Most of the haematological malignancies both inherited and acquired occur as a result of mutation of the proto-oncogenes or tumor suppress genes as show in table 1 above resulting in either gain-of-function and loss-of-function mutations respectively. P35 is a tumor suppressor gene that has been implicated in chronic lymphocytic leukaemia (CLL). Mutations that deactivate P53 in cancer usually occur in homo-oligomerisation
domain (OD) residues,207-355 of the gene. Most of these mutations destroy the ability of the protein to bind to its target DNA sequences and thus prevents transcriptional activation of these genes. Therefore OD mutations have a dominant negative effect on the function of P53. An example of a mutation in P53 is where arginine 248 is altered, sometimes causing a disruption in balance and making the protein unable to bind with the DNA.
Gene sequencing of P53 gene and other proto-onco-genes that are mutated in various cancers has enabled the correct sequence of these genes to be produced, thus transduction of the correct genes with viral vector is believed to cure the leukaemia and other forms of cancers.
A case study currently going on at memorial sloan-kettering cancer center in New York led by Michel Sadelain showed that gene therapy cures leukaemia within eight days.
The key to the new therapy is identifying a molecule unique to the surface of cancer cells, and then genetically engineering a patient’s immune cells to attack it. In acute lymphoblastic leukaemia, immune cells called β-cells become malignant. The team were able to target a surface molecule known as CD19 that is only present on β-cells. They extracted other immune cells called T-cells from the patients; the T-cells were treated with a harmless virus which installed a new gene redirecting them to attack all cells bearing CD19. When the engineered T-cells were reinfused into the patients, they rapidly killed all β-cells, cancerous or otherwise.
“The stemming finding was that in all five patients, tumors were undetectable after the treatment” says sadelain.He reckons that the body should replenish the immune system with regular T-cells and healthy β-cells after a couple of months. However, the patients received donated bone marrow to ensure they could regrow a healthy immune system. Fifty patients are currently waiting to undergo the same treatment. (Science Translational Medicine, doi.org/kwz). It is believed that gene therapy can provide a lasting solution to many cancers, either by genetically engineering T-cells to attack cancer cells with specific antigen or by obtaining the correct sequence of these proto-oncogenes and transducing them into bone marrow cells using gene therapy vectors.
Cancers can also be cured using gene therapy by:
- Stimulating the body’s immune cells to attack cancer cells.
- Insertion of genes into cancer cells to make them more sensitive to chemotherapy, radiotherapy or other treatment.
- Introducing suicide genes into a patient’s cells (pro-drug) which is activated in cancer cells thereby causing their destruction.
- Prevention of cancer cells from developing new blood cells(angiogenesis)
Treatment of Eptein Bar Virus Lymphomas in haematological
Gene transfer has also been proved useful for generating the LMP2 protein of Epstein Bar Virus which is expressed on some human lymphomas in antigen presenting cells in vitro. Development of lymphoid populations with LMP2 specificity has yielded T-cells with potent anti tumor reactivity against the lymphoma cells, in one recent study by the response of 5 of 6 patient with active, relapse lymphoma (Bollard et al., 2007 ).
Development of tumor vaccines
Much effort has also focused on the development of effective tumor vaccines by expressing genes that enhance the immune response at the vaccine injection site by releasing cytokines, costimulatory molecules or gene products which block inhibitor signals from tumor cells (Hedge et al.,2006 Hodi and Dramoff, 2006). These strategies have relied on the transduction of primary autologous tumor cells or the use of other autologous cells after genetic modification. Ten patients in a recent study (including 7 children with high-risk, acute myeloid or lymphoblastic leukaemia) received multiple injections of irradiated tumor cells mixed with autologous skin fibroblast expressing IL-2 and CD40L, immunization produced a substantial increase in frequency of T-cell reactive against recipients-derived blast cells. Eight patients remain disease free for up to 5 years after treatment (Rousseau et al., 2006).
Creation of chimeric antigen receptors (CARs)
This is another approach of gene therapy in which a single chain antibody with specificity for an antigen expressed on human tumor cells is linked to an internal kinase domain which mediates cell activation when the antibody engaged by the target antigen.
For example; CARs targeting CD19 expressed on human β-cells malignancies have been developed and shown to eradicate tumors in a mouse model (Brentjens et al., 2003). Because of the need for costimulation to achieve T-cell activation in vivo, the cytoplasmic signaling domain of the CD28 receptor has been added to CAR. Analogous approaches are being used to derive primary natural killer cells for use in treating childhood leukaemia. (Imaiciwamto and Campana, 2005).
Haemophillia:- There has been considerable interest in the development of gene therapy for haemophilia over the years, involving the use of oncoretroviral, lentiviral, adenoviral and adeno-associated viral vectors.These disorders have seemed particularly amenable to gene therapy approaches since small amounts of the clotting factor are sufficient to significantly correct the bleeding phenotype. The only obstacle to development of gene therapy for this disorder, is finding a suitable vector that will not elicit immune response.
Other novel gene therapy-based strategies that are previously being explored for the treatment of haematological malignancies include those that aim to restore the function of tumor suppressor genes (McNeish et al., 2004) enhance the immune response to leukaemic blasts and inhibit angiogenesis (Kyriakou et al.,2003).
GENE THERAPY FOR TREATMENT OF ADENOSINE DEAMINASE SEVERE COMBINED IMMUNODEFICIENCY (ADA-SCID)
Other recent success of gene therapy is the correction of SCID that results from Adenosine deaminase deficiency, an autosomal recessive condition with a complex phenotype (Aiuti ., 2002). In a departure from conventional gene transfer protocols, two patients (aged 6 and 30months, respectively), who were not on enzyme replacement therapy received non-myeloablative conditioning with busulphan prior to transplantation with oncoretrovirally transduced CD34+ cells (1-4x106 cells/kg). Transient myelo-suppression was observed; however, neither patients experienced serious toxicity nor required blood products. The transduction efficiency of the clonogenic progenitors was between 21% and 25%. Patient 1, who received the higher cell dose, has been followed for over 2 years and has normal peripheral T, B and NK cell numbers as well as normal immunoglobin levels.
Cell-medicated and humoral response to a variety of neoantigens, including tetanus toxoid, was comparable with those of age-matched controls. Immunological reconstitution in the second patient has been less impressive, although significantly improved on pre-transplant levels.
This patient was older at the time of transplantation and received a lower dose of gene-modified cells. ADA activity in the plasma increased in both patients after gene therapy with a parallel decline in the toxic adenine deoxyribonucleotide metabolities in red blood cells to levels found in patients after a successful allogenic transplantation. A third patient has been treated more recently using the same protocol and is showing a similar recovery to that of patient 1 (Aiuti, 2003).
Predictably, peripheral blood T, B and NK cells showed very high levels of gene marking (between 70% and 100%), when compared with circulating myeloid progenitor (5-20%). However, in this limited data set, marking was high in the myeloid compartment than observed in patients with SCID-XI described above; perhaps, a result of myeloablation at the time of transplantation, which would create an environment that would foster expansion of gene modified cells (Amit et al., 2004).