ETHICAL AND SAFETY ISSUES IN GENE TRANSFER

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ETHICAL AND SAFETY ISSUES IN GENE TRANSFER

• David Jay Weber, M.D., M.P.H.
• Professor of Medicine, Pediatrics Epidemiology
• Epidemiologist, GCRC
• University of North Carolina at Chapel Hill

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ACKOWLEDEGMENTS

• Special thanks to the following for kindly
  sharing their slides
• John Wiley and Sons
• Dr. Martin Evans, Professor University of
  Kentucky Medical Center

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INFECTION CONTROL

• Charge
• Prevent patient-to-patient, patient-to-staff,
  staff-to-patient transmission laboratory worker
  acquisition
• Biologically active agents
• BCG Used for bladder fulguration risk
  mycobacteriosis
• Non-sterile pharmaceuticals
• Blood (gt20 agents transmitted by needlestick),
  organs/tissue, leeches, other (e.g., human growth
  home risk CJD)
• Sterile pharmaceuticals
• Most drugs risk intrinsic or extrinsic
  contamination

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INFECTION CONTROL

• Surveillance (Comprehensive)
• Outbreak investigations
• Policies/procedures (gt70 policies)
• Education (Every employee, every year)
• Provision of personal protective equipment
• Infection control rounds (assure compliance)
• Monitoring Environment, equipment
• Occupational health

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PERSON-TO-PERSON ACQUISITION
Weber D, Rutala W. CID 200132446.
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LECTURE TOPICS

• Basics of gene transfer (gene therapy)
• Scope of gene therapy trials
• Ethical and safety issues
• Infection control and laboratory safety issues

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HUMAN EXPERIMENTATION CHALLENGES FOR THE 21ST
CENTURY

• Current
• Gene transfer
• Near future
• Xenotransplantation, biological and chemical
  agents of terrorism, cloning (reproductive,
  therapeutic), genetic screening, in utero gene
  transfer, stem cell transplantation
• More distant future
• Nanotechnology, life prolonging therapy (drugs
  altering program cell death)

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GENE TRANSFER COMPONENTS

• Gene transfer products
• Gene(s) encoding the therapeutic protein
• Vector that delivers gene into the cells
• Vectors
• Viral (e.g., retroviruses, adenoviruses)
• Non-viral (e.g., naked DNA)
• Cellular (e.g., allogenic, xenogenic, or
  autologous cells)

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STRATEGY OF GENE TRANSFER
Virus
VG 1
VG 2
VG 3
TG
Insert Transgene
Infect Host
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EX VIVO GENE TRANSFER
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IN VIVO GENE TRANSFER
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STRATEGIES FOR USE OF GENE THERAPY TRANSFER

• Supplementation
• Transfer a functional gene into cells that have a
  defective gene
• Example Transfer gene that produces factor IX
• Immunotherapy
• Deliver a gene that will elicit an immune
  response when the gene product is expressed
• Example Infect with vaccinia containing
  prostate-specific antigen gene

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STRATEGIES FOR USE OF GENE THERAPY TRANSFER

• Cancer therapy
• Deliver a therapeutic gene into cancer cell
• Example Infect cancer cells with adenovirus
  containing the gene for tumor necrosis factor
• Chemoprotection
• Transfer of a gene for drug resistance into
  normal cells to protect them from chemotherapy
• Example Transfer a multi-drug resistance gene
  into normal bone marrow cells transplant the
  cells and administer chemotherapy to kill
  unprotected tumor cells

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STRATEGIES FOR USE OF GENE THERAPY TRANSFER

• Ablative therapy
• Deliver a gene that will allow activation of a
  prodrug leading to cell death
• Example Insert the herpes simplex virus
  thymidine kinase gene into tumor cells and
  administer ganciclovir
• Antiviral therapy
• Deliver a gene into infected cells that
  interferes with viral replication
• Transfer a gene for hairpin ribozyme, which
  cleaves HIV-1 RNA in HIV-infected T cells

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STRATEGIES FOR USE OF GENE THERAPY TRANSFER

• Marking
• Insert a gene into cells to identify them when
  the gene is expressed
• Example Infect harvested bone marrow cells with
  a retrovirus containing neomycin
  phosphotransferase gene after transplantation,
  look for cells producing the enzyme as evidence
  of engraftment

