Building high-resolution synthetic lethal networks: a ‘Google map’ of the cancer cell

Paul, James; Templeton, Shaina D.; Baharani, Akanksha; Freywald, Andrew; Vizeacoumar, Franco J.

doi:10.1016/j.molmed.2014.09.009

Cited by 31 publications

(23 citation statements)

References 95 publications

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“…To show that our optimal conditions eliminate heteroduplex structures, we also used a pooled GeCKO (Genome-scale CRISPR Knock Out) library 19 that has recently become a powerful tool to query the genome for complete loss of function as opposed to shRNA libraries that function in hypomorphic context 4 . As these CRISPR libraries are originally designed from mixed-oligo PCRs, they also suffer from the formation of heteroduplexes structures (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…For example, large-scale, genome-wide screens using pooled shRNA or CRISPR libraries query the genome and subsequent sequencing identifies the unique shRNA or sgRNA sequences that affect cell viability 1–6 . Such methods are increasingly applied to identify therapeutically relevant synthetic lethal targets 4–11 or cancer-specific essential genes 2, 3, 12–20 . These novel interactions reveal potential targetable vulnerabilities of malignant cells and have resulted in the initiation of several clinical trials in the recent past (NCT01791309; NCT01750918; NCT01719380).…”

Section: Introductionmentioning

confidence: 99%

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Enhancing the throughput and multiplexing capabilities of next generation sequencing for efficient implementation of pooled shRNA and CRISPR screens

Islam

Watanabe

Wong

et al. 2017

Sci Rep

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Next generation sequencing is becoming the method of choice for functional genomic studies that use pooled shRNA or CRISPR libraries. A key challenge in sequencing these mixed-oligo libraries is that they are highly susceptible to hairpin and/or heteroduplex formation. This results in polyclonal, low quality, and incomplete reads and reduces sequencing throughput. Unfortunately, this challenge is significantly magnified in low-to-medium throughput bench-top sequencers as failed reads significantly perturb the maximization of sequence coverage and multiplexing capabilities. Here, we report a methodology that can be adapted to maximize the coverage on a bench-top, Ion PGM System for smaller shRNA libraries with high efficiency. This ligation-based, half-shRNA sequencing strategy minimizes failed sequences and is also equally amenable to high-throughput sequencers for increased multiplexing. Towards this, we also demonstrate that our strategy to reduce heteroduplex formation improves multiplexing capabilities of pooled CRISPR screens using Illumina NextSeq 500. Overall, our method will facilitate sequencing of pooled shRNA or CRISPR libraries from genomic DNA and maximize sequence coverage.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Enhancing the throughput and multiplexing capabilities of next generation sequencing for efficient implementation of pooled shRNA and CRISPR screens

Islam

Watanabe

Wong

et al. 2017

Sci Rep

Self Cite

View full text Add to dashboard Cite

show abstract

“…Previous study proposed to identify essential genes in human cancer cell lines according to the expression profile of genetic interaction neighbors [37]. Specifically, one gene becomes more essential if its negative genetic interaction neighbors are inactive and its positive genetic interaction neighbors are active [38]. In this paper, we studied more direct phenotypic growth fitness from chemical genomics assays, instead of gene expression data.…”

Section: Methodsmentioning

confidence: 99%

Network-assisted target identification for haploinsufficiency and homozygous profiling screens

Wang

Peng

2017

PLoS Comput Biol

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Chemical genomic screens have recently emerged as a systematic approach to drug discovery on a genome-wide scale. Drug target identification and elucidation of the mechanism of action (MoA) of hits from these noisy high-throughput screens remain difficult. Here, we present GIT (Genetic Interaction Network-Assisted Target Identification), a network analysis method for drug target identification in haploinsufficiency profiling (HIP) and homozygous profiling (HOP) screens. With the drug-induced phenotypic fitness defect of the deletion of a gene, GIT also incorporates the fitness defects of the gene’s neighbors in the genetic interaction network. On three genome-scale yeast chemical genomic screens, GIT substantially outperforms previous scoring methods on target identification on HIP and HOP assays, respectively. Finally, we showed that by combining HIP and HOP assays, GIT further boosts target identification and reveals potential drug’s mechanism of action.

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“…Development of mechanisms or tools to efficiently utilize these loss-of-function alterations for therapeutic purposes would dramatically expand our options in treatment personalization. In this context, the identification of synthetic lethal (SL) interactions, where suppression of one gene causes lethality only when another gene is also inactivated [2, 3], provides a unique opportunity to target these loss-of-function genetic defects. To exploit this model, we studied an unusual member of the Eph group of receptor tyrosine kinases (RTKs), the EPHB6 receptor.…”

Section: Introductionmentioning

confidence: 99%

Targeting synthetic lethality between the SRC kinase and the EPHB6 receptor may benefit cancer treatment

et al. 2016

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Application of tumor genome sequencing has identified numerous loss-of-function alterations in cancer cells. While these alterations are difficult to target using direct interventions, they may be attacked with the help of the synthetic lethality (SL) approach. In this approach, inhibition of one gene causes lethality only when another gene is also completely or partially inactivated. The EPHB6 receptor tyrosine kinase has been shown to have anti-malignant properties and to be downregulated in multiple cancers, which makes it a very attractive target for SL applications. In our work, we used a genome-wide SL screen combined with expression and interaction network analyses, and identified the SRC kinase as a SL partner of EPHB6 in triple-negative breast cancer (TNBC) cells. Our experiments also reveal that this SL interaction can be targeted by small molecule SRC inhibitors, SU6656 and KX2-391, and can be used to improve elimination of human TNBC tumors in a xenograft model. Our observations are of potential practical importance, since TNBC is an aggressive heterogeneous malignancy with a very high rate of patient mortality due to the lack of targeted therapies, and our work indicates that FDA-approved SRC inhibitors may potentially be used in a personalized manner for treating patients with EPHB6-deficient TNBC. Our findings are also of a general interest, as EPHB6 is downregulated in multiple malignancies and our data serve as a proof of principle that EPHB6 deficiency may be targeted by small molecule inhibitors in the SL approach.

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Building high-resolution synthetic lethal networks: a ‘Google map’ of the cancer cell

Cited by 31 publications

References 95 publications

Enhancing the throughput and multiplexing capabilities of next generation sequencing for efficient implementation of pooled shRNA and CRISPR screens

Enhancing the throughput and multiplexing capabilities of next generation sequencing for efficient implementation of pooled shRNA and CRISPR screens

Network-assisted target identification for haploinsufficiency and homozygous profiling screens

Targeting synthetic lethality between the SRC kinase and the EPHB6 receptor may benefit cancer treatment

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