Zhuang, Xiaoxuan, Long, Eric O, Long, Eric O., Xiaoxuan Zhuang, Eric O. Long
Abstract
Natural killer (NK) cells are an important component of the cancer immune surveillance system. They are regulated by germline-encoded receptors that activate and inhibit their effector function, such as secretion of cytokines and direct lysis of tumor cells and virus-infected cells. Without the need to be primed by prior exposure to tumor antigen, NK cells can detect ligands expressed on tumor cells and selectively kill these cells. NK cells are under strict control by inhibitory receptors that bind to HLA class I on target cells and block early activation signals, thus preventing lysis of target cells. The sensitivity to lysis by NK cells is therefore determined to a large extent by the expression of HLA class I molecules on tumor cells. In addition to receptor-ligand interactions that occur at NK-target cell synapses, many other factors determine the sensitivity of tumor cells to lysis by NK. Intrinsic properties of tumor cells, such as their metabolism and signaling networks establish a threshold above which they will succumb to the death pathways triggered by NK cell attack. Here we provide a protocol for a genome-wide CRISPR screen in tumor cells to identify factors that regulate their sensitivity to primary human NK cells. Tumor cells first transduced for expression of Cas9 are then transduced with a guide RNA (gRNA) library and co-cultured with NK cells. Deep sequencing of the library generated from the genome of tumor cells that survived the selection by NK cells and analysis of the distribution of guide RNAs is performed to identify genes that promote either sensitivity or resistance to NK-mediated killing. The contribution of individual genes to tumor sensitivity can be validated by knockouts using individual gRNAs. The techniques and workflow described here could be applied to primary tumors from cancer patients and reveal tumor-specific points of vulnerability that could be exploited for cancer immunotherapy, such as checkpoint blockade or expression of chimeric antigen receptors specifically designed to activate NK cell cytotoxicity.
This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 04 April 2022.
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Altmetric has tracked 23,479,361 research outputs across all sources so far. This one is in the 37th percentile – i.e., 37% of other outputs scored the same or lower than it.
So far Altmetric has tracked 13,156 research outputs from this source. They receive a mean Attention Score of 3.4. This one has gotten more attention than average, scoring higher than 67% of its peers.
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We're also able to compare this research output to 791 others from the same source and published within six weeks on either side of this one. This one has done well, scoring higher than 75% of its contemporaries.
