A Q methodology study on divergent perspectives on CRISPR-Cas9 in the Netherlands.

BACKGROUND: CRISPR-Cas9, a technology enabling modification of the human genome, is developing rapidly. There have been calls for public debate to discuss its ethics, societal implications, and governance. So far, however, little is known about public attitudes on CRISPR-Cas9. This study contributes to a better understanding of public perspectives by exploring the various holistic perspectives Dutch citizens have on CRISPR-Cas9. METHODS: This study used Q methodology to identify different perspectives of Dutch citizens (N = 30) on the use of CRISPR-Cas9. The Q-sort method aims at segmenting audiences based on the structural characteristics of their perspectives. Participants individually ranked 32 statements about CRISPR-Cas9 and discussed their rankings in small groups. By-person factor analysis was performed using PQMethod. Participants' contributions to the discussions were used to further make sense of the audience segments identified. RESULTS: Five perspectives on CRISPR-Cas9 were identified: (1) pragmatic optimism (2) concerned scepticism; (3) normative optimism; (4) enthusiastic support; and (5) benevolent generalism. Each perspective represents a unique position motivated by different ranking rationales. Sorting rationales included improving health, preventing negative impacts on society, and fear of a slippery slope. Overall, there is broad, but not universal support for medical uses of CRISPR-Cas9. CONCLUSIONS: Research on CRISPR-Cas9 should prioritise the broadly supported applications of the technology. Research and public debates on CRISPR-Cas9, its uses, its broader implications, and the governance of CRISPR-Cas9 are recommended. A discourse that includes all perspectives can contribute to the embedding of future uses of CRISPR-Cas9 in society. This study shows that Q methodology followed by group discussions enables citizens to contribute meaningfully to discourses about research.

CRISPR/Cas genome editing to optimize pharmacologically active plant natural products.

Since time immemorial, human use medicinal plants as sources of food, therapy and industrial purpose. Classical biotechnology and recent next-generation sequencing (NGS) techniques have been successfully used to optimize plant-derived natural-products of biomedical significance. Earlier, protein based editing tools viz. zinc-finger nucleases (ZFNs) and transcription activator-like endonucleases (TALENs) have been popularized for transcriptional level genome manipulation. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated9 (Cas9) endonuclease system is an efficient, robust and selective site-directed mutagenesis strategy for RNA-guided genome-editing. CRISPR/Cas9 genome-editing tool employs designed guide-RNAs that identifies a 3 base-pair protospacer adjacent motif (PAM) sequence occurring downstream of the target-DNA. The present review comprehensively complies the recent literature (2010-2020) retrieved from scientific-databases on the application of CRISPR/Cas9-editing-tools as potent genome-editing strategies in medicinal-plants discussing the recent developments, challenges and future-perspectives with notes on broader applicability of the technique in plants and lower-organisms. In plants, CRISPR/Cas-editing has been implemented successfully in relation to crop-yield and stress-tolerance. However, very few medicinal plants have been edited using CRISPR/Cas genome tool owing to the lack of whole-genome and mRNA-sequences and shortfall of suitable transformation and regeneration strategies. However, recently a number of plant secondary metabolic-pathways (viz. alkaloid, terpenoid, flavonoids, phenolic, saponin etc.) have been engineered employing CRISPR/Cas-editing via knock-out, knock-in, point-mutation, fine-tuning of gene-expression and targeted-mutagenesis. This genome-editing tool further extends its applicability incorporating the tools of synthetic- and systems-biology, functional-genomics and NGS to produce genetically-engineered medicinal-crops with advanced-traits facilitating the production of pharmaceuticals and nutraceuticals.

Public Acceptability of Gene Therapy and Gene Editing for Human Use: A Systematic Review.

Gene therapy and gene editing technologies are complex and it can be difficult for the public to understand their possible benefits or side effects. However, patient and public support is critical for the successful adoption of any new technology. Given the recent advances in gene therapy and gene editing, their potential clinical benefits, and the significant attention that has been given to the first-known successful attempt at permanent and heritable changes to the human genome, a systematic review was performed to assess beliefs and attitudes toward gene therapy and gene editing for human use, and to highlight the factors that influence acceptability. A systematic search following Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines was undertaken in April 2018 to identify articles examining opinions and attitudes regarding the acceptability of gene therapy and gene editing. Overall, 1,561 records were retrieved from 4 databases (Ovid Medline, PsycINFO, Scopus, and Web of Science). Duplicates were removed, and titles and abstracts independently screened, leaving 86 full-text articles assessed for eligibility. Following full-text review, 33 were included, with 5 articles added after forward/backward searching. An additional three articles were added following an updated search in March 2019 (total n = 41). Findings from the studies were integrated according to common themes: the impact of demographics; risks versus benefits of success; treatment specifics (e.g., medical vs. other reasons; disease severity and status; somatic vs. germ line; and mode of delivery); moral or ethical issues; and changes with time. In general, perceptions were positive, particularly for medical reasons and fatal diseases, but were also influenced by perceived risk. Somatic therapies had higher levels of acceptability than germ line therapies. While available in various forms, limitations exist in the measurement of perceptions of gene therapy and gene editing. Treatment acceptability is essential for future clinical trials, so it is important for scientists and clinicians to be clear about the risks and benefits of these technologies, and how these are communicated to the public, while encouraging education about genetic therapies to a broad range of individuals.

Designing Preclinical Studies in Germline Gene Editing: Scientific and Ethical Aspects.

Human germline gene editing is often debated in hypothetical terms: if it were safe and efficient, on what further conditions would it then be ethically acceptable? This paper takes another course. The key question is: how can scientists reduce uncertainty about safety and efficiency to a level that may justify initiation of first-time clinical trials? The only way to proceed is by well-designed preclinical studies. However, what kinds of investigation should preclinical studies include and what specific conditions should they satisfy in order to be considered well-designed? It is argued that multispecies and multigenerational animal studies are needed as well as human embryo editing without implantation. In order to be possible to translate to first-time clinical trials, animal studies need to satisfy strict conditions of validity. Moreover, embryo studies intended for translation to first-time clinical trials need to correspond to the animal studies in experimental design (with exception of implantation). Only in this way can uncertainty about risk for harm (safety) and prospect of benefit (efficiency) in first-time clinical trials be reduced to a modest level. If uncertainty is not reduced to such a level, first-time clinical trials in germline gene editing should not be initiated.

Reproductive medicine involving genome editing: clinical uncertainties and embryological needs.

Genome editing based on site-directed nucleases facilitated efficient and versatile genetic modifications in human cells. However, recent reports, demonstrating CRISPR/Cas9-mediated genome editing in human embryos have raised profound concerns worldwide. This commentary explores the clinical justification and feasibility of reproductive medicine using germline genome editing. Despite the perceived utility of reproductive medicine for treating intractable infertility, it is difficult to justify germline genome editing from the perspective of the prospective child. As suggested by the UK legalization regarding mitochondrial donation, the prevention of genetic disease in offspring by genome editing might be acceptable in limited cases of serious or life-threatening conditions, where no alternative medicine is available. Nonetheless, the mosaicism underlying human embryos as well as the off-target effect by artificial nucleases will likely hamper preimplantation genetic diagnosis prior to embryo transfer. Such considerations suggest that this type of reproductive medicine should not be developed toward a clinical application. However, the clinical uncertainties underscore the need for embryology that can address fundamental questions regarding germline aneuploidy and mosaicism using genome editing.