A Genome-wide siRNA Screen Identifies Proteasome Addiction as a Vulnerability of Basal-like Triple-Negative Breast Cancer Cells

Petrocca, Fabio; Altschuler, Gabriel; Tan, Shen Mynn; Mendillo, Marc L.; Yan, Haoheng; Jerry, D. Joseph; Kung, Andrew L.; Hide, Winston; Ince, Tan A.; Lieberman, Judy

doi:10.1016/j.ccr.2013.07.008

Cited by 156 publications

(169 citation statements)

References 45 publications

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“…more than MCF10A cells, 12% validated with 3 of 4 or 4 of 4 individual siRNAs, a validation rate comparable with other genome-wide RNAi studies (19,20).…”

Section: Resultssupporting

confidence: 76%

Synthetic Lethal Screens Identify Vulnerabilities in GPCR Signaling and Cytoskeletal Organization in E-Cadherin–Deficient Cells

Telford

Chen

Beetham

et al. 2015

Molecular Cancer Therapeutics

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The CDH1 gene, which encodes the cell-to-cell adhesion protein E-cadherin, is frequently mutated in lobular breast cancer (LBC) and diffuse gastric cancer (DGC). However, because E-cadherin is a tumor suppressor protein and lost from the cancer cell, it is not a conventional drug target. To overcome this, we have taken a synthetic lethal approach to determine whether the loss of E-cadherin creates druggable vulnerabilities. We first conducted a genome-wide siRNA screen of isogenic MCF10A cells with and without CDH1 expression. Gene ontology analysis demonstrated that G-protein-coupled receptor (GPCR) signaling proteins were highly enriched among the synthetic lethal candidates. Diverse families of cytoskeletal proteins were also frequently represented. These broad classes of E-cadherin synthetic lethal hits were validated using both lentiviral-mediated shRNA knockdown and specific antagonists, including the JAK inhibitor LY2784544, Pertussis toxin, and the aurora kinase inhibitors alisertib and danusertib. Next, we conducted a 4,057 known drug screen and time course studies on the CDH1 isogenic MCF10A cell lines and identified additional drug classes with linkages to GPCR signaling and cytoskeletal function that showed evidence of E-cadherin synthetic lethality. These included multiple histone deacetylase inhibitors, including vorinostat and entinostat, PI3K inhibitors, and the tyrosine kinase inhibitors crizotinib and saracatinib.Together, these results demonstrate that E-cadherin loss creates druggable vulnerabilities that have the potential to improve the management of both sporadic and familial LBC and DGC. Mol Cancer Ther; 14(5); 1213-23. Ó2015 AACR.

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“…more than MCF10A cells, 12% validated with 3 of 4 or 4 of 4 individual siRNAs, a validation rate comparable with other genome-wide RNAi studies (19,20).…”

Section: Resultssupporting

confidence: 76%

Synthetic Lethal Screens Identify Vulnerabilities in GPCR Signaling and Cytoskeletal Organization in E-Cadherin–Deficient Cells

Telford

Chen

Beetham

et al. 2015

Molecular Cancer Therapeutics

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“…Deletion of a catalytic residue confirmed mechanistically that antitumor activity was because of the enzymatic activity of the toxin and not merely disruption of receptor signaling (37). The genetic signatures of MDA-MB-468 and HCC70 cells show close similarities to the signatures of clinical basal-like TNBC cases (38), confirming a clinical relevance to our models.…”

Section: Discussionsupporting

confidence: 63%

“…Basal-like TNBCs have been shown to be sensitive to loss of the anti-apoptotic protein MCL-1 (38), which is rapidly turned over in many cell types (39). Immunotoxin-mediated inhibition of protein synthesis results in rapid MCL-1 depletion (40), which suggests that use of an immunotoxin as part of a combination therapy may uniquely sensitize TNBC to other treatments.…”

Section: Discussionmentioning

confidence: 99%

Targeting a Cancer-Specific Epitope of the Epidermal Growth Factor Receptor in Triple-Negative Breast Cancer

Simon

Antignani

Sarnovsky

et al. 2016

JNCI J Natl Cancer Inst

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Background: Triple-negative breast cancers (TNBCs) are typically more aggressive and result in poorer outcomes than other breast cancers because treatment options are limited due to lack of hormone receptors or amplified human epidermal growth factor receptor 2 (HER2). Many TNBCs overexpress the epidermal growth factor receptor (EGFR) or manifest amplification of the EGFR gene, supporting EGFR as a therapeutic target. While EGFR-directed small molecule inhibitors have shown limited effectiveness in clinical settings, use of EGFR as a mechanism of delivering enzymatic cytotoxins to TNBC has not been demonstrated. Methods: Using the single-chain variable fragment (scFv) of the 806 antibody that binds only cells with overexpressed, misfolded, or mutant variants of the EGFR, a recombinant immunotoxin was engineered through gene fusion with Pseudomonas aeruginosa Exotoxin A (806-PE38). The potency of 806-PE38 on reducing TNBC cell growth in vitro and in xenograft models (n 6) was examined for six TNBC cell lines. All statistical tests were two-sided. Results: 806-PE38 statistically significantly reduced the viability of all tested TNBC lines, with IC 50 values below 10 ng/mL for three of six cell lines, while not affecting cells with wild-type EGFR (IC 50 >300 ng/mL). Systemic treatments with 806-PE38 vs vehicle resulted in statistically significantly reduced tumor burdens (806-PE38 mean ¼ 128 mm 3 [SD ¼ 46 mm 3 ] vs vehicle mean ¼ 749 mm 3 [SD ¼ 395 mm 3 ], P ¼ .001) and increased median survival (806-PE38 median ¼ 82 days vs vehicle median ¼ 50 days, P ¼ .01) in a MDA-MB-468 TNBC mouse xenograft. Deletion of the catalytic residue eliminated both cytotoxic activity in vitro and the reduction in tumor burden and survival (P ¼ .52). Conclusions: These data support the further development of the 806-PE38 immunotoxin as a therapeutic agent for the treatment of patients with EGFR-positive TNBC. Follow-up experiments with combination therapies will be attempted to achieve full remissions.

