CRISPR-Cas9, originating from the adaptive immune response of prokaryotic cells, has transformed genome editing and pharmaceutical research. Researchers employed Rapidly Short Interspersed Repeats, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR), and Cas/CRISPR-associated proteins identified in the 1980s. Subsequent discoveries revealed that these bacterial defence mechanisms aimed at viral DNA were also present in eukaryotes¹. Jennifer Doudna and Emmanuelle Charpentier developed CRISPR-Cas9 for accurate, programmable DNA cleavage. This innovation initiated a new era in genetic engineering, resulting in their receipt of the 2020 Nobel Prize in Chemistry². Between 2011 and 2013, CRISPR-Cas9 employed a labelled guide RNA directly associated with the nuclease. This linkage enables the technology to guide Cas9 in cleaving specific DNA sequences. Consequently, researchers can accurately implement mutations, deletions, and insertions. CRISPR-Cas9 is recognised for its accurate modifications, yet TALENs and zinc-finger nucleases are also capable of rectifying DNA. CRISPR is distinguished by its simplicity and efficacy. Experimental genomics is an additional domain where this technology has proliferated^3,4.

CRISPR-Cas9, a premier pharmaceutical research instrument, targets genes associated with specific diseases and facilitates personalised medicine, transforming drug development. This technology has been extensively utilised in preclinical studies to develop more precise disease models, particularly those involving intricate genetic challenges. Researchers can now surmount these challenges with greater ease⁵. Utilising this technology, researchers have created innovative protein alternatives, accelerating cancer research. The ability to simultaneously manipulate multiple genes enables the evaluation of gene therapy in cancer treatment⁶. This gene therapy method is innovative for addressing genetic disorders such as sickle cell anaemia and beta-thalassemia. This technological advancement has facilitated the development and implementation of gene therapies. This technology is applicable in oncology and other diseases. Consequently, it has pinpointed essential gene targets implicated in cancer cell proliferation, facilitating targeted chemotherapies and more individualised cancer therapies⁷.

This review analyses the transformative influence of CRISPR-Cas9 on pharmacological research, emphasising its varied applications, intricate technical prerequisites and regulatory approvals, as well as the ethical dilemmas associated with human genome editing. The review examines these factors to comprehend the pharmaceutical opportunities and constraints of this technology. CRISPR-Cas9 demonstrates significant potential for enhancing human health; however, off-target effects and the possible misuse in germline editing present ethical dilemmas that necessitate additional investigation⁸.

2. Overview of CRISPR-Cas9 technology:

CRISPR-Cas9 revolutionises genome editing by offering unparalleled control and adaptability for accurate DNA sequence modifications. This innovative system employs the innate immune defence of bacteria to identify and eradicate viral DNA during an assault. The CRISPR-Cas9 system utilises guide RNA (gRNA) and the Cas9 nuclease as a molecular navigation and cleavage mechanism. This technology enables researchers to precisely target DNA modifications, rendering it remarkable.

2.1. Mechanism of action:

CRISPR-Cas9 editing commences when cells exhibit sensitivity to short gRNAs and Cas9. The gRNA comprises two components: a DNA-complementary sequence and a scaffold structure that binds to Cas9, enabling the complex to identify and attach to DNA³. Cas9 cleaves DNA at the designated site subsequent to its binding with a DNA sequence containing a Protospacer Adjacent Motif (PAM), a requisite short DNA sequence for Cas9 attachment. DNA double-strand breaks (DSBs) elicit nonhomologous end joining (NHEJ) and homologous directed repair (HDR)⁹.

NHEJ is a rapid, error-prone repair mechanism that produces minimal insertions but can impair gene function, rendering it advantageous for gene knockout investigations. Utilising an external DNA template, HDR is an infrequent yet exact mechanism that substantially alters genes. Prospective applications involve rectifying disease-associated mutations utilising this technique¹⁰. These two repair pathways thus grant the use of CRISPR to both the disruption of genes and the precise correction of genes.

