Transposons: Jumping from the Lab to the Clinic
Transposons, also known as transposable elements (TEs) and “jumping genes”, are sequences of DNA that can move locations within the cell’s genome. First discovered by Barbara McClintock in maize, transposons are present throughout the tree of life in both prokaryotic and eukaryotic cells – in fact, about half of the human genome is derived from transposons. The discovery of transposons introduced not only a new paradigm for understanding genetic evolution, but a powerful new tool for basic research and clinical treatments as well.
How do transposons work?
There are two main classes of transposons, defined by how they move around. Class I transposons, or retrotransposons, use an RNA intermediate that is reverse-transcribed into a cDNA copy, which integrates into the genome in a different place and essentially creates a copy of the transposon. Class II transposons, or DNA transposons, generally encode for a transposase, which can be used to excise the element and facilitate direct DNA movement. Only about 3% of the human genome comprises Class II transposons, with the other ~40-50% occupied by Class I elements.
Class I transposons can be divided into elements that contain long-terminal repeats (LTRs), and those that don’t (non-LTR). The most abundant Class I transposon is LINE-1, a non-LTR Long Interspersed Nuclear Element (LINE), followed by Alu. The most well-known LTR transposons are endogenous retroviruses (ERVs), which make up about 8% of the human genome.
The vast majority of Class II transposons contain terminal inverted repeats (TIRs), which are mirrored sequences that flank the element. These transposons can be subdivided into superfamilies based on shared features like TIRs and transposases. The most well-studied group of Class II transposons is the Tc1/mariner superfamily.
Are transposons beneficial or harmful?
Just as genetic mutations can have potentially positive or negative consequences on the host organism, transposons play a complex role in human evolution. Transposons can contribute to genomic instability and alter gene expression in ways that lead to disease states. Germline insertion mutations caused by transposons can result in a wide range of diseases, including hemophilia, β-thalassemia, neurofibromatosis, and many more.
Despite these risks, most transposons in the human genome don’t cause any trouble at all – this is because they are largely transcriptionally silenced by epigenetic mechanisms such as DNA methylation. Transposons offer a substantial source of genetic diversity that may have led to the development of evolutionarily beneficial genes, and may have also been repurposed as important regulatory elements.
How are transposons biomedically useful?
As a relatively stable and straightforward vector for safe, long-term DNA integration, transposons have been explored as a vector for mediating delivery of gene therapies. DNA, or Class II, transposons are predominantly used because their “cut and paste” mechanism of action is simpler to employ and has lower potential for downstream effects compared to retrotransposons. For one, the transposase enzyme necessary for the reaction is either encoded in the transposon or can easily be supplied secondarily. Second, this mechanism results in only one copy of the desired gene, rather than multiple at different parts of the genome.
Even in the same class of transposons, different systems come with their own benefits and challenges. Transposon systems can differ in their efficiency of insertion, preferred integration sites, tendency to leave a “footprint” sequence at the excision site, length of DNA sequence they can accommodate (cargo capacity), and propensity for inducing mutations or rearrangements.
Two of the most commonly used transposon systems in gene therapy research are known as Sleeping Beauty (SB) and piggyBac. SB was the first transposon system to be effectively used in mammalian cells, with piggyBac emerging as another candidate a few years later. Both of these systems have their own applications for which they are best suited, which can be explored in greater detail in this 2021 review by Hackett et al..
How do transposon systems compare to viral vector-mediated gene delivery?
Viral vectors like adeno- and lentiviruses have largely been the gene delivery system of choice for the last few decades. Viral vectors leverage the natural ability of viruses that can enter host cells and integrate their genetic material into the host genome in order to deliver desired gene cargo. Researchers insert their gene of interest (GOI) into stripped-down versions of the viral delivery vehicle, called a capsid, which lacks potentially dangerous virulent DNA and leaves only the genes necessary for effective delivery and integration of the GOI. The infected cells go on to replicate into cells that contain the desired gene.
Despite their popularity, viral vector-based systems come with several major challenges, including: a tendency to provoke negative immune responses, relatively limited cargo capacity (4-8kb), mutations at the insertion site, and difficulties with scaling up production.
Transposon-based systems show promise in overcoming several of these limitations. As these vectors do not contain viral components, they are safer to generate in the lab and may provoke fewer negative immune responses. They also naturally have large cargo capacities (>100kb), are easier and more cost-effective to produce at scale, and have more predictable integration patterns.
What is the state of clinical research using transposon-based therapies?
In the last 15 years, biomedical applications of transposon systems have migrated from preclinical to clinical territory. The SB system entered clinical trials in 2011, being evaluated as a tool for generating CD19-specific CAR-T cells to treat a subset of advanced non-Hodgkin lymphoma and acute lymphoblastic leukemia patients. In the published Phase I results, Kebriaei et al. successfully used SB to modify T cells to express a chimeric antigen receptor (CAR) specific to the CD19 protein, which is expressed on the surface of most malignant B-cells but not healthy ones. Treatment with these transposon-modified CAR-T cells proved to be safe, long-lasting, and effective. Since this study, CAR-T cells have been further improved as an effective treatment for multiple kinds of hematological cancer, with 6 FDA-approved therapies currently available.
While CAR-T cell therapy in cancer has been the most widely explored and successful application of transposon-based technology thus far, ongoing research in other areas such as rare genetic disorders and neurodegenerative diseases is also promising. In a clinical study examining encapsulated cell biodelivery of nerve growth factor (NGF) as a therapeutic for Alzheimer’s disease, Eyjolfsdottir et al. used the SB transposon system to improve NGF secretion by an implanted device. The therapy was well-tolerated, with SB modification successfully improving NGF release compared to a previous clinical trial, demonstrating an early application of transposon technology to treat diseases of the central nervous system.
Conclusion
Persisting over tens of millions of years and comprising half of the modern human genome, mobile genetic elements called transposons have long been a source of biological innovation. And while genomic re-shuffling can cause disease and dysregulation, it can also seed new regulatory networks and genes with new functions – as well as inspire the creation of profoundly impactful scientific tools. Transposon-based gene editing systems continue to advance both our understanding of basic biology and the development of successful treatments for complex diseases like cancer.