Recent research highlights the system’s effectiveness in identifying and validating genomic targets with tumorigenic potential, as well as its therapeutic applications (16, 17). Additionally, CRISPR/Cas9 shows promise in cancer immunotherapy, enhancing genome editing specificity and triggering a robust anti-tumor response (10). Moreover, its applications extend to breast cancer diagnostics, disease modeling, and treatment, including gene therapy development and malignant cell reprogramming (18). Collectively, these findings suggest the significant promise of CRISPR/Cas9 in breast cancer research and treatment.

CRISPR/Cas9 technology has shown promise in the treatment of lung cancer, with applications including gene therapy, identification of target genes, and construction of animal tumor models (19). The first human trial using CRISPR/Cas9-modified cells for lung cancer treatment is set to take place in China, with a similar trial planned in the US (20). The technology is also being investigated as a potential tool in cancer immunotherapy, with ongoing animal models and human clinical trials (10).

Recent studies have highlighted the potential of CRISPR/Cas9 in prostate cancer treatment. A genetic screen for prostate cancer performed to date has contributed to the identification of biologically critical and clinically relevant target genes (21). CRISPR/Cas9 system accompanied by single- stranded oligonucleotide (ssODN) was shown to effectively repair the TP53 414delC gene region and inhibit the cell proliferation of PC-3 cells (22). Knockout of NEAT1 gene using CRISPR/Cas9 technique caused significant changes in the expression of apoptosis-related genes in a way that the expression of P21, BCL2, and BIRC5 genes decreased (23). These studies collectively suggest that CRISPR/Cas9 holds potential for the treatment of prostate cancer.

CRISPR/Cas9 technology has shown promise in the treatment of colorectal cancer, with potential applications in basic research, diagnosis, and personalized treatment (24). It has been used to correct causative mutations and enhance the immunological response in cancer immunotherapy, with ongoing clinical trials in immune cell modification (10). The technology has also been utilized for genetic screens, identification of druggable targets, and cancer gene therapy, with some applications in clinical trials (25). However, the technology’s limitations, such as off-target mutations and ethical concerns, need to be addressed (10).

CRISPR/Cas9 technology has shown promise in the treatment of pancreatic cancer, with potential applications in gene editing and targeted therapy (26). This technology, when combined with nanocarriers, has the potential to improve clinical outcomes for patients with pancreatic ductal adenocarcinoma (PDAC) (27). Furthermore, CD73 deletion inhibited the ERK/STAT3 pathway and activated the E-cadherin pathway in human and murine cell lines (28). In preclinical studies, CRISPR/Cas9 has been used to induce pancreatic cancer in mice, demonstrating its potential for in vivo research (29).

A range of clinical trials have explored the use of various treatments for liver cancer. Heo et al. found that JX-594, an oncolytic and immunotherapeutic vaccinia virus, demonstrated dose-related survival benefits in individuals with advanced hepatocellular carcinoma (30). Chen et al. reported promising results for camrelizumab, an anti-PD-1 antibody, in the treatment of primary liver cancer (31). Baskin-Bey et al. investigated the use of the pan-caspase inhibitor IDN-6556 in reducing liver cell apoptosis and injury during liver transplantation, with positive results (32). However, Newsome et al. found that granulocyte colony-stimulating factor (G-CSF) with or without haemopoietic stem-cell infusion did not improve liver dysfunction or fibrosis in patients with liver cirrhosis, and may be associated with increased adverse events (33). These studies highlight the potential of immunotherapies and oncolytic viruses in liver cancer treatment, but also the need for further research to confirm their efficacy and safety.

The role of genes in ovarian cancer development has been investigated through the application of CRISPR/Cas9 technology. CRISPR/Cas9-mediated knockout of the OC-2 negatively regulated the ovarian cancer progression possibly by apoptosis activation and angiogenesis inhibition, demonstrating its potential as a therapeutic tool (34). However, the safe and effective delivery of CRISPR/Cas-9 remains a challenge, with lipid-mediated delivery systems showing promise (35). Despite these challenges, CRISPR/Cas-9 is considered a powerful tool for the development of novel diagnostic approaches and therapeutic targets in ovarian cancer (36).

