi: 10.1097/ICU.0000000000000426.

17,306 survivors of EVD

Post Ebola Virus Disease Syndrome (PEVDS):

  • Ocular disease – uveitis the most common - 13 – 34 % of survivors
  • Arthritis
  • Hearing loss
  • Abdominal pain
  • Neuropsychiatric disorders
  • Virus persistence in immune-privileged organs

Clinical Features of Ebolavirus infection

  • Filoviridae family: Ebola and Marburg
  • Enveloped, nonsegmented single stranded RNA virus
  • 5 different EBOV species:
  • Zaire – cause of West African epidemic – highest fatality rate, hemorrhagic fever (EBOV)
  • Sudan (SUDV)
  • Tai Forest (TAFV)
  • Reston – nonlethal in humans but lethal to NHP (RESTV)
  • Bundibugyo (BDBV)
  • Incubation period: 2 – 21 days
  • Sx: flu-like-illness, fever, headache, malaise, diarrhea
  • Animal-to-human transmission direct contact or consumption – animals: chimpanzees, gorillas, fruit bats, duikers
  • Exposure to body fluids: blood, saliva, sweat, urine, breast milk, feces, semen
  • Mucosal surfaces and open wounds
  • Macrophages and dendritic cells are infected →fibroblast, endothelial cells, hepatocytes, adrenal cortical cells, epithelial cells
  • Suppressed dendritic cell activation→ decreased cytokine release and T-cell activation
  • Leads to upregulation of macrophages with activation and release of cytokines to cytokine storm
  • Downregulates interferons
  • Clinically lead to increased vascular permeability, hypovolemic shock, multisystem failure, DIC, diffuse hemorrhage, maculopapular rash and death
  • Diagnosed in BSL 4 lab by RT-PCR or immunoglobulin M and G antibodies or specific antigen detection

Post Ebola Disease Syndrome

Fatigue

Vision loss

Headache, memory loss

Sleep disorders, amenorrhea, miscarriages

Enthesitis, arthralgias

Mechanism: elevated inflammatory cytokines, molecular mimicry, damage from virus and viral persistence in immune-privileged sites

Ocular Disease

Impacts ADLs and quality of life

Most common finding is uveitis – eye pain, redness and photophobia and may lead to acute or chronic vision loss

Conjunctival injection was seen in 48-58% patients with acute EVD and in one study predictive of acute EVD infection

Treatment was first 1% atropine and corticosteroids in previous epidemics (Kitwit)

Uveitis: posterior (57%) and panuveitis (29%) and anterior (14%)

Unilateral more common, vision worse than 20/400 in 39% affected eye

PREVAIL III:

  • NIH-sponsored study in Liberia longitudinally following EVD survivors and first degree contacts for eye disease.
  • 24% have uveitis
  • Younger age and more time in ETU greater risk

Repatriated healthcare workers

  • Physician 1
  • Unilateral anterior uveitis developed 40 days after initial EVD
  • Vision declined to 20/200
  • Resolved completely with topical homoatropine and oral corticosteroids
  • Labs suggested broad but transient systemic immune activation
  • Physician 2
  • Another physician had unilateral, hypertensive anterior uveitis that went to sight threatening panuveitis 10 weeks after illness
  • High levels of virus from anterior chamber paracentesis

Treatment:

Antivirals (one pt got favipiravir)

