The Role Of CRISPR Gene Editing Within Gene Therapy

Introduction

Gene editing focuses on inserting DNA, deleting it or modifying it, or in some cases even replacing it in the genome of the chosen organism (Richter, 2016). Clustered regular interspaced short palindromic repeats (CRISPR) is seen as an innovating piece of technology and a useful technique within the genetic editing field (Redman et al, 2016). CRISPR can identify DNA, which it will then lock on to and then it will start to make precision cuts, but these precision cuts are conducted by an enzyme (Zhang, 2019). This enzyme is known as Cas9, hence the name CRISPR-Cas9, Cas9 is also seen as a molecular scissor, as it behaves to that of a scissor due to the way it cleaves the target area of the DNA (Ishino,Krupovic and Forterre, 2019). Since Cas9 cleaves the foreign DNA, it is the CRISPR part of CRISPR/Cas9, which behaves like a guide and will guide the Cas9 enzyme to the part of the DNA it needs to cleave (Ishino,Krupovic and Forterre, 2019). So both CRISPR and Cas9 work together to disable a gene or place something new into the area where the Cas9 enzyme has cleaved within the DNA (Ishino,Krupovic and Forterre, 2019). In 1987, Yoshizumi Ishino had initially discovered CRISPR within E.coli. He observed an unusual repetitive DNA sequence within the E.coli genome (Cox, Prevett and Finch, 2019). Furthermore, similar sequence patterns had been seen within other bacteria as well as in halophilic archaea, Franciso Mojica observed this at the University of Alicante in Spain (Cox, Prevett and Finch, 2019). However, it was Jennifer Doudna, who suggested that CRISPR-Cas9 could be utilized for editing genomes (Cox, Prevett and Finch, 2019).CRISPR-Cas9, now has a vital role in gene therapy, as it is currently being used to develop cures for diseases such as cancer or HIV. This essay aims to examine the role of CRISPR within gene therapy by looking at the advantages and drawbacks of trying to use CRISPR/Cas9 to develop a cure for cancer and HIV.

Advantages of using CRISPR/Cas9 within gene therapy

CRISPR/Cas9 is a key tool in gene therapy as it can develop cures for cancer and HIV. In China, at the University of Sichuan, researchers had used CRISPR/Cas9 for a lung cancer sufferer(Castillo, 2019). They injected the patient with the T cells, which had been modified by CRISPR/Cas9 so that the PD-1(programmed cell death) protein had been disabled. This was done by using CRISPR/Cas9 to disable the PDCD1 gene, which codes for the PD1 protein (Castillo, 2019). This was an attempt to raise the immune system's response to the cancer so that it would attack the malignant tumour cells, the attempt here was to get the immune system to fight the cancer (Castillo, 2019).

Alongside this CRISPR/Cas9 is also being used to develop a cure for HIV, this is being done by disrupting the co-receptor known as CCR5, as HIV tends to enter the white blood cells known as CD4 by binding to the CCR5 co-receptor (Xiao et al, 2019). The CCR5 co-receptor is a protein which is found on the surface of the CD4 cells (Ishino, Krupovic and Forterre, 2019). Xiao et al, had utilized CRISPR/Cas9 to target and deactivate the CCR5 gene within human CD34+ HSPCs (Hematopoietic stem and progenitor cells) (Xiao et al, 2019). This had resulted in the CCR5 disruption, which in turn lead to HIV being inhibited. Furthermore, silencing the CCR5 within the secondary repopulating hematopoietic stem cells had proven to be stable, giving a framework to work from to develop an HIV cure by implementing CCR5 modified HSCs (Hematopoietic stem cells) (Xiao et al, 2019).

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Using CRISPR/Cas9 to develop a cure for cancer and HIV places the research and medical community one step forward, as CRISPR/Cas9 provides a framework for them to work from, which would increase the chances of a cure being produced such as the CCR5 modified HSCs. Therefore having an increased chance of a cure being produced due to the framework provided, could improve cancer and HIV patient lifestyles, as they would not have to rely on chemotherapy, anticancer drugs or antiretrovirals. As the side effects of these treatments are very inconvenient and painful at times, so a potential cure from the basis which has been provided by CRISPR/Cas9 can get rid of the pain a typical cancer patient will have to endure or the inconvenience of taking antiretrovirals a HIV patient has to endure.