2.2 Variants of CRISPR‒Cas systems:

CRISPR-Cas9 remains the predominant system; however, alternative CRISPR systems have been devised for various applications. Cas12 can differentiate PAM sequences and cleave DNA in a staggered manner, offering various gene-editing possibilities ¹¹. Cas12's ancillary activity can identify and analyse highly specific nucleic acids. These capabilities have facilitated the development of CRISPR-based diagnostics for viruses such as SARS-CoV-2. Cas13 employs a specific RNA-based mechanism rather than a DNA-based one. This distinctive property enables CRISPR to facilitate transcriptomic genome editing and gene expression modulation, which are crucial for addressing RNA-related disorders¹².

2.3 Recent technological advancements:

Since its inception, CRISPR-Cas9 technology has undergone enhancements via semantic refinement to augment precision. The Cas9 protein generates off-target effects that modify the genome, complicating the clinical application of CRISPR. To resolve this issue, sophisticated high-fidelity Cas9 variants such as eSpCas9 and HypaCas9 have been created. These versions enhance targeting precision and markedly diminish off-target effects, thereby mitigating the risks associated with genetic manipulation¹³. Base editors employ catalytically inactive Cas9 and a deaminase enzyme to modify DNA bases without inducing a double-strand break, thereby advancing this domain. Precise base editing can rectify mutations associated with sickle cell disease through single-base modifications¹⁴.

Additional advancements encompass engineered gRNAs that enhance binding specificity and dual-guide RNA techniques that facilitate target recognition. CRISPR-associated transposase systems, an advanced genome editing methodology, can incorporate extensive DNA sequences. This approach possesses significant potential in gene therapy and synthetic biology¹⁵.

3. Applications of CRISPR-Cas9 in Pharmaceutical Research:

The novel CRISPR-Cas9 system is indispensable in pharmaceutical laboratories. It facilitates the identification of drug targets, the development of therapeutic interventions, and the progression of personalised medicine. Its distinctive characteristics have transformed biopharmaceuticals, gene editing, and disease modelling.

Figure 1 Applications of CRISPR-Cas9 in pharmaceutical research

3.1. Drug Discovery:

1. Target identification:

CRISPR-Cas9 is primarily utilised in pharmaceutical research to pinpoint genetic targets implicated in disease pathogenesis. This technology enables scientists to conduct extensive genetic screenings to assess the impact of gene knockouts or modifications on disease phenotypes⁶. A CRISPR-based initiative identified genes essential for HIV replication, which are presently under investigation as potential tools for inhibiting viral replication¹⁶. In addition, CRISPR screens have identified genes that are involved in the survival of cancer cells, providing insights into potential targets for tumours that are resistant to chemotherapy¹⁷.

2. Functional Genomics:

Functional genomics examines the relationship between genes and phenotypes, particularly in the context of diseases. Notwithstanding the potential for off-target effects, CRISPR-Cas9 is a potent and accurate gene editing technique, especially adept at validating drug targets. In oncology, CRISPR-Cas9 has been employed to investigate tumour suppressor genes and oncogenes, pinpointing mutations that induce tumour proliferation and metastasis¹⁸. It is possible to predict future molecule interactions and test drugs on tumour suppressors in realistic cell environments⁴.

3.2. Gene therapy:

1. Treatment of Genetic Disorders:

CRISPR-Cas9 can rectify mutations and address genetic disorders through accurate gene editing. The technology can effectively rectify a single-nucleotide mutation in the β-globin gene, potentially curing sickle cell anaemia. A clinical study utilised CRISPR-Cas9 to modify haematopoietic stem cells for the expression and production of heterozygous haemoglobin¹⁹. This discovery is a significant step in gene therapy, as the edited cells were able to engraft and produce functional blood cells.

CRISPR is utilised to investigate hereditary cystic fibrosis. Mutations in the CFTR gene result in this pulmonary and gastrointestinal disorder. Researchers rectified CFTR mutations in cellular models utilising CRISPR, yielding insights for prospective clinical applications²⁰. Notwithstanding the difficulties in securely administering CRISPR components to human tissues, vector technology and delivery techniques hold promise for the elimination of numerous genetic disorders.

