1. Introduction

Due to its annual millions of fatalities, cancer remains a major global health problem.[3] Many cancers remain difficult to cure even with advancements in medical treatments including immunotherapy, chemotherapy, and surgery, especially in cases where the cancer was discovered late or did not respond to normal care. A promising new approach to treating cancer is made possible by gene editing techniques like HDR, Base Editing, and Prime Editing, which allow precise genetic alterations to repair mutations or boost the immune system. However, there are ethical and societal issues along with technical challenges with these technologies. To develop more effective and targeted clinical cancer methods, this review evaluates the potential benefits, drawbacks, and future directions of various technologies as they relate to cancer treatment.

2. Homology Directed Repair (HDR)

2.1. Invention History and Principle Elucidation

Cells have devised a complex method called homology-directed repair (HDR)[7] to fix double strand breaks in DNA. Although the cell's ability to repair DNA damage was discovered through early molecular biology studies, the specific mechanism of HDR was subsequently discovered. HDR is now highly important in gene therapy, especially when targeting certain genetic mutations for repair utilizing homologous arms, thanks to the development of DNA recombination technology. In the process of editing genes, HDR uses the CRISPR-Cas9 system to accurately insert or replace genes by inducing double-strand breaks at specified sites. This is done by using a pair template. The study conducted by Doudna and Charpentier has led to improvements in the efficiency and accuracy of this technology.

2.2. Efficiency indifferent Cancer/Tumor Treatments

HDR technology has showed great promise in the therapy of cancer, especially in the repair of genes linked to hereditary malignancies. For example, HDR technology can repair mutations in the BRCA1 and BRCA2 genes, which are intimately linked to the development of ovarian and breast cancers, helping to stop the cancer from progressing. Scientists are investigating the application of HDR technology in clinical research to alter particular genes in T cells with the goal of improving their capacity to identify and combat cancer cells. While HDR's editing efficiency may not always match that of the NHEJ pathway, its efficiency has been greatly increased by advancements in the CRISPR-Cas9 system's design[6] and the utilization of more efficient repair templates.[3]These advancements improve the viability of using HDR technology in cancer treatment.

2.3. Challenges

HDR technology faces major clinical challenges, including off-target effects that may lead to unintended genetic mutations. Its efficiency is also relatively low in non-dividing cells, restricting its utility in certain cancer treatments. Another obstacle is the possibility of triggering an immune response against CRISPR-Cas9 because it may have an impact on therapeutic results. Lastly, there's the challenge of safely and effectively delivering edited cells to patients, which is essential to the clinical use of HDR.

2.4. Development Prospects

Future research on HDR technology is expected to focus on enhancing its editing efficiency and precision. Scientists are studying new variants of CRISPR-Cas9,[4] such as high-fidelity Cas9, to reduce off-target editing. At the same time, improvements in the design of sgRNA and repair templates are expected to enhance the editing efficiency of HDR. Moreover, the use of advanced delivery systems, e.g. nanotechnology, can further increase the delivery performance of gene editing. With a deeper understanding of the HDR mechanism, new methods may be developed in the future to activate the HDR pathway and further increase gene-editing efficiency. Although HDR technology shows great potential in cancer treatment. it has yet to overcome existing problems in order to reach widespread clinical use.

3. Base Editing

3.1. Invention History and Principle Elucidation

Base Editing, introduced by David Liu's lab in 2017, expands on traditional CRISPR-Cas9 by enabling direct single-base conversions in the genome without double-strand breaks (DSBs).[2] This technology fuses a Cas9 variant with a cytosine or adenine deaminase, guided by sgRNA to specific DNA sequences. Cytosine Base Editing converts C to T, and adenine Base Editing changes A to G, with controlled deaminase activity to increase editing specificity. The introduction of point mutations that do not contain the DNA repair template provides an important advantage in the research and treatment of genetic diseases associated with single base mutations.

3.2. Efficiency in Different Cancer/Tumor Treatments

Base Editing is still in the early stages of cancer, but it has demonstrated the potential in vitro to repair cancerous mutations such as BRCA1 and BRCA2. It also helps in studying gene functions by precisely adding or removing base mutations, enhancing our understanding of their role in cancer. While its efficacy and accuracy remain to be verified in preclinical and clinical trials, Base Editing has the advantage of avoiding DNA double-strand breaks and relying on repair templates to reduce the risk of off-target editing.[5]

3.3. Challenges

Among the challenges that Base Editing technology is facing in the field of cancer therapy are increased editing efficiency, specificity, and reduction of off-target editing. Although Base Editing reduces the off-target effects of DSBs, the non-specificity of the deaminases can result in modifications to non-target sites, resulting in unexpected genetic mutations. Furthermore, the efficiency of Base Editing is influenced by various factors, including the design of sgRNA, the activity of deaminases, and the type of cell. Therefore, the optimization of the Base Editing system to increase its efficiency and specificity in certain cell types is a current research focus.