Steroids – topical and oral

More blindness in West African pts due to lack of timely intervention

Viral Persistence in Eye and immune-privileged organs

  1. Cultured from aqueous humor during the uveitis
  2. EVICT (Ebola Virus Persistence in Ocular Tissues and Fluids underway in Sierra Leone – evaluating prevalence of persistent EBOV in ocular fluids in survivors
  3. Also found EBOV RNA in CNS, placenta and semen up to 565 days post acute EVD
  4. Breastfeeding and sexual contact new transmission chains
  5. Identified in eye, brain and testes in rhesus monkeys (Zheng et al, 2017)
  6. Followed monkeys 43 days after EBOV exposure
  7. 9.8% detectable RNA in eye
  8. 1/11 in brain
  9. 1/11 in testes
  10. Capable of infecting human retinal pigment epithelial cells
  1. Smith JR, Todd S, Ashander LM, Charitou T, Ma Y, Yeh S, Crozier I, Michael MA, Apuukuttan B, Williams KA, Lyn DJ and Marsh GA. 2017. Retina, Pigment Epithelial Cells are a Potential Reservoir for Ebola Virus in the Human Eye. Transitional Vision Science & Technology 2017 6(4), Article 12.
  • Uveitis may be anterior or posterior
  • Anterior uveitis, painful, leads to glaucoma or cataract
  • Posterior leads to scarring in macular region of retina and visual disturbances
  • 75% of 277 cohort in Sierra Leone with uveitis developed visual loss and 26% went legally blind
  • Eye is an immune privileged site, i.e. limited inflammation – specificall the monolayers of pigment epithelial cells in posterior eye and iris and ciliary body in anterior eye
  • ocular pigment epithelial cells produce membrane-bou d ligans and soluble factors that inhibit inflammatory acitivities of leukocytes.
  • Smith et al grewEBOV in ARPE-19 human retinal pigment epithelial cells
  • Increased viral antigen and recoverable viral particles from the cytoplasm of the cell lines
  • EBOV increased interferon production by the infected cells perhaps modulating immune response
  • Ultimately, uveitis presents because the immune response overwhelms the immune privilege
  1. Steptoe PJ, Scott JT, Baxter, JM et al. 2017. Novel retina Lesion in Ebola Survivors, Sierra Leone, 2016. Emerging Infectious Diseases 23(7), July 2017, 1102-1109
  • Case-Control study in Freetown, Sierra Leone of 82 EVD survivors with 105 asymptomatic controls.
  • Novel retinal lesion found in 14.6% of EVD survivors and no controls CI (7.1 – 25.6%) that followed the anatomical distribution of the optic nerve axons, suggesting neuronal transmission as a route of ocular entry
  • Ocular disease occurs in 14 to 60% of EVD survivors
  • Acute uveitis 18 – 58%
  • Viable ZEBOV detected in aqueous humor 9 weeks after clearance of viremia
  • Figure 1 showed a significant difference in type 6 pigmented lesion
  • Figure 2 shows the lesions
  • Retinal lesion did not affect vision
  • Visual impairment was due white cataract in 7.3% of EVD survivors
  • No isolation of EBOV in aqueous humor in this study.
  • Uveitis accounts for 24% of blindness in Sierra Leone – 2nd most common cause, 1st being cataracts
  1. Racine T and Kobinger GP. 2017. Unlocking Ebola Persistence. Nature Microbiology 2, 17124

Report on Zeng et al, 2017 using rhesus macaques to isolate persisting EBOV from testes, brain and eye.

  1. Rojek A, Horby P and Dunning J. 2017. Insight from clinical research completed during the west Africa Ebola virus disease epidemic. Lancet 17: e80 – 92. 2017.

Clinical burden of disease

  • Median age 32 years (uncertain if report bias or true increased risk of exposure
  • No marked gender difference – 48.8% probable or confirmed cases were men
  • Young age predictor of death: Odds Ratio per year of life 0.91 (CI: 0.85 – 0.97)
  • For children, median time of 3 days from admission to ETU to death (study of 300 children)
  • Mortality higher in men > 45 years
  • Previous Case Fatality Rates (CFR) reported were maternal 90% and neonatal 100% -- possibly over-estimated.
  • Hemorrhagia – late finding; predominant finding of advanced disease was severe gastrointestinal disease
  • Sx: fever, fatigue, anorexia, vomiting, diarrhea, headache and abdominal pain
  • Volumes up to 10 L per day of stool output reported in repatriated patients.
  • Fever absent in 10% of cases (part of case definition)
  • Hiccups, confusion and conjunctivitis – good discriminatory importance (table 1)
  • 7.5% of pts in a study of 187 individuals were antibody positive (anti-EBOV glycoprotein)

but did not have EVD

CFR: 70% (CI 69 – 72) (WHO)

