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Tianzuo Zhan, Niklas Rindtorff, Johannes Betge, Matthias P. Ebert, and Michael Boutros

CRISPR/Cas9 technology has greatly accelerated genome engineering and allows for alterations to genomic sequences and transcriptional modifications. CRISPR/Cas9 promises to advance cancer research by enabling the efficient dissection of tumorigenesis mechanisms, identification of drug targets, and development of cell-based therapies. Large-scale CRISPR/Cas9 screens have identified over 1500 essential genes across many cancer cell lines and are working to determine gene essentiality in more cell lines. Combining CRISPR screening with drug treatment can also identify genes that act synergistically or show resistance, providing insights into cancer responses.

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32 views

Tianzuo Zhan, Niklas Rindtorff, Johannes Betge, Matthias P. Ebert, and Michael Boutros

CRISPR/Cas9 technology has greatly accelerated genome engineering and allows for alterations to genomic sequences and transcriptional modifications. CRISPR/Cas9 promises to advance cancer research by enabling the efficient dissection of tumorigenesis mechanisms, identification of drug targets, and development of cell-based therapies. Large-scale CRISPR/Cas9 screens have identified over 1500 essential genes across many cancer cell lines and are working to determine gene essentiality in more cell lines. Combining CRISPR screening with drug treatment can also identify genes that act synergistically or show resistance, providing insights into cancer responses.

Uploaded by

Miyuki Mey
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Claire Therese P.

Rico
2 BSMT 1
Tuesday 4:00 – 5:00 p.m.

CRISPR/Cas9 or Cancer Research and Therapy


Tianzuo Zhan, Niklas Rindtorff, Johannes Betge, Matthias P. Ebert, and Michael Boutros

SUMMARY
Through the development of CRISPR/Cas9 technologies, genome engineering was greatly accelerated.
CRISPR makes the alteration of the genomic sequence of cells and transcriptional modifications possible.
CRISPR/Cas9 promises to accelerate cancer research by providing an efficient technology to dissect
mechanisms of tumorigenesis, identify targets for drug development, and arm cells for cell-based therapies.
Clustered regularly interspersed short palindromic repeats (CRISPR) were found in E. coli in 1987 and
later on in many other bacteria species. For several years, the role of CRISPR was unknown until 2005 when a
hypothesis was raised that the sequences of CRISPR are part of an adaptive immune system in bacteria.
Further studies show that CRISPR and CRISPR-associated proteins (Cas9) are linked to the adaptive
immunity targeting foreign viral DNA.
CRISPR targeting RNA (crRNA) and the trans-activating RNA (tracrRNA) activate and guide Cas
proteins to bind viral DNA sequences which are then cleaved. Hence, CRISPR has the potential to be utilized
as a tool for genome editing. By 2013, the type II Cas protein from Streptococcus pyogenes was used for RNA
guided DNA cleavage in animal cells for the first time, which laid the foundation for using CRISPR/Cas9 as a
widely applicable genome-editing tool.
CRISPR/Cas9 screens can be used to discover novel targets for cancer therapy wherein a cell
population with a diversity of gene knockouts (a potent and irreversible means to inactivate a gene) needs to
be generated. Efficient single guide RNAs (sgRNAs) are selected for every target gene. The sgRNA library is
then synthesized as a pool of oligonucleotides and are cloned into a generation of viral particles which are
used to infect Cas9-expressing cells so that cells potentially carry a distinct sgRNA cassette and specific gene
knockout. The knockout cells are kept in culture for a fixed period of time and have their genomic DNA
extracted afterwards. Through this, it is possible to monitor the phenotypic effect of specific gene knockouts
within the cell population.
One main goal of pooled CRISPR/Cas9 screens in cancer research is to identify genotype-specific
vulnerabilities. Hence, large-scale CRISPR/Cas9 screens have been conducted to discover essential genes
across many cancer cell lines wherein approximately 1500 essential genes were identified. Since CRISPR
screens have revealed many unknown essential genes, there are ongoing research efforts that apply this
method to determine gene essentiality in a larger panel of cancer cell lines.
Another major application of CRISPR/Cas9 screening is the dissection of chemico-genetic interactions,
which enables insights into how cancer responds to drug treatment. Combining pooled CRISPR screening with
a drug can identify gene knockouts that act synergistically or show resistance to the agent.
Since its development into a genome editing tool, the CRISPR/Cas9 technology has revolutionized
biology by providing a simple and versatile method to manipulate the genome, transcriptome and epigenome
across a broad range of organisms. However the potential of CRISPR/Cas9 for both basic and translational
cancer research is yet beginning to unfold. Hopefully in the future, pooled CRISPR screens will provide a
comprehensive set of essential genes across most cancer cell lines which will enable the extensive
identification of synthetic lethal interactions and facilitate the discovery of novel drug targets.

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