CRISPR/Cas9 technology has shown promise in the field of leukemia research, particularly in the study of chronic lymphocytic leukemia (CLL) and chronic myeloid leukemia (CML). Oshima et al. and Hernández-Sánchez et al. highlight the potential of CRISPR screening and genome-editing systems in understanding the role of mutated genes in leukemia pathogenesis and advancing functional studies of CLL (37). A CRISPR-Cas9 deletion system efficiently interrupts BCR/ABL1 oncogene in primary leukemic stem cells leading to the restoration of normal hematopoiesis, demonstrating potential therapeutic benefit (38). The CRISPR/Cas9 system is employed to investigate drug resistance in Acute Myeloid Leukemia (AML), enhance prognosis, and potentially move towards curing AML. Additionally, it aids in identifying new therapeutic targets, including the FST gene, lysine acetyltransferase KAT7, and LSD1 (39). Furthermore, CRISPR-competent AML patient-derived xenograft models tractable for genome editing are valuable targets for translational value suggesting that CRISPR/Cas9 holds promise for both understanding the genetic basis of leukemia and potentially developing targeted therapies (40).

Recent studies have demonstrated the potential of CRISPR/Cas9 in treating melanoma. Wu et al. developed a light-inducible CRISPR-Cas9 system to inhibit the progression of melanoma cells by targeting the BRAF V600E mutation (41). This approach shows promise as a novel therapeutic strategy. On the contrary Giuliano et al. found that CRISPR/Cas9-induced mutations in the MELK gene did not result in a specific anti-proliferative phenotype, while Ercolano et al. study suggested that targeting the PTGS2 gene could be a potential strategy for melanoma treatment (42, 43). These findings underscore the complexity of using CRISPR/Cas9 in cancer treatment and the need for further investigation.

CRISPR/Cas9 technology has shown promise in the treatment of glioblastoma multiforme (GBM), a highly aggressive brain tumor with poor prognosis (44). This gene-editing tool has been successfully applied in glioma stem cells, leading to efficient gene knockouts and the potential for genome-wide CRISPR screening (45). The use of lipid-based nano-delivery systems for in vivo CRISPR/Cas9 gene editing in brain cancer stem cells has also been explored, with promising results in disrupting the epidermal growth factor receptor variant III (EGFRvIII) mutation (46). Multiple studies have investigated the role of genes in GBM cell biology that can impact multiple survival pathways in cancers (47, 48). Furthermore, CRISPR-Cas9 gene-edited CAR T cells demonstrated enhanced activity in preclinical glioma models (49). These studies collectively highlight the potential of CRISPR/Cas9 in GBM treatment and the need for further research and clinical trials to validate its efficacy.

Targeted treatments for cancer, which pinpoint specific genes essential for cancer cell survival, are limited for oral cancer. Understanding the genetic basis of oral cancer, particularly in Asian populations where most cases occur, remains a challenge. In the realm of CRISPR/Cas9 technology, research focused on oral cancer has gained momentum, aiming to elucidate the molecular mechanisms underlying oral carcinogenesis, identify potential therapeutic targets, and develop innovative treatment strategies. This technology has the potential to be a powerful tool in cancer therapy, with applications in gene editing, whole genome screening, and identifying cancer immunotherapy targets (50).

Pooled CRISPR screens are a robust method for elucidating the essential genes in cancer cell survival. These screens involve the introduction of a library of single guide RNAs (sgRNAs) into cancer cells as a collective population, followed by the analysis of resulting phenotypes to identify indispensable genes (54). For instance, a recent study employed a CRISPR screen to identify essential genes in the survival of pancreatic cancer cells. Utilizing a custom CRISPR library targeting 18,009 genes, this study discovered a subset of genes, including those associated with DNA repair and cell cycle regulation, that are crucial for the survival of pancreatic cancer cells (55).