Mortality was lower in pts admitted to hospital (CFR 61% CI 59-62) compared to not admitted (88% CI 86-90)

CFR in Europe and USA was 19%

Table 1: improved characterization of the broad spectrum of organ involvement – important contribution to knowledge about EVD from this epidemic

Clinical Research on Organ Systems:

  1. Abdominal:
  • reason for diarrhea unclear
  • Possible paralytic ileus by ultrasound identified
  • Bowel wall edema, hypoproteinemia
  • Inflammation leading to bacteremia from gut
  1. Renal:
  • Study of 150 pts: acute kidney injury in 50% pts and independent predictor of death (OR 5.84, CI 1.15 – 29.58)
  • Precedes GI symptoms, so possibly rhabdomyolysis (myalgias, Increased CK and K), decreased perfusion due to DIC
  • Hypoglycemia in children
  • Hyponatremia, hypocalcemia, hypomagnesemia
  1. Hepatic:
  • Transaminitis, AST > ALT
  • High AST in 1st week associated with fatal outcome – surrogate marker of viral load
  • Reason: liver damage, muscle damage or both
  1. Respiratory:
  • Dyspnea, tachypnea 41 – 50 % pts
  • Acute lung injury with 52% showing hypoxemia
  • Mechanism: metabolic acidosis (dehydration diarrhea) and pulmonary edema, direct viral pneumonitis (interstitial pulmonary infiltrates and detection of EBOV in bronchial aspirate fluid)
  1. Cardiovascular:
  • Inappropriate bradycardia (dehydration) along with CNS suggested neurological causes
  • Arrhythmias – possibly multifactorial – electrolyte imbalances, but viral myocarditis
  • DIC to due immune mechanisms, hypercoagulated state due to hemoconcentration
  1. Neurological:
  • Common: headache 61%, confusion 13% and coma/unconsciousness 6%
  • 1/3rd patients treated in Europe showed encephalopathy during illness with EBOV isolated from CSF
  • Meningoencephalitis
  1. Co-infection and sepsis:
  • 1182 pts with EVD found that pts with Plasmodium spp were 20% more likely to survive, independent of treatment with antimalarial drugs
  • Mechanism: possibly modulating exuberant cytokine response
  • Sepsis and septic shock was common in EVD

Treatment: Supportive Care

  • Oral and IV fluids, analgesia, antiemetics, antidiarrheal medications, empiric antibiotics and antimalarials
  • Loperamide contraindicated due to paralytic ileus
  • Gap in knowledge: an optimal package of supportive care measures to be used

Survivors

  • Neurological deficits important contributor to morbidity
  • Psychological distress → neurocognitive dysfunction
  • Poor social acceptance, stigmatization
  • Pathological mechanisms: not understood yet – possibly post-infectious inflammatory processes, viral replication in immune privileged sites
  • EBOV isolated from aqueous humor of survivor with panuveitis 14 weeks after diagnosis, total duration of viral sequestration unknown but less than 18 months
  • EBOV detected in CSF in a survivor with meningoencephalitis 9 months after acute illness
  • Follow up study of 151 survivors showed that late redescence (illness or death after full recovery) was estimated at 0.7%
  • EBOV detected in semen 18 months after discharge from ETC; one study reports prevalence in males at 49%

Treatment

  • Table 2 shows clinical trials conducted of therapeutics during the epidemic – none showed conclusive benefit.
  • Phase 2 clinical trial of favipiravir showed no survival benefit with high viral load but may help with lower viral load.
  • RNA lipid nanoparticle TMK130803 showed no benefit in a study in Sierra Leone
  • Ebola-Tx trial of convalescent plasma showed no survival benefit; antibody titers were low in the donor plasma
  • Multicenter, randomized trial of ZMapp lowered CFR from 37% (supportive care) to 22%, did not meet specified statistical threshold, posterior survival probability was 91%
  • Discrepancy: 100% survival seen in NHP not replicated in the clinical trials. Why?