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“…Overexpression in breast cancer is associated with a high tumor grade and poor survival, 21 and pre-clinical evidence suggests that Mcl-1 represents a promising target for the treatment of breast cancers. 19,22,23 Indeed, the MCL1 gene is the most common genetic amplification (after TP53) that occurs following neoadjuvent therapy in TNBC. 24 Further, Mcl-1 overexpression is implicated as a resistance factor for multiple therapies, including the widely prescribed microtubule-targeted agents paclitaxel and vincristine, 25 and compounds that inhibit the related family members Bcl-2 and Bcl-xL.…”

mentioning

confidence: 99%

Myeloid cell leukemia-1 is an important apoptotic survival factor in triple-negative breast cancer

et al. 2015

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Breast cancer is the second-most frequently diagnosed malignancy in US women. The triple-negative breast cancer (TNBC) subtype, which lacks expression of the estrogen receptor, progesterone receptor and human epidermal growth factor receptor-2, afflicts 15% of patients and is refractory to current targeted therapies. Like many cancers, TNBC cells often deregulate programmed cell death by upregulating anti-apoptotic proteins of the B-cell CLL/lymphoma 2 (Bcl-2) family. One family member, myeloid cell leukemia-1 (Mcl-1), is commonly amplified in TNBC and correlates with a poor clinical prognosis. Here we show the effect of silencing Mcl-1 and Bcl-2-like protein 1 isoform 1 (Bcl-xL) expression on viability in a panel of seventeen TNBC cell lines. Cell death was observed in a subset upon Mcl-1 knockdown. In contrast, Bcl-xL knockdown only modestly reduced viability, indicating that Mcl-1 is a more important survival factor. However, dual silencing of both Mcl-1 and Bcl-xL reduced viability in most cell lines tested. These proliferation results were recapitulated by BH3 profiling experiments. Treatment with a Bcl-xL and Bcl-2 peptide had only a moderate effect on any of the TNBC cell lines, however, co-dosing an Mcl-1-selective peptide with a peptide that inhibits Bcl-xL and Bcl-2 was effective in each line tested. Similarly, the selective Bcl-xL inhibitor WEHI-539 was only weakly cytotoxic across the panel, but sensitization by Mcl-1 knockdown markedly improved its EC 50 . ABT-199, which selectively inhibits Bcl-2, did not synergize with Mcl-1 knockdown, indicating the relatively low importance of Bcl-2 in these lines. Mcl-1 sensitivity is not predicted by mRNA or protein levels of a single Bcl-2 family member, except for only a weak correlation for Bak and Bax protein expression. However Breast cancer is the second-most frequently diagnosed malignancy in US women with 230 000 new cases and 40 000 deaths in 2011. The triple-negative breast carcinoma (TNBC) subtype, which does not express the estrogen receptor (ER) and progesterone receptor (PR) and lacks overexpression of human epidermal growth factor receptor-2 (HER2), afflicts nearly 15% of all breast cancer patients and remains refractory to currently available endocrine and HER2-directed therapies.1,2 The current standard of care for TNBC is radiation and neoadjuvant cytotoxic chemotherapy, and carries a poor clinical prognosis. [3][4][5] As with most cancers, TNBC cells are under metabolic and oncogenic stress and require inhibition of the intrinsic apoptotic pathway for survival. 6 Under normal physiological conditions, this pathway is tightly regulated by both pro-and anti-apoptotic members of the B-cell CLL/lymphoma 2 (Bcl-2) family. Stressors such as DNA damage, hypoxia or oncogenic signaling, cause increased expression or translocation of proapoptotic Bcl-2 family members, such as Bim, Bad and Noxa, to the mitochondria. 7 These proteins subsequently trigger pore formation in the mitochondrial outer membrane via induced multimerization of Bak or Bax, a proc...

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A Genome-wide siRNA Screen Identifies Proteasome Addiction as a Vulnerability of Basal-like Triple-Negative Breast Cancer Cells

Cited by 156 publications

References 45 publications

Synthetic Lethal Screens Identify Vulnerabilities in GPCR Signaling and Cytoskeletal Organization in E-Cadherin–Deficient Cells

Synthetic Lethal Screens Identify Vulnerabilities in GPCR Signaling and Cytoskeletal Organization in E-Cadherin–Deficient Cells

Targeting a Cancer-Specific Epitope of the Epidermal Growth Factor Receptor in Triple-Negative Breast Cancer

Myeloid cell leukemia-1 is an important apoptotic survival factor in triple-negative breast cancer

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