CSL Antivenom Manufacture Jonah Smith Manufacturing Manager Global Issues in Clinical Toxinology Nov

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CSL Antivenom Manufacture Jonah
SmithManufacturing ManagerGlobal Issues in
Clinical ToxinologyNovember 2008
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Agenda

• Why make antivenoms
• What products do we make
• How do we make them
• What are we doing with antivenoms
• What are the key challenges

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Why make antivenoms The Australian context

• Westgate fire to continue burning under bridge
  Metropolitan Fire Brigade hampered by tiger
  snakes

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Why make antivenoms The Australian context
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Why make antivenoms The Australian context

• Long history of service to treatment of venomous
  bites
• Antivenom development
• Pressure immobilisation technique (Sutherland)
• Manufacture of products specific for venomous
  creatures in Australia
• As local manufacturer and sole supplier
  responsibility to ensure ongoing supply
• Products of National Significance
• Antivenoms Q Fever Vaccine human immunoglobulins

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ANTIVENOMS at CSL
First Antivenom 1930
Tiger Snake AV in 1930
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Tick Antivenom 1938

• CSL was the first to make antivenom against
  creatures OTHER than snakes.

Taipan Antivenom 1955
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Brown Antivenom 1955
Death Adder Antivenom1958
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Black Snake Antivenom 1959
Sea Snake Antivenom 1961
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Red Back Spider Antivenom 1961
Stonefish Antivenom 1962
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Box Jellyfish Antivenom 1970
Funnel Web Spider Antivenom 1980
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Why make antivenoms The Australian context

• Agreement with Commonwealth Dept of Health
  Ageing
• Office of Health Protection
• Scope of office - Pandemic flu malaria
  antivenoms etc
• Intent CSL committed to medicines that improve
  and save lives
• Objective Uninterrupted supply of antivenoms
  to community (Australian and abroad)
• Costs Community purchases antivenoms govt
  subsidises cost of manufacture Normal
  commercial imperatives do not apply
• Cost recovery only through product sale and
  govt

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CSL Range of Antivenoms

• Equine (Horse-derived) 250 L starting plasma
• Brown Snake
• Black Snake
• Tiger Snake
• Taipan
• Death Adder
• ANG (Australia New Guinea) Polyvalent Snake
  (includes all of the above)
• Sea Snake
• Redback Spider
• Stonefish

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CSL Range of Antivenoms

• Box Jellyfish (Sheep-derived) 5 L starting
  plasma
• Small Scale Process - static dialysis (7 days)
  centrifugation
• Funnel Web Spider (Rabbit-derived) 1 L starting
  plasma
• Small Scale Process - Chromatography machine
  dialysis
• Funnel Web Spider AV is the only freeze dried AV-
  all others liquid

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CSL Range of Snake Antivenoms

• Terrestrial Snake Antivenoms
• Brown Snake Black Snake Tiger Snake Taipan
  Death Adder
• Polyvalent Snake (includes all of the above)
• Marine Sea Snake (from Enhydrina schistosa)

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Snake Venom

• Snakes milked at approved facilities
• Australian Reptile Park Gosford NSW
• Representation of the 5 prevalent immunotypes in
  Australia
• Tiger (species used Notechis scutatus)
• Brown (Pseudonaja textilis)
• Black (Pseudechis australis- King Brown Snake)
• Death Adder (Acanthophis antaraticus)
• Taipan (Oxyuranus scutellatus)
• Venom freeze-dried and stored at ambient
  temperature in dessicator

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Non-Snake Venoms

• Redback Spider venom (female Laterodectus
  hasselti)
• Supplied by Venom Supplies Pty. Ltd. South
  Australia from wild-captured and raised spiders
• Funnel Web Spider venom (male Atrax robustus)
• Supplied by Australian Reptile Park from
  wild-captured spider collection
• Sea Snake venom (Enhydrina schistosa)
• Expedition-based collection in the past
  (Malaysia)
• Box Jellyfish venom (Chironex fleckeri)
• Private supplier in Cairns previously- new
  supplier required
• Stonefish venom (Synanceia verrucosa)
• Expedition-based collection in the past (Papua
  New Guinea)

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Horse Hyper-immunisation

• Aim stimulate horse immune system -gt high
  concentrations of antibodies
• Prime (sensitise) horses immune system with very
  small doses of the 5 land snake venoms repeated
  at 12 weeks
• First course doses follow consisting of
  increasing venom amounts followed by routine
  dosing
• Test bleeds collected for EIA testing to assess
  horse response
• Sheep (for Box Jellyfish AV) and Rabbit (for
  Funnel Web Spider AV) hyper-immunisation follow
  similar principles without the first course doses

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Plasma Collection

• Horse Plasma sourced from two locations
• CSL Farm Woodend Victoria
• Plasvacc Boonah Queensland
• Sheep and Rabbit Plasma sourced from CSL Farm
• Automated Plasmapheresis Used for Horses and
  Sheep
• Collection cycles generally 5 weeks
• Horse Plasma stored 2-8 C
• Sheep Rabbit plasma stored -60 C

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Large Scale Antivenom Production Overview
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Determination of Venom Strength (LD50)

• Median Lethal Dose (LD50) is the quantity of
  venom that when injected subcutaneously will kill
  50 of Guinea Pigs or mice within 7 days
• Animals are administered serial dilutions of
  venom and observed over 7 days

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Determination of Antivenom Potency (ED50)

• 1 unit of AV will neutralise 0.01mg of dried
  venom
• Venom LD50 value of venom used in calculating the
  ED50.
• Median Effective Dose (ED50)
• Quantity of antivenom (in mL) that will protect
  50 of Guinea Pigs over 7 day period
• Measures the total ability of the antivenom to
  prevent DEATH from all venom components

