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A chemical-genetic screen reveals a mechanism of resistance to PI3K inhibitors in cancer.

Muellner MK, Uras IZ, Gapp BV, Kerzendorfer C, Smida M, Lechtermann H, Craig-Mueller N, Colinge J, Duernberger G, Nijman SM - Nat. Chem. Biol. (2011)

Bottom Line: However, there has been no systematic approach for analyzing gene-drug interactions in human cells.Applying this approach to breast cancer revealed various synthetic-lethal interactions and drug-resistance mechanisms, some of which were known, thereby validating the method.These data reveal a new mechanism of resistance to PI3K inhibitors with direct clinical implications.

View Article: PubMed Central - PubMed

Affiliation: CeMM-Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

ABSTRACT
Linking the molecular aberrations of cancer to drug responses could guide treatment choice and identify new therapeutic applications. However, there has been no systematic approach for analyzing gene-drug interactions in human cells. Here we establish a multiplexed assay to study the cellular fitness of a panel of engineered isogenic cancer cells in response to a collection of drugs, enabling the systematic analysis of thousands of gene-drug interactions. Applying this approach to breast cancer revealed various synthetic-lethal interactions and drug-resistance mechanisms, some of which were known, thereby validating the method. NOTCH pathway activation, which occurs frequently in breast cancer, unexpectedly conferred resistance to phosphoinositide 3-kinase (PI3K) inhibitors, which are currently undergoing clinical trials in breast cancer patients. NOTCH1 and downstream induction of c-MYC over-rode the dependency of cells on the PI3K-mTOR pathway for proliferation. These data reveal a new mechanism of resistance to PI3K inhibitors with direct clinical implications.

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NOTCH activation renders breast cancer cells resistant to PI3K/mTORC1 inhibition(a) Bar graph showing relative viability of ICN1 or control MCF10A cells treated with PP242 (3.0 μM) for 5 days. Shown is the mean of a representative experiment performed in triplicate and standard deviations. (b) Dose-response analysis of MCF10A cells treated with the indicated concentrations of Everolimus (Rad001) for 5 days. Mean and standard deviations are indicated (c) Box plots of GFP positive cells transduced with an ICN1-ires-GFP virus and treated with BEZ-235 (10 pg/μl) or DMSO for 7 days. Data from three replicates each are shown. (d) Oncomine analysis (see Methods) of NUMB expression in 274 PI3K/mTOR inhibitor sensitive or resistant cell lines. The red boxed area indicates cell lines with lower than median expression of NUMB.
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Figure 3: NOTCH activation renders breast cancer cells resistant to PI3K/mTORC1 inhibition(a) Bar graph showing relative viability of ICN1 or control MCF10A cells treated with PP242 (3.0 μM) for 5 days. Shown is the mean of a representative experiment performed in triplicate and standard deviations. (b) Dose-response analysis of MCF10A cells treated with the indicated concentrations of Everolimus (Rad001) for 5 days. Mean and standard deviations are indicated (c) Box plots of GFP positive cells transduced with an ICN1-ires-GFP virus and treated with BEZ-235 (10 pg/μl) or DMSO for 7 days. Data from three replicates each are shown. (d) Oncomine analysis (see Methods) of NUMB expression in 274 PI3K/mTOR inhibitor sensitive or resistant cell lines. The red boxed area indicates cell lines with lower than median expression of NUMB.

Mentions: To begin to uncover the mechanism whereby activation of NOTCH1 in cells confers resistance to PI3K inhibitors we analyzed one of the main downstream effector pathways of PI3K: the serine-threonine kinase mTOR, which resides in the two distinct protein complexes mTORC1 and mTORC2 29. We found that ICN1 expressing cells were also less sensitive to PP242, an mTOR kinase inhibitor, and Everolimus or Rapamycin, non-ATP competitive mTOR inhibitors that may affect mTORC1 more potently than mTORC2 (Figs. 3a,b, Supplementary Fig. 17) 30. Similarly, ICN1 cells were much less affected by mTOR knockdown than control cells (Supplementary Fig. 18). Together, this indicates that activation of NOTCH1 can bypass the cellular requirement for this growth pathway and that consistent with previous reports, in these cells PI3K inhibitors mainly exert their effect by acting on the mTOR pathway 31.


