Update - Gene Therapy for Patients with Head & Neck Cancer
George Yoo, M.D.
Head and Neck Cancer
Head and neck cancer affects 42,000 patients per year. The standard therapy for head and neck cancer is surgery, radiation and sometimes chemotherapy. In advanced head and neck squamous cell carcinoma (HNSCC), the five-year survival rate is less than 40%; which has not changed significantly in the past forty years. Therefore, research outside the standard treatment modalities is being studied. One biological approach, gene therapy, demonstrates tumor suppression in preclinical studies and is being tested in humans. Before the details of gene therapy are presented, background information on molecular genetics and tumor immunology will be reviewed.
Genetics of Cancer
Certain genes in cancer cells become altered, and these cancer cells gain a growth advantage over normal surrounding cells. In general genes can promote tumor growth (oncogenes) or genes can suppress growth (tumor suppressor genes). Cancer cells either knock out tumor suppressor genes or activate oncogenes so that they can proliferate without being checked. Gene therapy reengineers these genetic alterations so that cancer cell growth can be suppressed. After a gene is transfected into a cell, mRNA is transcribed and then its protein product is translated.
Tumor Immunology
The basis of tumor immunology is that the immune system can kill cancer cells. Cancer cells avoid this immune surveillance by suppressing the body's immune system. Immune cells, such as cytotoxic T-lympocytes, attack cancer cells. Cytokines, such as interleukin-2 (IL2) and interleukin-12 (IL-12), activate the immune system. Tumor specific antigens are expressed on tumor cells and help the body mount a specific immune response against these cells. The goal of gene therapy is to enhance the immune response against cancer and overcome the immune suppression so that the cancer cells are killed.
Approaches to Gene Therapy for Cancer
There are three main approaches to gene therapy: 1. augmentation of immunotherapy or chemotherapy, 2. gene replacement therapy and 3. adjuvant therapy (Table 1).
Table 1. Classification of Gene Therapy Approaches
I. Augmentation of immunotherapy or chemotherapy
A. Augmentation of immunotherapy
1. Cytokine gene transfer
i. In-vivo
ii. Ex-vivo2. Vaccination
i. Tumor specific antigen.
ii. Costimulatory molecule
iii. Foreign antigenB. Augmentation of chemotherapy
1. Drug sensitization
2. Drug resistance
II. Gene replacement
A. Replace tumor suppressor gene
B. Inhibit an oncogene1. Antisense cDNA
2. Gene that regulate oncogene
III. Adjuvant therapy
A. Adjuvant with chemotherapy
B. Adjuvant with radiation therapy
C. Adjuvant with surgery
Immunotherapy is augmented by cytokine gene transfer. Cytokine gene transfer is performed in-vivo where tumor cells or immune cells are transfected in the body which up regulates the inunune response. Ex-vivo cytokine gene transfer is performed after immune cells or cancer cells are removed from the body, and then these cells are placed back in the body. Gene therapy can also be used to vaccinate tire body against-tumors. A tumor specific antigen gene is injected into a cancer cell which helps the body recognize the tumor cell and reject it. The problem with head and neck cancer is that there are no reliably known tumor specific antigens. Another vaccination approach is to add a gene that can produce a costimulatory molecule. A costimulatory molecule will help aid the immune response against the tumor cell by helping immune cells recognize the tumor. A third approach is simply adding a gene that produces a foreign antigen. If a foreign antigen gene is added into a tumor cell, the body forms an immune response against the growth of the tumor cell.
Gene therapy can be used to augment chemotherapy by either drug sensitization or resistance approach. In the drug sensitization approach, a gene is transfected to convert a pro-drug into its active metabolite. This allows for drug conversion and a high level of active drug only in the tumor bed. One example is the herpes simplex virus thymadine kinase (TK) gene, which converts gangcyclovir into its cytotoxic triphosphate. Another way to augment chemotherapy is to use a drug resistance approach. A drug resistant gene is added into cells that are sensitive to chemotherapy, such as hematopoetic stem cells, so that they can resist chemotherapy. Therefore, higher dose; of chemotherapy can -be used since-the most sensitive cells are now resistant to the toxicity chemotherapy.
Gene therapy can be used as a replacement approach. The simplest approach is just to replace the wild type tumor suppressor gene. This results in suppression of tumor growth when this cell produces the gene product. Alternatively, an oncogene can be inhibited by either transfecting the antisnse cDNA so this binds to the mRNP of the oncogene or a gene can be added that regulates and inhibits the transcription of an oncogene.
Gene therapy can be used in adjuvant therapy with conventional therapy chemotherapy, radiotherapy or surgery Gene therapy using p53 has been studied as intraoperative therapy in head and neck cancer. After the p53 gene was injected in the tumor bed after resection no toxicities were noted. A 27% survival after 18 months was reported. Currently several protocols are being developed it which genes are transfected simultaneously with chemotherapy and radiation therapy to determine if an enhanced response occurs.
Genes Used in Therapy
There are many genes that can be used for gene therapy (Table 2). However, only genes that are currently being tested it HNSCC will be explained in detail Interleukin-2 (IL-2), interleukin-12 (IL12), interferon-y and GM-CSF are cytokines that enhance the immune response against tumors. The HLA-B7 gene is a costimulatory molecule injected into tumors that helps the immune system recognize antigens on the tumor cells and therefore kill the tumor cells.