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SCOPE OF GENE TRANSFER TRIALS
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HUMAN DISEASES TARGETED FOR GENE TRANSFER

• Monogenic diseases
• Adenosine deaminase severe immunodeficiency
• X-linked severe combined immunodeficiency
• Cystic fibrosis
• Hemophilia B
• Chronic granulomatous disease
• Thalassemia
• Sickle-cell anemia

• Cancers
• Head and neck
• Glioblastoma
• Lung
• Breast
• Liver
• Colon
• Prostate
• Cervix, ovary
• Hematologic

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HUMAN DISEASES TARGETED FOR GENE TRANSFER

• Infectious diseases
• Hepatitis B infection
• Influenza infection
• HIV
• Herpes virus infection
• CMV

• Other
• Coronary artery disease
• Peripheral artery disease
• Amyotrophic lateral sclerosis
• Rheumatoid arthritis

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CLINICAL TRIALSPROTOCOLS
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CLINICAL TRIALSPATIENTS
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ETHICAL AND SAFETY ISSUES
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OVERSIGHT OF GENE TRANSFER
FDA CBER
NIH/OBA RAC
Gene Transfer Safety Assessment Board
Principal Investigator
Institutional Biosafety Committee (IBC)
Institutional Review Board (IRB)
Infection Control Committee
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SAFETY ISSUES PATIENT

• Administration risks (e.g., bleeding,
  anaphylaxis)
• Risks due to vector
• Infection due to replication competent virus
• Inflammatory response
• Cancer (e.g., activation of oncogene)
• Injury to healthy cells or tissue

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SAFETY ISSUES INFECTION CONTROL

• Vector production Laboratory workers
• Vector transport Transportation personnel
• Vector administration Healthcare workers
• Post-administration care Healthcare workers,
  contacts

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ADVERSE EVENTS

• Death Pennsylvania, 1999
• Disease Partial ornithine transcarbamylase
  (OTC) defiency
• Benefit None demonstrated
• AER Death
• Mechanism Inflammatory reaction to adenovirus
  vector injected into hepatic artery

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ADVERSE EVENTS

• Cancer France, 2002
• Disease Severe combined immunodeficiency
  disorder (SCID)-XI (caused by mutations in
  X-linked gene IL2RG that encodes ?chain of the
  lymphocyte receptors for IL-2
• Benefit Immune recovery in gt90 of treated
  infants
• AE 2 children have developed T-cell leukemia
  (gt1 year after gene therapy)
• Mechanism Insertion of vector in chromosome in
  or near gene LM02 (insertion mutagenesis)

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ASSESSING RISK

• Principle Development can proceed using drug
  and vaccine approach
• History Extrapolation of no observed effect
  level (NOEL) and maximally tolerated dose (MTD)
  has been based on animal experiments
• Issue For gene therapy with human pathogens
  (e.g., adenovirus) animal data may not be
  relevant

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ASSESSING RISK

• Principle Level of toxicity is proportional
  to the gene transfer or that toxicity and
  administration dosage are at least in a constant
  relation
• In pharmacology, this principle is generally
  valid and is limited only by some mainly
  genetically based individual pre-dispositions
  that are statistically of low frequency
• In gene therapy the above may not be true. For
  example, 5 of the population are not immunized
  to Ad type 5. Such people may have an extremely
  divergent immunological pre-disposition.

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LOCAL GENE TRANSFER REVIEW COMMITTEES

• Standard IRB membership
• Community advocate
• Epidemiologists, statisticians
• Physicians, scientists
• Expert(s) in gene therapy
• Infection control, laboratory safety
• Vector expert (high level of technical expertise
  required)
• Animal model expert
• Disease specialist (Limited choices if disease is
  rare)

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KEY ISSUES TO REVIEW

• Benefits to the patient (if any)
• Risks to the patient
• Benefits to society
• Risks to society
• Informed consent
• Benefits not over emphasized
• Risks not minimized

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KEY ISSUES TO REVIEW

• Vector production
• Specific vector
• Replication competence (i.e., genes deleted)
• Virulence
• Susceptible cells
• Site of genome integration
• Adventitial material
• Gene(s) inserted
• Laboratory safety

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KEY ISSUES TO REVIEW

• Method of delivery
• In vivo versus ex vivo
• Source of target cells
• Route (e.g., inhalation, injection, etc.)
• Number of administrations
• Precautions to prevent exposure to healthcare
  workers