2. Clinical Trials:

CRISPR technologies are undergoing clinical trials to assess their safety and efficacy in treating genetic disorders. CRISPR Therapeutics and Vertex Pharmaceuticals are at the forefront of clinical trials employing CRISPR-Cas9 to address β-thalassemia, a haematological disorder akin to sickle cell anaemia²¹. CRISPR therapies for congenital blindness resulting from CEP290 gene mutations are currently undergoing trials. CRISPR therapy was initially employed to address LCA in 2020, and the administration of a CRISPR-based medication into the retina facilitated innovative treatment for congenital eye disorders²².

3.3. Disease Modelling:

Gene-edited models, such as "happy" mice, are essential for investigating disease pathogenesis and conducting drug screenings, a possibility facilitated by CRISPR-Cas9 technology. Researchers can "manipulate" critical defects in cell lines or animal models to investigate human diseases through novel methodologies. In oncological research, CRISPR and alternative techniques have been employed to develop murine models exhibiting mutations analogous to those found in cancer patients. Researchers can monitor tumour proliferation and evaluate pharmaceuticals aimed at particular molecules in a regulated setting²³. These models have been used to study cancer forms that exhibit highly variable genomic rearrangements, e.g., glioblastoma and pancreatic cancer.

Utilising CRISPR technology to develop cellular models with gene mutations linked to amyloid plaque formation is essential for investigating neurodegenerative diseases such as Alzheimer's. These models evaluate interventions that may influence disease pathology. In disease modelling, CRISPR surpasses conventional drug discovery techniques in efficiency and offers profound insights into disease mechanisms, enabling researchers to formulate targeted therapies²⁴.

3.4. Pharmacogenomics: Tailoring Medicines based on Genetic Profiles:

CRISPR-Cas9 is transforming pharmacogenomics, which customises treatment according to individual genetic profiles. CRISPR can be utilised to modify drug metabolism genes to investigate the impact of genetic polymorphisms on medication response. The effectiveness and toxicity of cancer therapies are contingent upon a patient's genetic profile, underscoring the significance of this application^14,25. An experiment employed CRISPR to modify metabolic CYP2C9 gene variants, yielding a more tailored anticoagulant therapy. Through CRISPR, pharmacogenomics can mitigate adverse effects, enhance drug efficacy, and elevate patient perception²⁶.

Moreover, CRISPR can facilitate the development of cell lines with varied genetic backgrounds. This advancement facilitates drug testing across a broader demographic, ensuring that all prospective medications are efficacious and safe for individuals with varying genetic profiles. It addresses a significant deficiency in pharmaceutical testing^26,27.

4. Challenges in the application of CRISPR-Cas9:

CRISPR-Cas9 has progressed considerably in genome editing; nonetheless, several challenges may limit its utilisation in pharmacology and clinical research. The demand for technical, clinical, regulatory, and commercial research and policy development creates these challenges.

Figure 2 Challenges in the application of CRISPR-Cas9

4.1. Technical Challenges:

1. Off-target effects:

A primary technical challenge of CRISPR-Cas9 is off-target editing, occurring when the Cas9 enzyme inadvertently induces DNA breaks at unintended locations. Off-target effects may arise from inaccuracies in the DNA modification process, potentially leading to significant issues, including unintended gene deletions or the activation of oncogenes. This may arise from unintentional gene disruption or the widespread activation of oncogene expression²⁸. Research demonstrates that the binding of guide RNA (gRNA) to DNA can occasionally be persistent, enabling Cas9 to function effectively at sequences that are similar yet not identical to the intended target. This transpires owing to the inaccuracy of the targeted incisions in specific cases. The consequences of this are significant, particularly in therapeutic settings, as it may lead to adverse effects. Consequently, CRISPR-based therapies may endanger their intrinsic dynamism¹³.

