New technology protects the authenticity of engineered cell lines

Dr. Leonidas Bleris (front) and Dr. Yiorgos Makris, preparing samples in the lab, uses a laminar flow tissue culture hood, a closed cabinet designed to avoid contamination, to work with mammalian cell lines. Bleris and Makris introduced technology to protect custom cell lines from misidentification, cross-contamination and illegal replication. Credit: University of Texas at Dallas

Advances in synthetic biology and genome editing have led to a growing industry developing custom cell lines for medical research. These engineered cell lines, however, can be vulnerable to misidentification, cross-contamination, and illegal replication.

A team of researchers at the University of Texas at Dallas has developed a first-of-its-kind method to create a unique identifier for each copy of a cell line to allow users to verify its authenticity and protect the manufacturer’s intellectual property (IP). Engineers demonstrated the method in a study published online May 4 and in the May 6, print edition. advances in science.

The patent-pending technology is the result of an interdisciplinary collaboration among UT Dallas faculty members. The corresponding authors of the study are Dr. Leonidas Bleris, a professor of bioengineering specializing in genetic engineering, and Dr. Yiorgos Makris, a professor of electrical and computer engineering who specializes in electronic hardware security.

Customized cell lines are used in the development of vaccines and targeted therapies for a range of diseases. The global cell culture market is projected to reach US$41.3 billion by 2026, up from US$22.8 billion in 2021, according to a forecast by market research firm MarketsandMarkets.

The research by UT Dallas engineers to develop unique identifiers for genetically modified cells was inspired by so-called physically non-clonable functions (PUFs) in the electronics industry. A PUF is a physical characteristic that can serve as a unique “fingerprint” for a semiconductor device such as a microprocessor. In semiconductors, PUFs are based on natural variations that occur during the manufacturing process and must meet three requirements: They must have a unique fingerprint, produce the same fingerprint with each measurement, and be virtually impossible to replicate.

To apply this concept to modified cells, the researchers developed a two-step process that takes advantage of the cell’s ability to repair damaged DNA, which is made up of sequences of small molecules called nucleotides.

First, they incorporated a five-nucleotide barcode library into a part of the cell’s genome called a safe harbor, where the modification will not harm the cell. Barcodes alone, however, do not satisfy all three properties of PUFs. In the second step, the researchers used the CRISPR gene editing tool to cut the DNA in the vicinity of the barcode. This action forces the cell to repair its DNA using random nucleotides, a process called nonhomologous error repair. During this repair process, the cell naturally inserts new nucleotides into the DNA and/or deletes others – collectively, these are called indels (insertions/deletions). These random corrections, in combination with barcodes, create a unique pattern of nucleotides that can help distinguish the cell lineage from any other.

“The combination of the barcode with the inherently stochastic cellular error repair process results in a unique, irreproducible fingerprint,” said Bleris, who is also the Cecil H. and Ida Green Professor of Systems Biology Science.

This first generation of CRISPR-designed PUFs provides the means for researchers to confirm that the cells were produced by a particular company or laboratory, a process called attestation of provenance. With more research, engineers aim to develop a method to track the age of a specific copy of a cell lineage.

“Companies that develop cell lines are making a big investment,” said Bleris. “We need a way to differentiate between 1,000 copies of the same product. Although the products are identical, each of them has a unique identifier that cannot be replicated.”

Makris said the business of developing engineered cells is so new that companies are focused on monetizing their investments rather than on attestations of safety and provenance. He said the semiconductor industry was the same in the beginning, until incidents of counterfeiting and tampering highlighted the need for security measures.

“We think this time maybe we can be ahead of the curve and have that capability developed when the industry realizes it needs it,” Makris said. “It will be too late when they realize they were hacked and someone monetized their IP.”

Other study authors include Dr. Yi Li, bioengineering research scientist; Mohammad Mahdi Bidmeshki Ph.D., former postdoctoral researcher in Makris’ lab; Taek Kang, PhD student in biomedical engineering and Eugene McDermott Graduate Fellow; and Chance M. Nowak, a graduate student in bioengineering.

Highly secure, physically non-clonable cryptographic primitives based on interfacial magnetic anisotropy

More information:
Yi Li et al, Genetic physical non-clonable functions in human cells, advances in science (2022). DOI: 10.1126/sciadv.abm4106

Provided by the University of Texas at Dallas

Quote: New technology protects the authenticity of engineered cell lines (2022, June 6) retrieved June 6, 2022 at

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