Suicide gene

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In the field of genetics, a suicide gene is a gene that will cause a cell to kill itself through the process of apoptosis (programmed cell death). Activation of a suicide gene can cause death through a variety of pathways, but one important cellular "switch" to induce apoptosis is the p53 protein. Stimulation or introduction (through gene therapy) of suicide genes is a potential way of treating cancer or other proliferative diseases.

Contents

Suicide genes form the basis of a strategy for making cancer cells more vulnerable or sensitive to chemotherapy. The approach has been to attach parts of genes expressed in cancer cells to other genes for enzymes not found in mammals that can convert a harmless substance into one that is toxic to the tumor. [1] Most suicide genes mediate this sensitivity by coding for viral or bacterial enzymes that convert an inactive drug into toxic antimetabolites that inhibit the synthesis of nucleic acid. Suicide genes must be introduced into the cells in ways that ensure their uptake and expression by as many cancer cells as possible, while limiting their expression by normal cells. Suicide gene therapy for cancer requires the vector to have the capacity to discriminate between target and non target cells, between the cancer cells and normal cells. [2]

Apoptosis

Cell death can majorly occur by either necrosis or apoptosis. Necrosis occurs when a cell is damaged by an external force, such as poison, a bodily injury, an infection or getting cut off from blood supply. When cells die from necrosis, it's a rather messy affair. The death causes inflammation that can cause further distress of injury within the body. Whereas, apoptosis causes degradation of cellular components without eliciting an inflammatory response. [3]

Many cells undergo programmed cell death, or apoptosis, during fetal development. A form of cell death in which a programmed sequence of events leads to the elimination of cells without releasing harmful substances into the surrounding. Apoptosis plays a crucial role in developing and maintaining the health of the body by eliminating old cells, unnecessary cells, and unhealthy cells. The human body replaces perhaps one million cells per second. When a cell is compelled to commit suicide, proteins called caspases go into action. They break down the cellular components needed for survival, and they spur production of enzymes known as DNase, which destroy the DNA in the nucleus of the cell. The cell shrinks and sends out distress signals, which are answered by macrophages. The macrophages clean away the shrunken cells, leaving no trace, so these cells do not damage surrounding necrotic cells do. Apoptosis is also essential to prenatal development. For example, in embryos, fingers and toes are initially connected to adjacent digits by tissue. The cells of this connecting tissue undergo apoptosis to produce separate digits. In brain development, initially millions of extra neurons are created. The cells that don't form synaptic connections undergo apoptosis. Programmed cell death is also necessary to start the process of menstruation. That's not to say that apoptosis is a perfect process. Rather than dying due to injury, cells that go through apoptosis die in response to signals within the body. When cells recognize viruses and gene mutations, they may induce death to prevent the damage from spreading. Scientist are trying to learn how they can modulate apoptosis, so that they can control which cells live and which undergo programmed cell death. Anti-cancer drugs and radiation, for example, work by triggering apoptosis in diseased cells. Many diseases and disorders are linked with the life and death of cells—increased apoptosis is a characteristic of AIDS, Alzheimer's, and Parkinson's disease, while decreased apoptosis can signal lupus or cancer. Understanding how to regulate apoptosis could be the first step to treating these conditions. [4]

Too little or too much apoptosis can play a role in many diseases. When apoptosis does not work correctly, cells that should be eliminated may persist and become immortal, for example, in cancer and leukemia. when apoptosis works overly well, it kills too many cells and inflicts grave tissue damage. This is the case in strokes and neurodegenerative disorders such as Alzheimer's, Huntington's, and Parkinson's disease. Also known as programmed cell death and cell suicide. [5]

Applications

Cancer suicide gene therapy

The ultimate goal of cancer therapy is the complete elimination of all cancer cells, while leaving all healthy cells unharmed. One of the most promising therapeutic strategies in this regard is cancer suicide gene therapy (CSGT), which is rapidly progressing into new frontiers. The therapeutic success, in CSGT, is primarily contingent upon precision in delivery of the therapeutic transgenes to the cancer cells only. This is addressed by discovering and targeting unique or / and over-expressed biomarkers displayed on the cancer cells and cancer stem cells. Specificity of cancer therapeutic effects is further enhanced by designing the DNA constructs, which put the therapeutic genes under the control of the cancer cell specific promoters. The delivery of the suicidal genes to the cancer cells involves viral, as well as synthetic vectors, which are guided by cancer specific antibodies and ligands. The delivery options also include engineered stem cells with tropisms towards cancers. Main mechanisms inducing cancer cells' deaths include: transgenic expression of thymidine kinases, cytosine deaminases, intracellular antibodies, telomeraseses, caspases, DNases. Precautions are undertaken to eliminate the risks associated with transgenesis. Progress in genomics and proteomics should help us in identifying the cancer specific biomarkers and metabolic pathways for developing new strategies towards clinical trials of targeted and personalized gene therapy of cancer. By introducing the gene into a malignant tumor, the tumor would reduce in size and possibly disappear completely, provided all the individual cells have received a copy of the gene.