Vaccines

  • 4 met WHO criteria for fast-track assessment (summarized in Table 3):
  • Replication competent rVSV expressing EBOV Zaire glycoprotein – Ebola ca Suffit trial ring vaccination showed 100% CI 75-100 protection
  • Replication-deficient ChAd3-ZEBOV trial in USA and Mali showed single dose protected, but for long term protection, a booster with MVA would be required
  • Ad26-ZEBOV with booster with MVA. Similar to 3 above; trials ongoing
  • Novavax – nanoparticle vaccine – phase I trial still ongoing
  • Cases were waning when the vaccines went to trial and safety and immunogenicity data are scarce

Conclusion:

  1. The west African epidemic saw several studies done rapidly
  2. Vaccines went to trial in record times
  3. Lack of evidence and studies to standardize supportive care
  4. Disappointing results with therapeutics
  5. Ring vaccination may be the strategy for future vaccine trials
  1. Mire CE and Giesbert TM. Neutralizing the Threat: Pan-Ebolavirus Antibodies Close the Loop. Trends in Molecular Medicine. 23*8): 669 – 671. August 2017.
  • WHO released guidance on the use of convalescent whole blood or plasma – immunotherapy.
  • Used in 1976 and 1995 outbreaks
  • Clinical trials initiated in Sierra Leone, Liberia and Guinea. 1 trial showed no benefit
  • NHP studies showed no benefit
  • 2012: purified polyclonal antibodies from sera of NHPs against EBOV GP was used: treatment 2 days post-challenge resulted in 100% survival
  • Also in 2012, 2 labs identified monoclonal antibody combinations: ZMab (2G4, 4G7 and 1H3) and MB-003 (6D8, 13F6 and 13C6) which showed efficacy. These were combined in ZMapp (2G4, 4G7 and 13F6).
  • ZMApp completely protected NHPs as late as 5 days after exposure.
  • Figure 1 shows the antibodies
  • Desire an antiviral or vaccine that is efficacious against minimally EOV, SUDV and BDBV – the mythical ‘rainbow unicorn’.
  • Ebolaviruses diverge by 32 – 41% at nucleotide and amino acid level in GP.
  • GP is cleaved to GPCLwhich then goes through a fusion loop stage for insertion into the host-cell endosomal membrane
  • In figure, CA45 and ADI15878 158742 bind to more conserved regions shared between the Ebolavirus species
  • Has been protective in small animal models: mouse and guinea pigs (EBOV), mouse (SUDV) and ferret (BDBV)
  1. Li T, Yao H-W, Liu, D et al. 2017. Mapping the clinical outcomes and genetic evolution of Ebola virus in Sierra Leone. JCI Insight 2017; 2(14):e88333
  • Li et al analyzed the spatial-temporal and haplotype pattern of 514 EBOV genomes and clinical outcomes in Sierra Leone and identified 11 different lineages with 372 unique haplotypes that had different CFRs. Figure 1 and figure 2. They showed that certain SNPs in noncoding regions that correlated with different CFRs
  1. Hayden EC. Ebola Outbreak Halted. Nature 547, 6 July 2017
  • DRC detected an Ebola outbreak on May 11. It was detected using the GeneXpert Ebola assay
  • Outbreak declared ended on July 2 by WHO – 4 deaths
  • April 2017: 39 year old man had vomiting and bleeding after handling a carcass of a dead wild boar in a forest in Likati in Bas Uele in DRC.
  • He died en route and 2 transporters fell ill and their contacts. 2 of these tested positive for Ebola and ultimately 5 tested positive
  • Genetic analysis showed the strain was similar to the 1995 Kitwit epidemic strain
  1. Sulaiman WAW, Mat, LNI, Hoo FK et al. 2017. Is Ebola a sexually transmitted disease? Implication of Ebola RNA persistence in the semen. Journal of Infection and Public Health.
  • During the west Africa Ebola epidemic, the primary mode of transmission was through direct contact
  • The isolation of EBOV RNA from semen led to revised recommendation by WHO to abstain from sexual contact for 6 months and to be tested at 3 months post acute illness and regularly

EBOV Diagnostics