Guanarito Buruli Crimean-Congo Dengue TB · Hookworm Vaccine: Human Trials. Human Hookworm Vaccine Initiative (Peter Hotez and David Diemert) Na-ASP-2 Hookworm vaccine. 21.3 kDa recombinant

Tropical Disease Vaccines

HIV Malaria

TB

Schistosomiasis

Leischmaniasis

Ascariasis

Hookworm

Dengue

Trachoma ChagasLeprosy

Buruli

Trypanosomiasis

Yellow Fever

Ebola

Marburg

Lassa

Crimean-Congo

Rift Valley Fever

Junin

Machupo

Hantaan

Guanarito

Ebola

Neglected Tropical Diseases

1in 6have a

disease you have never heard of…


Disease and Poverty linked

NTDs = Poverty Promoting

60% of Global Disability linked to Infectious Diseases

No Market for drugs


13 Poverty Promoting Illnesses

3 Protozoans- Leishmanisis- Trypanosomiasis- Chagas

3 Bacterial-Trachoma- Leprosy- Buruli Ulcer

7 Helminths- Schistosomiasis- Hookworm- Ascariasis

Schistosomiasis

Viral Hemorrhagic FeversNBC News

Background: Viral Hemorrhagic Fevers

FeverMyalgiaHeadacheProstrationHemorrhageCapillary leak

Background: Viral Hemorrhagic Fevers

Filoviridae: Flaviviridae

Ebola HF – Ebola Virus Mosquito-borneMarburg HF – Marburg Virus - Yellow Fever

- Dengue HF – Den 1,2,3,4Tick-borne- Kyasanur Forest Disease- Omsk HFBunyaviridae

- - Hataan- Puumala- Sin Nombre (4 Corners)

Distribution: Yellow Fever Virus

Distinctly lacking in Asia

Flavivirus: Located in Africa and the Americas

Origins: W. Africa Americas

Vector: Aedes aegypti spp.

Transmission: Eggs dessicate and travel across borders

Types: Sylvan (jungle) and Urban (outbreaks)

WHO data

10% of Philadelphia perished

Background: Yellow Fever Virus

1878: New Orleans - 20,000 deaths

“The American Plague”

Background: Yellow Fever Virus

1898: Spanish American War- 989 War related fatalities

- > 5000 deaths to disease

- Walter Reed put in charge of Yellow Fever Commission

Walter Reed

Disease: Yellow Fever Virus

Classic Yellow Fever – 3 periods

Infection: Fever, headache, Faget’s Sign, black emesis (Day 3), nausea, bleeding

Remission: Symptom resolution - Hours to days

Intoxication: Fever, epigastric tenderness, bleeding, hypotension, death

Councilman body in liver

Treatment and Prevention: Yellow Fever Virus

No specific treatment exists. Mortality: 5-50% depending on severity of illness. 200,000 cases with 30,000 deaths.

Vaccine- Live attenuated1951 Nobel Prize - Max Thieler

www.rayur.com

RNA enveloped virus

Vaccine: Yellow Fever Virus

Yellow Fever Vaccine – Based on 17D strain and administered since the 1930’s

Safety Profile – 600 million doses administered: 12 post-vaccination VF cases identified

Contraindications – Egg allergy, < 6 mo or > 59 years, thymus disorders, primary immunodeficiences or on immunodeficiency drugs.

