Novel Technique for Human Artificial Chromosomes

Human artificial chromosomes (HACs), with the capacity to function within human cells, hold promise for advanced gene therapies targeting certain cancers and various laboratory applications. However, formidable technical challenges have impeded their progress. Now, a team led by researchers at the Perelman School of Medicine at the University of Pennsylvania has achieved a significant breakthrough in this domain, effectively overcoming a common obstacle.

In a study published today in Science, the researchers elucidated how they developed an efficient technique for creating HACs from single, elongated constructs of designer DNA. Previous methods for HAC production were hampered by the tendency of DNA constructs to “multimerize,” forming unpredictably long series with unpredictable rearrangements. The novel approach enables the crafting of HACs with greater speed and precision, thereby accelerating DNA research. Over time, with an effective delivery system, this technique could lead to improved cell therapies for diseases like cancer.

“We essentially revolutionized the conventional approach to Human Artificial Chromosomes design and delivery,” remarked Ben Black, PhD, the Eldridge Reeves Johnson Foundation Professor of Biochemistry and Biophysics at Penn. “The HAC we developed holds significant promise for applications in biotechnology, particularly in large-scale genetic engineering of cells. Additionally, they coexist with natural chromosomes in the cell without necessitating alterations to the natural chromosomes.”

The advent of HACs dates back 25 years, with artificial chromosome technology well-established for simpler chromosomes of lower organisms. However, human chromosomes present unique challenges due to their larger sizes and more complex centromeres. The researchers devised improved Human Artificial Chromosomes with several innovations, including larger initial DNA constructs containing more complex centromeres. They also employed a yeast-cell-based delivery system capable of transporting larger payloads to cells.

Rather than attempting to inhibit multimerization, the researchers circumvented the issue by enlarging the input DNA construct, ensuring it naturally remained in predictable single-copy form.

The researchers demonstrated that their method was significantly more efficient in forming viable HACs compared to standard techniques, producing HACs capable of self-replication during cell division.

The potential advantages of artificial chromosomes are numerous, offering safer and more effective platforms for therapeutic gene expression compared to virus-based delivery systems. Moreover, artificial chromosomes enable the expression of large gene ensembles and facilitate the construction of complex protein machines.

Black anticipates that the approach employed in this study will be instrumental in developing artificial chromosomes for other organisms, including agricultural applications in plants.

Researchers from the J. Craig Venter Institute, the University of Edinburgh, and the Technical University Darmstadt also contributed to the study.

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