CRISPR, the gene-editing technology and acronym for clustered regularly interspaced short palindromic repeats, and CRISPR-associated protein9 (Cas9) have created much enthusiasm among the scientific community lately. Although not the first tool in the field of gene editing, higher specificity, ease of use and the requirement of inexpensive reagents to selectively modify gene(s) of interest have made the system a potential game changer for research in genome engineering. Despite several applications, from microbial engineering to agriculture, its potential to correct genetic defects in human embryos and implanting healthy embryos in the wombs is why it’s drawing wide interest.
In a nutshell
The latest trigger is a report in the science journal Nature. In the study, researchers claim to have used CRISPR-Cas9 to correct a heterozygous mutation (a type of genetic change where only one copy of the gene is defective) in the MYBPC3 gene for a patient with hypertrophic cardiomyopathy, a type of heart condition in which the muscle wall of the heart thickens abnormally.
Gene editing with CRISPR-Cas9 starts with the introduction of circular DNA plasmids encoding components of the editing machinery into the recipient cells. The editing machinery consists of two parts, the DNA that codes for the Cas9 protein and another one for a specific RNA, called guide RNA or gRNA. In the recipient cell, the gRNA guides the machinery to the site of the genome where the Cas9 protein makes a double-stranded break next to the site of editing, which is subsequently repaired using the recipient cell’s endogenous DNA repair mechanisms. As Cas9 protein is produced from the introduced plasmid DNA, it stays in the recipient cell for a long time, resulting in the protein making cuts at additional unintended sites of the genome, causing ‘off-target’ effects or sometimes undesirable mutations.
In the current study, the researchers’ key innovation involved using ways to obviate such mutations by using a pre-assembled system and ensuring that the Cas9 didn’t stay too long in the cell to stoke damage.
Second, the study reduced the number of mosaics or mixed cell population; where some cells carry the defective gene while others the corrected ones, by injecting the Cas9 protein-gRNA complex into the healthy recipient egg cell while it is being fertilized by the sperm carrying the MYBPC3 gene mutation rather than waiting for a few hours to inject the editing components post fertilization.
Finally, the study showed better overall gene targeting efficiency through microinjection of the editing machinery into the embryos (with 72.2% efficiency) compared to a method where the editing components are introduced into the skin-derived induced pluripotent stem cells (with 27.9% efficiency).
As news on the study’s therapeutic potential spreads, it is important to curtail the noise. Safety, efficacy along with the complex legal, societal and ethical issues need to get sorted before gene editing involving germ cells goes mainstream. India — like the rest of the world — has to pay special attention as both biological and surrogate women can be used to produce healthier babies.
Patients with common genetic disorders in India, like, cystic fibrosis, sickle-cell anaemia, Duchenne muscular dystrophy and with rare genetic diseases like Hirschsprung’s disease and Gaucher’s disease will potentially benefit from the gene editing technology. However, proper policy framework and guidelines need to be in place involving research on genetic modifications in germ cells. We need to make sure that gene-editing in the human embryo does not culminate in live pregnancies but on the other hand encouraging ethical research involving unviable human embryos. In making such policy document, we can take cues from the recent policy statement, published in the 03 August 2017 issue of the American Journal of Human Genetics, and endorsed by 11 genetics organisations around the world.
In a country with mushrooming unregulated clinics practising in vitro fertilization (IVF) and pre-implantation genetic diagnosis (PGD) that often engage in unethical practices and commercial exploitation of potential parents, we need to be vigilant. In this direction, the introduction of The Surrogacy (Regulation) Bill, 2016 in Parliament is timely. It will be prudent to include stringent guidelines to eliminate germ line gene editing for human use altogether by the IVF and PGD clinics, at least for the time being.
Although research related to germ line genetic engineering or reproductive cloning is prohibited in India, putting specifics on gene editing in the proposed National Ethical Guidelines for Biomedical and Health Research involving Human Participants, 2016 — prepared by the Indian Council of Medical Research (ICMR) — and in the Biomedical and Health Research Regulation Bill, 2015 will help strengthen safety measures in research involving human germ cells.
Binay Panda is with Ganit Labs, Bengaluru. The views expressed are personal