Disadvantages of CRISPR/Cas9 within gene therapy

However, using CRISPR/Cas9 to develop a cure for cancer can cause cancer instead, which is a major drawback. A study published by Haapaniemi et al, found that using CRISPR/Cas9, had led to the activation of the P53 gene, which is a tumour suppressor (Happaniemi et al, 2019). Novartis had found that using CRISPR/Cas9, to edit the genome of the cells, had led to cells having a dysfunctional P53 (Schmierer, 2019). This meant the proliferation of the cells would not be controlled, leading to the uncontrolled proliferation of cells, forming a tumour and which then becomes cancerous (Schmierer, 2019). A dysfunctional P53 is now responsible for over 50 per cent of ovarian cancer, 43 per cent of colorectal cancer and 38 per cent of lung cancer (Schmierer, 2019). On the other hand, Novartis and Koralinska medical institute found using CRISPR/Cas9 to develop a cure for cancer is inefficient (Schmierer, 2019). As when the cell's genome had been successfully edited by CRISPR/Cas9, the P53 would cause this cell to self-destruct, which is a part of the P53s defence mechanism, making CRISPR/Cas9 inefficient (Haapaniemi et al, 2018). As the edited cell would self-destruct after giving no time to study it. This shows that the P53 makes editing difficult which delays the production for a cure. Furthermore, P53 makes the cell harder to study delaying the possibility of a cure further, as the P53 self-destructs the cell. Studying the cell would indicate how CRISPR/Cas9 edits the cell and how to inhibit the P53, which would make CRISPR/Cas9 more efficient to develop a cure faster. Furthermore, even when CRISPR/Cas9 does make a successful edit, the P53 of the cell becomes faulty causing cancer, making CRISPR/Cas9 infective as well as inefficient. This puts CRISPR/Cas9 in a tough spot, as it has the potential to cure cancer but to cause it too, which shows CRISPR/Cas9 could be doing more harm than good. This puts the usage of CRISPR/Cas9 in a dilemma.

Furthermore using CRISPR/Cas9 to disrupt the CCR5 co-receptor in an attempt to develop a cure for HIV has drawbacks. For example, it does not have an impact on the X4 virus it promotes it, X4 is a strain of HIV, which enters through the co-receptor known as the CXCR4(Ebina, Gee and Koyanagi, 2015). HIV is dual tropic so it can bind to either the CCR5 or the CXCR4 co-receptors in order to enter and infect the CD4 cells (Ebina, Gee and Koyanagi, 2015).However, the X4 virus tends to occur in the later stages of HIV, where the CD4 cell count is lower, having a low CD4 cell count is AIDS(Perelson and Weinberger, 2011).Since CRISPR/Cas9 can inhibit the HIV entry by inhibiting the CCR5 co-receptors, there will be a sharp increase in the X4 virus and AIDS, as HIV will bind to the CXCR4 receptor if the CCR5 receptor is inhibited(Perelson and Weinberger, 2011).Allowing for the X4 virus to occur earlier as CCR5 is inhibited. This shows that trying to inhibit HIV by CRISPR/Cas9 in an attempt to develop a cure for HIV is ineffective and unsafe. As HIV can bind to other receptors such as the CXCR4 on the CD4 cells, this allows for the entry of HIV into the CD4 cells, so the entry of HIV is not entirely inhibited by CRISPR/Cas9 making it ineffective as HIV still enters, infecting the CD4 cell, so HIV does not have much effect in preventing the entry for HIV. Furthermore, as the CCR5 co-receptor is inhibited by CRISPR/Cas9, this leads to the X4 virus becoming more prominent as HIV will start to bind to the CXCR4 receptor more. This would lead to AIDS at a faster rate harming the human body further as it would lead to complications such as being susceptible to more diseases such as toxoplasmosis. Being susceptible to more diseases leads to more damage to the human health making it unsafe as well as unreliable.

Conclusion

Using CRISPR/Cas9, within gene therapy, to develop a cure for HIV and cancer promises a huge benefit, but due to the drawbacks of trying to use CRISPR/Cas9 to develop a cure places the usage of CRISPR/Cas9 within the neutral area in gene therapy. For example, using CRISPR/Cas9 provides a framework for a cure which can then eventually lead to a definitive cure, leading to an improvement of many cancer and HIV patients lives as they would not have to rely on treatment such as chemotherapy and antiretrovirals, and their side effects. So having CRISPR/Cas9 provide this framework to cure diseases is hugely beneficial, as it would lead to a cure, improving lives. However, using CRISPR/Cas9 to develop a cure for these diseases are accompanied by drawbacks. For example, trying to develop a cure for cancer leads to cancer being caused as the P53 gene would be dysfunctional every time a cell is edited. Alongside this using CRISPR/Cas9 to develop a cure for HIV, leads to the same result which is HIV entering and infecting the CD4 cells as HIV enters via alternative co-receptors such as the CXCR4, but as well as entering through other receptors, CRISPR/Cas9 promotes the X4 virus and AIDS due to the inhibition of the CCR5 co-receptor, which leads to further complications. So, in this case, CRISPR/Cas9 is ineffective, inefficient and unreliable in developing a cure for both cancer and HIV. As it just leads to the disease being caused again, doing more harm than good, as it is doing more harm than good, CRISPR/Cas9 can be regarded as dangerous and possibly not suited for gene editing. But using CRISPR/Cas9 to develop cures promises a massive and a long term benefit, but the drawbacks hold CRISPR/Cas9 back to achieve its full potential within gene therapy to produce a cure.