Several high-fidelity Cas9 variants, including eSpCas9 and HypaCas9, have been engineered to minimise off-target activity to mitigate the aforementioned concerns. These engineered allele variants exhibit a substantial reduction in off-target interactions while maintaining high on-target specificity²⁹. Short guide RNAs with reduced complementary sequences can also improve targeting precision and reduce off-target effects³⁰. Despite their status as rapidly moving targets, the total eradication of off-target risks remains a formidable challenge, particularly within the intricate human genome.

2. Delivery Mechanisms:

Administering CRISPR components to targeted cells and tissues poses significant challenges. The Cas9 protein and guide RNA must remain unaltered in target cells without causing damage or eliciting an immune response. Adeno-associated viruses (AAVs) are the most prevalent delivery system, recognised for their superior transfection efficiency and capacity to target various tissues³¹. However, viral vectors have drawbacks like limited cargo capacity and immunogenicity, which can cause immune reactions and limit repeat dosing.

Nonviral delivery techniques such as lipid nanoparticles and electroporation diminish immune responses. Lipid nanoparticles can transport CRISPR components in vivo, particularly to hepatic tissues. Challenges persist in tissue-specific organ delivery and the stability of small molecules. Furthermore, a secure equilibrium of targeting and delivery mechanisms is essential for the extensive clinical application of CRISPR technology³².

4.2. Clinical challenges:

1. Efficacy across Genetic Diversities:

Genetic diversity hinders the uniform provision of therapeutic benefits across genetically varied populations, complicating research efforts. Single-nucleotide polymorphisms (SNPs) can modify target sequences among individuals, resulting in off-target effects or diminishing the efficacy of the CRISPR biosystem. The genetic diversity present at both the periphery and core of the population underscores the necessity for robust governance mechanisms to avert unauthorised interference in human subject experiments ³³.

2. Safety concerns:

The primary concern is the long-term safety of CRISPR-Cas9 genetic modifications, as the immune system may respond to genomic alterations. Exogenous CRISPR proteins, including viral delivery vectors, may elicit innate and adaptive immune responses, resulting in inflammation or other adverse effects. A further concern is preexisting immunity to viral vectors, as individuals previously infected by pathogens can neutralise them with antibodies³⁴.

Safety concerns encompass inadvertent gene activation and the elimination of genomic regulatory regions. Minor effects resulting from random CRISPR interventions can disrupt cellular homeostasis, potentially leading to cancer. Consequently, additional research is required to evaluate the long-term safety of CRISPR-based therapies in humans³⁵.

3. Scalability:

A significant barrier to the translation of CRISPR therapies from research to clinical application is scalability. The production of clinical-grade CRISPR components and large-scale delivery systems necessitates sophisticated technology for purification, stability, and efficacy. For example, the production of viral vectors is intricate and costly, potentially rendering CRISPR-based therapies inaccessible in the future ³⁶. Bioprocessing techniques and infrastructure must improve to ensure high-quality production for clinical trials and commercial use.

5. Ethical Considerations in CRISPR-Cas9 Research:

The swift advancement of CRISPR-Cas9 technology has elicited ethical apprehensions, especially regarding human and animal research. As molecular genome editing advances, ethical discourse is essential to responsibly manage and mitigate harm in CRISPR-related research and therapies³⁷.

Figure 3 Ethical considerations in CRISPR-Cas9 research

5.1. Human Germline Editing:

Human germline editing—altering DNA in reproductive cells to produce inheritable changes across generations—represents one of the most compelling ethical dilemmas associated with CRISPR. It has the potential to eliminate genetic disorders and produce ideal offspring with superior physical or cognitive traits, which induces fear in some individuals. Consequently, proponents of CRISPR assert that it has the potential to avert hereditary diseases such as Huntington's and cystic fibrosis, thereby alleviating suffering. Nonetheless, germline editing may engender social disparities, such as a genetic divide, if access to genetic enhancement is limited to the affluent ³⁸.

