The once and future gene therapy

Gene therapy is at an inflection point. Recent successes in genetic medicine have paved the path for a broader second wave of therapies and laid the foundation for next-generation technologies. This comment summarizes recent advances and expectations for the near future.

have a tremendous impact on gene therapy in the future by building on the clinical success of nanoparticle-delivery of siR-NAs and the first approval of an siRNA-based drug, Onpattro for the treatment of hereditary ATTR amyloidosis, in 2018 11 . One possible advantage of nanoparticles is the potential to circumvent detection by the immune system that limits viral delivery. Additionally, chemically defined nanoparticle formulations present unique opportunities for functionalization and tissue targeting that may ultimately be critical to success of in vivo gene transfer outside of the retina and liver.
Beyond the first generation of gene therapies that have focused on delivery of transgenes, gene editing technologies are enabling an entirely new modality for treatments based on precise modification of human genome sequences. While gene editing therapies first entered clinical trials in 2010 as an approach to prevent HIV infection of T cells 12 , the first example of disease-modifying efficacy was demonstrated only in the past year in clinical trials of CRISPRbased gene editing for sickle cell disease and beta-thalassemia (CTX001) 13 . This pioneering success, combined with a promising safety record thus far for gene-edited T cells and HSCs in human trials [13][14][15] , has set the stage for highly anticipated results from ongoing and imminent clinical trials of in vivo genome editing, including a current trial of AAV-based gene editing in the retina (EDIT-101) 16 and a planned trial for non-viral nanoparticle based delivery of CRISPR to the liver (NTLA-2001) 13,17 . Nevertheless, expanding to target tissues outside of the retina and liver comes with many challenges. To this point, the National Institutes of Health has announced a commitment of $190 million over six years to support a Somatic Cell Genome Editing Consortium that will directly address the challenges of delivery, safety, and modeling of systems to advance in vivo genome editing to broadly address human health disorders across diverse tissue types and disease conditions. Assuredly, the products of this Consortium will significantly accelerate the progression of gene editing therapies over the next ten years and beyond.
Current gene editing technologies use nuclease-based systems to cut DNA strands and stimulate DNA repair pathways to introduce desired sequence changes. While these technologies are only currently beginning to be tested clinically, multiple waves of next-generation editing technologies are lined up at the heels of    these efforts to improve specificity, accuracy, efficiency, and applicability to different classes of disease 18,19 . For example, the inventions of base editing and prime editing have enabled the precise alteration of genomic sequences in the absence of DNA breaks and without the reliance on the activity of endogenous DNA repair pathways 18 . RNA-targeted editing technologies allow for transient and reversible modification of gene expression without necessitating permanent changes to genome sequences, potentially leading to greater efficiency and safety 19 . Finally, epigenome editing technologies have the advantage of tunability, reversibility, and the potential for sustained outcomes after transient editor activity that are heritable through cell division 20 .
In parallel to these advanced editing modalities, the roster of possible DNA-targeting systems continues to expand, particularly with the exponentially increasing diversity of CRISPR-Cas systems derived from engineered variants, various bacterial species, and distinct classes of CRISPR targeting mechanisms 19 . The rapid pace of technological innovation in these editing fields is certain to both transform how we currently think about gene therapies but also dramatically broaden the scope of human disease to which these approaches can be applied. Another area of innovation that will dramatically impact the gene therapy field in the near future is functional genomics and our understanding of the regulation of the human genome. For example, the function of~6,000 of the~20,000 human genes is currently unknown 21 . In parallel to enabling gene editing therapies, CRISPR technologies are also facilitating the functional dissection of these gene sequences 22 . Moreover, scientific studies and therapeutic interventions have traditionally focused almost exclusively on genes, even though 98% of our genome consists of non-coding DNA that harbors epigenetic regulators responsible for >90% of susceptibility for common disease 23 . In fact, the first example of therapeutic efficacy of a CRISPR gene editing approach (CTX001) involves the editing of a distal gene regulatory element to alter gene expression, rather than the editing of the underlying genetic mutation, as a strategy to compensate for lost beta globin function in hemoglobinopathies 24 . While efforts such as the NIH ENCODE Consortium have mapped more than two million of these gene regulatory elements across hundreds of human cell types and tissue samples, the function of very few of these sites is known 25 . Annotating this dark matter of the genome will lead to whole new areas of disease biology and classes of therapeutic targets that will enable attacking human disease from an entirely new angle with gene therapy, gene editing, and other modalities.
Remarkably, the rate of technological innovation of gene and cell therapy is significantly outpacing the ability to safely and expeditiously move promising candidates forward in order to benefit patients. Current regulatory models that require large numbers of patients to establish safety and efficacy are not applicable to curative technologies that address a mutation that is found in a single patient or very few patients. One promising strategy is to create a single composition that can treat a larger patient population. Universal cell therapies, which are generated by applying gene editing to engineer "immune stealth" allogeneic donor cells that evade the detection of the host immune system, can be used in both regenerative medicine and adoptive cell immunotherapy 26 . Several clinical trials are currently underway to investigate therapies of this design 13 , and the readout of results from those trials in the near future will significantly shape the future of gene and cell therapy. Despite the promise of this approach, it does not address correction of genetic mutations in vivo and does not leverage the tremendous opportunity and increasing efforts in developing transformative technologies such as base editing and prime editing, which have the potential to correct individual private mutations. In a similar vein, the recent report of an oligonucleotide-based therapy targeted to a private genetic mutation and successful treatment of a human Batten disease patient provides both a potential blueprint and motivation for these efforts 27 . Consequently, one of the greatest changes in the field of cell and gene therapy in the near future will be in the area of regulatory sciences and accommodating the unique challenges posed by these innovative technologies as we move toward personalized therapies.
Gene therapy is arguably the most exciting area of biotechnology at this moment -both due to recent progress and because of the possibilities on the horizon. Unprecedented levels of control over nucleic acid delivery, modulation of the immune system, and precise manipulation of the human genometechnologies not imaginable ten years agowill certainly unlock new areas of medicine over the next ten years. At the same time, this nascent glimpse of a new world of technical capabilities has inspired whole new areas of research, such as synthetic biology, cell reprogramming, and highthroughput functional genomics, which will undoubtedly continue to reshape the face of biomedical research.