Diphtheria, Tetanus, Pertussis

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Diphtheria, Tetanus, Pertussis

• MedCh 401
• Lecture 3

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Adjuvant

• Substances that enhance the immune response
• Two categories
• vehicles
• immunomodulators

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Adjuvants functioning as vehicles I

• Human use
• Alum compounds
• Aluminum hydroxide and phosphate
• the only licensed adjuvants in U.S.
• MF59
• Oil and water emulsion
• Marketed in Europe

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Adjuvants functioning as vehicles II

• Animal use
• Freunds Complete Adjuvant (CFA)
• dessicated Mycobacterium butyricum, mineral oil
  and an emulsifying agent, mannide monooleate
• causes potentially secere local inflammatory
  lesions, chronic granulomas, abscesses, and
  tissue sloughs. Injected into the murine
  footpad, it can cause chronic lameness and
  arthritis injected intraperitoneally, it can
  cause peritonitis
• Freunds Incomplete Adjuvant
• Mineral oil and Mannide monooleate
• Fewer side effects, adequate for boosting

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Immunomodulatory Adjuvants

• Purified Protein Derivative (PPD)
• Lipopolysaccharide (LPS bacterial endotoxin)
• Lipid A - lipid portion of LPS
• Cholera toxin B subunit
• CpG

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Diphtheria vaccine

• Detoxified bacterial, protein toxin
• Injectable, IM administration
• Toxigenic Corynebacterium diphtheriae (infected
  with b phage)
• Primarily a childhood disease

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Diphtheria Transmission

• Person-to-person by respiratory droplet
• No animal reservoir

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Manufacturing Process

• Toxigenic strain of C. diphtheriae grown in
  Fenton medium with a bovine extract
• After suitable growth, toxin purified from cells
  by centrifugation
• Toxoided by incubation with formaldehyde for
  several weeks
• Concentrated with ultrafiltration
• Purified by precipitation, dialysis and sterile
  filtered
• Adsorbed onto aluminum hydroxide, Al(OH)3

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Excellent Vaccine Efficacy

• Mortality
• 1860-1897 46-196/100,000 people
• Case fatality rates gt50 during outbreaks
• 1920 15/100,000 people
• Cases
• 1971 - 1981 - 1,288 in U.S.
• 1980 - 1995 - 41
• occasional in developing countries and Native
  populations in U.S. and Canada

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Tetanus Vaccine

• Detoxified bacterial, protein toxin
• Clostridium tetani
• Injectable dosage form
• IM

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Tetanus Transmission

• Not a communicable disease
• The only vaccine-preventable infection that is
  not communicable
• Disease acquired through exposure to bacterial
  spores in the environment
• inoculation of bacterial spores into body by
  puncture or deep cut

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Transmission II

• Neonatal tetanus most common worldwide
• Case fatality increases with age (50 for 80
  years of age)
• Disease reservoirs
• Soil
• Animal feces

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Tetanus toxins

• Tetanolysin - possible role in establishing
  infection at inoculation site
• Tetanospasm
• accumulates intracellularly during log-phase
  growth
• released into medium upon autolysis
• Minimum human lethal dose 2.5 ng/kg

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Tetanus disease

• Tetanospasms
• localized - spasm of muscles close to site of
  injection weeks to months duration rare but may
  precede generalized symptoms
• generalized - 80 of cases
• Complications of the spasms
• fractures of the long bones and vertebrae
• asphyxia from glottic obstruction

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Nervous system effects

• Toxin travels up nerve endings by intra-axonal
  transport
• Gains entry to neuromuscular junctions by binding
  to gangliosides
• Interferes with release of neurotransmitters from
  presynaptic inhibitory fibers
• Excitatory reflexes multiply unchecked, causing
  spasms

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Tetanus

• Toxin-mediated disease
• Symptoms may progress clinically despite use of
  parenteral antibiotics
• Treatment
• hyperimmune serum
• supportive care
• Recovery may depend on development of new
  functional nerve connections

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Manufacturing Process

• Growth of C. tetani in modified Latham broth in
  fermenters
• Harvest extracellular toxin by filtration
• Purify
• Detoxify with formaldehyde for 3 weeks
• Adsorb with Alum adjuvant
• Diafiltration

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Pertussis (Whooping Couth)

• Bordetella pertussis
• Whole cell and acellular vaccines
• traditional and recombinant
• Injectable dosage form
• IM

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Pertussis Transmission

• Person-to-person
• Respiratory droplets

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Pertussis (whooping cough)

• Killed Whole cell -
• old, not licensed in U.S. or Europe
• still used in developing countries
• relatively cheap
• Acellular (aP) -
• currently licensed in U.S., Japan and Europe
• some are recombinant
• expensive

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B. pertussis virulence factors

• Pertussis toxin (PT)
• Filamentous hemagglutinin (FHA)
• Fimbriae (fimbrial agglutinogens) (Fim)
• Pertactin (PRN)
• Adenylate cyclase
• Tracheal cytotoxin
• Dermonecrotic or heat-labile toxin
• BrkA
• Endotoxin (LPS)

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Vaccine Efficacy

• 1914 (Pre-vaccine)
• 270,000 cases, 10,000 deaths annually
• killed gt5 of every 1000 children born in U.S.
• 1976 - 1,010 cases
• 1996 - 7,796
• 2002
• 8,296 cases
• 3.01/100,000 pop.

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Vaccine History

• 1914 First whole-cell vaccine (P)
• 1948 DTP
• 1991 - 2002 DTaP for children
• 2005 Tdap for adults 19-64

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Manufacturing

• B. pertussis cultures grown in modified
  Stainer-Scholte broth
• Acellular antigens
• purified from culture medium (PT and FHA) or
• extracted from cells (PRN, FIM)
• Formaldehyde and/or glutaraldehyde detoxification
  of PT
• Aluminum adjuvants

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Manufacturing II

• Novartis-Behring (Chiron) - Recombinant
• Genetically detoxified PT
• Two other components
• Not available in U.S. yet

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DTaP Vaccine Formulations
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Licensed DTaP Vaccines