Upload
chalverson
View
1.718
Download
0
Embed Size (px)
DESCRIPTION
Citation preview
Global Adventitious Agent Regulation of Raw Materials Used in
Biopharmaceutical ManufacturingBarbara J. Potts, Ph.D. Biologics Consulting Groups, Inc. USA
T.W. Tanaka Biologics Consulting Group, Inc. Japan
1
Bio Process International ConferenceSeptember 20-24th 2010
Rhode Island Convention CenterProvidence, RI
2
USDA 9 CFR 113.53 – Requirements for Ingredients of Animal Origin Used for Production of Biologics and 113.47 – Detection of Extraneous Viruses by the Fluorescent Antibody Technique
ICH Q 5A CBER Guidance – Cell Substrates and Biomaterials USP <1046> Cell and Gene Therapy Products USP <1043> Ancillary Materials for Cell, Gene and Tissue-
Engineered Products
3
Global Regulations that apply to the Control of Adventitious Agents In Raw Materials
European Medicines Agency – Note for Guidance on Minimizing the Risk of Transmitting Animal Spongiform Encephalopathy Agents via Human and Veterinary Medicinal Products (EMEA/410/01 Rev.2, 2004). Rev.4 has been released for consultation 2008.
WHO Recommendations on the Evaluation of Cell Substrates Evaluation. Released for public consultation 2010.
Japanese Pharmacopeia 210 European Pharmacopeia 2.6.7
4
Global Regulations that apply to the Control of Adventitious Agents In Raw Materials
5
Risk Assessment for Adventitious Agent Control Identify and Assess
Agent Likelihood of Contamination
Consequences Impact
Prions Very Low Recall of productsLong term shut down of manufacturingReduced inventory of drugs for patients Reduced public trust
Very high
Viruses Medium Short term shutdown of manufacturing
High
Mycoplasma Low - Medium Rejection of impacted lots Medium – low (depends on frequency)
Bacteria Medium - high Depending on level - may lead to rejection of impacted lots
Medium – low (depends on frequency)
Mold-Yeast Low Depending on level – may lead to rejection of impacted lots
Medium
6
Diagram from Scientific American, July 2004, p.88 in article by Stanley Prusiner
7
Disease Causing Prion (PrPsc)
TSE
Builds up, and if not removedby cell’s machinery, illness can result.
(Transmissible Spongiform Encephalopathy)
Deer, Elk Cows Cats Sheep, Goats Humans
Chronic WastingDisease
BSE Feline SE Scrapie Kuru, Creutzfeldt-Jakob Disease (CJD)vCJD, Fatal Familial Insomnia
8
Viruses
Parasites at the genetic level Virus reproduction is strictly dependent on host cell’s biochemical machinery. This parasitism separates viruses from free – living parasites such as some Mycoplasma Viruses are made up of:
• Genome (RNA or DNA)• Few proteins• Some have a lipid envelope acquired from host cells
A successful viral infection requires a “team” of virions for “infectious unit” to infect a host cell. >Like a touchdown in Football.< Cannot be seen using light microscopy
• Must use transmission electron microscopy to see virions.
9
Major Groups of Viruses
Bacterial Viruses(Bacteriophage)
Host range does notcut across taxonomic boundaries between plants or animals
Animal Viruses
Replicate only in bacteria
Replicate in animal cells
Plant Viruses
Replicate in plant cells
Some viruses may replicate in plants and animals.
10
Mycoplasma
Smallest free – living microorganisms Pleomorphic
• Vary in size from 0.12 to 0.25 µm• Pliability allows Mycoplasma to pass through
small pore filtration devices. Derived from ancestral anaerobic bacteria (clostridia)
by gene deletion. Contains no cell wall hence, they do not take up Gram
stain Assume various shapes from round to filamentous. Can be seen by dark – field and phase – contrast light microscopy. One Mycoplasma can initiate an infection resulting in 109 CFU/mL within 48 – 72 hours
11
Bacteria Prokaryotic
*Absence of a membrane surrounding the nuclear region.*Absence of sub cellular organelles. Much smaller dimensions than most eukaryotic cells, (plants,
animals) which have a membrane bounded nucleus. (1µm compared to 10µm). Thrive in a range of environments that are extreme.*Can use an infinite array of energy sources. Can be seen by using light microscopy Have a cell wall surrounding the cytoplasmic membrane.*Gram Stain of cell wall will identify
> Gram positive bacteria (e.g.. Bacillus)> Gram negative bacteria (e.g.. E.coli) One bacteria can initiate an infection.
