1. Ashish A. Deshpande

2. Spongiform = Sponge like
Encephalopathy= Brain Disease
[encephalon + pathy= the technical name of brain +
Pathy = disease]

3. Disease name Natural host Prion name PrP isoform
Non-human mammals
Scrapie Sheep and goats Scrapie prion OvPrPSc
Transmissible mink encephalopathy (TME) Mink TME prion MkPrPSc
Chronic wasting disease (CWD)
Elk, White-tailed
deer, Mule Deer and
Red Deer
CWD prion MDePrPSc
Bovine spongiform encephalopathy (BSE)
commonly known as "Mad Cow Disease"
Cattle BSE prion BovPrPSc
Feline spongiform encephalopathy (FSE) Cats FSE prion FePrPSc
Exotic ungulate encephalopathy (EUE)
Nyala and greater
kudu
EUE prion NyaPrPSc
Human diseases
Kuru
Humans
Kuru prion
HuPrPSc
Creutzfeldt-Jakob disease (CJD) CJD prion
(New) Variant Creutzfeldt-Jakob disease
(vCJD, nvCJD)
vCJD prion
Gerstmann-Sträussler-Scheinker syndrome
(GSS)
GSS prion
Fatal familial insomnia (FFI) FFI prion
Known Spongiform Encephalopathies

4. In general, it is a disease. A group of rare degenerative brain disorders
characterized by tiny holes that give the brain a "spongy" appearance.
Occurs in Humans and Vertebrate animals (Vertebrate = Animals having backbone)
TSE = A disease capable of being transmitted by infection and gives the
appearance of sponge like tiny holes in the brain. There are many types of Human
TSEs.
The literal meaning of TSE is as below
Transmissible = Capable of being transmitted by infection
Spongiform = Sponge like
Encephalopathy= Brain Disease
[encephalon + pathy= the technical name of brain + Pathy = disease]
TSE - Transmissible Spongiform Encephalopathy ?

5. It is a brain disease that occurs in bovines; generally known as “Mad Cow Disease”
BSE = A disease capable of transmitted by infection and gives a appearance of
sponge like tiny holes in the brain of bovines.
The literal meaning of BSE is as below
Bovine = Characteristic of or resembling cows or cattle.
Spongiform = Sponge like
Encephalopathy= Brain Disease
[encephalon + pathy= the technical name of brain + Pathy= disease]
BSE - Bovine Spongiform Encephalopathy ?

7. Prion
• Smaller than smallest known virus
• Not yet completely characterized
• Most widely accepted theory
– Prion = Proteinaceous infectious particle
• Normal Protein
– PrPC (C for cellular)
– Glycoprotein normally found at cell surface inserted in
plasma membrane

8. Normal protein
• Secondary structure dominated by alpha
helices
• Easily soluble
• Easily digested by proteases
• Encoded by PRNP gene (in humans)
– Located on human chromosome 20

9. Abnormal Protein
• PrPSc (Sc for scrapie)
– Same amino acid sequence and
primary structure as normal
protein
– Secondary structure dominated
by beta conformation
• When PrPSc contacts PrPC
– Converts it to the abnormal form

10. Abnormal Protein
• Insoluble in all but strongest solvents
• Highly resistant to digestion by proteases
– Survives in tissues post-mortem
• Extremely resistant
– Heat, normal sterilization processes, sunlight
• No detectable immune response

11. o No antibiotics can cure disease caused by prions
o They are not typical of a prokaryotic organism or a eukaryotic
organism
o All that is present in this pathogen is the protein PrPSc, the
mutation of PrPC
o PrPSc is resistant to any form of digestion
o Prions are non immunogens and do not induce an immune
response
o Prions are not easy to decompose biologically
o They are resistant to high temperatures & disinfectants

13. Center for Food Security and Public Health
Iowa State University - 2004
Kuru
• New Guinea in early 1900’s
– People practicing
cannibalism
– 1957-1968
• Over 1,100 people died
– Majority of deaths
• Women, children and
elderly
– Incubation period >30 days

14. Creutzfeldt-Jakob disease (CJD)
• Sporadic (Scattered) human encephalopathy
• Worldwide 1-2 cases/million people
• Different forms
– Spontaneous (85%)
– Genetic (10-15%)
– Iatrogenic/ consequence of medical treatment
(<1%)
• Average age of onset 65 years
• Duration of illness, 4.5 months

15. Bovine Spongiform Encephalopathy
• 1986, First confirmed case in United Kingdom (UK)
• 1988, UK bans meat and bone meal from ruminants in cattle feed
• 1989, USDA bans importation of ruminants from countries with
BSE
• 1993, Peak of BSE in UK
– 1,000 new cases reported weekly
• 1997, US & Canada ban ruminant products fed back to ruminants
– US importation ban extended to all of Europe
• 2001, European Union ordered mandatory tests on cattle
– Older than 30 months destined for slaughter
• May 2003, BSE diagnosed in 6 yr. old Angus beef cow in Alberta,
Canada
– Herd mates all tested negative
– Single case in 1993 was cow imported from UK