When the DNA sample in the virus is taken from the patient's own healthy cells, the virus does not need to be able to differentiate between cancer cells and healthy ones. In addition, the advantage is that it is also able to prevent metastasis upon the death of a tumor.

As a cancer treatment

Indirect gene therapy Indirect gene therapy.png
Indirect gene therapy

One of the challenges of cancer treatment is how to destroy malignant tumors without damaging healthy cells. A new method that shows great promise for accomplishing this employs the use of a suicide gene. A suicide gene is a gene which will cause a cell to kill itself through apoptosis. Suicide gene therapy involves delivery of a gene which codes for a cytotoxic product into tumor cells. [6] This can be achieved by two approaches, indirect gene therapy and direct gene therapy. Indirect gene therapy employs enzyme-activated prodrug, in which the enzyme converts the prodrug to a toxic substance and the gene coding for this enzyme is delivered to the tumor cells. For example, a commonly studied strategy based on transfection of herpes simplex virus thymidine kinase (HSV-TK) along with administration of ganciclovir (GSV), in which HSK-TK assists in converting GCV to a toxic compound that inhibits DNA synthesis and causes cell death. [7] [6] [8] Whereas, direct gene therapy employs a toxin gene or a gene which has the ability to correct mutated proapoptotic genes, which can in turn induce cell death via apoptosis.  For instance, the most researched immunotoxin for cancer therapy is the diphtheria toxin as it inhibits protein synthesis by inactivating elongation factor 2 (EF-2) which in turn inhibits protein translation, [6] [9] Moreover, p53 is identified to be frequently abnormal in human tumors and studies show that restoring function of p53 can cause apoptosis of cancer cells. [6] Suicide gene therapy is not necessarily expected to eliminate the need for chemotherapy and radiation treatment for all cancerous tumors. The damage inflicted upon the tumor cells, however, makes them more susceptible to the chemo or radiation. This approach has already proven effective against prostate and bladder cancers. The application of suicide gene therapy is being expanded to several other forms of cancer as well. Cancer patients often experience depressed immune systems, so they can suffer some side effects of the use of a virus as a delivery agent.[ citation needed ]

Direct gene therapy Direct gene therapy.png
Direct gene therapy
Bystander effect Bystander effect.png
Bystander effect

Improved vectors

Suicide gene delivery can be broadly classified into three groups which include viral vectors, synthetic vectors and cell-based vectors. [6] The most efficient vehicles for gene delivery are viral vectors. Widely used viruses for gene therapy include retrovirus, adenovirus (Ads), lentivirus and Aden-associated viruses (AAVs). Non-viral vectors like synthetic vectors were used to combat certain disadvantages of viral vectors like immunogenicity, insertional mutagenesis to name a few. Synthetic vectors refer to use of nanoparticles, like gold nanoparticles, to delivery genes to target cells. [10] Lastly, cell-based vectors employ stem cells as carriers of suicide genes. In the last few years, cell-mediated gene therapy for cancer using mesenchymal stem cells (MSCs) was patented. [11]

Bystander effect

The bystander effect (BE) is phenomenon as a result of which it is possible to kill untransfected tumor cells located adjacent to transduced cells in suicide gene therapy. [8] [12] [13] As hundred percent transduction of all tumor cells is very difficult to achieve, BE is critical feature of suicide gene therapy.

Limitations

The drug is supposed to show high specificity towards cancer in order to effective, but studies have shown this to be rarely achieved. Moreover, expression of suicide gene was under control of tumor-specific promoters like human telomerase (hTERT), osteocalcin, carcinoembryonic antigen; however, only hTERT promoter was found to enter clinic trials. [14] This is majorly because of the low transcriptional power of these tumor-specific promoters for suicide gene expression. Additionally, poor accessibility to target cells is an important limitation of suicide gene therapy. Another major hurdle of suicide gene therapy is partial vector specificity to target affected cells. Finally, lack of specific animal models to predict the clinical outcome and other effects of SGT.

Biotechnology

Suicide genes are often utilized in biotechnology to assist in molecular cloning. Vectors incorporate suicide genes for an organism (such as E. coli). The cloning project focuses on replacing the suicide gene by the desired fragment. Selection of vectors carrying the desired fragment is improved since vectors retaining the suicide gene result in cell death.