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KEY ISSUES TO REVIEW

• Study design
• Trial phase
• Clear hypothesis
• Patient selection (minimize risks, therapy
  warranted)
• Adequate sample size
• Appropriate outcome measures
• Appropriate safety monitoring
• Adequate duration of monitoring

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KEY ISSUES TO REVIEW

• Previous studies
• Animal studies
• Appropriate animals
• Appropriate doses
• Sufficient numbers
• Safety analysis (e.g., pathology of organs)
• Adequate analysis of adverse events
• Human studies
• Appropriate doses
• Sufficient number
• Adequate analysis of adverse events

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INFECTION CONTROL

• MANAGING RISKS TO HEALTHCARE WORKERS, LABORATORY
  PERSONNEL, AND PATIENT CONTACTS

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GENE TRANSFER HOSPITAL CARE

• Isolation precautions
• Standard
• Airborne?, droplet?, contact?
• Visitor guidelines
• Travel outside research/hospital room
• Disinfection Surfaces, equipment
• Restrictions on HCWs

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GENE TRANSFER HOSPITAL CARE

• Laboratory testing risks
• Percutaneous injury (bloodborne vector)
• Aerosolization (body secretions containing
  vector)
• Monitoring
• Research subject/patient
• Medical staff
• Environment
• Household contact (visitor)

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INFECTION CONTROL ISSUES

• Replication competency of vector
• Method of vector administration
• Ability of vector to integrate into host genome
• Ability of vector to establish latent or long
  lived infection
• Mode of transmission (i.e., contact, droplet,
  airborne)
• Infectivity (communicability)
• Environmental survival of vector
• Susceptibility to disinfectants

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LIMITATIONS OF RECOMMENDATIONS

• Limited data published on which to base
  recommendations
• Data often considered proprietary
• Focus on efficacy not potential infection control
  problems
• Limitations of data obtained from current trials
• Small number of patients in current trials
• Highly selected patients
• Special handling of vectors
• Few phase III trials
• Major problem difficulty of assessing rare
  events

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BASIS OF INFECTION CONTROL RECOMMENDATIONS

• Infection control recommendations based on
  microbiology and epidemiology of vector used in
  gene therapy protocol
• Recommendations altered after scientific studies
  provide data to liberalize or alter
  recommendations

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GENERAL RECOMMENDATIONS

• Laboratory
• Manage vector as per biosafety guidelines (NIH)
• Research personnel should use appropriate
  personnel protective equipment (PPE)
• Dispose of vectors and contaminated material as
  infectious waste
• Decontamination using EPA registered disinfectant
• Pharmacy
• As per laboratory

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GENERAL RECOMMENDATIONS

• Patient management
• Isolation precautions based on vector, mode of
  transmission, and risk of transmission
• Manage all patients using Standard Precautions
  (PPE to prevent contact with all secretions and
  excretions except sweat)
• Healthcare workers should use appropriate PPE
• Avoid sharps injuries

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CDC ISOLATION PRECAUTIONS
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GENERAL RECOMMENDATIONS

• Healthcare workers
• Appropriate training in use of PPE
• May require screening and restriction
• Example Do not allow HCW with possible
  adenovirus infection to work with patient
  receiving aerosolized adenovirus vector
• Protocol for dealing with sharp injuries

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SPECIFIC RECOMMENDATIONS
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NIH BIOHAZARDOUS AGENTS AND GENE TRANSFER VECTORS

• Risk group 1 Not associated with human disease
• Adeno-associated virus, murine leukemia virus
• Risk group 2 Associated with human disease which
  is rarely serious and for which therapeutic or
  preventative interventions are often available
• Adenovirus, poxviruses, herpes viruses,
  laboratory attenuated strains of vesicular
  stomatitis virus

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NIH BIOHAZARDOUS AGENTS AND GENE TRANSFER VECTORS

• Risk group 3 Associated with serious or lethal
  human disease for which therapeutic or
  preventative interventions may be available (high
  individual but low community risk)
• Retroviruses (HTLV, HIV, SIV), Semliki forest
  virus
• Risk group 4 Likely to cause serious or lethal
  human disease for which therapeutic or
  preventative interventions are not usually
  available (high individual and high community
  risk)
• Ebola

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PERSONAL PROTECTIVE EQUIPMENT

