Gene Therapy

1. Gene Therapy - Problems and
Challenges
Alison M. Beaney
Regional Quality Assurance
Specialist
North-East and Yorkshire
Helapet Aseptic Study Day 2008

2. 2
Gene Therapy
•Background to Gene Therapy
•Potential Benefits
•Perceived Hazards and Risks
•Regulations
•Implications for Pharmacy Aseptic Units
•Future?

3. 3
Gene Therapy
•Definition
The deliberate introduction of genetic material into
human somatic cells for therapeutic, prophylactic
or diagnostic purposes
Addition of EXTRA genes
Aim is to cure disease (or at least help the patient)
First introduction of gene-modified cells into a patient
was in 1989
First gene therapy product approved for market in 2004
•Still very experimental and early in its development

4. PTQA April 2008 4

5. 5
Gene Therapy Vectors
•Vectors deliver genes to cells
Therapeutic gene
(Transgene)
Therapeutic
proteinVector for efficient gene delivery
Transcription
Translation

6. 6
Types of Gene Therapy Vectors
•Non-viral vectors
Naked DNA
Liposomes/DNA
Polymer/DNA complex
(polyplex)
Liposome/Polymer/DN
A (lipopolyplex)
•Viral vectors
DNA viruses
Adenovirus
Herpes Simplex
Virus
RNA viruses
Retrovirus

7. 7
Gene Therapy Strategies
1)Gene Replacement
•Replace ‘faulty’ genes with normal genes
•Corrects inherited genetic errors
•Provides a missing function
•Monogenic diseases e.g. cystic fibrosis,
haemophilia, X-SCID
2)Gene Addition
•Delivers genes to provide a new function
•Polygenic diseases e.g. cancer

8. •Were trying to make a mouse
contraceptive vaccine for pest control
•Used modified mousepox virus as
vehicle for transporting antibodies
into mice
•Inserted gene to create ↑ IL-4
(interleukin 4) to boost production
•Surprise !!
totally suppressed the "cell-mediated
response“ which combats viral
infection
• Mousepox 100% lethal
2001
8

9. December 19, 2007
Boy gets leukaemia after gene treatment
to cure ‘bubble baby syndrome’
•3 year-old with X-linked severe combined immunodeficiency
(X-SCID) - immune system fails to develop
•Treated with genetically modified virus to correct the faulty DNA
that causes X-SCID
•Inserting the replacement DNA activated another gene that
promotes cancer
•Now an acknowledged risk of gene therapy
Also seen in 4 / 11 patients in a French trial
One has died while 3 are in remission
Retrovirus vector

10. 10
Regulations governing the handling
of gene therapy vectors
•No additional regulations governing the
handling of Non-Infectious vectors
Non-viral & Non-bacterial
•Viral vectors are Genetically Modified
•Genetically Modified Organisms
(GMOs)

11. 11
Genetic Modification
•Genetic modification is officially defined
as ‘the alteration of genetic material
(DNA or RNA) of an organism by
means that could not occur naturally
through mating and/or recombination’
A guide to Genetically modified organisms (Contained Use)
Regulations 2000. Health and Safety Executive

12. 12
Regulations governing the handling
of gene therapy viral vectors
Two sets of Regulations:
GMO (Contained Use) Regs 2000, HSE
All possible barriers (physical, biological or chemical) are
in place to limit contact of the GMOs with humans and the
environment
GMO (Deliberate Release) Regs 2002, DEFRA
All appropriate measures are taken to avoid damage to
the environment from the escape or release from human
control of GMOs
aimed at laboratories (difficult to interpret clinically)
no reference to product or patient safety

13. 13
Additional Regulations that apply to
Gene Therapy Clinical Trials
Protection of the Patient
Gene Therapy Advisory Committee (GTAC)
Established 1993, Department of Health
UK national research ethics committee (REC) for
gene therapy
Ethical acceptability for human gene therapy
Scientific merits
Potential benefits and risks
Patient flagging and long term monitoring
Advice to UK health Ministers on developments in
gene therapy research
Applies to ALL GENE THERAPY CLINICAL
TRIALS using viral and non-viral vectors

14. Containment Measures Required
Isolatable
Lab Suite
Microbiological
Safety Cabinet
Gloves Protective
Clothing
Class 1
Level 1
NO NO NO YES
Requires first use of
premises notification to
HSE
Class 2
Level 2
NO Risk
Assessment
R/A
R/A YES
Minimum requirement for
any human blood or
clinical samples.
Requires HSE notification
Class 3
Level 3
YES YES YES YES
+ Footwear
Requires HSE notification
Class 4
Level 4
YES YES YES YES
Complete
change of
clothing and
footwear on
entry and exit
Requires HSE notification
Containment Levels for GMOs

15. 15
Guidelines on Handling GMOs
in Pharmacy
•QA of Aseptic Preparation Services (4th Ed
n.
)
Appendix 6 Gene Therapy
•Scientific Advisory Committee on Genetic
Modification (SACGM), Part 6,
Guidance on the use of genetically modified
micro-organisms in a clinical setting
•European Association of Hospital Pharmacists (EAHP)
Guidance on the Pharmacy Handling of Gene
Medicines
•Rules and Guidance for Pharmaceutical
Manufacturers and Distributors 2007
– No Specific Guidance

