CRISPR in medicine: applications and challenges

Jin-Yu Sun, Hai-Bo Hu, Yan-Xiang Cheng, Xiao-Jie Lu, CRISPR in medicine: applications and challenges, Briefings in Functional Genomics, Volume 19, Issue 3, May 2020, Pages 151–153, https://doi.org/10.1093/bfgp/elaa011

Initially discovered in bacteria and archaea as adaptive immune strategies against invading nucleic acids, the clustered regularly interspaced short palindromic repeats-associated protein (CRISPR-Cas) system has now been repurposed as a practical tool for genome editing and other applications [ 1]. Owing to its flexibility, simplicity, efficiency and precision, the application of CRISPR systems in medicine has attracted significant attention and experienced rapid development. Currently, CRISPR systems have been used to screen novel drug targets [ 2], identify pathogenic genes [ 3], establish disease research modes [ 4], develop therapies against various diseases [ 5, 6] and so forth.

CRISPR-based genome editing tools provide effective and diverse strategies to prioritize candidate drug targets or validate putative targets [ 2, 7]. With CRISPR-Cas9 genetic screening in a diverse collection of cancer cell lines, statistical methods were developed to identify core and context-specific fitness genes systematically [ 8]. Based on the results from CRISPR-Cas9 screens and related genetic biomarkers, further assessments can be made to decide target genes with the most potential for novel anti-cancer drugs [ 8]. CRISPR systems can be also used to validate putative targets of new drugs. Lin et al. [ 9], for example, used CRISPR-Cas9 mutagenesis to investigate a serious of cancer drug targets in different stages of clinical trials. Upon the silencing of these putative targets, the anti-cancer effectiveness of some of these drugs remained, indicating the existence of unexpected targets.

Besides drug target identification or validation, CRISPR systems can also be used as diagnostic tools, which facilitates accurate detection and thus early intervention for infectious/noninfectious diseases [ 10–12]. Gootenberg et al. [ 13] developed a platform called SHERLOCKv2 (specific high-sensitivity enzymatic reporter unlocking version 2), which can detect one dsDNA and three ssRNA targets in a single reaction. The multitarget DNA and RNA detection by SHERLOCKv2 can be applied as a portable, rapid and quantitative diagnostic test for Dengue or Zika virus as well as pneumonia pathogens [ 10, 13].

Furthermore, the CRISPR-based protective and therapeutic strategies are probably the most promising applications of CRISPR system. CRISPR-Cas9 is revolutionizing the area of gene therapy, and advances have been achieved in Duchenne muscular dystrophy [ 14], α1-antitrypsin deficiency [ 15], hearing loss [ 16] and hematopoietic diseases [ 17], human immunodeficiency virus type 1 (HIV-1) infection [ 18, 19] as well as cancers [ 20, 21]. Several clinical trials are underway to evaluate the feasibility and safety of CRISPR-based treatments (e.g. NCT02793856, NCT03081715, NCT03398967, NCT03655678 and NCT03399448). For example, transplantation of cells with CCR5 gene artificially disrupted showed a remarkable therapeutic effect against HIV-1 infection [ 22]. Xu et al. [ 23] established a CCR5-ablating system by CRISPR-Cas9 tool in hematopoietic stem and progenitor cells (HSPCs), which conferred HIV-1 resistance in vivo. In their recent clinical study, the investigators used CRISPR-Cas9 technology to knock out the CCR5 gene of the donor stem cells and performed a successful transplantation of the CCR5-ablated HSPCs into a patient with HIV-1 infection and acute lymphoblastic leukemia [ 18]. After 19 months, the acute lymphoblastic leukemia was in complete remission, edited stem cells were engrafted with full donor chimerism and no apparent adverse event was observed [ 18]. Despite these promising results, further researches remain required to demonstrate the safety and feasibility before clinical practice.

Apart from application in basic and clinical medicine, substantial effort has been paid to enhance the efficiency and precision of CRISPR-based genome editing tools in the last 2 years [ 24–26]. Anzalone et al. [ 27] improved the CRISPR system by developing a precise ‘search-and-replace’ genome editing method (called prime editing), which can induce targeted insertions, deletions and point mutation without DSBs or donor DNA templates. They successfully use prime editing to correct primary mutations that caused sickle cell disease or Tay-Sachs disease in human embryonic kidney cells, with high efficiency and few byproducts [ 27]. Although this improved editing tool and delivery protocol reduced the off-target activity, concerns remain on the large deletions and complex rearrangements generated by on-target cutting and repair, which may incur potential pathogenic consequences [ 28, 29].