High-content CRISPR screens represent a potent approach to identify genes that are pivotal for cancer cell survival. These screens offer remarkable possibilities to conduct mechanistic investigations on a large scale, employing advanced models like organoids and whole organisms, versatile perturbations such as gene activation and repression, diverse biological challenges, and data-rich readouts like single-cell sequencing and imaging (56). For instance, by performing a genome-wide CRISPR screen utilizing a T cell clone with a specific recognition mechanism for cancer cells independent of HLA, Holcomb et al. unveiled various genes crucial for tumor immunity, including those involved in antigen presentation and T-cell activation (57).

Genome-wide CRISPR-Cas9 knockout screens hold significant potential in the identification of genes essential for the survival of cancer cells. These screens serve the purpose of elucidating the intricate genotype-phenotype relationship through the comprehensive disruption of gene expression on a genome-wide scale, enabling the examination of resulting phenotypic alterations. The technique leverages the CRISPR-Cas9 gene editing system in conjunction with libraries of single guide RNAs (sgRNAs) meticulously designed to target each gene within the genome (58). For instance, utilizing CRISPR-Cas9 in patient-derived GBM stem cells (GSCs), significant pathways for GBM growth are identified. Key genes such as those in the SOX transcription factor family, SOCS3, USP8, DOT1L, and protein ufmylation are implicated. Insights into temozolomide resistance suggest potential combination therapies. This functional approach uncovers genetic dependencies, enhancing comprehension of GBM treatment resistance (59).

The CRISPR/Cas9 system comprises two crucial components: a guide RNA that directs the Cas9 enzyme to the specific genomic location requiring genetic modification, and the Cas9 enzyme itself, acting as a molecular pair of scissors that cleaves DNA at the precise site guided by the RNA. This level of precision enables the cell’s natural repair mechanisms to make targeted and controlled alterations to the genome (60). Cancer, characterized by uncontrolled cell growth and proliferation, arises from genetic mutations and other genomic changes. While conventional therapies such as chemotherapy and radiation therapy effectively eliminate cancer cells, they also inflict damage to normal cells and entail adverse side effects. In contrast, CRISPR/Cas9 offers the ability to specifically target and modify cancer-related genes, thus providing safer and more precise therapeutic approaches (60).

In the field of biomedical research, CRISPR/Cas9 finds broad applications encompassing the study of gene function, the engineering of genetically modified organisms, and the exploration of innovative therapies for genetic disorders. In the context of cancer research, CRISPR/Cas9 has played a pivotal role in uncovering new avenues for cancer treatment and investigating the function of genes implicated in the disease. Multiple studies have showcased the remarkable diagnostic and therapeutic potential of this technology in the realm of cancer. It has demonstrated exceptional efficacy in targeting and disrupting oncogenes, tumor suppressor genes, and other genes involved in cancer development and progression. Moreover, CRISPR/Cas9 has shown promise in the engineering of T cells for immunotherapy, exhibiting encouraging outcomes in preclinical trials (61).

Oral cancer represents a complex and heterogeneous form of head and neck cancer affecting various structures within the oral cavity, such as the tongue, lips, and larynx. It is estimated that in 2020, over 377,000 new cases and 177,000 deaths worldwide will be attributed to oral cancer, ranking it as the sixth most prevalent cancer globally. Current standard treatments for oral cancer involve surgery, radiation therapy, and chemotherapy, all of which are associated with significant morbidity and mortality rates. Although CRISPR/Cas9 holds immense potential for advancing cancer research, its application to oral cancer is still in its nascent stages (62). Given the transformative potential of CRISPR/Cas9 technology in overcoming the limitations of existing oral cancer therapies, a comprehensive exploration of its utility in this context is imperative.

