An integrative genomic and proteomic analysis of PIK3CA, PTEN and AKT mutations in breast cancer

Phosphatidylinositol-3-kinase (PI3K)/AKT pathway aberrations are common in cancer. By applying mass spectroscopy-based sequencing and reverse phase protein arrays to 547 human breast cancers and 41 cell lines, we determined the subtype specificity and signaling effects of PIK3CA, AKT and PTEN mutations, and the effects of PIK3CA mutations on responsiveness to PI3K inhibition in-vitro and on outcome after adjuvant tamoxifen. PIK3CA mutations were more common in hormone receptor positive (33.8%) and HER2-positive (24.6%) than in basal-like tumors (8.3%). AKT1 (1.4%) and PTEN (2.3%) mutations were restricted to hormone receptor-positive cancers with PTEN protein levels also being significantly lower in hormone receptor-positive cancers. Unlike AKT1 mutations, PIK3CA (39%) and PTEN (20%) mutations were more common in cell lines than tumors, suggesting a selection for these but not AKT1 mutations during adaptation to culture. PIK3CA mutations did not have a significant impact on outcome in 166 hormone receptor-positive breast cancer patients after adjuvant tamoxifen. PIK3CA mutations, in comparison with PTEN loss and AKT1 mutations, were associated with significantly less and indeed inconsistent activation of AKT and of downstream PI3K/AKT signaling in tumors and cell lines, and PTEN loss and PIK3CA mutation were frequently concordant, suggesting different contributions to pathophysiology. PTEN loss but not PIK3CA mutations rendered cells sensitive to growth inhibition by the PI3K inhibitor LY294002. Thus, PI3K pathway aberrations likely play a distinct role in the pathogenesis of different breast cancer subtypes. The specific aberration may have implications for the selection of PI3K-targeted therapies in hormone receptor-positive breast cancer

Miz1 and HectH9 regulate the stability of the checkpoint protein, TopBP1

The Myc-associated zinc-finger protein, Miz1, activates transcription of the p21cip1 gene in response to UV irradiation. Miz1 associates with topoisomerase II binding protein1 (TopBP1), an essential activator of the Atr kinase. We show here that Miz1 is required for the recruitment of a fraction of TopBP1 to chromatin, for the protection of TopBP1 from proteasomal degradation and for Atr-dependent signal transduction. TopBP1 that is not bound to chromatin is degraded by the HectH9 (Mule, ARF-BP1 and HUWE1) ubiquitin ligase. Myc antagonizes the binding of TopBP1 to Miz1; as a result, expression of Myc leads to dissociation of TopBP1 from chromatin, reduces the amount of total TopBP1 and attenuates Atr-dependent signal transduction. Our data show that Miz1 and Myc affect the activity of the Atr checkpoint through their effect on TopBP1 chromatin association and stability

ARID1A mutant ovarian clear cell carcinoma: A clear target for synthetic lethal strategies

SWI/SNF chromatin remodeling complexes play an important role in the epigenetic regulation of chromatin structure and gene transcription. Mutual exclusive subunits in the SWI/SNF complex include the DNA targeting members ARID1A and ARID1B as well as the ATPases SMARCA2 and SMARCA4. SWI/SNF complexes are mutated across many cancer types. The highest mutation incidence is found in ARID1A, primarily consisting of deleterious mutations. Current advances have reported synthetic lethal interactions with the loss of ARID1A in several cancer types. In this review, we discuss targets that are only important for tumor growth in an ARID1A mutant context. We focus on synthetic lethal strategies with ARID1A loss in ovarian clear cell carcinoma, a cancer with the highest ARID1A mutation incidence (46-57%). ARID1A directed lethal strategies that can be exploited clinically include targeting of the DNA repair proteins PARP and ATR, and the epigenetic factors EZH2, HDAC2, HDAC6 and BRD2

A functional genetic approach identifies the PI3K pathway as a major determinant of trastuzumab resistance in breast cancer

A large-scale RNA interference screen to discover genes involved in trastuzumab resistance in breast cancer identified only PTEN as a modulator of drug sensitivity. Oncogenic mutants of PIK3CA (activator of the same pathway and frequently mutated in breast cancer) also conferred resistance to trastuzumab in cell culture. In a cohort of 55 breast cancer patients, activation of the PI3K pathway, as judged by the presence of oncogenic PIK3CA mutations or low PTEN expression, was associated with poor prognosis after trastuzumab therapy, and the combined analysis of PTEN and PIK3CA identified twice as many patients at increased risk for progression compared to PTEN alone. Thus, assessment of PI3K pathway activation may provide a biomarker to identify patients unlikely to respond to trastuzumab-based therap

ARID1A mutation sensitizes most ovarian clear cell carcinomas to BET inhibitors

Current treatment for advanced stage ovarian clear cell cancer is severely hampered by a lack of effective systemic therapy options, leading to a poor outlook for these patients. Sequencing studies revealed that ARID1A is mutated in over 50% of ovarian clear cell carcinomas. To search for a rational approach to target ovarian clear cell cancers with ARID1A mutations, we performed kinome-centered lethality screens in a large panel of ovarian clear cell carcinoma cell lines. Using the largest OCCC cell line panel established to date, we show here that BRD2 inhibition is predominantly lethal in ARID1A mutated ovarian clear cell cancer cells. Importantly, small molecule inhibitors of the BET (bromodomain and extra terminal domain) family of proteins, to which BRD2 belongs, specifically inhibit proliferation of ARID1A mutated cell lines, both in vitro and in ovarian clear cell cancer xenografts and patient-derived xenograft models. BET inhibitors cause a reduction in the expression of multiple SWI/SNF members including ARID1B, providing a potential explanation for the observed lethal interaction with ARID1A loss. Our data indicate that BET inhibition may represent a novel treatment strategy for a subset of ARID1A mutated ovarian clear cell carcinomas