Ethical and safety concerns on the application of CRISPR system have received much attention in the past years. Since the first CRISPR-Cas9-based human embryo genome editing in 2015, the speculation over embryo gene editing has never stopped [ 30]. Recently, the medical genomes editing on twin baby girls by Jiankui He caused fierce controversy in this area [ 31]. Without considering the long-term adverse consequences, it was irresponsible to inactivate the CCR5 gene, since this alteration may increase the risk of complications from other viral infections [ 32]. After in-depth discussions, Lander and colleagues called for a temporary global moratorium on all clinical uses of human germline editing before the establishment of an international framework [ 33, 34]. Clinical application of germline editing should not be permitted unless the long-term consequences are fully understood.

In this special issue, we included nine review articles to summarize and discuss new trends and current challenges in the application of CRISPR systems in medicine.

Zhou et al. [ 35] focused on the ethical controversies and current regulations on the CRISPR-Cas9-based human embryo gene editing. They reviewed the technical defects of this tool and provided suggestions for the development trend. Generally, they hold a cautiously optimistic attitude towards the development and application of embryo gene editing in the premise of strict supervision. In the article by Sun et al. [ 36], the authors introduced the mechanisms of CRISPR-Cas9 tool and its development evolution. They summarized the therapeutic applications of CRISPR-Cas9 in genetic diseases, cancers and infectious diseases (such as oncogenic virus, HIV, influenza virus, respiratory syncytial virus, herpesvirus and human neurotropic polyomavirus).

In the three papers by Li et al. [ 37], Chen et al. [ 38] and Ureña-Bailén et al. [ 39], they reviewed the applications of CRISPR-Cas9 in cancer immunotherapy and revealed the promising potential of CRISPR-Cas9 tool in improving current or creating the next-generation treatment strategy. For example, the advances in CRISPR-Cas9 technology open a window to accelerate the development of the next-generation chimeric antigen receptor (CAR)-T cell-based cancer immunotherapy, including universal CAR-T cells, more potent CAR-T cells and improved controllable CAR-T cells. The article by Chen et al. [ 38] also introduced the applications of CRISPR-Cas9 in various cancer immunotherapies besides CAR-T cell therapy and highlighted the application in screening novel drug targets.

Herrera-Carrillo et al. [ 40] focused on the novel CRISPR-based anti-HIV strategies, which could eliminate HIV infection by interrupting the viral components or host cell functions that play critical roles during HIV replication. They discussed the advantages and limitations of the CRISPR-based HIV therapies from the aspects of efficiency, specificity, off-target effect and delivery methods.

In the article by Chen et al. [ 41], they introduced the CRISPR-Cas classification and the mechanisms of CRIPR-based gene editing. The authors reviewed the preclinical application of CRISPR-Cas9 in cancer treatment (e.g. cancer immunotherapy, cancer-related genome and epigenome manipulation, and elimination or inactivation of carcinogenic viral infections). Pei et al. [ 42] introduced several EpiEffectors used in epigenome editing, including transcriptional activators, transcriptional repressors, histone modification enzymes and DNA modification enzymes. They reviewed the application of CRISPR-Cas9 in epigenome editing, which might offer novel therapeutic avenues against various diseases (e.g. cancer, central nervous system disorders, blood system diseases and metabolic diseases). In the article by Xu et al. [ 43], the authors focused on animal cancer models generated by CRISPR-Cas9. They also highlighted the current shortages of the CRISPR-Cas9 tool, including fitness of editing cells, delivery methods, off-target effects, immune response, ethical problems and so forth.

We are happy to bring these papers together into this special issue, which can provide readers a comprehensive view of current applications and challenges of CRISPR systems in medical research.

Conflict of Interest

Funding

This work was supported by the National Natural Science Foundation of China (Grant No. 81772596 to X.-J.L.).

Jin-Yu Sun, Department of General Surgery, The First Affiliated Hospital of Nanjing Medical University; Sparkfire Scientific Research Group, Nanjing Medical University, Nanjing, China.

Hai-Bo Hu, Department of Thoracic Surgery, Huai'an Second People's Hospital, The Affiliated Huai'an Hospital of Xuzhou Medical University, Huai'an, China.

Yan-Xiang Cheng, Department of Obstetrics and Gynecology, Renmin Hospital of Wuhan University, Wuhan, Hubei 430060, China.

Xiao-Jie Lu, Department of General Surgery, The First Affiliated Hospital of Nanjing Medical University; Sparkfire Scientific Research Group, Nanjing Medical University, Nanjing, China.