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IMMUNOTHERAPY
History
Immunotherapy, or the concept of boosting the immune system to target and destroy cancer cells, has been a goal of cancer treatment for over 100 years. However, limited success has been achieved with traditional immunotherapy, as cancer cells tend to evolve mechanisms that evade immune detection. A wide array of gene therapy techniques are being used to overcome this limitation.19
Currently gene therapy is being used to create recombinant cancer vaccines. Unlike vaccines for infectious agents, these vaccines are not meant to prevent disease, but to cure or contain it by training the patient’s immune system to recognize the cancer cells by presenting it with highly antigenic and immunostimulatory cellular debris. Initially cancer cells are harvested from the patient (autologous cells) or from established cancer cell lines (allogeneic) and then are grown in vitro. These cells are then engineered to be more recognizable to the immune system by the addition of one or more genes, which are often cytokine genes that produce pro-inflammatory immune stimulating molecules, or highly antigenic protein genes. These altered cells are grown in vitro and killed, and the cellular contents are incorporated into a vaccine (figure 1A▶).20 Immunotherapy is also being attempted through the delivery of immunostimulatory genes, mainly cytokines, to the tumor in vivo. The method of introducing a gene to the tumor varies and is discussed in more detail in the gene transfer section of this review. Once in the cancer cell, these genes will produce proteins that unmask the cells from immune evasion and encourage the development of antitumor antibodies (figure 1B▶).
other ways to defeat cancer:-
Vaccines using engineered cells are showing great promise for the treatment of many cancers that respond poorly to conventional therapy.
The next generation of vaccines is already in clinical trials for several cancer types. Table 1▶ provides a list of the more advanced clinical trials in this field, including phase, the type of cell used and the gene used to create a better immune response. These trials were picked to illustrate the fact that there are wide ranges of trials in different stages of efficacy testing using a variety of vectors for many cancer types.
| Cancer | Stimulating genes | ClinicalTrials.gov47 identifier # | Description | Phase |
| Prostate | Murine α(1,3)- galactosyltranferase | NCT00105053 | Mouse protein-sugars are expressed on allogeneic prostate cells to induce a hyperacute rejection response | II |
| Pancreatic | CEA and MUC-1 | NCT00088660 | Replication incompetent vaccinia and fowlpox viruses engineered to produce CEA and MUC-1 given subcutaneously to produce an immune response to pancreatic cancer | III |
| Prostate | GM-CSF | NCT00122005 | Allogeneic prostate cells expressing the GM-CSF gene are used to induce immune response following chemotherapy and peripheral blood mononuclear cells infusion | I/II |
| Lymphoma | GM-CSF and CD40L | NCT00101101 | Autologous tumor cells are combined with allogeneic cells that express GM-CSF and CD40L and incorporated into a vaccine with low doses of IL-2 | II |
| Melanoma | IL-2 | NCT00059163 | Autologous tumor cells engineered to express IL-2 are incorporated into a vaccine. | II |
| Kidney | CD-80 | NCT00040170 | A modified replication incompetent adenovirus containing the tumor antigen CD-80 is injected subcutaneously along with the cytokine IL-2 to produce an immune response to the prostate cancer |
Future Directions
Because oncolytic virotherapy is not yet a mature technology, there is plenty of room for improved treatment vectors. In order for virotherapy to be successful, viral particle production rates in the infected cancer cells must outstrip the growth rate of the uninfected cancer cells. This may be difficult to achieve with large established tumors54 and may mean that virotherapy must be combined with an existing therapy, such as surgery, to decrease the number of cancer cells in the initial treatment. In addition, the most effective treatment delivery method is yet to be determined. In preliminary studies, systemic injection required 1000x the viral load necessary to achieve results than injection intratumorally.55
However, once these factors are overcome, there are many benefits to oncolytic therapy. The selective nature of the virotherapy ensures that healthy tissue will be minimally impacted. In addition, when combined with cytotoxic gene expression, this therapy can affect not only rapidly dividing cells, but those in the surrounding tissue making the microenvironment less favorable for cancer growth. The combination of the powerful killing nature of these vectors combined with the selectivity makes them an exciting avenue for lowering the number of cancer deaths.