Yellow Book – Rare cases of Neurotropic or Viscerotropic disease in travelers (8 died – 1.2-2.2 per 100,000)

2013 – WHO concluded that lifelong immunity results after 1 dose

Background: Dengue Virus

Most important arboviral infection: 4 distinct dengue virus serotypes (DENV-1-4)

Antigenically distinct with no cross-reactivity (single-stranded RNA virus)

Transmitted: Aedes aegyptii

DENV-2 is most distinct with little homology 1, 3, 4

Pathology: Disseminates to RE system and into blood. Endothelial swelling and perivascular edema, extravasation of blood

Incubation: 3-14 days

390 million cases/year(95% CI 284-588)

500,000 severe cases/year

1970 – 9 countries

2015- 110 countries

Disease: Dengue Virus Symptoms

Rash: 1. Transient flush, mottling, fleeting pinpoint eruption

2. Maculopapular/scarlatiniform (d 3-4), pruritic

3. Convalescent rash: Confluent petechial rash with scattered pale rounded areas of normal skin

20 petichiae/inch2 after tourniquet deflated (5 min)

Disease: Dengue Hemorrhagic Fever

Dengue Shock Syndrome (WHO)

- Fever or history of fever 2-7 days- Any hemorrhagic manifestation - Thrombocytopenia (platelet count of

<100,000/mm3) - Increased vascular permeability

Can develop Dengue to each strain (up to 4 episodes)

DHF risk increases if you have “seen” a previous strain (ADE)

Dengue Virus: Antibody-dependent Enhanced Immunity

Initial Infection

Invade Cell and replicate

B cell (ab) and T cell immunity develops

After virion release, ab coat and MΦ engulf

Alternate DenV Infection

Memory B cells produce ab

Poor binding to alternate DENV strain

Escape MΦ and replicate

Vaccine Development: Tetravalent Dengue Virus

Live attenuated

Inactivated

Sub-Unit

Viral-like particles and DNA

Most advanced in testing – Phase 3Yellow Fever and JE models workSanofi Pasteur**, NIAID, WRAIR

More expensive to manufactureNo risk of transmission to mosquitoesCan use in immunocompromised WRAIR/GSK – TDEN-PIV

Phase 1 testing

Pre-clinical and Phase 1

Vaccine Development: Dengue Virus

Sanofi-Pasteur (CYD-TDV) tetravalent chimeric vaccine

Efficacy –

Dengue ~60%

Phase III VE Asia S. America1- 50% 50.3%2- 35% 42.3%3- 78.4% 74.0%4- 75.3% 77.7%

http://cvi.asm.org/content/16/12/1709/F3.large.jpgCapeding, Lancet, 2014, 384: 1358

http://cvi.asm.org/content/16/12/1709/F3.large.jpg

Vaccine Development: Sanofi Chimeric Vaccine

Phase 3 - > 35,000 Children aged 2-16 years, 10 countries

Year 3 – Children under 5 had 5X risk of dengue hospitalization (0.99 vaccinated vs 0.2% controls = 20 kids (5 with DHF) vs. 2 kids)

By Year 3 – VE in children < 9 yr was 44.6% (95% CI 31.6%-55.0) whereas >9 yr was 65.6% (95% CI 60.7-69.9)

Children seronegative at enrollment had strong nAb responses but poor protection compared to seropositive children

Strong evidence of antibody-dependent enhancement Year 3

Ebola

Background: The ebola virus

• Enveloped, host cell membrane with Ebola glycoprotein• Non-segmented, negative sense, ssRNA• 7 genes, 19kb• Family: Filoviridae (filum, Latin, thread)• Genus: Ebolavirus• Species:

– Zaire (EBOV)– Sudan (SUDV)– Ivory Coast or Tai Forest (TAFV)– Bundibugyo (BDBV)– Reston (RESTV)

• BSL-4, Bio-threat category ACourtesy – James Campbell, MD

Background: Virus Presentation

1967: Marburg, Germany- Imported Ugandan Green Monkeys

- Laboratory workers fall ill

- 7 of 31 die.

- Virus later identified and named Marburg.

- Predilection for children – 75% < 5 years of age

- High case mortality – 89%

- Egyptian Fruit Bat – Rousettus aegyptiacus


1976: Yambuku, Congo-“Green Monkey Fever”

-Villagers fall ill – yellow fever?

