LAUSR

Development of resistance and insensitivity to target therapy is a growing challenge during clinical treatment for patients with cancer that may require solutions via genome medicine (Gu & Wang 2017). Gene changes such as mutations and heterogeneity could foster cancer evolution and reduce optimal efficacy of single or combinational therapies (Wang, 2016a). The current situation in clinical practice is that gene sequencing is performed to screen whether there are mutated genes which may be associatedwith the occurrence of drug resistance when target drugs fail to show effects in patients (Wu et al. 2017). More recently, gene mutation or heterogeneity are measured in tumor tissues harvested from cancer patients during surgery or biopsies, to select therapeutic strategies of target drugs and indicate the potential occurance of drug resistance. A number of obstacles limit efficient and precise detections of genes responsible for drug resistance during clinical therapy, e.g., intraor inter-tumor heterogeneity between molecules, cells, and locations. Single-cell gene sequencing has been proposed as an alternative to detect intercellular heterogeneity of cancer (Qian et al., 2016; Wang 2015). Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated (Cas9) is considered as an efficient and critical method and solution to modify expression and function of target genes for screening and evaluating target genes or to treat cancer (Fang & Wang, 2016). One of the most important functions is to in parallel measure the large-scale perturbations and phenotypes of individual cells by the integration of single-cell RNAseq with CROP-seq, CRISP-seq, Perturb-seq, and CRISPR/Cas9 as a single-cell CRISPR screen. Pooled CRISPR screening is a system where lentiviral transduction delivers the Cas9 nuclease and single guide RNAs (gRNA) into cells and knock out thousands of individual genes. As an important part of the system, the next-generation sequencing data from cells collected at the start of the screen are analyzed and compared with those at the end of a screen. Pooled CRISPR screening is an important tool and paradigm to identify functionand target-oriented genes in molecular characterization and mechanism of biological process, sensitivity and resistance against drugs, and screening of therapeutic strategies, rather than as a simple readout. The pooled CRISPR screening in mammalian cells was reported as a pooled, loss-of-function genetic screening approach and a powerful tool for systematic genetic analysis using a genome-scale lentiviral single gRNA library (Wang et al., 2014; Shalem et al., 2014). Such an approach can be used for tracking a complex mutant pool by massively parallel sequencing on the basis of the integration of single gRNA expression cassettes into the genome. Dixit et al. (Dixit et al., 2016a, b) developed Perturbseq integrating single-cell RNA-seq and CRISPR-based perturbations to detect complex transcriptional profiles at scale and understand gene function and regulation of transcription factors. This is a milestone study to Cell Biol Toxicol (2017) 33:207–210 DOI 10.1007/s10565-017-9396-7