GENE TRANSFER
History
One of the most exciting treatments to emerge from the concept of gene therapy is that of gene transfer or insertion. This is a radically new treatment paradigm involving the introduction of a foreign gene into the cancer cell or surrounding tissue. Genes with a number of different functions have been proposed for this type of therapy, including suicide genes (genes that cause cellular death when expressed), antiangiogenesis genes and cellular stasis genes (figure 3▶). A number of different viral vectors have been used in clinical trials to deliver these genes, but most commonly have used a replication incompetent adenovirus. Nonviral methods, including naked DNA transfer and oligodendromer DNA coatings, as well as electroporation are also viable modes of gene delivery.56 The type of delivery vehicle chosen depends on the desired specificity of the gene transfer therapy, as well as the length of time the gene must be expressed in order to be effective. For instance, a replication incompetent adenoviral vector containing the herpes simplex virus thymidine kinase (HSVtk) gene needs only transient expression to accomplish cell death and is generally delivered via an adenoviral vector.57 However, antiangiogenesis genes, such as sFLT-1 and statin-AE, need continuous expression for therapeutic effect and have been delivered using plasmids that contain a transposon to insert the gene into the cellular DNA
| Cancer | Transferred genes | ClinicalTrials.gov47 identifier # | Description | Phase |
| Pancreatic | Rexin-G | NCT00121745 | A cytocidal cyclin G1 construct accumulates preferentially in the tumor cells to block the action of cyclin G1 and initiate cell death | I |
| Glioblastoma | HSVtk | NCT00001328 | The HSVtk gene is introduced into glioblastoma cells via a mouse retrovirus. Glioblastoma cells with the HSVtk gene are then sensitive to the drug glanciclovir which is administered | I |
| Head and neck | p53 | NCT00041613 | Transfer of the p53 gene via a replication incompetent adenovirus to tumor cells to inhibit cell growth and induce apoptosis | III |
| Melanoma | MDA-7 | NCT00116363 | MDA-7 a novel tumor suppressor molecule is introduced into the melanoma cells and overexpression inhibits cellular proliferation and induces apoptosis | II |
| Pancreatic | TNF-α | NCT00051467 | The TNF-α gene under the control of a radiation inducible promoter is introduced into tumor cells and in combination with the radiation therapy induces cell death |
Another exciting gene therapy treatment agent is Rexin-G, the first injectable gene therapy agent to achieve orphan drug status from the Food and Drug Administration for treatment of pancreatic cancer.68 This gene therapy agent contains a gene designed to interfere with the cyclin G1 gene and is delivered via a retroviral vector. The gene integrates into the cancer cell’s DNA to disrupt the cyclin G1 gene and causes cell death or growth arrest. In a phase I trial, 3 of 3 patients experienced tumor growth arrest with 2 patients experiencing stable disease. These results have led to larger phase I and II trials.69 Rexin-G is also being evaluated for colon cancer that has metastasized to the liver.
Another exciting gene therapy treatment
A gene transfer technology that shows great promise is the replication incompetent adenovirus delivering the HSVtk gene to a tumor followed by ganciclovir treatment. Ganciclovir is not toxic unless metabolized by the HSVtk gene,70 and therefore only the cancer cells that are treated with the gene and the surrounding cells will be affected by treatment. In a large phase I study involving glioblastoma patients, the HSVtk-engineered viral treatment increased median survival from 39 weeks to 70.6 weeks and was the first glioblastoma gene therapy trial to show any measurable improvement in survival.71
Several agents that use a replication incompetent adenoviral vector to deliver the p53 gene to cancer cells are also currently in phase II and III trials. The p53 gene is an important cell cycle regulator that has been extensively studied and is mutated in 50% to 70% of human tumors.72 Mutations in this gene are often linked to aggressiveness. It has been shown that restoration of a functional p53 gene in cancer cells results in tumor cell stasis and often apoptosis.72 Using this information, INGN 201, an adenoviral vector containing p53 for gene transfer, is in current phase III testing for squamous cell carcinoma of the head and neck, and has completed phase I studies on prostate, ovarian, glioma and bladder cancer.
Future Directions
Gene transfer, while a radical new type of treatment, is also the only gene therapy product to obtain regulatory approval in any global market, as demonstrated by China’s 2003 approval of Gendicine for clinical use.76 Gendicine is a modified adenovirus that delivers the p53 gene to cancer cells and is approved for the treatment of head and neck squamous cell carcinoma. Since approval, thousands of patients have been treated in China; some with repeated injections. As yet, large-scale efficacy trial results have not been published; the results of which are eagerly awaited.
Gene transfer technology allows an incredible diversity of treatment possibilities. This diversity can be used to complement traditional therapies, as well as provide radically new frontiers for treatment. Gene transfer therapy can rely on the current information known about the genetics of cancer formation, bringing a more sophisticated and personalized approach to therapy. Current gene transfer trials have demonstrated statistically significant survival improvements for cancers such as glioblastoma and pancreatic cancer, as discussed previously. These studies have provided very encouraging signs that current research is on the right path.
ONCOLYTIC AGENTS
History
Another growing area of gene therapy treatment for cancer is the use of oncolytic vectors for cancer destruction. Like immunotherapy, this is a concept that has been around for almost a century and, like immunotherapy, it is undergoing a renaissance due to gene therapy.38 Oncolytic gene therapy vectors are generally viruses that have been genetically engineered to target and destroy cancer cells while remaining innocuous to the rest of the body. Oncolytic vectors are designed to infect cancer cells and induce cell death through the propagation of the virus, expression of cytotoxic proteins and cell lysis (figure 2▶).39 A number of different viruses have been used for this purpose, including vaccinia, adenovirus, herpes simplex virus type I, reovirus and Newcastle disease virus.38 These viruses have been chosen, in many cases, for their natural ability to target cancers, as well as the ease at which they can be manipulated genetically.
