What are the strategies for gene therapy in cancer?

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What are the strategies for gene therapy in cancer?

What are the strategies for gene therapy in cancer?

Gene therapy involves modifying gene expression or regulating biological features of tissues through specific genetic material.

When peptides cannot be used for recombinant therapy due to issues like bioavailability, stability, toxicity, or production costs, gene therapy may present a viable solution.

Since its emergence in the early 1990s, gene therapy has rapidly gained ground among today’s prevalent treatment methods.

While initial research was specifically aimed at treating hereditary diseases, today, various conditions such as neurodegenerative diseases, rheumatoid arthritis, cardiovascular diseases, and infectious diseases are also under investigation.

The primary goal of gene therapy is to provide a copy of one or more impaired genes, either enhancing the therapeutic gene expression for a disease or suppressing the impaired gene expression.

With the identification of genes that contribute to cancer formation, research and clinical studies on gene therapy for cancer are gaining momentum. For many different diseases, gene therapy could be an alternative to current treatment approaches.

Hence, gene therapy products with safe delivery pathways ensuring genetic material stability will play a crucial role in the prevention and treatment of various diseases.

Gene therapy involves delivering genes (DNA, RNA, mRNA, siRNA, etc.) to patients to repair damaged genes causing diseases. The drugs used in gene therapy are advanced technical products with therapeutic, preventive, or diagnostic purposes. They repair tissue damage, supplement deficiencies to maintain bodily functions, and prevent unwanted gene expressions. According to the European Medicines Agency (EMA), gene therapy medicinal products (GTMP) are classified as advanced therapy medicinal products. Gene therapy aims to change gene expression, altering the biological characteristics of living cells to achieve therapeutic goals.

Gene therapy can operate through various mechanisms:

Replacing mutated disease-causing genes with healthy copies
Inhibiting the expression of mutated genes
Silencing excess genes
Replacing missing genes
Delivering therapeutic genes to target tissues

Cancer is a significant public health issue globally. It’s reported that around 10 million deaths in 2020 were related to cancer. Cancer is a heterogeneous and complex disease. It involves uncontrollable proliferation of some cells in the body, invading other tissues, and presents significant individual differences. Treatment plans depend on the type, staging, prognosis, and the individual, including chemotherapy, surgery, radiotherapy, hormone therapy, photodynamic therapy, hyperthermia, immunotherapy, stem cell transplantation, and targeted therapy. Gene therapy differs from traditional treatments and tends to have fewer side effects. Strategies for gene therapy in cancer can be classified into suicide gene therapy, tumor suppressor gene activation, immunotherapy, inhibition of oncogene activation, and anti-angiogenesis gene therapy.

What are the strategies for gene therapy in cancer?

Figure 1: Application of Gene Therapy in Cancer Treatment

Suicide Gene Therapy
Tumor Suppressor Gene Activation
Immunotherapy
Inhibition of Oncogene Activation
Anti-Angiogenesis Gene Therapy

1) Suicide gene therapy

Toxic chemotherapeutic agents are administered to cells in the form of prodrugs, and the genes encoding the enzymes that activate the prodrugs are transferred into the cancer cells, thus killing the cancer cells using the active drug with toxic effects. The most studied suicide gene/prodrug system in this field is the herpes simplex virus thymidine kinase/ganciclovir system. This enzyme converts ganciclovir into ganciclovir monophosphate by catalyzing the phosphorylation of ganciclovir. The product is converted into ganciclovir triphosphate by cellular enzymes. The final product causes chain termination by binding to DNA. Cell death. Unlike ganciclovir, ganciclovir triphosphate cannot pass through the cell membrane but remains within the cell. In recent studies, herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/GCV) gene therapy combined with radiofrequency hyperthermia stands out as a new treatment technology, demonstrating its efficacy in rats and mice. Has good anti-cancer effect.

Suicide gene therapy using mesenchymal stem cells (MSCs) as a carrier has received widespread attention in tumor treatment, especially in the treatment of solid tumors, demonstrating significant migration properties against tumors and their metastasis. Mesenchymal stem cells can be easily isolated from bone marrow and adipose tissue, and transduced mesenchymal stem cells produce cytotoxic metabolites that can kill cancer cells. In order to obtain transduced cells expressing high levels of therapeutic genes, MSCs must be modified. Finally, after the MSCs are injected and transduced into the cancer cells, an inactive prodrug is given. This is a promising step for larger clinical studies, but it also needs to be considered that the active drug may also affect neighboring cells.

2)Tumor suppressor gene activation

Other genes responsible for converting healthy cells into cancer cells are recessive tumor suppressor genes, which, as their name suggests, play a role in the inactivation of cancer pathogenesis. Rb gene, p53 gene, cdk inhibitor and BRCA-1/2 are major tumor suppressor genes, which prevent uncontrollable proliferation of cells by inhibiting the cell cycle or triggering apoptosis. p53 is the most studied tumor suppressor gene, responsible for detecting DNA damage and inducing apoptosis. Inhibition of tumor growth and tumor regression following p53 transfection have been observed in animal models. There are a few studies that have evaluated the efficacy and safety of recombinant human adenovirus-mediated p53 in combination with chemoradiotherapy and reported promising results in terms of survival time.