16. • December 2003, BSE diagnosed in 6½ yr old Holstein
cow in Washington State
– Possibly imported from Canada
• DNA testing being conducted
– Complications following calving
– Sent to slaughter
– Brain tissue sent to NVSL–per FSIS protocol
• Presumptive positive by NVSL
– Definitively positive by UK lab

17. vCJD
• Variant Creutzfeldt-Jakob disease (vCJD) is a rare and fatal
human neurodegenerative condition.
• The consumption of food of bovine origin contaminated with
the agent of Bovine Spongiform Encephalopathy (BSE), a
disease of cattle, has been strongly linked to the occurrence
of vCJD in humans.
• 1995, UK: First confirmed case
• Incubation period not known
• Mean age at death- 28 years old
• Mean duration of infection : 1- 4 Months
• 175 cases of vCJD were reported in the UK , and 49 cases in
other countries from October 1996 to March 2011.
• Following the successful containment of the BSE epidemic in
cattle, the number of cases of vCJD in the United Kingdom has
declined since 2000.

18. Characteristic Classic CJD Variant CJD
Median age at death 68 years 28 years
Median duration of illness 4–5 months 13–14 months
Clinical signs and symptoms
Dementia; early
neurologic signs
Prominent psychiatric/behavioral symptoms;
painful dysesthesias; delayed neurologic
signs
Periodic sharp waves on
electroencephalogram
Often present Often absent
Signal hyper intensity in the caudate
nucleus and putamen on diffusion-
weighted and FLAIR MRI
Often present Often absent
Pulvinar sign-bilateral high signal
intensities on axial fluid attenuated
inversion recovery (FLAIR) MRI. Also
posterior thalami involvement on sagittal
T2 sequences
Not reported Present in >75% of cases
Immunohistochemical analysis of brain
tissue
Variable accumulation.
Marked accumulation of protease-resistant
prion protein
Presence of agent in lymphoid tissue Not readily detected Readily detected
Increased glycoform ratio on immunoblot
analysis of protease-resistant prion
protein
Not reported
Marked accumulation of protease-resistant
prion protein
Presence of amyloid plaques in brain
tissue
May be present May be present
Clinical and Pathologic Characteristics:

20. Transmission

21. Human Transmission
• Humans consuming cattle products infected
with BSE can develop vCJD
– Brain and spinal tissue
• Dose required not known
• Genetic susceptibility
– All human cases have been homozygous for
methionine at codon 129 of PrPC
• Possible modes
– Transmission from surgical instruments used on tonsils,
appendix, or brain tissue
– Growth hormone injections
– Vaccines

22. Human Transmission
• Unlikely modes
– Blood transmission
– Consumption of milk and milk products
– Gelatin products (when manufacturing process is done
correctly)

23. Transmission: Animals
• BSE
– Cattle feed
• Sheep carcasses infected with scrapie
• Cattle carcasses infected with unknown TSE
– Maternal transmission occurs at very low levels
• Scrapie
– Spread from ewe to offspring through placenta and
placental fluids
– Genetic susceptibilities
• FSE and exotic zoo animal forms
– Feed contaminated with BSE infected cattle products
• CWD: Environmental, other?
• TME: Not known at this time
• Number of animals infected experimentally
– Hamsters, mice, rats, voles, gerbils, some species of
monkeys

24. Diagnosis: vCJD
• U.K. criteria for antemortem diagnosis
– Neuropsychiatric disorder
with duration
longer than 6 months
– Specific clinical signs
– Abnormal EEG
– Tonsilar biopsy with detectionof prion protein
• Post mortem definitive diagnosis
– Amyloid plaques surrounded by vacuoles
– Prion protein accumulation in cerebellum
– Spongiform appearance in gray matter

25. Treatment: vCJD
• No effective treatment available
– Experimental drugs under investigation
• Quinicrine
• Symptomatic treatment
• Supportive care

27. Incubation Period
• Scrapie: Sheep 2-5 years
• BSE: Cattle 2-8 years
• CWD: Deer and elk 18 months
• TME: Mink 7+ months
• FSE: Feline unknown, most 4-9 years of age

28. There is a possible risk of contamination of infected animal
derived products in the pharmaceutical finished dosage form for
human consumption leads to transmission of TSE/BSE to human
beings.
In some cases, the Pharmaceutical preparations like Finished
Dosage Forms, Active Pharmaceutical Ingredients and their
Starting Materials, and Primary Packaging Materials involves the
use of products/materials derived from animals. For example,
use of proteins, enzymes, amino acids from animals used in the
manufacturing of API and API starting materials.
The primary packaging materials like gelatins capsules derived
from the fat of animals also increases the possibilities of
transmission of TSE/BSE.
Risk in Pharmaceuticals

29. There is a high risk in the case of biotechnological products like
serums, blood products and vaccines where the source material
is derived from animals and animal derived products
There is also a possible risk of TSE/BSE through the
equipments/utilities where in biologically-derived products
and/or products of animal origin is handled. For example, Culture
media used in reactors for media fill studies.
Ideally, the use of such animal derived product/material should
be avoided in the pharmaceutical preparation. However, during
unavoidable circumstances, the use of such animal derived
products is accepted, provided that the manufacturing process
and procedures complies to the applicable regulations set by
WHO, European Commission and USFDA
Risk in Pharmaceuticals