Initial trials of oncolytic therapies have highlighted both its incredible power, as well as unique obstacles to treatment implementation. Mammalian models of oncolytic gene therapy have worked remarkably well. In murine models, both colon and bladder cancer have shown survival benefits and reduced metastasis using oncolytic viral agents.40,41 In a canine model, using an oncolytic virus designed to destroy osteosarcoma, survival was prolonged even in immunocompetent dogs with syngenic osteosarcoma.42 However, there are several unique stumbling blocks for oncolytic virotherapy in humans. Most people have antibodies to the common viruses used for therapy development which often leads to an immune response that clears the viral agent before it has had time to infect cells. In addition, the use of replication competent viral particles often calls for increased safety precautions, making clinical trials more expensive and cumbersome.43 In a trial using a modified vaccinia virus to treat breast and prostate cancer, patients were required to be isolated in a specialized hospital facility for a week to ensure that the virus had completely cleared before being allowed back into the general population.18 Because of these limitations, there have been relatively few trials with oncolytic therapy. However, new vectors are being created and past experience is being incorporated into current trials to enhance results so that they mimic those in animal studies.
Current Clinical Trials
Even in this early stage, oncolytic viral therapy has demonstrated some success. Both adenovirus and herpes virus agents have ongoing clinical trials for intractable cancers. The most notable adenoviral therapy is the ONYX-015 viral therapy. ONYX-015 is an adenovirus that has been engineered to lack the viral E1B protein.44 Without this protein, the virus is unable to replicate in cells with a normal p53 pathway. In addition, the E1B protein is essential for RNA export during viral replication.45 Cancer cells often have deficiencies in the p53 pathway due to mutations and thus, allow ONYX-015 to replicate and lyse the cells.44 Cancer cells also exhibit altered RNA export mechanisms that allow for the export of viral RNA even in the absence of the E1B protein.45 ONYX-015 has been tested in phase I and II trials on squamous cell carcinoma of the head and neck that resulted in tumor regression which correlated to the p53 status of the tumor. Tumors with an inactive pathway demonstrated a better response.46 Phase II trials of ONYX-015, in combination with chemotherapy, demonstrated even better tumor response and have led to a phase III study.47 In addition to squamous cell carcinoma, ONYX-015 is currently being tested as a preventative treatment for precancerous oral tissue, the theory being that even in the precancerous state, there are p53 pathway inactivating mutations that will allow the oncolytic adenovirus to replicate and eliminate the cells before they become cancerous.48
The second type of oncolytic virotherapy undergoing clinical trials uses herpes simplex virus type 1 (HSV-1). Two vectors, G207 and NV1020, are currently in phase I and phase II trials for treatment of intractable cancers. Mutations in several genes of these herpes viruses ensure that they replicate efficiently only in cancerous cells. G207 is mutated so that it has attenuated neurovirulence and cannot replicate in nondividing cells.41 NV1020, a derivative originally used for vaccine studies, has multiple mutations, including a deletion in the thymidine kinase region and a deletion across the long and short components of the genome, and an insertion of the thymidine kinase gene under the control of the α4 promoter.41 These viral vectors have two distinct cell killing mechanisms. The lytic portion of the life cycle directly kills cells and the thymidine kinase that is expressed from the viral genes sensitizes cells to ganciclovir. These viral therapy vectors have been used with great success in vitro and in model animals against a wide number of solid cancers.49–51 Clinical trials using these vectors include a phase I trial of G207 for treatment of malignant glioma52 and a phase I/II trial of NV1020 for treatment of colorectal cancer metastases to the liver.53 In addition, NV1020 has also been tested for treatment of glioblastoma.53
Future Directions
Because oncolytic virotherapy is not yet a mature technology, there is plenty of room for improved treatment vectors. In order for virotherapy to be successful, viral particle production rates in the infected cancer cells must outstrip the growth rate of the uninfected cancer cells. This may be difficult to achieve with large established tumors54 and may mean that virotherapy must be combined with an existing therapy, such as surgery, to decrease the number of cancer cells in the initial treatment. In addition, the most effective treatment delivery method is yet to be determined. In preliminary studies, systemic injection required 1000x the viral load necessary to achieve results than injection intratumorally.55
However, once these factors are overcome, there are many benefits to oncolytic therapy. The selective nature of the virotherapy ensures that healthy tissue will be minimally impacted. In addition, when combined with cytotoxic gene expression, this therapy can affect not only rapidly dividing cells, but those in the surrounding tissue making the microenvironment less favorable for cancer growth. The combination of the powerful killing nature of these vectors combined with the selectivity makes them an exciting avenue for lowering the number of cancer deaths.
New delivery methods and more sophisticated gene expression cassettes will create better therapeutic alternatives to make the goal of cancer treatment and eradication achievable.
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