CRISPR/Cas9 has emerged as a powerful tool for investigating the mechanisms of tumorigenesis and development in oral cancer. It has been used to identify cancer-specific vulnerabilities, generate organoid and mouse models of cancer, and track carcinogenic progression (64). In particular, CRISPR/Cas9 has been effective in identifying genes associated with oral cancer pathobiology, such as the CD147, and in exploring their potential as therapeutic targets (65). Furthermore, CRISPR/Cas9 has been used to establish a p75 neurotrophin receptor (p75NTR)-knockout cell line, which has led to a reduction in pro-tumorigenic behavior in oral cancer cells (66). These studies collectively demonstrate the potential of CRISPR/Cas9 in advancing our understanding of oral cancer and in developing targeted therapies.

CRISPR-Cas9 can be utilized to identify genetic vulnerabilities in OSCC cell lines. By conducting genome-wide CRISPR-Cas9 screens in OSCC cell lines, researchers can identify genetic vulnerabilities that can be explored as potential therapeutic targets. This approach has already been employed to uncover new therapeutic targets for OSCC and develop targeted therapies for some of the genes identified in these studies (59).

CRISPR-Cas9 unveils novel therapeutic targets for OSCC, enabling the development of tailored therapies. CRISPR-Cas9 technology has been utilized to identify essential genes, druggable targets, and biomarkers for cancer research, aiding in understanding genotype-phenotype interactions and providing potential approaches for cancer treatment or diagnosis. Additionally, immune cells edited with CRISPR-Cas9 have been developed for cancer immunotherapies, with some already in phase I/II clinical trials (67). This approach has already led to the discovery of new therapeutic targets and the development of specific therapies (68).

CRISPR-Cas9 offers an effective method for precise gene modification in immune cells and oncolytic viruses, enhancing their potential for cancer therapeutic purposes. Through precise editing of genes in these entities, researchers can pioneer innovative therapies that specifically target cancer cells while safeguarding healthy ones. CRISPR/Cas9 has been extensively applied in cancer research for various purposes, and its effectiveness makes it a valuable tool for producing allogeneic CAR-T cells, potentially overcoming barriers in CAR-T cell therapy (69).

CRISPR-Cas9 technology has revolutionized cancer research and therapy by enabling the establishment of comprehensive genomic libraries for cancer patients. Studies have highlighted the potential of CRISPR-Cas9 for identifying cancer-specific vulnerabilities and drug targets (70, 71). This is further supported by Pacini et al., who integrated large-scale CRISPR-Cas9 viability screens to identify genetic dependencies in cancer, and Michels et al., who used pooled CRISPR-Cas9 screening to identify tumor suppressors in human colon organoids (72, 73). These studies collectively underscore the potential of CRISPR-Cas9 for personalized cancer treatment through the establishment of comprehensive genomic libraries.

CRISPR-Cas9 presents a potent tool in precision dentistry, particularly for targeted oral cancer treatment. By precisely modifying genes in oral cancer cells, researchers can develop innovative therapies that selectively target malignancies while safeguarding healthy tissues. Moreover, harnessing CRISPR-Cas9’s capabilities in elucidating tumorigenesis mechanisms aids in identifying novel therapeutic targets and advancing treatment strategies (74). Notably, preclinical and clinical investigations have demonstrated promising outcomes with this approach in developing new cancer therapies. Additionally, recent studies underscore CRISPR-Cas9’s potential in identifying oral cancer-associated genes and its broader applications in precision dentistry, affirming its significant promise in oral cancer management (75).

CRISPR/Cas9 technology has experienced rapid advancements since its initial discovery. Scientists have made significant modifications to enhance its efficiency, specificity, and accuracy. Notable progress includes the development of base editors, which enable precise modification of a single nucleotide in the DNA sequence without necessitating DNA cleavage. Additionally, the application of CRISPR/Cas9 for epigenetic editing has emerged, allowing for gene expression modulation without altering the DNA sequence. Moreover, researchers have devised novel delivery methods, such as nanoparticles and viral vectors, to enhance the efficiency and specificity of CRISPR/Cas9. These advancements have significantly expanded the potential applications of CRISPR/Cas9 technology across diverse domains, including medicine, agriculture, and biotechnology (50).

