Researchers advance CRISPR technology to replicate human disease with unprecedented accuracy

研究人員推進 CRISPR 技術以前所未有的準確性複制人類疾病

Generation and validation of the pdCas9a-SAM construct in the Eµ-Myc lymphoma cell line. Schematic representation of the pdCas9a-SAM lentiviral structure. b Western blot analysis of BCL-2 in treated Eµ-Myc lymphoma cell line. EMRK-1184 or MRE-721 Eµ-Myc lymphoma-derived cell lines were transduced with pdCas9a-SAM only or pdCas9a-SAM plus Bcl-2 sgRNA. Cell lysates were harvested from transduced cell lines before or after puromycin selection and examined for expression of the indicated proteins by western blotting. Heat shock protein 70 (HSP70) was probed as a loading control. Two independent experiments were performed with similar results. c, d Viability of Eµ-Myc lymphoma cell lines transduced with pdCas9a-SAM alone or pdCas9a-SAM plus Bcl-2 sgRNA. Cells were treated with the MCL-1 inhibitor S63845 or the BCL-2 inhibitor venetoclax at the indicated drug concentrations. Cell viability was determined at 24 hours by propidium iodide (PI) staining and subsequent flow cytometry analysis. Data are presented as mean ± SD, n = 3 independent experiments. Statistical significance was determined by two-way ANOVA test using EMRK-1184 pdCas9a-SAM or MRE-721 pdCas9a-SAM cells as controls. credit: Nature Communications (2022). DOI: 10.1038/s41467-022-32485-9

A powerful new genome editing technique that allows researchers to replicate human disease with unprecedented accuracy promises to revolutionize the drug discovery process for a range of cancers.

The technology, advanced by the WEHI team, can activate any gene – including those that have been silenced – to explore new drug targets and causes of resistance at an unparalleled level.

For the first time, researchers have replicated an aggressive lymphoma using the unique technique they used to identify a gene responsible for triggering resistance to a new blood cancer treatment currently in use in Australia.

Lymphoma is the most common blood cancer in Australia, with about 6,500 Australians diagnosed each year. Double hit lymphoma (DHL) is an aggressive subtype that affects white blood cells called B lymphocytes or B cells.

First, the research team was able to enhance a genome editing technique, called CRISPR activation, to accurately mimic DHL. Project leader Professor Marco Herold said the team focused on DHL because the disease is difficult to treat, in part due to the lack of effective preclinical modelling.

“Without the ability to model disease, there is limited opportunity to correctly test which drugs work against it in the clinic,” said Professor Herold, who established and leads one of Australia’s most advanced CRISPR laboratories at WEHI.

“This technology is a game-changer for the scientific community and clinical staff as it allows us to model diseases such as DHL and correctly test drug treatments against them for the first time.

“That’s important when you think about the vast array of human diseases that can be better modeled by using this tool.”

The research attracted international attention, and the WEHI team worked closely with researchers at Nanjing University (China) and Genentech (USA), a member of the Roche Group, to develop the technology.

The findings were published in Nature Communications.

engineering resistance

Venetoclax is the result of a research collaboration between WEHI and Roche, Genentech (a member of the Roche Group) and AbbVie, and is based on the Institute’s more than three decades of groundbreaking scientific discoveries. It was developed by Roche, Genentech and AbbVie.

The cancer drug is based on a discovery at WEHI in the late 1980s that a protein called BCL-2 helps cancer cells survive indefinitely. A1 is a pro-survival protein of the BCL-2 family. Activation of this gene has been reported in various forms of cancer, including leukemia, lymphoma, melanoma, gastric cancer and breast cancer.

While A1 is thought to play an important role in cancer progression, Dr. Student and lead author Deng Yexuan said that until now, this had not been confirmed. “If the DHL lymphoma in our model could be killed with venetoclax, we were able to use this to show for the first time that A1 is a major factor in developing resistance to this drug,” Deng said.

While cancers are often triggered by turning on genes, researchers have largely only been able to turn them off in previous models of the disease. Project leader Associate Professor Gemma Kelly said the team was able to engineer resistance because their model could activate any gene – even those that had been silenced.

“Our unprecedented ability to turn on A1 using this model allowed us to identify this protein as a resistance driver,” Associate Professor Kelly said. “Our study will allow more genes to be activated in other models to better understand cancer drivers and, crucially, identify other causes of drug resistance.”

“First” trio

Co-lead author, Ph.D. Sarah Diepstraten said the findings suggest that A1 is a promising drug target for DHL.

“This discovery came about because we were able to create a model for DHL that would allow us to turn on any gene,” said Dr. Deepstraten. “This demonstrates an unparalleled level of power of our technology in modeling human disease and exploring why drug targets work or fail.”

The study, “Generation of a CRISPR activation mouse that enables the modeling ofgressive lymphoma and interrogation of venetoclax resistance,” was published in Nature Communications.

Suspects as therapeutic target for B-cell lymphoma have been ruled out

More information:
Yexuan Deng et al., Generation of CRISPR-activated mice capable of modeling aggressive lymphomas and interrogating venetoclax resistance, Nature Communications (2022). DOI: 10.1038/s41467-022-32485-9

Courtesy of the Walter and Eliza Hall Institute of Medicine

Citation: Researchers advance CRISPR technology to replicate human disease with unprecedented accuracy (September 21, 2022) Retrieved September 21, 2022 from technology-replicate-human.html

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