A loss-of-function RNA interference screen for molecular targets in cancer.

Published

Journal Article

The pursuit of novel therapeutic agents in cancer relies on the identification and validation of molecular targets. Hallmarks of cancer include self-sufficiency in growth signals and evasion from apoptosis; genes that regulate these processes may be optimal for therapeutic attack. Here we describe a loss-of-function screen for genes required for the proliferation and survival of cancer cells using an RNA interference library. We used a doxycycline-inducible retroviral vector for the expression of small hairpin RNAs (shRNAs) to construct a library targeting 2,500 human genes. We used retroviral pools from this library to infect cell lines representing two distinct molecular subgroups of diffuse large B-cell lymphoma (DLBCL), termed activated B-cell-like DLBCL and germinal centre B-cell-like DLBCL. Each vector was engineered to contain a unique 60-base-pair 'bar code', allowing the abundance of an individual shRNA vector within a population of transduced cells to be measured using microarrays of the bar-code sequences. We observed that a subset of shRNA vectors was depleted from the transduced cells after three weeks in culture only if shRNA expression was induced. In activated B-cell-like DLBCL cells, but not germinal centre B-cell-like DLBCL cells, shRNAs targeting the NF-kappaB pathway were depleted, in keeping with the essential role of this pathway in the survival of activated B-cell-like DLBCL. This screen uncovered CARD11 as a key upstream signalling component responsible for the constitutive IkappaB kinase activity in activated B-cell-like DLBCL. The methodology that we describe can be used to establish a functional taxonomy of cancer and help reveal new classes of therapeutic targets distinct from known oncogenes.

Full Text

Duke Authors

Cited Authors

  • Ngo, VN; Davis, RE; Lamy, L; Yu, X; Zhao, H; Lenz, G; Lam, LT; Dave, S; Yang, L; Powell, J; Staudt, LM

Published Date

  • May 4, 2006

Published In

Volume / Issue

  • 441 / 7089

Start / End Page

  • 106 - 110

PubMed ID

  • 16572121

Pubmed Central ID

  • 16572121

Electronic International Standard Serial Number (EISSN)

  • 1476-4687

Digital Object Identifier (DOI)

  • 10.1038/nature04687

Language

  • eng

Conference Location

  • England