-Women >> Men

-Ground Zero - Yambuku Mission Hospital – 200 dead

Mission Nuns and a Priest Self Quarantined – 4 colleagues had died

Please stop, anybody who crosses here may die….


- Two vials of blood sent to Belgium – 1 broke

- Peter Piot examined

- WHO ordered vials sent to CDC

- Electron microscopy image of virus

- CDC confirmed it was not Marburg

- New virus – Named it Ebola


- International team arrived to help quell the outbreak and study the virus

- Yambuku Mission – 5 syringes

- Pre-natal care and deliveries common

- Also associated with funerals

- Outbreak lasted 3 months and ~300 dead (88% mortality)

- Simultaneous outbreak –Sudan 1976

- Disappeared until 1994

The Ebola River

Epidemiology: Transmission

- Bushmeat is a primary protein source in West/Central Africa

- Accounts for ½ of meat at market –20% are primates

- Humans exposed during slaughter

Epidemiology: Transmission

Downstream Effects- 95% mortality in Great

Apes, 77% Chimps

- 1/3 Great Apes Dead

- Desperately require a vaccine as well

Epidemiology: Reservoir

- Primates are not a reservoir

- Bats long suspected

- Marburg – R. aegyptiacus

- Ebola - Little collared and hammerhead fruit bats

- Bats are a delicacy in W. Africa

Marburg and Ebola Vaccine:

GP = Glycoprotein – basis of vaccine

Viral Membrane – May contain HLA or other human surface receptors

VP40 and VP24: Ebola matrix proteins critical for budding

Ebola RNA: Packed with NP protein for nucleocapsid

Ebola Polymerase: Synthesizes positive sense virus RNA

VP35 and VP30: IFN antagonist ebola proteins

Lipid membrane wraps viral particle

Pathogenesis: Virulent Factors

Marburg and Ebola Vaccine:

Viral Pathology:Single-strand RNA virus. Evades type I IFN’s by VP35 blockade of induction and VP24 blockade of IFN action.

Circulating infected monocytes express tissue factor leading to DIC

Endothelial and parenchymal cell infection, likely mediated by glycoprotein results in further tissue damage.

Pathogenesis: Virulent Factors

Outbreak: Ebola Epidemic

December 6, 2013: Guéckédou, Guinea- Child Zero dies – linked to fruit bat

-Confirmed by WHO- March 2014

-Previous to this – only W. African Ebola outbreak seen was in Cote D’Ivoire

-Lassa Fever suspected

Guéckédou

NEJM 10.9.2014

Outbreak: Ebola Epidemic

Ebola deathsFigures up to 26 April 2015

10,899Deaths - probable, confirmed and suspected(Includes one in the US and six in Mali)

4,608 Liberia

3,899 Sierra Leone

3,584 Guinea

8 Nigeria Source: WHO Getty Image

Treatment: Ebola Vaccines

All depend upon a viral platform and ebola genes - 3

C. Ad26-EBOV/MVA EBOV– J&J, Janssen, and Bavarian Nordic - Adenovirus 26-EBOV prime boosted by MVA

NEJM Oct 2014

VRC, UMB, Emory, Oxford UK, Mali

Canada, US, Gabon, Switzerland, Germany




NEJM Oct 2014






NEJM Oct 2014




Lancet ID 2016,16(1):31-42


Treatment: Phase 2 Ebola Vaccines

Sierra Leone Guinea

Design- Head to head

GSK vs. Merck vs. None

9000 volunteers

Design- Wedge

HCW primarily

6000 volunteers

Design- Ring Design

Vacc ~50 surrounding contacts of each case

9000 volunteers in two arms (early vs. late)

Liberia

Treatment: Phase 2 Ebola Vaccines

Guinea

Design- Ring Design

Vacc ~50 surrounding contacts of each case

9000 volunteers in two arms (early vs. late)

Lancet 2015, 386:857

Helminth Vaccines

Blood flukes:Schistosoma mansoni, hematobium, etc.

