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GENE THERAPY

Primary Immune Deficiencies – these are a subset of rare, inherited monogenic disorders
...
SCID-X1 is due to a deficiency of common interleukin receptor gamma-c (common)
chain, which is required for IL-2, -4, -7, -9, -15, and -21 receptors
...
Without the presence of this molecule, there can be no T or NK cells, and thus can
be lethal at 4 months if left untreated but is curable with successful bone marrow
transplantation
...
Patients can survive without bone marrow transplantation for ~10 years but under
sterile conditions – “bubble babies”
...
With human leukocyte antigen (HLA)-identical donors, HSCT treatment now allows
over 90% long-term survival with sustained immune recover
...
Transplantation from unrelated or haplo-identical (siblings) donors result in high
mortality rates of 70 and 37% respectively
...
Successful gene therapy for SCID-X1 in 1990 resulted in 9 children cured of the
disease and off any other treatment
...
Additionally, another fatal genetic disorder, ADA deficiency (SCID), which is due to
the absence of the adenosine deaminase (ADA) and results in a similar phenotype
(i
...
no T cells) to SCID but remains molecularly distinct, is curable by gene therapy
...
Usually, ADA is treated with enzyme replacement therapy (ERT)
...
However, despite the success in 25 patients, there were 5 cases of T cell leukaemia
and resulted in one death
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Therefore, gene therapy is not a cure
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SCID-X1 and ADA can be treated with two types of lentiviruses:
- Gamma retroviruses, which result in the development of leukaemia in SCID-X1
only and myeloid leukaemia in CGD
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• Severe combined immune deficiency (SCID-X1 or ADA-SCID):
1
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2
...


• Chronic Granulomatus Disease (CGD):
1
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2
...

3
...


• Wiscott-Aldrich Syndrome (WAS):
1
...




















2
...

•

Lipid metabolism disorders:
1
...


•

Haemophilia:
1
...

2
...

3
...


•

Other genetic disorders:
1
...


•

Cancer:
1
...

2
...

3
...


•

Neurodegenerative diseases:
1
...


•

Other acquired diseases:
1
...
e
...

2
...

- This is done via CRISPR, whereby the mutation is introduced to stop HIV entry
into the T cells

Retroviral Life Cycle:









These viruses have pseudo-random integration techniques into the host cell
genome (3)
...
Insertional inactivation – whereby, the provirus integrates into the coding sequence
of the gene and thus increases the risk of developing cancer
...
Insertional activation – whereby, the pro-virus is integrated upstream of the
regulatory gene sequence
...
Insertional truncation – whereby, the gene undergoes a gain-of-function due to the
loss of self-regulating proteins and the proto-oncogene is transformed into an
oncogene
...
Replication defective (helper dependent) retro-viral vectors:
- Viral components have very few genes in order to survive but encode for
multiple proteins
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-









These helper-dependent adenoviral vectors are devoid of all viral coding
sequences, possess a large cloning capacity, and can efficiently transduce a wide
variety of cell types from various species independent of the cell cycle and can
result in long-term transgene expression
...


5
...

- The chances of these two insertions – LMO2 and ll2rg – occurring together is very
small and thus suggests that these two genes are deregulated together to
induce leukaemia
...
Other possible factors contributing to development of T cell leukaemia in the
gammaC clinical trial (SCID-X1):
- Use of retroviral vectors – hence inherent risk of insertional mutagenesis
- Selective growth advantage T cells expressing gc
- The inherent anti-apoptotic effect of gc gene expression
- Genetic modification of haematopoietic stem cells
- Genetic modification of large numbers of cells – hence increased numbers of
cells at risk for mutagenesis
- The immune suppressed status of the host – lack of tumour immune response
(immune surveillance is present in ADA-SCID)
- Reduced endogenous numbers of competing T cells
- Potential predisposing cytogenetic abnormalities
7
...
e
...
e
...


Gene therapy strategies:
1
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There is less control in vivo than ex vivo, except in terms of the lipid disorder
...
Non-viral gene therapy strategies are inefficient and potentially dangerous
...
Cancer is an age-dependent disease and therefore, risk of insertional mutagenesis is
not a big concern for these patients
...
However, having said that, gene correction in cancer is ineffective because one cell
missed by this therapy will lead to the recurrence of the cancer and perhaps in a
more aggressive form
...
Some barriers to successful gene therapy include:
-

Poor uptake, transport and uncoating – endosomal degradation
Short-term persistence of the vector genome – episomal
maintenance/chromosomal integration
Poor transcriptional activity and inadequate transgene expression
Immunological responses to the vector, or the transduced cells, hence loss of the
transduced cells
...
Gendicine was the 1st licensed gene therapy product (2004) that targeted p53 in
solid tumours
...


2
...


Licensed Drugs (West):
1
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2
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3
...

4
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5
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Chemotherapy to induce cancer cell destruction
...

T cells isolated from donor, and gene edited to avoid graft vs host disease
...




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