Table 2. Genes used for therapy in head and neck cancer
Augmentation of Immunotherapy
B7-1
IL-2
IL-12
IFN-y
GM-CSF
Augmentation of Chemotherapy
(Prodrug)
HSV TK
Replacement Therapy
p53
EIA
Adjuvant Therapy to Chemotherapy
p53
The herpes simplex virus-thymadine kinase (TK) gene converts the anti-viral agent, gangcyclovir, into its toxic triphosphate metabolite. After TK is transfected into tumor cells and gangcylclovir is given to a patient, the activated drug kills not only the tumor cells, but also allows for a killing of surrounding tumor cells, "bystander effect", because of high levels of the activated drug is produced locally.
Two genes have been used in replacement therapy, p53 and ElA. P53 is a tumor suppressor gene. The p53 gene has been transfected into tumor cells and has been shown to suppress growth. The EIA adenovirus gene functions to inhibit tumor growth by several pathways. In HER-2/ neu overexpressing tumors, ElA transcriptionally down-regulates HER-2/ neu expression. Stable transfection of ElA in malignant cells expressing basal levels of HER-2/neu results in reversion to an epithelial phenotype, loss of anchorage independent growth, and decreased tumorgenicity in nude mice.
Vector to Transfect Genes into Cells
A vector is used so that a gene can be transfected into a cell and the gene can
produce its protein product. The idea vector would have a high efficiency (100% of cells get transfected), a high specificity, (only tumor cells receive the gene) and low toxicity. No known vector meets all of these criteria. Vectors classified as viral and non-viral. Although many viral vectors are currently being studied adenoviruses and retroviruses are most commonly used. These viruses are attenuated to transfect genes, but they cannot replicate nor cause an infection Non-viral vectors, such as liposomes protein and liposome-protein combinations, are easy to manufacture and use. Furthermore, non-viral vector; avoid the biohazard risks associated with viruses. However, non-viral vectors have no cell specificity and usually lower transfection efficiency.
Multiple gene delivery methods are being investigated. The majority of studies have performed intratumoral injections However, intravascular delivery, such as hepatic artery injections, systemic deliver; and immunogenic site injections are also being studied. Since head and neck cancer is a local regional problem and access to most lesions is a relatively simply procedure using intratumoral injections the head and neck area is a common site studied using gene therapy.
Clinical Trials in Gene Therapy
Currently, 173 gene therapy trials are approved in the United States in which 138 are approved for cancer. There are 15 trials using eight different genes that are being tested in head and neck cancel (Table 3).
Table 3. Clinical gene therapy trials in head and neck cancer.
I. p53/adenovirus
Phase 1: MD Anderson (Introgen)
Phase I: Univ Pitt (Shering-Plough)
Phase II: multicenter & WSU
(RPR-Gencell)
Phase III: Multicenter and WSU
(p53 vs. methotrexate)
II. BL7/liposome
Phase I: Univ. Cincinnati
(Vical)
Phase II: multicenter (Vical)
III. EIA/liposome
Phase 1: WSU, Rush, (Targeted Genetics Corp)
Phase II: multicenter (Targeted
Genetics Corp)
IV IL-2/liposome
Phase I: Johns Hopkins
Phase II: ongoing
V TK/liposome
Phase I: Johns Hopkins
VI. IL-12/fibroblast
Phase I: Univ of Pitt
Phase II: multicenter
VII. IFN-y/PVP
Phase I: Univ Penn (GeneMedicine)
VIII. GM-CSF/Adenovirus
Phase 1: ongoing
All of these trails are examining recurrent HNSCC using intratumoral injections. Over one billion dollars have been invested in gene therapy to date by pharmaceutical and bio-technical companies. The gene therapy approaches have been tested in-vitro and in-vivo using animal models and have demonstrated tumor suppression or killing.
Nine phase 1 and six phase 2 trials have bee completed or are in progress in head and neck cancer. The Department of Otolaryngology has participated in four of these trials. A review of all gene therapy trials can be found at the Office of Recombinant DNA Activities/NIH (www.nah.gov/od/orda/docs.htm) and www.wiley.com/genmed/clinical. The first reported phase 1 trial in head and neck cancer utilized the p53 gene and an attenuated adenovirus vector. This agent showed low toxicity and some anti-tumor activity. The BL7/liposome, EIA/ liposome, IL-2/adenovirus and IL-12/fibroblast phase 1 trials demonstrated a low toxicity and some responses in patients with recurrent HNSCC. The maximum tolerated dose was not achieved in many of these trials since no significant toxicity was observed at the highest dose tested. TK, interferon-y and GM-CSF trials are currently in the phase 1.
Two p53-adenovirus phase 2 trial demonstrated a 6% response rate with a 30% anti-tumor effect. Anti-tumor effect was designated as stabilization of disease. Four other phase 2 trials using the BL7/ liposome, ElA/liposome, IL-2/adenovirus and EL- 12/fibroblast are ongoing. The EIA phase 2 trial was developed at Wayne State University.
Pitfalls of Gene Therapy
Gene therapy has many limitations that will have to be addressed with further research. Transfection efficiency can be increased by developing improved vectors. The dose and dosing schedule also needs to be optimized. Since intratumoral delivery treats only gross disease, systemic delivery methods have to be developed to treat microscopic and metastatic tumors.
Summary
Gene therapy is in its infancy and further phase 2 & 3 testing needs to be completed. Pharmaceutical companies have invested nearly two billion dollars in gene therapy because it has shown activity in both invitro and in-vivo models. It is not yet known whether gene therapy will provide the ultimate benefit, which is improved survival. Gene therapy may produce other effects, such as stabilization of disease and improved quality of life. Although promising data has been reported, no agent has received FDA approval to date.