Knock in of the AKT1 E17K mutation in human breast epithelial cells does not recapitulate oncogenic PIK3CA mutations

Lauring, Josh; Cosgrove, David; Fontana, Stefani C.; Gustin, John P.; Konishi, Hiroyuki; Abukhdeir, Abde M.; Garay, Joseph P.; Mohseni, Morassa; Wang, Grace M.; Higgins, Michaela J.; Gorkin, David U.; Reis, Marcelo; Vogelstein, Bert; Polyák, Kornélia; Cowherd, Meredith; Buckhaults, Phillip; Park, Ben Ho

doi:10.1038/onc.2009.516

Cited by 52 publications

(78 citation statements)

References 30 publications

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“…Past studies comparing overexpression of a mutant cDNA to single nucleotide knockin of mutant oncogenes have shown dramatic differences in signaling and transformed phenotypes (11)(12)(13). Additionally, we have shown KRAS G12V mutations, as single copies, have relatively little effect in breast epithelial cells, but have a profound effect on cancerous phenotypes and tumorigenicity when coupled with a PIK3CA oncogenic hotspot mutation (14).…”

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confidence: 65%

HER2 missense mutations have distinct effects on oncogenic signaling and migration

Zabransky

Yankaskas

Cochran

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Recurrent human epidermal growth factor receptor 2 (HER2) missense mutations have been reported in human cancers. These mutations occur primarily in the absence of HER2 gene amplification such that most HER2-mutant tumors are classified as "negative" by FISH or immunohistochemistry assays. It remains unclear whether nonamplified HER2 missense mutations are oncogenic and whether they are targets for HER2-directed therapies that are currently approved for the treatment of HER2 gene-amplified breast cancers. Here we functionally characterize HER2 kinase and extracellular domain mutations through gene editing of the endogenous loci in HER2 nonamplified human breast epithelial cells. In in vitro and in vivo assays, the majority of HER2 missense mutations do not impart detectable oncogenic changes. However, the HER2 V777L mutation increased biochemical pathway activation and, in the context of a PIK3CA mutation, enhanced migratory features in vitro. However, the V777L mutation did not alter in vivo tumorigenicity or sensitivity to HER2-directed therapies in proliferation assays. Our results suggest the oncogenicity and potential targeting of HER2 missense mutations should be considered in the context of cooperating genetic alterations and provide previously unidentified insights into functional analysis of HER2 mutations and strategies to target them. A great success in the treatment of breast cancer has come from the identification of human epidermal growth factor receptor 2 (HER2)/Neu (ERBB2) amplification/overexpression as a targetable driver in ∼20% of breast cancers (1). HER2 is a member of the ErbB family of transmembrane receptor tyrosine kinases, which includes the epidermal growth factor receptor (EGFR/ErbB1), HER3 (ErbB3), and HER4 (ErbB4) (2). Activation of ErbB signaling causes receptor tyrosine autophosphorylation and induces interactions with cytoplasmic signal transduction partners that promote a wide variety of cellular processes including proliferation, motility, and escape from apoptosis. In addition to their key role in normal cellular growth and maintenance, the dysregulation of ErbB receptors has been extensively implicated in the development of numerous cancers (1).Whereas overexpression or amplification of HER2 has been well described to deregulate ErbB signaling, cancer genome sequencing studies have demonstrated that somatic point mutations in the HER2 gene occur in a number of cancers, including 2-4% of breast cancers (3-6). Importantly, these mutations are most often found in patients as single copies without amplification/overexpression of HER2 (HER2-"negative" breast cancers), though HER2 protein expression is often still present. Overexpression studies have implicated a number of these mutations as activating and oncogenic (4, 7-9). Additionally, one mutation, L755S, has been described to be associated with lapatinib resistance when overexpressed (4, 10).Past studies comparing overexpression of a mutant cDNA to single nucleotide knockin of mutant oncogenes have shown dramatic differences ...

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mentioning

confidence: 65%

HER2 missense mutations have distinct effects on oncogenic signaling and migration

Zabransky

Yankaskas

Cochran

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…It is possible that AKT1 or β-catenin mutation alone is able to produce a benign tumor but not a malignant tumor. AKT1 mutation itself has a weak transforming activity (35), which might make the cells proliferate and change morphology but not acquire the capability to further progress to malignancy.…”

Section: Discussionmentioning

confidence: 99%

Whole-exome sequencing identifies recurrent AKT1 mutations in sclerosing hemangioma of lung