The enhancement and application of germline editing for therapeutic purposes present ethical dilemmas. Therapeutic applications rectify or alter detrimental mutations, whereas human enhancement may result in advantageous traits such as intelligence or athletic prowess. Social groups may grapple with the concept of normality, resulting in discrimination, genetic trait segregation, and the stigmatisation of individuals with disabilities³⁹. Alongside the deliberate annihilation of the germline, off-target effects have induced harmful mutations, the point mutations of which persist and are transmitted to subsequent generations, exacerbating the patient's issue⁴⁰.

5.2. Equity and Accessibility:

The exorbitant expense of CRISPR therapies, encompassing both development and implementation, has elicited concerns regarding equity and accessibility. To customise technology-specific hardware for particular applications, CRISPR therapies enlist top engineers, training institutions, and operational specialists. These advanced facilities necessitate costly equipment that developing nations and their middle classes cannot finance. This results in global healthcare disparities: affluent nations possess CRISPR technology, whereas impoverished nations rely solely on conventional genetic therapies⁴¹.

Ethical discourse underscores the necessity for equitable funding and advancement of CRISPR therapies to ensure their clinical approval and affordability for a broader patient population. If we fail to address these disparities, we jeopardise the establishment of a bifurcated healthcare system wherein only the affluent can access advanced genetic therapies, thereby contravening healthcare equity⁴².

5.3. Informed Consent and Risk:

An additional ethical concern regarding CRISPR-based therapies is the acquisition of genuine informed consent, particularly due to the experimental characteristics of gene editing and associated risks. Patients undergoing this novel gene editing procedure must be informed about off-target effects, unintended immune responses, and long-term safety concerns. Although gene editing is straightforward, elucidating its associated risks can be challenging, and patients may lack comprehension⁴³.

The long-term safety of human CRISPR trials is influenced by the potential for edited genes to induce delayed or unforeseen health complications. Consequently, participants must be apprised of both immediate and potential risks and provide informed consent. This engenders ethical dilemmas as patients' eagerness for innovative therapies may lead them to undervalue or overlook the associated risks, thereby questioning the validity of their consent⁴⁴.

5.4. Animal Welfare in Research:

The augmented application of CRISPR-Cas9 in animal research for the establishment of disease models accelerates therapeutic development. Gene-editing experiments may induce significant health issues in animal models, prompting a consideration of the equilibrium between scientific advancement and ethical responsibilities. Genetically modified animals exhibiting human-like ailments may experience pain, distress, or diminished quality of life, prompting ethical enquiries regarding the justification of scientific knowledge⁴⁵.

CRISPR, an innovative laboratory technology characterised by its efficiency and specificity, is now extensively utilised in research, augmenting the number of animals subjected to genetic modifications and their consequent effects. The ethical guidelines in animal research advocate the "3Rs" principle—replacement, reduction, and refinement—to minimise animal suffering. This schema enables CRISPR applications to utilise substitute models, decrease animal usage, and enhance suffering protocols, thereby rendering research more ethical⁴⁶.

6. Case studies of CRISPR-Cas9 in pharmaceutical research:

CRISPR-Cas9 technology has transformed pharmaceutical research, particularly in the treatment of genetic disorders and cancer. Although the technology has limitations, it also encounters challenges that are ultimately advantageous. This section of the report examines CRISPR's achievements in medicine and its challenges, including the necessity for enhanced technical precision in the technology's adaptation.

6.1. Success-stories:

1. Sickle Cell Disease and Beta-Thalassemia:

Haematological disorders represent one of the most compelling applications of CRISPR-Cas9. CRISPR Therapeutics and Vertex Pharmaceuticals executed a distinctive clinical trial to modify the BCL11A gene in haematopoietic stem cells to enhance foetal haemoglobin, thereby addressing both diseases. Preliminary findings from 2021 indicated that patients with beta-thalassemia have attained stable normal haemoglobin levels, thereby diminishing their reliance on transfusions and alleviating painful crises in individuals with sickle cell syndrome. This achievement demonstrates that CRISPR can detect and rectify genetic anomalies in chronic diseases, enhancing its therapeutic efficacy^21,47.