In addition, according to the results of a meta-analysis, p53 is an effective therapy for cervical cancer, and its cell cycle and apoptosis functions play an important role in preventing tumor development. The tumor suppressive mechanisms of p53 include cell cycle arrest and apoptosis, which play an important role in preventing tumor progression. The development of gene therapies targeting p53 expression and regulation is a promising cancer treatment strategy. In cancer, in addition to p53, the role of tumor suppressor genes in gene therapy is also under investigation. After transfection of the p21 tumor suppressor gene, the tumor volume of breast cancer rats was significantly reduced. In another study, the P14ARF tumor suppressor gene prevented apoptosis in prostate cancer cells. In addition, the anti-tumor effect of cisplatin combined with p53, p16, and PTEN gene therapy for the treatment of bladder cancer is better than classic cisplatin treatment. In recent years, coadministration of functional complementary tumor suppressor genes has been shown to have an important role in triggering apoptosis.

3) Immunotherapy

To date, most cancer gene therapy research has been conducted with immunotherapy. Cancer immunotherapy relies on a patient’s immune system to recognize and attack cancer cells. Cancer cells are immunogenic, and tumor antigens are intracellular molecules. Therefore, T cell-mediated cellular immunity is more important than B cell-mediated humoral immunity. However, conventional immune responses are insufficient to eliminate tumor cells. The ability of cancer cells to evade the immune system relies on the secretion of immunosuppressive factors, expression of antigens, and downregulation of major histocompatibility complex molecules. With these in mind, gene therapy approaches have been developed that target the synthesis of multiple genes encoding immunostimulatory factors, major histocompatibility complexes, and costimulatory molecules. Cytotoxicity is thought to play an important role in anticancer immunity. In cancer, different immune molecules are used to trigger anti-tumor immune responses, which are effective in gene immunotherapy. For example, genes encoding various cytokines are transferred into cancer cells in vivo or in vitro. Cancer cells synthesize proteins encoded by genes that are transferred to the tumor microenvironment. These immune system stimulating factors modulate the tumor microenvironment.

In the first protocols for cancer gene therapy, newly isolated tumor cells from patients were modified with cytokines. IL-2 has been used to treat head and neck cancer. IL-2 produced by activated T lymphocytes stimulates natural killer cells and T lymphocytes. This method provides tumor inhibition in rat models. In recent years, with the advancement of immunotherapy technology, the determination of tumor-associated antigens and the mechanism of anti-tumor immune responses have been gradually clarified. Cloning tumor-associated antigens from human tumor cells is one of the most important steps in developing cancer gene therapy. In this way, tumor cells can be recognized by CD8+ or CD4+ T lymphocytes. This development supports the idea that T lymphocyte responses to tumor cell antigens can be molecularly enhanced.

Chimeric Antigen Receptor-T Cell Therapy

Chimeric antigen receptor (CAR)-T cell therapy is a promising method for tumor immunotherapy using genetically modified leukocyte T cells. Chimeric antigen receptors are synthetic receptors designed for their cytotoxic effects on tumor cells and are composed of four main components, namely, the extracellular target antigen-binding domain, the hinge region, the transmembrane domain and one or more cellular Inner signal domain.

What are the strategies for gene therapy in cancer?

Figure 2 Structure of chimeric antigen receptor

Each of these elements has a different function. According to the structure of the intracellular domain, CAR can be divided into five generations. The first generation contains only one intracellular signaling domain, CD3ζ, which is insufficient to trigger CAR-T cell expansion and produce sustained anti-tumor activity. The second and third generations use first-generation CARs as the backbone and contain one or two additional costimulatory signaling domains, such as CD28 or 4-1BB (CD137), CD27 and OX40 (CD134). Fourth-generation CAR uses second-generation constructs modified to contain inducible expression cassettes containing transgenic proteins such as IL-12 cytokines. The fifth-generation CAR consists of a new costimulatory domain that can activate some specific signaling pathways, such as STAT3/5.

What are the strategies for gene therapy in cancer?

Figure 3 Structure of fifth-generation CAR-T cells

Unlike T cell receptors, CAR recognizes and binds to specific proteins or antigens on tumor cells in the major histocompatibility complex and is not restricted by tumor cell killing. It allows T cells to bind to the surface of target cells through single-chain variable fragments. Antigens that effectively prevent immune evasion by downregulating the expression of major histocompatibility complex in tumor cells.

Each CAR-T cell therapy is designed to fight a specific cancer antigen, by collecting T cells from a patient and re-engineering them in the lab, converting them into CARs that are cytotoxic to cancer cells. The T cells are then returned to the patient.