Jung

Kim

Lee

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Pulmonary sclerosing hemangioma (PSH) is a benign tumor with two cell populations (epithelial and stromal cells), for which genomic profiles remain unknown. We conducted exome sequencing of 44 PSHs and identified recurrent somatic mutations of AKT1 (43.2%) and β-catenin (4.5%). We used a second subset of 24 PSHs to confirm the high frequency of AKT1 mutations (overall 31/68, 45.6%; p.E17K, 33.8%) and recurrent β-catenin mutations (overall 3 of 68, 4.4%). Of the PSHs without AKT1 mutations, two exhibited AKT1 copy gain. AKT1 mutations existed in both epithelial and stromal cells. In two separate PSHs from one patient, we observed two different AKT1 mutations, indicating they were not disseminated but independent arising tumors. Because the AKT1 mutations were not found to cooccur with β-catenin mutations (or any other known driver alterations) in any of the PSHs studied, we speculate that this may be the singlemost common driver alteration to develop PSHs. Our study revealed genomic differences between PSHs and lung adenocarcinomas, including a high rate of AKT1 mutation in PSHs. These genomic features of PSH identified in the present study provide clues to understanding the biology of PSH and for differential genomic diagnosis of lung tumors.pulmonary sclerosing hemangioma | whole-exome sequencing | AKT1 mutation | copy number alteration

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“…Moreover, the effects of pharmacologic inhibition of the protein in models of breast and urothelial carcinoma with endogenous AKT1 E17K mutation have not been reported. Lauring and colleagues reported that AKT1 E17K had minimal phenotypic consequences and failed to induce ongogenic transformation of MCF10 breast epithelial cells, whereas PIK3CA E545K and PIK3CA H1047R mutations were transforming in the same cells, leading them to suggest that these mutations may not be functionally equivalent, and that cooperating genetic changes are required for AKT1 E17K to transform these cells (17). In breast cancer, PIK3CA and AKT1 mutations tend to be mutually exclusive, which suggests that either these mutations do have some redundancy of function, or that PIK3CA is not cooperative with AKT1 for oncogenic transformation of breast epithelial cells, or indeed progression of breast tumors to malignancy.…”

Section: Discussionmentioning

confidence: 99%

“…The E17K mutation has been shown to constitutively activate AKT1 by increased localization to the plasma membrane, where it stimulates downstream signaling and can transform Rat1 fibroblasts and induce leukemia in mice (15). Although knock-in of AKT1 E17K into isogenic MCF-7 breast cancer cells depleted of endogenous PIK3CA E545K restored proliferation in vitro and tumor growth in vivo (16), knock-in of E17K into the AKT1 gene of nontransformed MCF10A mammary epithelial cells had minimal phenotypic consequences and did not recapitulate the transforming properties induced by somatic cell knock in of PIK3CA hotspot mutations (17). In addition, the consequences of directly inhibiting this target in endogenous models of breast and urinary bladder cancer have not been reported.…”

Section: Introductionmentioning

confidence: 99%

Tumors with AKT1E17K Mutations Are Rational Targets for Single Agent or Combination Therapy with AKT Inhibitors

Davies

Guan²,

Logié

et al. 2015

Molecular Cancer Therapeutics

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AKT1E17K mutations occur at low frequency in a variety of solid tumors, including those of the breast and urinary bladder. Although this mutation has been shown to transform rodent cells in culture, it was found to be less oncogenic than PIK3CA mutations in breast epithelial cells. Moreover, the therapeutic potential of AKT inhibitors in human tumors with an endogenous AKT1 E17K mutation is not known. Expression of exogenous copies of AKT1 E17K in MCF10A breast epithelial cells increased phosphorylation of AKT and its substrates, induced colony formation in soft agar, and formation of lesions in the mammary fat pad of immunodeficient mice. These effects were inhibited by the allosteric and catalytic AKT inhibitors MK-2206 and AZD5363, respectively. Both AKT inhibitors caused highly significant growth inhibition of breast cancer explant models with AKT1 E17K mutation. Furthermore, in a phase I clinical study, the catalytic Akt inhibitor AZD5363 induced partial responses in patients with breast and ovarian cancer with tumors containing AKT1 E17K mutations. In MGH-U3 bladder cancer xenografts, which contain both AKT1 E17K and FGFR3 Y373C mutations, AZD5363 monotherapy did not significantly reduce tumor growth, but tumor regression was observed in combination with the FGFR inhibitor AZD4547. The data show that tumors with AKT1 E17K mutations are rational therapeutic targets for AKT inhibitors, although combinations with other targeted agents may be required where activating oncogenic mutations of other proteins are present in the same tumor.

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Knock in of the AKT1 E17K mutation in human breast epithelial cells does not recapitulate oncogenic PIK3CA mutations

Cited by 52 publications

References 30 publications

HER2 missense mutations have distinct effects on oncogenic signaling and migration

HER2 missense mutations have distinct effects on oncogenic signaling and migration

Whole-exome sequencing identifies recurrent AKT1 mutations in sclerosing hemangioma of lung

Tumors with AKT1E17K Mutations Are Rational Targets for Single Agent or Combination Therapy with AKT Inhibitors

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