3.4. Development Prospects

Future developments in Base Editing might include: improving the efficiency and specificity of Base Editing through protein engineering and optimization of sgRNA; secondly, developing new deaminases to expand the editing range of Base Editing, such as achieving the conversion of other base pairs; and thirdly, exploring the application of Base Editing in cancer immunotherapy, such as enhancing the recognition and killing ability of T cells against tumors by editing T cell receptors; Further preclinical and clinical trials were conducted to evaluate the safety and effectiveness of Base Editing for cancer therapy. As these challenges are overcome, Base Editing is expected to become an important tool in cancer treatment.

4. Prime Editing

4.1. Invention History and Principle Elucidation

Prime Editing is a genetic modification technique developed by David Liu's lab in 2019. This is a major innovation in addition to the traditional CRISPR-Cas9 technology, which is designed to overcome editorial constraints, e.g. dependence on DNA double-strand breaks (DSBs), and relatively poor editing performance. Prime Editing is based on a specific variant of Cas9 called nCas9-HF1 that is designed to cut only one strand of DNA and is fused with reverse transcriptase (RT). This complex is guided to the target DNA sequence and targeted by a sgRNA containing an RNA primer.[1] Once nCas9-HF1 has severed the target DNA strand, the RNA primer is used as a template by RT to reverse the transcription of new DNA sequences into the target site, allowing precise insertion, deletion, or substitution. A major advantage of Prime Editing is that it can perform editing without inducing DNA double-strand breaks, DSBs associated with reducing off-target effects, such as insertion and deletion caused by non-homologous end joining (NHEJ).

4.2. Efficiency in Different Cancer/Tumor Treatments

Prime Editing is still in its early stage of development, but it has demonstrated the potential to repair genetic mutations associated with cancer. For example, researchers have been able to repair in vitro models of certain genetic mutations leading to hereditary cancer using Prime Editing. In addition, Prime Editing has been used to investigate the function of cancer-related genes, enabling researchers to better understand their role in cancer development by accurately inserting or deleting certain DNA sequences. Although Prime Editing is a promising candidate for cancer therapy, but its efficacy and specificity need to be validated by more pre-clinical and clinical trials. At present, the effectiveness and specificity of Prime Editing are affected by a variety of factors, such as sgRNA design, reverse transcriptase activity and cell type.

4.3. Challenges

Among the challenges that Prime Editing is facing in the field of cancer therapy are further improvements in editing efficiency, specificity, and reduction of off-target editing. Although Prime Editing reduces DSBs related off-target effects, non-specific reverse transcriptase activity can result in modifications in non-target sites, resulting in unintended gene mutations. Furthermore, the efficiency of Prime Editing is influenced by various factors, including the design of sgRNA, the activity of reverse transcriptase, and the type of cell. Therefore, optimizing the Prime Editing system to enhance its editing efficiency and specificity in specific cell types is a current research focus.

4.4. Development Prospects

Future development of Prime Editing can be based on the following aspects: first, improving efficiency and precision in Prime Editing by protein engineering and optimization of the design of sgRNA; second, the development of new reverse transcriptases to extend the modification capability of Prime Editing, including more subtle insertion and deletion of DNA sequences; third, using Prime Editing to explore improvements in cancer immunotherapy, such as enhancing the ability of T cells to recognize and destroy by modifying the T cell receptors; lastly, extended preclinical trials and clinical studies for safety and effectiveness in cancer therapy. As these technological challenges slowly but surely work their way into existence, Prime Editing has the potential to become a serious player in the treatment of cancer.

5. Conclusion

HDR, Base Editing, and Prime Editing are breakthroughs within gene editing that may define a new frontier in cancer therapy.[8] While HDR is precise but bounded by the problems of efficiency, Base Editing is accurate but narrower in the scope of genetic variations. Prime Editing, although more complicated and expensive, can extend editing capabilities. Future improvements should enhance the efficiency and precision of all those systems and reduce undesired impacts, hence furthering cancer therapy.[9]Ultimately, the integration of these methods might well provide novel and individualized therapeutic strategies for the patients.