12
Fungi / Mold Eukaryotic* Membrane bound nucleus* Presence of sub cellular organelles
Develops branching filaments Includes yeast, filamentous fungi, and fungi that produce lichens and slime molds Are saprobes (feeds on dead and putrefying tissues)
orMutualists (association between two different species of organism in which there is
mutual aid and benefit) orParasites (lives in or on another living organism with no benefit to the host)
Distribution of most fungi is restricted to a host, mutualist or special substrate* However, there are common and widespread “weedy” saprobe species associated
with human activities, buildings and soils
13
Morphological Examination of Viruses, Mycoplasma, Bacteria and Fungi/Mold
Specimen Size MicroscopicRange
Resolution
Fungi / Mold 100 µm
Red Blood Cells 10 µm
Bacteria 1 µm
Mycoplasma 0.120 – 0.25 µm
Viruses 200 nm - 17 nm
Brightfield
Fluorescence
TEM SEM
Reference: Microbiology and Microbial Infections Vol. 2 Systematic Bacteriology Ed. A. Balows, B.I. Duerden. 1998, Oxford University Press Inc. N.Y.
14
Control of Adventitious Agents in Raw Materials
Control sourcing of raw materials of animal origin (Prions)
Testing of bovine serum (fetal, calf) for viruses, Mycoplasma and bacteria
Cleaning and decontamination of raw material process equipment
Filtration Heat treatment pH treatment Gamma irradiation
15
Increasing Resistance to Physical and Chemical Inactivation
Least Resistant Lipid or Medium-Size Viruses, Mycoplasma
Vegetative Bacteria
Fungi
Non-Lipid or Small Viruses
Mycobacteria
Most Resistant Bacterial Spores
Enveloped Viruses,
MycoplasmaVegetative Bacteria
Non-Enveloped
Small Viruses
Fungi
Spore Forming Bacteria Prions
70% Alcohol + + +/− − − −
Phenolic based disinfectants + + +/− − − −
10% Bleach + + + +/− +/− −
2.5% Acidified Bleach and Peracetic acid and Hydrogen Peroxide and Acetic Acid
+ + + + + −
0.1N NaOH + + + + + −
16
Chemical Inactivation
Pore Size Rating (µm)
Bacteria & Fungi Mycoplasma Viruses Prions
0.1 + +/− − −
0.2 + +/− − −
17
Filtration
Enveloped Viruses &
Mycoplasma Vegetative Bacteria
Non Enveloped Small Viruses Fungi
Spore Forming Bacteria Prions
90°C-100°C 5-10 seconds
+ + + + − −
121°C 20 minutes
+ + + + + −
18
Heat Inactivation
Enveloped Viruses Mycoplasma
Non-Enveloped
Small Viruses Prion
pH 2.0, Short Time
+/− + − −
pH 3.0*, 30 Hours
+ + + −
pH 12.5Short Time
+ + + −
19
pH Treatment
* Will not eliminate viral genome detected by PCR (circovirus)
Dose dependant decline in virus survival Varies with different viruses
◦ e.g. 25 kGy will inactivate below detection Bovine Viral Diarrhea Virus (Flavivirus) Infectious Bovine Rhinotracheitis Virus (Herpes Virus) Porcine Parvovirus (Parvovirus)
◦ But it takes 35 kGy to inactivate below detection Canine Adenovirus (Adenovirus) Bovine Reovirus (Reovirus)
BVDV is generally used to validate gamma irradiation of FBS Dose is generally reported as a target of 25 kGy with an accepted range
between15-30 kGy Gamma irradiation of viruses is not a sterilizing procedure Gamma irradiation of Mycoplasma in plant peptones has been validated at
84 kGy Gamma irradiation may not eliminate detectable viral genome by PCR
Gamma Irradiation Inactivation of Viruses and Mycoplasma
20
21
Examples of Rigorous Processes ThatInactivate Prions
• Cleaning process* Sodium hydroxide or chlorine releasing disinfectants (e.g. 20,000ppm. chlorine for 1 hour) Note: Studies have demonstrated infectious TSE bound to the surface of stainless steel• Tallow derivatives processed by* Trans – esterification or hydrolysis at not less than 200°C for 20 minutes under pressure. (glycerol, fatty acids, fatty acids esters production)* Saponification with NaOH 12M (glycerol and soap production)
> batch process at 95°C for 3 hours> continuous process at 140°C under pressure for 8 minutes