What are the strategies for gene therapy in cancer?

Figure 4 Schematic diagram of CAR-T cell preparation and processing

These approaches facilitate harnessing the therapeutic potential of CAR-T cells by blocking negative factors, such as immune checkpoint molecules, in CAR-T cells to provide effector responses. Among other things, increasing the complexity of CAR design and gene editing of T cells may amplify the risks associated with CAR-T cell therapy, and modifications of CAR-T cells may increase the impact of these adverse events. Therefore, more research is needed in the future. In 2017, the U.S. Food and Drug Administration (FDA) approved chimeric antigen receptors using two synthetically designed receptors, Tisagenelecleucel (Kymriah) and Aksicabtageneeciloleucel (Yescarta), as a new type of cellular immunotherapy. With the advancement of biotechnology and medicine, CAR-T cell therapy will become a treatment method for many types of cancer.

4) Inhibit oncogene activation

Cancer is a disease caused by damage to genes encoding molecules involved in intracellular signaling pathways. There is a physiological balance in the human body. In cancer, this balance is disrupted and cells begin to proliferate uncontrollably. If proto-oncogenes mutate for any reason, they are converted into oncogenes. In addition, some genes prevent tumor formation and are called tumor suppressors or suppressor genes.

Tumor suppressors can also be thought of as braking systems that provide balance. With the onset of mapping of the human genome, many oncogenes and tumor suppressor genes have been detected. These genes have been implicated in the development of various tumors of epithelial origin, such as lung, colon, and breast cancers. They influence cell division cycle genes and are active in intracellular pathways involved in cell growth control. Oncogene products may mimic the effects of growth factors or their receptors.

Because oncogenes are overexpressed in tumors, studies have been conducted to suppress oncogenes. In cases where tumor cells cannot be eliminated, continued use of oncogene antisense molecules may be required to suppress tumors. Nucleic acid-based therapeutic strategies have received significant attention over the past two decades and have become the mainstay of treatment for antisense oligonucleotide-mediated regulation of gene expression.

5) Anti-angiogenic gene therapy

Tumor tissue proliferation is related to angiogenesis. Angiogenesis provides necessary nutrients for the growth of cancer cells. One of the most important characteristics of tumor tissue growth is the increase in blood flow through angiogenesis. Therefore, one of the main goals of anti-angiogenic cancer therapy is the inhibition of angiogenesis inducers (VEGF, angiopoietin) or the use of angiogenesis inhibitors such as angiostatin, endostatin, IL-12 and p53.

Over the past two decades, several anti-angiogenic monoclonal antibodies (mAbs) that block VEGF signaling have been approved by the FDA, the most commonly used being bevacizumab. Although important results have been achieved with angiopoietin-inhibiting monoclonal antibodies, their short biological half-lives and high costs pose significant challenges to clinical practice. Although exosomes were originally considered a cellular waste product, they are beginning to be used as a system of nanoscale homogeneous vesicles secreted by cells. Exosomes may be important in future gastric cancer treatments as a drug or gene delivery system. meaning.

The antitumor effect of angiostatic drugs is closely related to several factors, including the angiogenic status of the tumor, endothelin heterogeneity, and the potential and biochemical activity of the drug used. Taking these factors into account, a study of a combination of drugs and genes in the treatment of anti-angiogenic cancers was designed, using a polyethylenimine delivery system to achieve effective results. The main advantage of this cancer treatment strategy is that vascular endothelial cells are easily accessible and applicable to different types of cancer.

Figure 5 Timeline of global approval of gene therapy drugs for cancer treatment

Cancer is a complex disease that is difficult to treat, and with the development of new technologies such as gene therapy, cancer will eventually cease to be a problem. Although the results of pure gene therapy such as tumor suppressor gene replacement and oncogene inhibition with antisense oligonucleotides have not yet reached the ideal level, this approach is being combined with existing treatments, especially suicide genes and chemotherapy. The combination offers great hope for the future. As the mechanisms of cancer are better understood to identify the right treatments and targets, new approaches such as gene therapy will become more popular than traditional treatments.

Gene therapy has made good progress in cancer treatment over the years, with many drugs approved and others still in trials. Relatively speaking, gene therapy for cancer treatment is safer and has more tolerable side effects. In the future, molecular analysis will be used for gene therapy in cancer treatment.

As genes are screened and the molecular basis of disease is better elucidated, it is thought that gene therapy will soon become a viable approach for clinical cancer treatment. In the future, the widespread application of genomic analysis and cellular immune assessment will help to better select the most appropriate gene therapy for patients, which will change the future of cancer treatment from broad cancer treatment strategies based on tumor size, nature and location. to strategies based on patient-specific genomic composition, immune status, and genetic profile. To achieve gene therapy efficiency, i.e., therapeutic benefit with low side effects, it is also necessary to fully understand the barriers to therapeutic intervention and develop methods to inhibit these barriers.