The use of recombinant genetic information (DNA, RNA, hybrid molecules) under different forms of pharmaceutical preparations as therapeutic agent.
Can offer promise for treating currently intractable diseases.
Can be used for both inherited monogenic disorders and acquired diseases.
Take a gene from a patient - EX VIVO.
Isolate the target cells.
Transfer the gene to them.
Reinfuse the gene-modified cells into the patient.
Infuse the therapeutic gene inside a vector into the patient - IN VIVO.
Haematopoietic system: stem cells, progenitor cells, mature cells
Muscle: myoblasts, myofibres
Vasculature: endothelium, smooth muscle
Skin: keratinocytes, fibroblasts
CNS: neurons, glia, microglia
IDEAL TARGET CELL: pluripotent, self-regenerating stem cells from the patient
VIRAL GENE THERAPY VECTORS
ADENO-ASSOCIATED VIRUS: non-integrating
HUMAN GENE THERAPY SUCESSES VIA EX VIVO PROCEDURE:
Severe combined immune deficiencies SCID - haematopoietic stem cells X-linked chronic granulomatous disease CGD - haematopoietic stem cells
Junctinoal epidermiolysis bulls JEB - epidermal stem cell
X-linked adrenoleukodystrophy ALD - haematopoietic stem cells
Metachromatic leukodystrophy MLD - haematopoietic stem cells
B-thalassemia/sicke cell disease SCD - haematopoietic stem cells
HUMAN GENE THERAPY SUCESSES VIA IN VIVO PROCEDURE
Haemophilia B - AAV vector
Parkinson’s disease - AAV vector
Leber’s congenital amaurosis - AAV vector
EX VIVO PROCEDURES:
X-linked severe combined immunodeficiency
Complete lack of NK and T-cells in the peripheral blood.
Mutation of gamma chain LR2RG a subunit of haematopoietic cytokine receptors IL2, IL4, IL7, IL9, IL15, IL21: result, block in T-cell development.
Children with XSCID usually die in their first month of life
EX VIVO gene devilry protocol
bone marrow harvested, CD34 positive stem cells are isolated
cells activated to grow for 24h with cytokines: need to activate mitosis as retroviruses will only transduce dividing cells
cells traduced by three rounds of culture int he presence of the supernatant containing IL2RG retroviral vector in sterile bags
transducer CD34+ cells infused back into the patient
WISKOTT - ALDRICH SYNDROME WAS
Infections, microthrombocytopenia, eczema, autoimmunity lymphoid malignancies.
X-linked, mutations in the WAS gene, codes for WASP a protein that regulates the cytoskeleton.
WASP-defective immune cells, alterations in proliferative responses after activation, cell migratino, immunological synapsis formation, cytotoxicity.
Regular treatment is allogenic haematopoietic stem cell transplantation.
Require the introduction of a normal functioning B-globin transcription unit in to the haematopoietic stem cells of the patient. A minimum of 25% of normal levels of B-globin are required for the therapeutic effect; 50% are curative.
Ex vivo protocol.
No current lentiviral vector capable of curing thalassemia major.
IN VIVO PROCEDURES:
LEBER’S CONGENITAL AMAUROSIS
In vivo delivery to the retina of AAV vector with RPE65 gene
X linked bleeding disorder, the royal disease, defect in coagulation factor IX.
Administratin of a AAV vector into peripheral veins targeting liver.
Results in transgene expression sufficient to improve bleeding phenotype over at least 3 years.