A chemical-genetic screen reveals a mechanism of resistance to PI3K inhibitors in cancer.

Muellner MK, Uras IZ, Gapp BV, Kerzendorfer C, Smida M, Lechtermann H, Craig-Mueller N, Colinge J, Duernberger G, Nijman SM - Nat. Chem. Biol. (2011)

NOTCH activation renders breast cancer cells resistant to PI3K/mTORC1 inhibition(a) Bar graph showing relative viability of ICN1 or control MCF10A cells treated with PP242 (3.0 μM) for 5 days. Shown is the mean of a representative experiment performed in triplicate and standard deviations. (b) Dose-response analysis of MCF10A cells treated with the indicated concentrations of Everolimus (Rad001) for 5 days. Mean and standard deviations are indicated (c) Box plots of GFP positive cells transduced with an ICN1-ires-GFP virus and treated with BEZ-235 (10 pg/μl) or DMSO for 7 days. Data from three replicates each are shown. (d) Oncomine analysis (see Methods) of NUMB expression in 274 PI3K/mTOR inhibitor sensitive or resistant cell lines. The red boxed area indicates cell lines with lower than median expression of NUMB.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3306898&req=5

Figure 3: NOTCH activation renders breast cancer cells resistant to PI3K/mTORC1 inhibition(a) Bar graph showing relative viability of ICN1 or control MCF10A cells treated with PP242 (3.0 μM) for 5 days. Shown is the mean of a representative experiment performed in triplicate and standard deviations. (b) Dose-response analysis of MCF10A cells treated with the indicated concentrations of Everolimus (Rad001) for 5 days. Mean and standard deviations are indicated (c) Box plots of GFP positive cells transduced with an ICN1-ires-GFP virus and treated with BEZ-235 (10 pg/μl) or DMSO for 7 days. Data from three replicates each are shown. (d) Oncomine analysis (see Methods) of NUMB expression in 274 PI3K/mTOR inhibitor sensitive or resistant cell lines. The red boxed area indicates cell lines with lower than median expression of NUMB.
Mentions: To begin to uncover the mechanism whereby activation of NOTCH1 in cells confers resistance to PI3K inhibitors we analyzed one of the main downstream effector pathways of PI3K: the serine-threonine kinase mTOR, which resides in the two distinct protein complexes mTORC1 and mTORC2 29. We found that ICN1 expressing cells were also less sensitive to PP242, an mTOR kinase inhibitor, and Everolimus or Rapamycin, non-ATP competitive mTOR inhibitors that may affect mTORC1 more potently than mTORC2 (Figs. 3a,b, Supplementary Fig. 17) 30. Similarly, ICN1 cells were much less affected by mTOR knockdown than control cells (Supplementary Fig. 18). Together, this indicates that activation of NOTCH1 can bypass the cellular requirement for this growth pathway and that consistent with previous reports, in these cells PI3K inhibitors mainly exert their effect by acting on the mTOR pathway 31.

Bottom Line: However, there has been no systematic approach for analyzing gene-drug interactions in human cells.Applying this approach to breast cancer revealed various synthetic-lethal interactions and drug-resistance mechanisms, some of which were known, thereby validating the method.These data reveal a new mechanism of resistance to PI3K inhibitors with direct clinical implications.

View Article: PubMed Central - PubMed

Affiliation: CeMM-Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria.

ABSTRACT
Linking the molecular aberrations of cancer to drug responses could guide treatment choice and identify new therapeutic applications. However, there has been no systematic approach for analyzing gene-drug interactions in human cells. Here we establish a multiplexed assay to study the cellular fitness of a panel of engineered isogenic cancer cells in response to a collection of drugs, enabling the systematic analysis of thousands of gene-drug interactions. Applying this approach to breast cancer revealed various synthetic-lethal interactions and drug-resistance mechanisms, some of which were known, thereby validating the method. NOTCH pathway activation, which occurs frequently in breast cancer, unexpectedly conferred resistance to phosphoinositide 3-kinase (PI3K) inhibitors, which are currently undergoing clinical trials in breast cancer patients. NOTCH1 and downstream induction of c-MYC over-rode the dependency of cells on the PI3K-mTOR pathway for proliferation. These data reveal a new mechanism of resistance to PI3K inhibitors with direct clinical implications.

Show MeSH
Related in: MedlinePlus