• Animal charcoal* Carbonization of bone by using high temperature >800°C• Amino acids from hides or skin* pH 1 to 2 followed by pH of > 11, followed by 140°C for 30 minutes
Reference: Note for guidance on minimizing the risk of transmitting animal spongiform encephalopathy agents via human and veterinary medicinal products EMEA/410/01 Rev. 2, July 1, 2004
22
Viruses That May Contaminate Biopharmaceutical Processes
Virus Family Genome Envelope Size Resistance a
PoxviridaeRhabdoviridaeParamyxoviridae HerpesviridaeOrthomyxoviridaeRetroviridaeBunyaviridae*CoronaviridaeAdenoviridaeReoviridae*ArteriviridaeTogaviridaeFlaviviridaePapovaviridaeCaliciviridae*Picornaviridae*Parvoviridae*Circoviridae*
DNARNARNADNARNARNARNARNADNARNARNARNARNADNARNARNADNADNA
YesYesYesYesYesYesYesYesNoNoYesYesYesNoNoNoNoNo
250 X 450 nm70 X 175 nm150 – 300 nm120 – 200 nm80 – 120 nm80 – 120 nm80 – 100 nm75 – 160 nm65 – 80 nm65 – 75 nm50 – 65 nm40 – 70 nm50 – 70 nm45 – 55 nm35 – 40 nm25 – 30 nm18 – 24 nm17 nm
Medium – HighLowLowLow – MediumLowLowMediumLowMediumHighLowLowLowMedium – High MediumMediumVery HighVery High
a Resistant to physicochemical treatments
23
Viruses That Replicate In CHO CellsVirus Family Virus
(host range)Genome Envelope Size Resistance*
Paramyxoviradae PI 1 (not known) PI 2 (human), PI 3 (human and cattle) Simian virus 5 (monkeys), Mumps virus (humans), Bovine Respiratory Syncytial virus (cattle)
RNA Yes 150-300 nm Low
Orthomyxoviradae Influenza A (human)RNA
Yes80-120 nm Low
Bunyaviridae Cache Valley Fever Virus (mice, mosquitoes, cattle)RNA
Yes80-100 nm Medium
Rhabdoviridae Vesicular Stomatitis Virus (cattle)RNA
Yes70 x 175 nm Low
Reoviridae Reovirus 1,Reovirus 2,Reovirus 3 (human) RNA No 65-75 nm Low
Togaviridae Sindbis (mosquitoes, mice, birds, humans)RNA Yes
40-70 nmHigh
Reoviridae Bluetongue virus(sheep, cattle, bitingmidge) RNA Yes 40 nm Low
Caliciviridae Vesivirus RNA No 35-40 nm Medium
Picornaviridae Encephalomyocarditis virus (mice, chimpanzees pigs, rats, rabbits, guinea pigs, humans)
RNANo 25-30 nm Medium-High
Parvoviridae MVM (mice) DNA No 18-24 nm Very High
* Resistance to physical and chemical inactivation
24
Virus Contamination Virus Family Enveloped Size (nm) SourceCache Valley Fever Virus
Bunyaviridae Yes 80-100 Fetal Bovine Serum
Blue Tongue Virus Reoviridae No 65-75 Fetal Bovine Serum
Blue Tongue Virus Reoviridae Pseudo-enveloped 40 Possible insect transmission in testing lab
Bovine Viral Diarrhea Virus
Flaviviridae Yes 40-70 Fetal Bovine Serum
Vesivirus Caliciviridae No 35-40 UnknownPorcine raw material ?
Equine Picorna Virus Picornaviridae No 25-30 Equine Serum
Minute Virus of Mice
Parvoviridae No 18-24 Non-Animal Raw Material
Circoviridae Circovirus Type I No 17 Porcine Trypsin
25
Source of Virus Contaminations in Raw Materials
Use raw materials that can withstand heat treatment (e.g. 90-100°C – 10 seconds), gamma irradiation (e.g. 15-30 kGy) or low pH (e.g. pH 3.0 for 30 hours)
If using raw materials of animal origin◦Use only animals designated for human consumption◦A quality assurance system must be in place for monitoring
the production process and for batch delineation of animal raw materials
◦Traceability, self-auditing of supplies Procedures in place to audit supplies of raw/starting materials◦Audit for mouse presence at facility, shipping, storage ◦Detect rodent urine with florescent light
26
Minimizing The Risks of Transmission of Viruses
Use no raw materials of animal origin If must use animal raw materials, use porcine or equine sources If bovine raw materials must be used
◦ Source from country with negligible BSE risk (e.g. Australia & New Zealand)◦ Use only animals designated for human consumption◦ Use tissues with no detectable TSE infectivity, e.g.