Related Research Articles

<span class="mw-page-title-main">Apoptosis</span> Programmed cell death in multicellular organisms

Apoptosis is a form of programmed cell death that occurs in multicellular organisms and in some eukaryotic, single-celled microorganisms such as yeast. Biochemical events lead to characteristic cell changes (morphology) and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, DNA fragmentation, and mRNA decay. The average adult human loses between 50 and 70 billion cells each day due to apoptosis. For an average human child between eight and fourteen years old, each day the approximate loss is 20 to 30 billion cells.

<span class="mw-page-title-main">Gene therapy</span> Medical field

Gene therapy is a medical technology that aims to produce a therapeutic effect through the manipulation of gene expression or through altering the biological properties of living cells.

<span class="mw-page-title-main">Tumor suppressor gene</span> Gene that inhibits expression of the tumorigenic phenotype

A tumor suppressor gene (TSG), or anti-oncogene, is a gene that regulates a cell during cell division and replication. If the cell grows uncontrollably, it will result in cancer. When a tumor suppressor gene is mutated, it results in a loss or reduction in its function. In combination with other genetic mutations, this could allow the cell to grow abnormally. The loss of function for these genes may be even more significant in the development of human cancers, compared to the activation of oncogenes.

Gene silencing is the regulation of gene expression in a cell to prevent the expression of a certain gene. Gene silencing can occur during either transcription or translation and is often used in research. In particular, methods used to silence genes are being increasingly used to produce therapeutics to combat cancer and other diseases, such as infectious diseases and neurodegenerative disorders.

<span class="mw-page-title-main">Small interfering RNA</span> Biomolecule

Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded RNA at first non-coding RNA molecules, typically 20–24 base pairs in length, similar to miRNA, and operating within the RNA interference (RNAi) pathway. It interferes with the expression of specific genes with complementary nucleotide sequences by degrading mRNA after transcription, preventing translation. It was discovered in 1998, by Andrew Fire at Carnegie Institution for Science in Washington DC and Craig Mello at University of Massachusetts in Worcester.

An oncolytic virus is a virus that preferentially infects and kills cancer cells. As the infected cancer cells are destroyed by oncolysis, they release new infectious virus particles or virions to help destroy the remaining tumour. Oncolytic viruses are thought not only to cause direct destruction of the tumour cells, but also to stimulate host anti-tumour immune system responses. Oncolytic viruses also have the ability to affect the tumor micro-environment in multiple ways.

Virotherapy is a treatment using biotechnology to convert viruses into therapeutic agents by reprogramming viruses to treat diseases. There are three main branches of virotherapy: anti-cancer oncolytic viruses, viral vectors for gene therapy and viral immunotherapy. These branches use three different types of treatment methods: gene overexpression, gene knockout, and suicide gene delivery. Gene overexpression adds genetic sequences that compensate for low to zero levels of needed gene expression. Gene knockout uses RNA methods to silence or reduce expression of disease-causing genes. Suicide gene delivery introduces genetic sequences that induce an apoptotic response in cells, usually to kill cancerous growths. In a slightly different context, virotherapy can also refer more broadly to the use of viruses to treat certain medical conditions by killing pathogens.

<span class="mw-page-title-main">TRAIL</span> Mammalian protein

In the field of cell biology, TNF-related apoptosis-inducing ligand (TRAIL), is a protein functioning as a ligand that induces the process of cell death called apoptosis.

<span class="mw-page-title-main">Short hairpin RNA</span> Type of RNA

A short hairpin RNA or small hairpin RNA is an artificial RNA molecule with a tight hairpin turn that can be used to silence target gene expression via RNA interference (RNAi). Expression of shRNA in cells is typically accomplished by delivery of plasmids or through viral or bacterial vectors. shRNA is an advantageous mediator of RNAi in that it has a relatively low rate of degradation and turnover. However, it requires use of an expression vector, which has the potential to cause side effects in medicinal applications.

<span class="mw-page-title-main">Viral vector</span> Biotechnology to deliver genetic material into a cell

Viral vectors are tools commonly used by molecular biologists to deliver genetic material into cells. This process can be performed inside a living organism or in cell culture. Viruses have evolved specialized molecular mechanisms to efficiently transport their genomes inside the cells they infect. Delivery of genes or other genetic material by a vector is termed transduction and the infected cells are described as transduced. Molecular biologists first harnessed this machinery in the 1970s. Paul Berg used a modified SV40 virus containing DNA from the bacteriophage λ to infect monkey kidney cell maintained in culture.