2. Cancer Research: T-Cell Engineering for Immunotherapy:

CRISPR-Cas9 can genetically alter T cells to enhance their efficacy in targeting tumours, thereby influencing cancer immunotherapy. A pioneering study from the University of Pennsylvania employed CRISPR to modify T cells by removing inhibitory receptors, thereby augmenting their capacity to target tumour cells in patients with advanced cancer. The preliminary trial outcomes indicated that modified T cells remained viable in patients and did not induce significant adverse effects, representing a significant advancement for CRISPR-based immunotherapy and a prospective pathway for personalised cancer therapies. This study demonstrates the versatility of CRISPR in enhancing treatment methodologies^25,48.

6.2. Emerging Applications:

1. Ongoing Research in Huntington’s Disease:

Although Huntington's disease is curable, CRISPR technology is currently limited to research on neurodegenerative disorders. Huntington's disease, a monogenic mutation, is the primary neurodegenerative disease target for CRISPR technology. Researchers at University College London (UCL) employed CRISPR to inhibit the mutant huntingtin (HTT) gene responsible for Huntington's disease⁴⁹. CRISPR reduces mutant HTT protein levels in animal models, enhancing symptoms in preclinical studies. The nascent application of CRISPR has unveiled opportunities for addressing various intricate neurological disorders and instilling optimism for previously incurable ailments⁵⁰.

2. CRISPR Diagnostics for Infectious Diseases:

Diagnostics now encompass biotechnology and CRISPR, a non-invasive method for diagnosing the majority of infections. CRISPR-based diagnostic systems such as SHERLOCK (specific high-sensitivity enzymatic reporter unLOCKing) can accurately and precisely detect viral RNA in patients with Zika and COVID-19⁵¹. Diagnostic and disease management devices utilise CRISPR effector proteins such as Cas12 and Cas13 to swiftly and economically identify pathogen DNA or RNA. Adolescents affected by pandemics such as COVID-19 and older individuals with prior experience have benefited from enhanced instrumentation. Resource limitations, particularly in low-income nations, enable these instruments to revolutionise diagnostic physicians ^52,53.

6.3. Failure and Lessons Learned:

1. Off-target effects in cancer studies:

Although CRISPR-based studies have potential, they have faced significant setbacks. Early studies have shown that oncogene activation causes unintended off-target effects, making CRISPR editing risky in precision. When genes that are likely to provide strong immune responses against cancer cells are knocked out, researchers have observed unexpected genetic changes that have rendered a gene nonfunctional, endangering cell viability and raising safety concerns. The rapid evolution of high-fidelity CRISPR variants like eSpCas9 has highlighted the need for thorough verification before clinical use^11,54.

2. Immune Responses in Gene Therapy Trials:

CRISPR-based gene therapy trials have been plagued by patient immune responses due to the Cas9 protein. This reaction has mostly been seen in patients with bacterial Cas9 protein immunity. Immune responses can reduce gene therapy efficacy and cause side effects. Research on animal models has shown these limitations, so researchers now use humanised or engineered Cas9 proteins to avoid immune detection. It makes us think about the lessons we can learn, and they point to the need to humanise CRISPR⁵⁵.

7. Future Directions in CRISPR-Cas9 and Pharmaceutical Research

The integration of novel technologies, innovative applications, and intricate regulatory frameworks is increasingly facilitating the use of CRISPR-Cas9 in pharmaceutical research. By resolving certain existing challenges associated with CRISPR, these advancements will enhance its application in therapeutic development, precision medicine, and regenerative therapies.