Liver flukes:Fasciola hepaticus, Clonorchis, Opisthorchis, etc.

Intestinal flukes:Fasciolopsis, Echinostoma , etc.

Lung flukes:Paragonimus

Enterobius vermicularis

Ascaris lumbricoides

Hookworm spp.

Trichuris trichiura

Strongyloides stercoralis

Taenia spp. Echinococcus granulosis

Diphyllobothrium latum

Hymenolepsis diminuta

Hymenolepsis nana

2 billion people infected

300 million suffer severe morbidity

400 million school-aged children infected

40% of all infectious disease burden (minus malaria) derives from STH

The BIG Three: Hookworm, Roundworm, Whipworm

Soil Transmitted Helminths: Epidemiology

Rapid Impact Package: Praziquantel, albendazole, ivermectin and azithromycin = US$ 0.40. 7 of NTDs cured and 7 of 8 Millenium Development Goals addressed

No commercially available human helminth vaccines exist

One veterinary vaccine exists: Huskvac (oral lungworm vaccine for calves) produced from radiation-attenuated L3 Dictyocaulus viviparus

Soil Transmitted Helminths: Epidemiology

STH associated DALYs

Soil Transmitted Helminths: Hookworm

Ancyclostoma duodenale(0.15 mL per worm per day)

Necator americanus(0.03 mL per worm per day)

Adult hookworms: 1 cm in length40-160 hookworms sufficient to cause anemia/malnutrition. Lifespan: ~5 (Ad) and ~1 years (Na)Children sustain physical and intellectual impairmentTreatment: Albendazole or mebendazoleReinfection occurs within 4 months

Soil Transmitted Helminths: Hookworm

740 million infectedHeavy infections (>5000 eggs/gm stool) associated w/ 9 mL loss/day73% of worldwide severe anemia assoc. with hookworm

Hookworm Vaccine: Rationale

STH prevalence (a) globally and; (b) in China

Antihelminthic drugs failing

No immunity garnered. Adults have high hookworm prevalence.

Very high reinfection rates documented.

Emerging albendazole resistance – single mutation in tubulin allele

Albendazole therapy failure - Mali and China

Hookworm Vaccine: Feasibility

Stage Development of HookwormEnvironmental Parasitic

Egg L2 L3 (infective) L4 L5 (adult) Egg

Feasibility of Vaccine Approach 1 (Larval L3 proteins):Live L3 inocula elicit immunity (Johns Hopkins – 1930s-40s)

Attenuated (irradiated) L3 inocula in dogs cause 60-80% reduction of infection upon challenge – led to a licensed vet vaccine in 1973

Anti-ASP-2 IgE antibodies associated with reduced risk of heavy hookworm burden in Brazil and China (Bethony, 2005, FASEB J, 19:1743)

L3 proteins (cloned and expressed)45 kDa Ancyclostoma secreted protein 1 (ASP-1)23 kDa Ancyclostoma secreted protein 2 (ASP-2)62 kDa Ancyclostoma secreted metalloprotease (MTP)

Hookworm Vaccine: Human Trials

Human Hookworm Vaccine Initiative (Peter Hotez and David Diemert)

Na-ASP-2 Hookworm vaccine

21.3 kDa recombinant antigen cloned from N. americanus larvae

Expressed in yeast (Pichia pastoris) adsorbed to Alhydrogel®

Phase I study in healthy human volunteers – Immunogenic and well tolerated IM dosing on Days 0, 56, and 112. (D.Diemert, Sabin Institute)

Phase I study in healthy Brazilan adults conducted – reactogenicity noted

(Immediate-type hypersensitivity - Generalized urticaria within 1-2 hours)

All larval-stage antigen-based vaccines halted

Hookworm Vaccine: Adult worm secreted proteins

Na-GST-1

Aim - Want high IgG but

LOW IgE

Hotez et al., Nat Reviews 2010

IgE levels

Hookworms express proteases (cysteine, aspartic, and metallo-proteases). Localize to brush border of worm intestine. Digest Hb and serum proteins.