Retinal gene therapy holds a promise in treating different forms of non-inherited and inherited blindness.

<span class="mw-page-title-main">Vectors in gene therapy</span>

Gene therapy utilizes the delivery of DNA into cells, which can be accomplished by several methods, summarized below. The two major classes of methods are those that use recombinant viruses and those that use naked DNA or DNA complexes.

Anticancer genes exhibit a preferential ability to kill cancer cells while leaving healthy cells unharmed. This phenomenon is achieved through various processes such as apoptosis following a mitotic catastrophe, necrosis, and autophagy. In the late 1990s, extensive research in the field of cancer cells led to the discovery of anticancer genes. Mutations in these genes due to base substitutions leading to insertions, deletions, or alterations in missense amino acids can cause frameshifts, thereby altering the protein. A change in gene copy number or rearrangements is also essential for deregulating these genes. The loss or alteration of these anticancer genes due to mutations or rearrangements may lead to the development of cancer.

<span class="mw-page-title-main">Necroptosis</span> Programmed form of necrosis, or inflammatory cell death

Necroptosis is a programmed form of necrosis, or inflammatory cell death. Conventionally, necrosis is associated with unprogrammed cell death resulting from cellular damage or infiltration by pathogens, in contrast to orderly, programmed cell death via apoptosis. The discovery of necroptosis showed that cells can execute necrosis in a programmed fashion and that apoptosis is not always the preferred form of cell death. Furthermore, the immunogenic nature of necroptosis favors its participation in certain circumstances, such as aiding in defence against pathogens by the immune system. Necroptosis is well defined as a viral defense mechanism, allowing the cell to undergo "cellular suicide" in a caspase-independent fashion in the presence of viral caspase inhibitors to restrict virus replication. In addition to being a response to disease, necroptosis has also been characterized as a component of inflammatory diseases such as Crohn's disease, pancreatitis, and myocardial infarction.

Adenovirus varieties have been explored extensively as a viral vector for gene therapy and also as an oncolytic virus.

Directed enzyme prodrug therapy (DEPT) uses enzymes artificially introduced into the body to convert prodrugs, which have no or poor biologically activity, to the active form in the desired location within the body. Many chemotherapy drugs for cancer lack tumour specificity and the doses required to reach therapeutic levels in the tumour are often toxic to other tissues. DEPT strategies are an experimental method of reducing the systemic toxicity of a drug, by achieving high levels of the active drug only at the desired site. This article describes the variations of DEPT technology.

<span class="mw-page-title-main">Lentiviral vector in gene therapy</span>

Lentiviral vectors in gene therapy is a method by which genes can be inserted, modified, or deleted in organisms using lentiviruses.

DNA-directed RNA interference (ddRNAi) is a gene-silencing technique that utilizes DNA constructs to activate an animal cell's endogenous RNA interference (RNAi) pathways. DNA constructs are designed to express self-complementary double-stranded RNAs, typically short-hairpin RNAs, that bring about the silencing of a target gene or genes once processed. Any RNA, including endogenous messenger RNA (mRNAs) or viral RNAs, can be silenced by designing constructs to express double-stranded RNA complementary to the desired mRNA target.

Anti-miRNA oligonucleotides have many uses in cellular mechanics. These synthetically designed molecules are used to neutralize microRNA (miRNA) function in cells for desired responses. miRNA are complementary sequences to mRNA that are involved in the cleavage of RNA or the suppression of the translation. By controlling the miRNA that regulate mRNAs in cells, AMOs can be used as further regulation as well as for therapeutic treatment for certain cellular disorders. This regulation can occur through a steric blocking mechanism as well as hybridization to miRNA. These interactions, within the body between miRNA and AMOs, can be for therapeutics in disorders in which over/under expression occurs or aberrations in miRNA lead to coding issues. Some of the miRNA linked disorders that are encountered in the humans include cancers, muscular diseases, autoimmune disorders, and viruses. In order to determine the functionality of certain AMOs, the AMO/miRNA binding expression must be measured against the expressions of the isolated miRNA. The direct detection of differing levels of genetic expression allow the relationship between AMOs and miRNAs to be shown. This can be detected through luciferase activity. Understanding the miRNA sequences involved in these diseases can allow us to use anti miRNA Oligonucleotides to disrupt pathways that lead to the under/over expression of proteins of cells that can cause symptoms for these diseases.

Adeno-associated virus (AAV) has been researched as a viral vector in gene therapy for cancer treatment as an oncolytic virus. Currently there are not any FDA approved AAV cancer treatments, as the first FDA approved AAV treatment was approved December 2017. However, there are many Oncolytic AAV applications that are in development and have been researched.