Reproductive tissues Musculo-skeletal tissues Trachea Skin Adipose tissue
Source from young animals (e.g. < 30 months of age) A Quality Assurance system must be in place for monitoring the production process
and for batch delineation of bovine raw materials Traceability, self-auditing of suppliers Procedures in place to audit suppliers of raw/starting materials
27
Minimizing the Risks of Transmission of TSE
Use raw materials that can be heat treated (60°C, 10 minutes), gamma irradiated or low pH (< pH 4.0) treated
Assume that plant or fish sourced raw materials have a Mycoplasma risk◦ Plants are a common host for Mycoplasma◦ There are many Mycoplasma that infect fish
Do not assume sterilizing filtration for Mycoplasma with 0.1µm or 0.2µm filters
Remember that Mycoplasma are very stable when in dried state (Mycoplasma stocks are stored in lyophilized state)
Remember that some Mycoplasma are not detected in the traditional culture methods
Implement a rapid detection system to limit impact of a Mycoplasma contamination
28
Minimizing the Risks of Transmission of Mycoplasma
Raw Material Mollicutes Genus/Species Detected1 Tryptic Soy Broth
(animal origin)A. laidlawii
2 Tryptic Soy Broth (plant origin)
A. laidlawii
3 Complex Media A. laidlawii4 Bovine Serum A. laidlawii5 Peptones-animal
sourceA. laidlawii
6 Peptone - plant source
A. laidlawii
7 Water A. laidlawii
29
Recent Mycoplasma Contamination of Raw Materials
Parental Drug Association Technical Report #50 “Alternative Methods for Mycoplasma Testing”◦To be released in 2010
Special Section in the Journal Biologicals on Mycoplasma March 2010 – Contains nine publication on risks, testing and control of Mycoplasma Vol.38, Number 2, March 2010
30
New Reports on Mycoplasma Risks, Testing and Control
Raw Material Documented Vendor Activity
Vendor Risk Documented Consumer Activity
Overall Risk
1. Bovine Serum Validated sterile filtration for bacteria, gamma irradiation for some viruses (BVDV)9 CFR testing for virus, Mycoplasma and bacteria, fungi
High-Medium 9 CFR testing with addition of production cells in virus testing, 0.1µm filtration
Medium-low
2. Recombinant Proteins, e.g. insulin
Virus, Mycoplasma testingViral clearance studies Validated sterile filtration for bacteria
Low Sterile filtration Low
3. Bovine, Porcine and Equine peptones , Plant peptones, Fish peptones
Heat treatments, pH, some filtration, bioburden monitoring, endotoxin test
High Validated heat inactivation, gamma irradiation, pH, 0.1 or 0.2µm filtration
Low
31
Risk Assessment: Adventitious Agents in Raw Materials (Bacteria, Fungi, Mycoplasma and Viruses)
Raw Material Documented Vendor Activity
Vendor Risk DocumentedConsumer Activity
Overall Risk
4. Trypsin, Pepsin Validated low pH treatment (pH 3.0 for 30 hours)Porcine virus testing (9 CFR) Post pH treatment Validated sterile filtration
Medium-low Porcine 9 CFR tests with addition of production cells in test
Low
5. Salts, amino acids, sugars, peptones, other raw materials used in large quantities
Rodent and insect control in manufacturing storage and shipping facilities
High if no quality control
Low if there is documented quality control
Routine quality audit of raw material vendors, manufacturing storage and shipping facilities, check for rodent infestation by using fluorescent light
Medium-Low
32
Risk Assessment: Adventitious Agents in Raw Materials (Bacteria, Fungi, Mycoplasma and Viruses)
TSE Transfection reagents made from lamb’s wool must come from live lambs Selection antibiotics are often made from bacteria. Make sure bacteria
process does not include bovine raw materialsViruses Make sure trypsin is low pH treated (pH 3.0/30hrs) to inactivate circovirus
and parvovirus. Inactivated viral genome may remain in the raw material and can be detected by PCR
Fetal bovine serum – may be contaminated with a virus that is not inactivated by gamma irradiation and is not detected in 9CFR virus testing.
Decision Making Have senior management review and approve the use of any animal raw
materials used to prepare Master Cell Banks.
Raw Material Watch Outs for Master Cell Bank Preparation
33