Cancer is the 2nd leading cause of death across the globe. It is the cause of around 9.6 million deaths in 2018. Lung, prostate, colorectal, stomach and liver cancer are the most common types of cancer in men, while breast, colorectal, lung, cervical and thyroid cancer are the most common among women. Immunotherapy is effective for cancer treatment that helps your immune system fight cancer. Immune system is our protector as it helps our body fight infections and other diseases. Our Immune system is made up of white blood cells and organs and tissues of the lymph system.

How effective immunotherapies are in treating diseases is dependent on the functionality of T cells, which in turn is determined by the specific genes expressed. Recently, researchers at Duke University developed a screening platform based on CRISPR to identify key epigenetic regulators of human T cell function. They discovered that the transcription factor Basic leucine zipper transcription factor ATF-like 3 (BATF3) plays a central role in reprogramming the expression of several genes. By improving the efficacy of CAR T cells in eliminating cancer cells, BATF3 may aid in the development of more effective T cell-based immunotherapies. The study was published in Nature Genetics.

“It’s a very elegant study. It’s really interesting to see how this field of CRISPR screen is developed here using primary human T cells, which are not the easiest to work with,” said Fredrik Wermeling, an immunologist at the Karolinska Institute who was not involved in the research.

Biomedical engineer and study author Charles Gersbach from Duke University, along with his team, have been developing technologies for years to manipulate the expression of genes in cells. They have previously used these tools to transform fibroblasts into neuronal cells and control cell differentiation in human neuronal and pluripotent stem cell populations.

In their latest study, the researchers focused on exploring the therapeutic application of epigenome editing tools in T cell-based immunotherapies, specifically CAR T cells. These tools may help expand the use of T cell-based therapies beyond the cancer types, such as blood cancers, in which they have been effective.
To achieve this, the team developed a CRISPR-based screening approach in primary human CD8+ T cells. They fused a catalytically inactive Cas9 nuclease with a transcriptional repressor for CRISPR interference (CRISPRi) or an activator for CRISPR activation (CRISPRa). Using guide RNAs, they directed the inactive Cas9 to either silence or activate gene expression.

The scientists analyzed public datasets describing changes in chromatin accessibility as T cells differentiate and identified 120 transcription factors enriched in chromatin regions that change accessibility during T cell differentiation. They then directed CRISPRi and CRISPRa to the genes encoding each of these transcription factors to determine which proteins would act as a master regulator.

By coupling loss-of-function and gain-of-function perturbations, the team identified several potential modulators of T cell function, with BATF3 being one of the most promising. Overexpressing BATF3 regulated a broad network of genes, with some of the upregulated genes relating to T cell survival and some of the downregulated genes associated with T cell exhaustion.

The researchers also found that overexpressing BATF3 enhanced the ability of CAR T cells to eliminate cancer cells in vitro. They then tested whether this strategy worked against tumors using a humanized breast cancer mouse model. The results showed that CAR T cells engineered to overexpress BATF3 reduced tumor size more than standard CAR T cells.

“We knew that BATF3 was important to T cells. What we didn’t know is that by overexpressing it, we could profoundly change their phenotypes and make them incredibly more potent against these tumor models,” Gersbach said.

The team aimed to determine whether overexpression of BATF3 gene leads to positive clinical outcomes. To do this, they compared the transcriptional signatures of T cells with or without BATF3 overexpression with the transcriptional profiles of patients who received CAR T cell therapy in a recent clinical trial. They observed that BATF3 overexpression silenced over 30 percent of the genes associated with nonresponse and activated 20 percent of the genes associated with a positive response to CAR T cell therapy. The researchers believe that establishing these gene expression patterns could prove useful for not only CAR T cells but also cancer vaccines and other T cell-based therapies. The next step for this research is to translate these findings into clinical trials, which Gersbach and his team are already pursuing. “We are very excited that we could actually impact cancer patients’ outcomes with these types of enhancements and interventions,” he said.

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