References

  1. ↑ Karjoo Z, Chen X, Hatefi A (April 2016). "Progress and problems with the use of suicide genes for targeted cancer therapy". Advanced Drug Delivery Reviews. 99 (Pt A): 113–128. doi:10.1016/j.addr.2015.05.009. PMC   4758904 . PMID   26004498.
  2. ↑ "Suicide gene". MedicineNet. Retrieved 15 November 2014.
  3. ↑ Fink SL, Cookson BT (April 2005). "Apoptosis, pyroptosis, and necrosis: mechanistic description of dead and dying eukaryotic cells". Infection and Immunity. 73 (4): 1907–1916. doi:10.1128/IAI.73.4.1907-1916.2005. PMC   1087413 . PMID   15784530.
  4. ↑ "What is apoptosis?". HowStuffWorks. 2010-03-31. Retrieved 15 November 2014.
  5. ↑ "Apoptosis definition". Medical Dictionary: Definitions of Popular Terms Defined on MedTerms. Medicinenet.com. Retrieved 15 November 2014.
  6. 1 2 3 4 5 Navarro SA, Carrillo E, Griñán-Lisón C, Martín A, Perán M, Marchal JA, Boulaiz H (September 2016). "Cancer suicide gene therapy: a patent review". Expert Opinion on Therapeutic Patents. 26 (9): 1095–1104. doi:10.1080/13543776.2016.1211640. PMID   27424657. S2CID   1495377.
  7. ↑ Düzgüneş N (2019). "Origins of Suicide Gene Therapy". In Düzgüneş N (ed.). Suicide Gene Therapy: Methods and Protocols. Methods in Molecular Biology. Vol. 1895. New York, NY: Springer. pp. 1–9. doi:10.1007/978-1-4939-8922-5_1. ISBN   978-1-4939-8922-5. PMID   30539525. S2CID   54473687.
  8. 1 2 Vassaux G, Martin-Duque P (April 2004). "Use of suicide genes for cancer gene therapy: study of the different approaches". Expert Opinion on Biological Therapy. 4 (4): 519–530. doi:10.1517/14712598.4.4.519. PMID   15102601. S2CID   29442217.
  9. ↑ Lin B, Gao A, Zhang R, Ma H, Shen H, Hu Q, et al. (August 2015). "Use of a Novel Integrase-Deficient Lentivirus for Targeted Anti-Cancer Therapy With Survivin Promoter-Driven Diphtheria Toxin A". Medicine. 94 (31): e1301. doi:10.1097/MD.0000000000001301. PMC   4616595 . PMID   26252309.
  10. ↑ Vago R, Collico V, Zuppone S, Prosperi D, Colombo M (September 2016). "Nanoparticle-mediated delivery of suicide genes in cancer therapy". Pharmacological Research. 111: 619–641. doi:10.1016/j.phrs.2016.07.007. PMID   27436147.
  11. ↑ Moradian Tehrani R, Verdi J, Noureddini M, Salehi R, Salarinia R, Mosalaei M, et al. (May 2018). "Mesenchymal stem cells: A new platform for targeting suicide genes in cancer". Journal of Cellular Physiology. 233 (5): 3831–3845. doi:10.1002/jcp.26094. PMID   28703313. S2CID   46865654.
  12. ↑ Hossain JA, Marchini A, Fehse B, Bjerkvig R, Miletic H (January 2020). "Suicide gene therapy for the treatment of high-grade glioma: past lessons, present trends, and future prospects". Neuro-Oncology Advances. 2 (1): vdaa013. doi:10.1093/noajnl/vdaa013. PMC   7212909 . PMID   32642680.
  13. ↑ Singhal, Sunil; Kaiser, Larry R. (1998-07-01). "Cancer Chemotherapy Using Suicide Genes". Surgical Oncology Clinics of North America. Cancer Gene Therapy. 7 (3): 505–536. doi: 10.1016/S1055-3207(18)30258-8 . ISSN   1055-3207. PMID   9624215.
  14. ↑ Saeb, Sepideh; Assche, Jeanne Van; Loustau, Thomas; Rohr, Olivier; Wallet, Clémentine; Schwartz, Christian (2022-03-01). "Suicide gene therapy in cancer and HIV-1 infection: An alternative to conventional treatments". Biochemical Pharmacology. 197: 114893. doi: 10.1016/j.bcp.2021.114893 . ISSN   0006-2952. PMID   34968484. S2CID   245548079.
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