Viral Vectors

416521 Medical DNA Technology

Graduate Course, First Semester, 2009

Assist. Prof. Somchai Saengamnatdej, Ph.D.

Thursday 25th June 2009
 Outline (4 periods)
1. Introduction and Concepts
 Definition & related terms
 Non-viral/ Viral vectors
 Systemic/ Local gene deliveries
 Gene delivery barriers
 Appropriate vectors: Factors to be considered
 The potential use of viral vectors
 Rationale for some experiments

2. Types of viral vectors

 Viruses : Review of features & their replication cycle
 Viral vectors ; structure, experiments, advantages, and limitations

3. Summary
4. Perspectives
CONCEPTS
 Vector Definition
 Vehicle for delivery of genetic material
to host cells
Replication-Deficient Viruses
DNA Plasmids.

Other Non-viral vectors

Virosome
 Gene Delivery
1. Systemic gene delivery.
 Via bloodstream.
 Distribute throughout the body.
 For target tissue that cannot be reached directly.
 Often low specificity of gene expression.
 Require large concentration.
 May influence & damage the function of normal
healthy tissues.
 High titre could carry immunologic or toxicity
concerns.
 Inappropriate for many musculoskeletal system
injuries due to lack of blood supply to various
tissues (such as cartilage or meniscus).
 Gene delivery
2. Local gene delivery.
 In vivo: direct injection of vector/ DNA
complex into the host tissue.
• Ex vivo:
Growing cells in culture

delivery gene (alone or with growth factors,

differentiation signals) into the cells
or with other chemical or physical alterations to
the culture environment

Transplant into patients.

EXAMPLE: Ex vivo TKD/IL-2 activated, patient-derived autologous NK cells.
(Note: Hsp70-peptide TKD, a 14-mer immunogenic peptide derived from
the C-terminal domain of Hsp70 protein, in combination with IL-2 or IL-15.)

(http://www.onkologie-regensburg.de/Forschung/Englisch/Multhoff/Immunotherapy_neu.htm 13 Nov2006)
 Terms
In vivo

Ex vivo
Autologous
Xenogeneic
Allogeneic
Syngeneic
 Gene delivery barriers

1. systemic (pre-existing immunity)

2. cellular (binding to cell surface)
 Factors: to select appropriate vectors.
 Duration of expression required (transient or long term.)
2. Nature of target cells
 ease of transduction
 receptor expression
 dividing (cancer) or non-dividing (neuron, islets, muscle cell).
 Route (Ex vivo or In vivo).
 Maximum threshold of vector-induced immune response
acceptable for the host.
 Level of expression.
 Desired temporal regulation of transgene expression
 Inducible or Constitutive.

[One vector unlikely appropriate for all]

 Non-viral vectors
Delivery:
 Direct injection:
simple & safe (dividing & non-dividing cells).
 Electroporation.
 Encapsulated within liposomes.
 Gene-activated matrices.
 Natural/ Synthetic polymers.
 Attached to gold microparticles for gene gun.
 Non-viral vectors
 Limitation:
Low transfection efficiency.
Transient expression.
Non-selective cell targeting.

 Advantages:
Easy to manufacture.
Able to incorporate large sequences of DNA.
Cost-effective.
Display low immunogenicity in vivo.
 Viral Vectors
 Engineering the virus to express the gene
of interest.

Advantages:
 Prolonged transgene expression.
 Markedly higher transduction efficiency

(more efficient than physical methods).

 Treatment
 Inherited congenital disorders :
 Cystic fibrosis
 Phenylketonuria
 Adenosine deaminase deficiency
 Haemophilia B.
 Inherited retinal degenerations
 Mutations in genes expressed in photoreceptors or the
retinal pigment epithelium
 Acquired diseases:
 AIDs, Cancer, Cardiopathy.
 Tissue engineering:
 Trauma, burns, degenerative diseases, and
diseases that produce tissue or organ failure.
 Rationale for Some Experiments (1)
 To deliver growth factors that control the
formation of new vasculature to ischemic
tissues in order to restore the blood supply
that has been pathologically disrupted or
reduced [therapeutic angiogenesis].

 Bone fractures: [ bone healing].

 non-healing or
 ~10%
 delayed union defects caused by
Require
 age-related bone deterioration
 traumatic injury
some forms
 tumour resection, or of orthopaedic
graft
 osteolytic disease
Rationale for Some Experiments (2)
3. Use of tumour-selective replicating viral vector
may enable the targeting of the gene-modified
viruses to malignant tissue without toxic effect
(happened with standard approaches) [anti-cancer].

4. To introduce genes designed to specifically block

or inhibit the gene expression or function of gene
products [intracellular anti-microbial].

5. To inhibit the spreading by expression of a

secreted inhibitory protein or stimulation of a
specific immune response [extracellular anti-
microbial]