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It is crucial in the development of cell and gene therapies to understand the impact of the gene editing process on chromosomal structure. When cells are edited, differentiated, modified, or engineered for therapeutic applications such as CAR-T, accurately measuring chromosomal stability and mutation is critical for evaluating genotoxicity. KROMATID’s KROMASURE™ platform of advanced cytogenetic tools enriches the dataset available beyond classic cytogenetic techniques and offers a custom suite of solutions for in-depth single cell analysis of genomic targets of interest.
Understanding risk to patients means evaluating gene therapies for genotoxic effects, alongside testing for efficacy. Because these medicines involve making changes to the genome, the potential for unintentional alterations is top of mind for both developers and regulators.
Standard cytogenetic technology relies on labor-intensive and largely manual processes that analyze only a few cells, usually 20, and produces data that is low-resolution. “Instead, we analyze up to thousands of cells at high-resolution providing key analysis of chromosomal mutations,” says Erin Cross, VP of platform for KROMATID.
Fully analyzing a gene therapy requires information from different scales. “You often want to look at the whole genome, (Figure 1, Figure 2), and then you want to look at the targets—what you’re actually targeting in the genome,” Cross says. “So, I think of coming at it with a set of advanced cytogenetic solutions that are looking at chromosomal mutations across the genome and ones that involve your edit site.”
Analyzing the genomes of edited cells is crucial to understanding the impacts of the process. To do so, scientists often use labeling techniques, such as fluorescence in situ hybridization (FISH). With classical FISH, however, the size of the target is usually hundreds of kilobases or more in a chromosome’s DNA. To address this limitation KROMATID developed a method to analyze genomic targets and their orientation even as small as about 2 kilobases. “The KROMASURE platform becomes impactful for developers assessing the effects of their editing including the transgene knock-in and potentially unintended small structural changes (Figure 3),” says Cross. The same method can also be used to analyze a cell’s entire genome.
In addition, KROMATID’s technology addresses the 20-cell count limitation of conventional cytogenetics and can analyze up to thousands of cells in one sample. This capability can be applied in many ways, including tracking low-prevalence structural variations and the distribution of abnormalities in a batch of edited cells.
This combination of capabilities offers crucial benefits to research scientists or companies that want to develop a novel cell or gene therapy. “These projects include multiplexed strategies for cellular engineering and gene editing, and the scientists want to make sure that the treatments are safe, as well as potent and effective,” Cross says. If a developer connects with KROMATID early in a project, she adds, “they will have confidence they’ve made the right measurements along the way, and several developers have used this set of measurements to support their submission to the FDA for an Investigational New Drug.”
Including cytogenetic characterization in the discovery and development phase of therapeutic development is important for generating robust and redundant data sets around genomic and edit integrity (Figure 4). “When developers engage with us early on, they develop an understanding of their process and outcomes, which provides a comprehensive data set that aligns with the regulatory recommendations and guidelines,” Cross says.
As Cross summarizes KROMATID’s technology, “It is a critical cell-based set of solutions for detecting chromosomal abnormalities and understanding genomic instability in cell and gene therapy development.”
Learn more www.kromatid.com.