Integrating CRISPR-Cas9 with existing or emerging therapies, such as immunotherapy and personalized medicine, represents a promising frontier in the battle against oral cancer. By harnessing the precision of CRISPR-Cas9 gene editing alongside the targeted mechanisms of immunotherapy, researchers aim to develop synergistic treatment modalities that enhance the body’s immune response to cancer cells while precisely targeting genetic alterations driving tumor growth. This approach is employed in non-small cell lung cancer, and it is under clinical trials to investigate safety and feasibility (76, 77). Between 2017 and the beginning of 2022, six CAR-T immunotherapy drugs received approval from both the U.S. Food and Drug Administration (FDA) and the European Commission (EC) for B-cell lymphoma, Mantle cell lymphoma and B-cell precursor acute lymphoblastic leukemia, and relapsed refractory multiple myeloma (78). Furthermore, personalized medicine approaches, guided by genomic profiling, enable tailored treatments that consider individual variations in tumor biology, treatment response, and potential side effects. However, challenges such as off-target effects and delivery efficiency remain significant hurdles that need to be addressed for the successful translation of CRISPR-Cas9-based therapies into clinical practice.

CRISPR/Cas9 technology has shown significant potential in preclinical applications, particularly in genome editing and gene therapy. Multiple studies have underscored the versatility and efficiency of CRISPR/Cas9 in these areas, noting its use in controlling transcription, modifying epigenomes, and conducting genome-wide screens (79, 80). Targeted genome engineering enables precise modification of genetic information for studying gene function and disease pathology. The CRISPR/Cas9 applications in biomedical discoveries have been broad and successful, despite some off-target effects (81). Furthermore, CRISPR/Cas9 has played a pivotal role in the engineering of cells for immunotherapy, such as chimeric antigen receptor T-cell (CAR-T) therapy for cancer treatment (82). CRISPR/Cas9 technology combined with stem cells is a powerful tool for generating various cell types for disease modeling, drug screening, toxicology, and targeted therapies. The system has been used for genetic modification of stem cells, leading to the differentiation of specific cell types for functional analysis and potential clinical transplantation. Recent advancements in CRISPR/Cas9 technology have expanded the possibilities for stem cell research and therapeutic applications (83). These preclinical endeavors have demonstrated the tremendous potential of CRISPR/Cas9 technology in the development of innovative therapies for a wide spectrum of diseases.

CRISPR/Cas9 technology has made remarkable strides in gene therapy research, disease modeling, and precision medicine. However, there remain several challenges that necessitate attention before its safe and effective implementation in clinical settings. One of the foremost challenges pertains to the potential for off-target effects, which may result in unintended genomic mutations. Off-target effects arise when Cas9 acts upon non-targeted genomic sites, inducing cleavages that can yield adverse outcomes. This concern is of paramount significance as it has the potential to inflict enduring harm upon the genome, leading to the development of malignancies (84).

Another challenge lies in the delivery of CRISPR/Cas9 to target cells, which can prove arduous in certain cases. Delivery to target cells stands as a pivotal step in the gene editing process, necessitating an efficient, safe, and specific delivery system. Various delivery methods exist, encompassing viral and non-viral vectors. However, each method harbors its own limitations, and the selection of an appropriate delivery approach hinges upon the specific application at hand (85).

The landscape of programmable nucleases, including meganucleases (MNs), zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeats-Cas (CRISPR-Cas), has also contributed to genome editing from research to clinical and industrial domains. Each technology exhibits unique modes of action impacting their applicability across the genome and is subject to distinct testing conditions. While CRISPR-Cas9 leads the field owing to its versatility and rapid adoption, it is not without limitations. Contextual considerations dictate the selection of the most suitable approach for genetic modification within a given organism. For instance, the precision of meganucleases may suffice for high-precision applications where efficiency is not paramount, while ZFNs and TALENs, despite their efficiency, face challenges in retargeting. CRISPR-Cas stands out for its versatility, allowing for straightforward retargeting via guide RNA modification and compatibility with various delivery methods. Safety considerations, particularly in clinical applications, require thorough assessment of off-target effects and other potential risks (86).

