Researchers at the University of Pennsylvania have made a major breakthrough in human artificial chromosome technology, developing a new method to simplify the construction of human artificial chromosomes. This innovation is expected to accelerate DNA research and will have a major impact on gene therapy and biotechnology, providing a reliable alternative to current gene delivery systems and broadening the application potential of genetic engineering in various fields. The researchers pointed out that this technology will improve the efficiency of laboratory research and expand the scope of gene therapy.
Artificial human chromosomes that function within human cells have the potential to revolutionize gene therapy, including treatments for certain cancers, and have many laboratory uses. However, huge technical challenges hinder their development.
Now, a team led by researchers at the University of Pennsylvania Perelman School of Medicine has made a major breakthrough in this area, effectively bypassing a common stumbling block.
In a recent study published in the journal Science, researchers explain how they devised a highly efficient technique to create HACs using a single, long designer DNA construct. Previous methods of making HACs have been limited by the fact that the DNA constructs used to make HACs tend to join together in unpredictable long sequences and unpredictable rearrangements - "multimerization." The new method makes HAC faster and more precise, directly speeding up DNA research. Over time, and coupled with effective delivery systems, this technology could lead to better engineered cell therapies for diseases such as cancer.
Comprehensive revamp of HAC design
"Essentially, we revolutionized the old approach to HAC design and delivery," said Ben Black, Ph.D., the Eldridge-Reeves-Johnson Foundation Professor of Biochemistry and Biophysics at Penn. "The HACs we made are very attractive for eventual deployment in biotech applications, such as those that require large-scale genetic engineering of cells. An added benefit is that they exist alongside the native chromosomes without altering the native chromosomes in the cell."
The first artificial genomes were developed 25 years ago, and artificial genome technology has become very advanced in the smaller, simpler chromosomes of lower organisms such as bacteria and yeast. Human chromosomes are a different story, mainly because they are larger and have more complex centrioles (the central region where the X-shaped chromosome arms connect). Researchers have been able to form small artificial human chromosomes from self-ligating lengths of DNA added to cells, but these DNA-length polymers have unpredictable organization and copy numbers—complicating their therapeutic or scientific use—and the resulting HACs sometimes end up binding to native chromosomal sites in the host cell, making their editing unreliable.
In their study, the Penn Medicine researchers engineered improved HACs through several innovations: These included larger initial DNA constructs containing larger, more complex centrioles, which allowed HACs to form from single copies of these constructs. When delivering to cells, they used a yeast cell-based delivery system that is capable of carrying larger payloads.
"For example, instead of trying to inhibit multimerization, we circumvented the problem by increasing the size of the input DNA construct so that it would naturally tend to maintain a predictable single-copy form," Black said.
The researchers' study shows that their method forms viable HAC more efficiently than standard methods and produces HAC that can reproduce itself during cell division.
Advantages and future applications
The potential advantages of artificial chromosomes are many - assuming they can be easily delivered into cells and function like natural chromosomes. Artificial chromosomes will provide a safer, more efficient, and more durable platform for expressing therapeutic genes than virus-based gene delivery systems, which may trigger immune responses and involve harmful viral insertion into native chromosomes. Normal gene expression in cells also requires many local and long-range regulatory factors that are nearly impossible to replicate artificially outside a chromosome-like environment. Furthermore, artificial chromosomes, unlike relatively narrow viral vectors, allow the expression of large, cooperative gene combinations, such as the construction of complex protein machines.
Black anticipates that the same broad approach his research group used in this study will help create artificial chromosomes for other higher organisms, including plants for agricultural applications such as insect-resistant, high-yielding crops.
Compiled from:ScitechDaily