• Use gloves that protect against viruses
• Use handwashing agents effective against viruses
• Alcohol not effective against non-enveloped
  viruses
• Prevent mucous membrane exposure with masks and
  when necessary eye protection
• Minimize risks of sharp injuries

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DISINFECTION

• Use appropriate disinfectant (EPA or FDA cleared)
• Use appropriate use dilution and contact time
• Thorough cleaning prior to disinfection critical
• Efficacy depends on microbe (hydrophobic viruses
  less susceptible to disinfectants)
• Hydrophobic viruses Coxsackie B1, polio virus 1
• Lipophilic viruses Herpes viruses, vaccinia

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DISINFECTANTS
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ETHICL AND SAFETY ISSUESCONCLUSIONS

• Risks and benefits of gene therapy unclear
• Review of proposals requires a committee with
  expertise in vector production, disease to be
  studied, animal experimentation, infection
  control and laboratory safety
• Better access to adverse events associated with
  clinical trials subdivided by transgene, vector,
  disease, delivery method is critical

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INFECTION CONTROLCONCLUSIONS

• Recommendations based on vector, route of
  transmission, environmental survival
• Recommendations limited by lack of published data
  on vector survival, persistence of vector, risk
  of transmission, and risk of development of
  replication competent vector
• Risk of wide spread use of gene therapy unknown
  since use confined to research trials (highly
  selected patients, research pharmacies, therapy
  in research units)

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SPECIFIC RECOMMENDATIONSBY VECTOR
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COMMON VECTORS

• Adenoviruses
• Retroviruses
• Adeno-Associated viruses
• Pox viruses
• Herpes virus
• Virus-like particles

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VECTORS OF CONCERN

• Replication competent
• Or able to combine with endogenous virus leading
  to competency
• Ability to establish latent infection
• Able to integrate into host genome
• Transmission via airborne/droplet or contact
  routes
• High infectivity
• Vector able to produce disease
• Transgene potentially detrimental to normal host

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RETROVIRUS EPIDEMIOLOGY

• Murine retroviruses
• Are not known to cause human disease
• Are inactivated by human complement
• Have a lipid envelope and do not withstand
  dessication
• Lentiviruses (HIV) are bloodborne human pathogens

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MURINE RETROVIRUSES

• Advantages
• Does not cause human disease
• Elicits little host immune response
• Transgenes may be expressed for life

• Disadvantages
• Oncogenic potential because integrates into the
  host genome
• Inactivated by human complement
• Must be given ex vivo
• Only infects dividing cells

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RECOMMENDATIONS FOR MURINE RETROVIRAL VECTORS

• Standard Precautions
• Manage blood spills with 110 final dilution of
  household bleach (like HIV or HBV)

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LENTIVIRUSES

• Advantages
• Can be given in vivo
• Not inactivated by human complement
• Infects dividing and non-dividing cells
• Transgenes may be expressed for life

• Disadvantages
• Oncogenic potential because integrates into the
  host genome
• May cause human disease

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RECOMMENDATIONS FORLENTIVIRAL VECTORS

• Standard Precautions
• Avoid sharps injuries
• Manage spills with 110 dilution of household
  bleach
• Role of anti-retrovirals post-exposure?

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ADENOVIRUSES

• Cause epidemic keratoconjunctivitis (pink eye),
  pharyngoconjunctival fever (head cold),
  gastroenteritis, acute hemorrhagic cystitis,
  pneumonia, meningoencephalitis
• Mortality 18-20 in liver and kidney transplant,
  and 10-60 in BMT patients

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ADENOVIRUS EPIDEMIOLOGY

• Can be recovered from infected eyes for up to 14
  days, and in the stool of AIDS patients for up to
  2 years
• Transmission by close personal contact
• Attack rates in hospitals range from 16 to 50
  with up to 300 secondary cases

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ADENOVIRUS

• Advantages
• Infects a wide variety of dividing and
  non-dividing cells
• Little oncogenic potential because does not
  integrate into the host genome
• Can carry larger transgenes

• Disadvantages
• Causes human disease
• Host immune response may limit use

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ADENOVIRUS INFECTION CONTROL ASPECTS

• The viruses are extremely hardy
• They persist on surfaces for up to 30 days
• Are not eliminated by soap and water, ethanol,
  chlorhexidine gluconate
• Quaternary ammonium compounds have variable
  efficacy