31. U.S. Government Precautions
• 1989: Import restrictions
– Live ruminants and ruminant products
– From countries known to have BSE
• 1997
– Import restrictions expanded
to include all European countries
– FDA “animal feed rule”
• Banned most mammalian proteins as
food source for ruminants

32. U.S. Government Precautions
• Targeted surveillance
– High risk animals
• Adult animals with neurological signs
• Non-ambulatory “downer” cows
• Rabies- negative cattle
• Cattle dying on farms
– 2003: 20,000 animals tested for BSE
• 47 times the number required by the OIE
– 2004: Enhanced surveillance
• Test maximum number of animals possible

33. Specific Guidelines for Hunters Regarding CWD
• Public health officials recommend avoiding human exposure
to CWD agent
• Harvest only animals that look and behave normally
• Do not eat any animal products of sick or infected elk and
deer
• Do not eat brain, spinal cord, eyes, spleen, tonsils and lymph
nodes
• Dress the deer/elk properly
– Minimize handling of brain and spinal tissues
– Wear rubber gloves when field dressing
– Use strong household bleach for cleaning knifes and saws
• Any suspicious elk and deer should be reported to
health officials
• Testing of elk and deer available in many states

34. Prevention and Control
• Surveillance for CJD in US
– CDC
• Blood/plasma donation restrictions
– Persons who have traveled or resided in the U.K. for 3 or
more cumulative months from 1980 to 1996
– FDA Website www.fda.gov/cber/gdlns/cjdvcjd.pdf
• Scrapie Flock Certification Program
– Voluntary
– Producers-industry-states-APHIS
– Certify origin in scrapie-free flock
• National Accelerated Scrapie Eradication Program
– Live animal testing and active slaughter
– Animal tracing/animal identification
– Clean-up strategies including genotyping

36. Regulations
• Annex I to European Parliament and council directives for
Veterinary 2001/82/EC and for Human medicinal products
[2001/83/EC (as amended by commission directive
2003/63/EC)] describes Regulatory compliance through Risk
Assessment, Legal Aspects and General Monographs.
• European Pharmacopeia:
Chapter identical with the Note for Guidance on Minimising
the Risk of Transmitting Animal Spongiform Encephalopathy
Agents via Human and Veterinary Medicinal Products –
Revision 3, (EMA/410/01 rev. 3).” 5.2.8. Minimising the risk of
transmitting animal spongiform encephalopathy agents via
human and veterinary medicinal products

37. Regulations
• European Pharmacopeia:
Has classified a group of animals as “TSE -relevant animal
species” (TSE-RAS).
It has been classified that Cattle, Sheep, Goats and animals
that are naturally susceptible to TSE agents or susceptible to
infection through oral route other than humans and non-
human primates are defined as “TSE relevant animal species”.
Any product/material derived from TSE-RAS like active
substances, excipients, adjuvant, raw and starting materials
and reagents used in the production should meet the
requirements of the Note for Guidance published by EC.
Example: Bovine Serum, Enzymes

38. Regulations
• In unavoidable circumstances, TSE-RAS shall be used in the
production for pharmaceutical preparation but it must be fully
justified by the applicant and necessary requirement of the
note for guidance (EU) should be met.

39. Regulatory Compliance
• The applicant/MA holder shall consider all the scientific
Principles for Minimizing the risk in case use of a TSE-relevant
animal species
• Based on the risk assessment strategies/methods applied by the
applicant and if it is found adequate, regulatory compliance is
certified by EDQM through Certificate of Suitability. However, the
final determination of regulatory compliance rests with the
competent authority. The application must encompass (along
with other requirements) as below
– All TSE-risk factors considered for the assessment study
– Control measures taken into account
– Risk minimized by applicant
– Risk Various material derived from TSE-RAS
– TSE reduction or inactivation techniques /steps employed
– Justification for the selection of the source
– Justification for the selection of tissues/body parts/fluids

40. Regulatory Compliance
In case of use of animal origin material, the following shall be
verified in general
• Type of tissue/body part/fluid used
• Type of animal from which the material is required (Ruminant/non-
ruminant/bovine/ Caprine /Ovine/Porcine)
• Age of animals/health status from which the material is required
• Geographical origin of the animal (Country/Continent)
• A detailed risk assessment and expert certification
• Type of stunning/slaughtering method employed
• Type of certification like ISO, HACCP or GMP of the manufacturer of animal
product manufacturer
• Traceability of animal slaughtering source
• Methods of segregation of tissues during slaughtering
• Potential of cross contamination during slaughtering/Packing/handling
• Name, complete address of the supplier
• Prior reduction claims from the manufacturer of animal derived products
• CEP certification of the pharmaceutical product
• Information regarding the Facility of manufacturing animal derived material
• Information regarding Products derived from elk, deer