Recent advancements in drug delivery systems have focused on overcoming challenges such as off-target effects and improving delivery mechanisms. Zhao et al. and Singh et al. both highlight the importance of tissue-specific drug delivery and the use of computational models to design and deliver drugs across biological barriers (87, 88). Nanomaterial-based drug delivery systems have also shown promise in addressing these challenges, with Chatterjee et al. discussing their applications and advancements, including stimulus-responsive and multifunctional nanocarriers (89). There is emphasis on the potential of nanoplatforms for targeted stimuli-responsive drug delivery, particularly in cancer therapy (90). These studies collectively underscore the potential of targeted drug delivery systems in minimizing off-target effects and improving therapeutic efficacy. Further, CRISPR/Cas9 off-target effects can be predicted online in-silico tools (91). These tools primarily rely on sgRNA sequences, potentially overlooking intracellular nuclear milieu. Strategies like enhancing Cas9 mutants such as eSpCas9 and SpCas9-HF1, exploring Cas9 homologs with rarer PAM sequences, and improving delivery methods significantly impact CRISPR/Cas9 gene editing fidelity. For in vivo gene therapy, CRISPR/Cas9 editing of somatic cells is a major application. To ensure safety, it’s crucial to directly measure off-target effects in tissues and living organisms. Techniques like Discover-seq and GUIDE-tag offer promising avenues for such assessments. Notwithstanding these challenges, CRISPR/Cas9 has showcased its potential utility in treating various diseases, including cancer, and holds promise for the development of novel therapies targeting hereditary conditions.

The emergence of CRISPR/Cas9 technology has engendered significant ethical and societal implications. Chief among the concerns is the potential for inadvertent consequences, encompassing off-target effects and unintended mutations. Another apprehension revolves around the possibility of unethical utilization of the technology, such as human germline editing. Furthermore, equitable distribution of the benefits arising from CRISPR/Cas9 technology, particularly in developing countries, raises concerns. These implications necessitate careful consideration as the technology continues to advance (92).

In addition to technical advancements, a comprehensive review of CRISPR-Cas9’s medical deployment necessitates a thorough examination of its ethical implications and societal impact. Ethical considerations surrounding germline editing pose fundamental questions regarding the manipulation of heritable traits in future generations, raising concerns related to consent, autonomy, and human dignity. Safety concerns also extend to the possibility of off-target effects and the long-term ramifications of gene editing upon the genome (93). Establishing robust ethical and regulatory frameworks is imperative to navigate these complex ethical landscapes and ensure the responsible application of CRISPR-Cas9 technology.

Preclinical studies have demonstrated the tremendous potential of CRISPR/Cas9 technology in treating diverse genetic disorders, including sickle cell anemia, cystic fibrosis, and Huntington’s disease. The system has also been instrumental in engineering cells for immunotherapy, exemplified by the utilization of CAR-T cells for cancer treatment. Furthermore, CRISPR/Cas9 can be employed to develop novel diagnostic tools for infectious diseases (82). The scope of potential applications in medical condition treatment using CRISPR/Cas9 technology is vast, and researchers continue to explore innovative avenues. Nonetheless, several challenges demand resolution before CRISPR/Cas9 can be safely and effectively employed in clinical settings. One notable challenge pertains to the possibility of off-target effects, which may induce unintended mutations in the genome. Delivery of CRISPR/Cas9 to target cells poses another challenge, particularly in certain scenarios. Additionally, the employment of CRISPR/Cas9 raises safety and ethical concerns, particularly concerning human germline editing (94). Addressing these challenges is imperative to ensure the secure and efficacious application of CRISPR/Cas9 technology in clinical settings. Notwithstanding these challenges, the potential of CRISPR/Cas9 technology in the development of innovative therapies for a wide array of diseases is immense, and researchers are diligently striving to overcome these obstacles to deliver these therapies to patients.