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EARLY ADENOVIRUS VECTOR TRIALS

• Low vector doses (105-109 pfu)
• Patients hospitalized 2 days before receiving
  vector
• Negative pressure isolation
• Isolated until cultures for RCAs and vector were
  negative for 3 days

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EXPERIENCE WITH ADENOVIRUS VECTORS

• 12 trials involving more than 300 patients
• Vectors with deletions in E1 and E3
• Doses ranging from 105 to 1013 pfu
• Multiple routes of administration
• RCAs and vectors looked for with culture and PCR
  in most studies
• --------------------------------------------------
  --------------------------------------------------
  ----------------------
• No RCAs isolated in any study
• Vector recovered in 5 studies

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RECOMMENDATIONS FOR ADENOVIRAL VECTORS

• Private room
• Standard Precautions if inoculum lt1013 particles
• Droplet Precautions if inoculum larger or for
  aerosol administration
• Clean with quaternary ammonium compounds if data
  supports efficacy or 110 dilution of household
  bleach solution
• Monitor RCA and vector shedding

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ADENO-ASSOCIATED VIRUS

• Single stranded DNA parvovirus
• Provirus integrated into the host-cell genome
• Remains latent until helper virus supplies genes
  for replication
• Helper viruses include adenovirus, herpesvirus,
  vaccinia

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AAV EPIDEMIOLOGY

• No known clinical illness
• 80 of adults have antibody
• Only isolated in patients with adenoviral
  infection
• Presumed route of transmission is respiratory or
  gastrointestinal

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ADENO-ASSOCIATED VIRUS

• Advantages
• Does not cause a known human disease
• High rates of transduction of muscle, brain, and
  liver cells
• May intercalate into the human genome at a
  predictable location

• Disadvantages
• Small size (can only carry genes up to 5 kb)
• Oncogenic potential due to integration

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RECOMMENDATIONS FOP AAV VECTORS

• Same as those for adenovirus
• Prevention of infections with helper viruses may
  be of value

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Portrait of Edward Jenner (1749-1823)
Ann Intern Med 1997127635-42
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ADVERSE REACTION RATES
Adapted from CDC.Vaccinia (smallpox vaccine)
recommendations of the ACIP, 2001. MMWR
200150(RR-10)
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VACCINIA VIRUS

• Advantages
• Can carry large transgenes (lt25kb)
• Genetically stable
• Infect a wide variety of cell types
• No oncogenic potential
• Easy to store as a freeze-dried preparation

• Disadvantages
• Not engineered to be replication-incompetent
• Many potential side effects
• Less effects with an immune response

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INFECTION CONTROL CONCERNS FOR VACCINIA VECTORS

• A replication-competent virus
• Shedding begins with appearance of the papule,
  reaches a maximum 4 to 14 days after inoculation,
  and persists until the lesion scabs over
• Cross-transmission occurs by close personal
  contact

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RECOMMENDATIONS FOR VACCINIA

• Screen for susceptible close contacts
• Young children, immunocompromised (HIV), pregnant
• Immunize in a private room with Standard
  Precautions
• Cover vaccination site with gauze and
  semipermeable transparent dressing
• Consider immunization for healthcare workers

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PRECAUTIONS BASED ON VECTOR AND ROUTE OF
ADMINISTRATION
Precautions A, airborne C, contact D, droplet
S, standard NA, not appicable
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CONCLUSIONS

• Recommendations based on vector, route of
  transmission, environmental survival
• Recommendations limited by lack of published data
  on vector survival, persistence of vector, risk
  of transmission, and risk of development of
  replication competent vector
• Risk of wide spread use of gene therapy unknown
  since use confined to research trials (highly
  selected patients, research pharmacies, therapy
  in research units)

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REFERENCES

• Weber DJ Rutala WA. Gene therapy. ICHE
  199920530-32.
• Evans ME, et al. Clinical infection control in
  gene therapy A multidisciplinary conference.
  ICHE 200021659-73.
• Evans ME Lesnaw JA. Infection control for gene
  therapy. CID 200235597-605.
• Dettweiler U, Simon P. Points to consider for
  ethics committees in human gene therapy trials.
  Bioethics 200115number 5/6.
• Preclinical evaluation of gene transfer products
  safety and immunologic aspects. Toxicology
  200217413-19.