16. APPENDIX 6 -
GENE THERAPY
•Facilities
•Documentation
•Labelling
•Training
•Aseptic
processing
•Cleaning
•Storage
•Transport
•Waste
Disposal
•Spillage
16

17. 17
Facilities
Gene therapy should not be manipulated in
clinical areas
Basic Principles - Containment
- Knowledge / understanding / skill
- Validated procedures
Persons handling the product should be masked and gloved
All disposable equipment and materials used for prep & admin
- handled as biohazardous
•Dedicated facilities required
-ve pressure isolators or Class II BSC
+ve pressure room or lobby
Containment level > 2

18. Clean room suite
designed to
provide protection
to the cleanroom

19. 19
Aseptic Manipulation
Doses
Calculation / dilutions / multiple dilutions
Needle stick injury risk
Units
Particle Units/ml (PU/ml)
Plaque Forming Units/ml (PFU/ml)
Infectious particle Units/ml (IU/ml)
Gene Transfer Units/ml (GTU/ml)
Stability
Container compatibilities - Plastic/glass
adhesion
Expiry date - Time to administration from thawing

20. 20
Decontamination
•Cleaning
Virucidal detergents (validated against GT vectors)
Cleaning Validation
Specific Detection methods needed for viruses
that are virus specific and highly sensitive
•Waste Disposal
On site validated autoclave for re-usable
equipment
Inactivation on-site for Class 3 vectors
Validated autoclave
Incineration
Disinfectant treatment

21. 21
•Spillage
Specific to GT vector
Spillage kit
Contents ( gloves, masks, aprons, goggles,
disposable shoe covers, virucidal detergents,
absorbent material, disposable forceps &
biohazard incineration bag)
Positioned in all GT handling areas
Notification to HSE
Accidental Exposure

22. 22
SOPs needed
Safe handling & protection
Storage
Operators
(Not pregnant, breastfeeding or immunosuppressed)
Training
Facilities
Spillage, contamination & needle stick
Waste disposal, cleaning and transport

23. 23
Risk Assessment
•Assess each product individually
Cytolytic viruses
Non-cytolytic viruses
Replication competent
Replication deficient
Class I, II or III

24. 24
What will the Future bring?
Dedicated facilities
Automation?
The first gene medicine in Europe could be
licensed in 2008
Licensed closed-system gene therapy products
Use of gene therapy as an adjunct to standard
therapy e.g. Radiotherapy & Chemotherapy
Vector development e.g.
Targeted vectors (viral & non-viral)
Bacterial vectors

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Additional Information
Gene Therapy Advisory Committee (GTAC) http://www.advisorybodies.
doh.gov.uk/genetics/gtac/index.htm
Gene therapy trials worldwide. Provided by the Journal of gene medicine
http://82.182.180.141/trials/index.html
A guide to Genetically modified organisms (Contained Use) regulations
2000. Health and Safety Executive
Genetically Modified Organism (Deliberate Release) Regulations 2002
[GMO(DR)]. Department for the Environment, Food and Rural Affairs (DEFRA)
http://www.opsi.gov.uk/si/si2002/uksi_20022443_en.pdf
Quality Assurance of Aseptic Preparation Services Fourth Edition. A.M.
Beaney. Pharmaceutical Press 2006. Appendix 6. Gene Therapy.
EU Clinical Trials Directive. http://www.wctn.org.uk
/downloads/EU_Directive/Directive.pdf
Implications of gene therapy for hospital pharmacists. Simpson.J, Stoner.
N. www.pjonline.com/pdf/articles/ pj_20030726_genetherapy.pdf

26. 26
Additional Information
Cancer gene therapy: from science to clinical trials. Searle. P.F, Spiers. I,
Simpson. J, James. J.D. Drug Delivery Systems and Sciences 2002, 2 (1),
5-13.
Standards for gene therapy clinical trials based on pro-active risk
assessment in a London NHS Teaching Hospital Trust. Bamford, K.B.,
Wood, S., Shaw, R.J. QJM 2005, 98, 75-86. www.qjmed.oupjournals.org
Progress in Gene Therapy – are hospital pharmacies the next barrier?
Simpson, J. Hospital Pharmacist, 2006, 13 (8), 266 http://www.pjonline.com/
pdf/hp/200609/hp_200609_comment.pdf
Cancer Biotherapy. An Introductory guide. Young, A. Rowett, L. Kerr, D.
Oxford University Press 2006
•Scientific Advisory Committee on Genetic Modification (SACGM), Part 6,
Guidance on the use of genetically modified microorganisms in a clinical
setting. http://www.hse.gov.uk/biosafety/gmo/acgm/acgmcomp/part6.pdf
•European Association of Hospital Pharmacists (EAHP) Guidance on the
Pharmacy Handling of Gene Medicines. http://www.ejhp.eu/