Rationale: Neutralize proteases involved in parasitic feeding and “starve” the worm.

Na- glutathione S-transferase-1 (GST-1)

Critical enzyme that enables parasitic feeding

Phase I testing in humans

Infect Immun, 2010

Hookworm Vaccine: Adult worm secreted proteins

Schistosomiasis (Bilharzia)

Water-borne blood fluke: S. mansoni, S. mekongi, S. haematobium, S. japonicum, S. intercalatum

200 million infected worldwide in 77 countries

20 million have severe disease

Each species has a predilection for a different venous plexus

1.

5 minutes after skin penetration, cercariae penetrates epidermis

2.

10 minutes after penetration, the cercariae has penetrated basementmembrane

3.

20 minutes after penetration, the cercariae has penetrated dermis

Mating pairs are egg production factories, generally living 5-8 years but potentially surviving for 30 years. End-stage sequelae of schistosomiasis occurs over decades of infection with large socio-economic tolls but still accounts for only a fraction of those w/ dz.

Male

Female

Chronic Schistosomiasis

S. mansoni/intercalatum: Inferior mesenteric venous plexus

Hepatosplenomegaly, granuloma/fibrosis, pancytopenia, GI ulcers, periportal fibrosis (4-8%) and portal hypertension. Rare cor pulmonale and neuroschistosomiasis

S. haematobium: Vesicular venous plexus. GU pathology, bladder calcifications and cancer.

Schistosomiasis Vaccine: Rationale

1. Vaccines provide cost-effective control for infectious diseases

2. High level protection is noted with irradiated cercariae injection

3. Chemotherapy has proven ineffective due to rapid reinfection

4. Drug resistance has formed and infrastructures in the developing world have proved inadequate for repetitive drug therapy

Vaccine Development:First attempts began 50 yrs ago: Crude worm extracts

Attenuated cercariae vaccination (1960s): >90% mice, 86% primates Theory: a proportion of larvae enter skin-draining lymph nodes inducing intense immune response.

Science – April 26, 2013

Sh Phase I vaccine-28 kDa GST-1

Completed 1989 –published 2012 (Phase II and III nearly complete)

Other candidates -Sm-14 used for fatty acid synthesis (Brazil – good abs but results not replicated)

Sm-TSP-2 (US – Sabin Institute)

Schistosomiasis Vaccine: Tetraspanins

Proteins prominent in outer tegument of schistosomes – function unknown

TSP-1 and TSP-2 (antibodies to TSP-2 are very high in putatively resistant and non-existent in chronically infected individuals) (Tran, Nat Med, 2006, 12:835)

Mice vaccinated with TSPs had reduced worm and egg burden:

TSP-1 - 34% adult worms53% egg burden

TSP-2 - 57% adult worms 64% egg burden

(Tran, Nat Med, 2006, 12:835)

Further helminth vaccine efforts are underway……….

Trichuris trichiura (whipworm)

Trichinella spiralis (trichinosis)

Onchocerca volvulus (river blindness)

Echinoccocus granulosus** (hydatid disease)

Cysticercosis** (Taenia saginata, T. ovis, T. solium) – application primarily directed as veterinary vaccines using DNA technology

Funding is a major obstacle to vaccine researchLarge philanthropies have not yet funded vaccine-related efforts.

Public-private partnerships (PPPs) are essential for accelerating NTD research towards vaccine development.

Acknowledgements:

SANARIAMALARIA ERADICATION THROUGH VACCINATION

Documents

Guanarito Buruli Crimean-Congo Dengue TB · Hookworm Vaccine: Human Trials. Human Hookworm Vaccine Initiative (Peter Hotez and David Diemert) Na-ASP-2 Hookworm vaccine. 21.3 kDa recombinant