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Antivenom Fill Determination

• Potency of the bulk antivenom determines the fill
  level with small overages for
• extractable volume
• assurance of the required antivenom units being
  available at product expiry (3 years except Box
  Jellyfish AV 2 years) as determined by
  stability trials
• Fill level will vary depending on potency of the
  bulk antivenom batch to batch

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Typical Fill Volumes for CSL Antivenoms

• Funnel Web Spider AV
• Freeze dried powder plug reconstituted with 10
  mL sterile water

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What are we doing
Horse/Sheep Breed Selection Automated
Plasmapheresis Single Venom Dose Program
Adjuvant Selection / Venom Reduction Immunoglobuli
n Recovery Production Campaign Optimisation
Caprylic Acid Purification
Antivenom Facility Upgrade Equine Box
Jellyfish Process Validation Vial Dispensing on
Rota Peristaltic Filler Antivenom Testing BP
Compliance Funnel Web Process Validation
Dispensing
Govt Agreements AVAT 2007-2011 Freeze Drier
Upgrade Box Jellyfish Process Validation Venom
Supply (Redback Sea Snake Box Jellyfish
Stonefish) Antivenom Filtration Upgrade
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What are we doing
Horse/Sheep Breed Selection Automated
Plasmapheresis Single Venom Dose Program
Adjuvant Selection / Venom Reduction Immunoglobuli
n Recovery Production Campaign Optimisation
Caprylic Acid Purification
Antivenom Facility Upgrade Equine Box
Jellyfish Process Validation Vial Dispensing on
Rota Peristaltic Filler Antivenom Testing BP
Compliance Funnel Web Process Validation
Dispensing
Govt Agreements AVAT 2007-2011 Freeze Drier
Upgrade Box Jellyfish Process Validation Venom
Supply (Redback Sea Snake Box Jellyfish
Stonefish) Antivenom Filtration Upgrade
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What are we doing
Horse/Sheep Breed Selection Automated
Plasmapheresis Single Venom Dose Program
Adjuvant Selection / Venom Reduction Immunoglobuli
n Recovery Production Campaign Optimisation
Caprylic Acid Purification
Antivenom Facility Upgrade Equine Box
Jellyfish Process Validation Vial Dispensing on
Rota Peristaltic Filler Antivenom Testing BP
Compliance Funnel Web Process Validation
Dispensing
Govt Agreements AVAT 2007-2011 Freeze Drier
Upgrade Box Jellyfish Process Validation Venom
Supply (Redback Sea Snake Box Jellyfish
Stonefish) Antivenom Filtration Upgrade
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Viral Clearance Validation/Viral Testing

• Small scale production models spiked with 9
  challenge
• Four Viral Models Used
• Canine distemper virus (CDV)
• large enveloped DNA virus 250-300 nm
• Canine adenovirus II (CAV2)
• large enveloped DNA virus 150-200 nm
• Infectious bovine rhinotracheitis (IBR)
• small non-enveloped DNA virus 60-70 nm
• Poliovirus type 1 (PV1)
• small non-enveloped RNA virus 28-30 nm

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Viral Clearance Validation
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Viral Clearance Validation Summary

• Pepsin treatment effective against enveloped
  virus only
• Heating step effective against enveloped virus
  only
• Filtration steps effective against both virus
  types
• Viral clearance rates higher than the minimum
  required 4 log reduction rate
• Viral clearance may be higher as some steps not
  analysed
• Viruses have not been detected in routine product

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Virus Testing What to do

• Hot topic British Pharmacopoeia WHO TGA
• Where to test
• Animals collected plasma pooled plasma
• What to test for
• Regio-specific virus selection
• Local recommendations
• Virus testing capabilities eg PCR cell culture
• How often
• What cost

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What are we doing
Horse/Sheep Breed Selection Automated
Plasmapheresis Single Venom Dose Program
Adjuvant Selection / Venom Reduction Immunoglobuli
n Recovery Production Campaign Optimisation
Caprylic Acid Purification
Antivenom Facility Upgrade Equine Box
Jellyfish Process Validation Vial Dispensing on
Rota Peristaltic Filler Antivenom Testing BP
Compliance Funnel Web Process Validation
Dispensing
Govt Agreements AVAT 2007-2011 Freeze Drier
Upgrade Box Jellyfish Process Validation Venom
Supply (Redback Sea Snake Box Jellyfish
Stonefish) Antivenom Filtration Upgrade
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What are we doing
Horse/Sheep Breed Selection Automated
Plasmapheresis Single Venom Dose Program
Adjuvant Selection / Venom Reduction Immunoglobuli
n Recovery Production Campaign Optimisation
Caprylic Acid Purification
Antivenom Facility Upgrade Equine Box
Jellyfish Process Validation Vial Dispensing on
Rota Peristaltic Filler Antivenom Testing BP
Compliance Funnel Web Process Validation
Dispensing
Govt Agreements AVAT 2007-2011 Freeze Drier
Upgrade Box Jellyfish Process Validation Venom
Supply (Redback Sea Snake Box Jellyfish
Stonefish) Antivenom Filtration Upgrade
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Adjuvant Selection The F-word

• Antibody titre animal health or both
• The F-word Freunds Adjuvant
• Historically used aluminium gels
• Freunds clear advantages but dirty word
• Educate regulators Animal Ethics committees
  with data clear benefits

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Challenges of Antivenom Manufacture

• On-going supply of GMP-compliant raw materials
  (venom plasma)
• GMP Requirements of Australian Therapeutic Goods
  Administration (TGA)
• Requirements of British Pharmacopoeia how
  products are tested
• Animal Ethics Considerations- reducing animal use
  and improving animal husbandry