Figure 4 Future directions for CRISPR-Cas9 and pharmaceutical research

7.1. Next-Generation CRISPR Technologies:

Next-generation CRISPR technologies, such as prime editing and base editing, which accurately modify DNA, represent the most promising developments. Prime editing employs CRISPR and reverse transcriptase to precisely modify a specific DNA sequence at the target site without inducing double-strand breaks⁵⁶. This DNA editing technology is safer due to its reduced off-target effects and diminished likelihood of inducing unintended alterations in vivo. Base editing can rectify point mutations in diseases by altering a single nucleotide without inducing double-strand breaks (DSBs). These tools may enhance the safety and acceptability of gene therapies, thereby broadening the spectrum of genetic disorders addressed⁵⁷.

7.2. AI integration for precision enhancements:

Artificial intelligence is anticipated to enhance the precision of CRISPR by enabling researchers to accurately forecast off-target effects. Algorithms trained on genomic data can detect DNA sequence patterns and potential mislocated sites. DeepCRISPR and similar AI-driven tools utilise deep learning to model and forecast guide RNA designs, enhancing the safety of gene editing ⁵⁸. AI integration technology might be the next step in developing CRISPR treatments that are tailored to the genetic variations of individuals (‘personalized’ genomes), thus making therapies more effective and less risky^59,60.

7.3. Regenerative medicine and stem cell therapies:

Current research is broadening the unexplored potential of gene-editing CRISPR-Cas9 in regenerative medicine, including stem cell therapies and tissue reconstruction. CRISPR enables personalised stem cell therapies by targeting mutations in stem cells, facilitating the replacement of aged cells with new ones. Recent research employs CRISPR-modified stem cells to regenerate cardiac tissue following myocardial infarction and to restore functionality in degenerative conditions such as Parkinson's disease⁶¹. Furthermore, CRISPR may significantly aid in tissue engineering by enabling precise cellular interventions for organ regeneration, potentially resolving the persistent problem of organ scarcity⁶².

7.4. Evolution of Regulatory Frameworks and Ethical Standards:

CRISPR-based therapies are gaining widespread acceptance. Regulatory frameworks must evolve to accommodate gene-editing technologies. Conventional drug regulations do not address issues related to permanent genetic modification, including safety, ethics, and societal implications. Future regulation of CRISPR-based therapies by the FDA and EMA may entail more

stringent long-term safety and efficacy monitoring⁶³. Despite advancements in international CRISPR research, human germline editing necessitates uniform ethical standards. Consequently, an international regulatory entity or framework is essential to govern ethical standards and oversee gene-editing practices, thereby ensuring global ethical compliance. This policy may avert malpractices, enhance transparency, and guarantee equitable CRISPR utilisation without genetic discrimination or eugenics.

CONCLUSION:

CRISPR-Cas9 has emerged as the most significant instrument in pharmaceutical research. It offers the utmost precision and efficacy in genome editing. The diverse applications of CRISPR-based technology, including drug discovery, gene therapy, diagnosis, and medical regeneration, demonstrate its significant potential to address formidable diseases, such as rare genetic disorders and various cancers. CRISPR enables targeted genetic interventions, thereby facilitating the development of personalised and potent therapies.

Nonetheless, CRISPR technology has not effectively addressed the challenges it faces. Off-target effects, immune responses, and challenges in efficient delivery remain the three principal technical obstacles, while ethical considerations surrounding human germline editing, equitable access, and informed consent necessitate meticulous assessment. The application of CRISPR in animal research presents additional ethical dilemmas that must be addressed through a balance of scientific and ethical considerations.

In the future, advanced CRISPR systems, including prime and base editing, along with the integration of AI for enhanced precision, are essential for improving the safety and accessibility of CRISPR-based therapies. However, the realisation of these aspirations will depend on continuous innovation, the establishment of strong regulatory frameworks, and a worldwide agreement on ethical issues. Mindful development has ushered in a novel realm propelled by CRISPR–Cas9 for pharmaceutical research and a transformative epoch of genetic medicine.

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Received on 26.01.2025 Revised on 17.02.2025

Accepted on 03.03.2025 Published on 08.07.2025

Available online from July 12, 2025

Asian J. Pharm. Tech. 2025; 15(3):296-304.

DOI: 10.52711/2231-5713.2025.00045

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