The role of insulin-like growth factor-binding protein (IGFBP)-5 in human breast cancer cell growth is unclear. We determined the effects of IGFBP-5 expression on the growth of human breast cancer cell lines in vivo and in vitro. Expression of IGFBP-5, both by stable transfection and adenoviral-mediated infection, was inhibitory to the growth of MDA-MB-231 and Hs578T human breast cancer cells over a 13-day period. IGFBP-5 expression resulted in a G 2 /M cell cycle arrest and the induction of apoptosis in both cell lines, an effect that was abrogated in the presence of the broad-spectrum caspase inhibitor, z-VAD-fmk. IGFBP-5-induced apoptosis was associated with a transcriptional increase in expression of the proapoptotic regulator bax and decrease in the anti-apoptotic bcl-2 compared with vector controls. Secreted IGFBP-5 when added exogenously to breast cancer cells was not internalized and had no effect on cell growth or apoptosis, suggesting that IGFBP-5 may elicit its inhibitory effects via a novel, intracrine mechanism. In athymic nude mice, stable expression of IGFBP-5 significantly inhibited both the formation and growth of tumors derived from MDA-MB-231 cells. IGFBP-5-expressing tumors also had a significantly elevated level of bax mRNA and decreased levels of bcl-2 mRNA compared with vector tumors. These data suggest that IGFBP-5 is a potent growth inhibitor and proapoptotic agent in human breast cancer cells via modulation of cell cycle regulation and apoptotic mediators.The insulin-like growth factor-binding proteins (IGFBPs) 1 are a family of proteins that bind with high affinity to IGFs and modulate their mitogenic actions by regulating their ability to interact with their signaling receptor, the type I IGF receptor (IGFRI, reviewed in Ref. 1). However, it is now becoming clear that many IGFBPs have direct roles in the regulation of cell growth and cell death. For example, we and others have demonstrated the antiproliferative and proapoptotic effects of IGFBP-3 in breast (2-4) and prostate (4) cancer cell lines.The role of IGFBP-5 in cell growth is complex, with reports that it can either stimulate or inhibit cell proliferation in various experimental systems (5-7). There is also evidence that IGFBP-5 expression is up-regulated by antiproliferative agents such as retinoic acid (8), vitamin D-related compounds (9), and antiestrogen ICI 182780 (10), with some evidence that it may mediate their growth inhibitory effects (8, 10). Similarly, growth stimulation of human breast cancer cells by estradiol is associated with a down-regulation of IGFBP-5 expression (10), although exogenous IGFBP-5 had no effect on IGF-I-stimulated DNA synthesis in the breast cancer cell line, MCF-7 (11). They demonstrated that addition of exogenous IGFBP-5 to Hs578T cells protects these cells from ceramide-induced apoptosis, suggesting IGFBP-5 may have a survival function in response to apoptotic stimuli (20,21). A similar conclusion was reached by Roschier et al. (22) following their demonstration that induction of apoptosis...

IGF binding protein (IGFBP)-3 has antiproliferative and proapoptotic effects on the growth of human breast cancer cells in vitro. However, clinical studies suggest that high levels of IGFBP-3 in breast tumor tissue are associated with large, highly proliferative tumors. In this study, we examined the effects of stable transfection with human IGFBP-3 cDNA on the growth of T47D human breast cancer cells in vitro and in vivo. Expression of IGFBP-3 initially inhibited the growth of T47D in vitro but was associated with enhanced growth in vivo. Furthermore, IGFBP-3-expressing cells in vitro became growth stimulated at higher passages post transfection, suggesting breast cancer cells may switch their response to IGFBP-3 with increasing tumorigenicity. These stimulatory effects observed in IGFBP-3-expressing cells were associated with an enhanced responsiveness to the proliferative effects of epidermal growth factor (EGF). When EGF receptor (EGFR) kinase activity was blocked using PD153035, high passage IGFBP-3 transfectants were growth inhibited compared with controls treated with inhibitor. These findings suggest that the interaction between IGFBP-3 and the EGFR system is central to whether IGFBP-3 acts as a growth stimulator or inhibitor in breast cancer cells and that therapies targeting EGFR may have increased efficacy in breast tumors expressing high levels of IGFBP-3.

Expression of IGF-binding protein-3 (IGFBP-3) and IGFBP-5 in human breast cancer cells induces apoptosis and is associated with modulations in Bcl-2 proteins, suggesting that these IGFBPs induce an intrinsic apoptotic pathway. In this study we demonstrate that although both IGFBPs induced the activation of caspase-8 and caspase-9, the expression of IGFBP-5, but not IGFBP-3, sensitized MDA-MB-231 breast cancer cells to the inhibitory effects of TNFalpha. This sensitivity to TNFalpha was associated with a block in nuclear factor-kappaB-mediated cell survival signals. IGFBP-5 expression was also associated with a caspase-8-independent activation of Bid, increased levels of cytosolic second mitochondria-derived activator of caspase (Smac)/direct inhibitor of apoptosis proteins (IAP) binding protein with low pI (DIABLO), and an enhanced phosphorylation of c-Jun N-terminal kinase, both basally and in response to TNFalpha. These results suggest that IGFBP-5 expression may influence extrinsic apoptotic pathways via a differential modulation of downstream cell survival and cell death pathways. Furthermore, although IGFBP-3 and IGFBP-5 share much structural and functional homology, they can modulate distinct apoptotic pathways in human breast cancer cells.

Somatic mutation of the tumor suppressor gene TP53 is reported in at least 50% of human malignancies. Most high-grade serous ovarian cancers (HGSC) have a mutant TP53 allele. Accurate detection of these mutants in heterogeneous tumor tissue is paramount as therapies emerge to target mutant p53. We used a Fluidigm Access Array™ System with Massively Parallel Sequencing (MPS) to analyze DNA extracted from 76 serous ovarian tumors. This dataset has been made available to researchers through the European Genome-phenome Archive (EGA; EGAS00001002200). Herein, we present analyses of this dataset using HaplotypeCaller and MuTect2 through the Broad Institute’s Genome Analysis Toolkit (GATK). We anticipate that this TP53 mutation dataset will be useful to researchers developing and testing new software to accurately determine high and low frequency variant alleles in heterogeneous aneuploid tumor tissue. Furthermore, the analysis pipeline we present provides a valuable framework for determining somatic variants more broadly in tumor tissue.

Background There are limited treatments for triple negative breast cancer (TNBC) patients and an unmet need for targeted approaches in these patients. In the last 4-5 years, the prevalence of high androgen receptor (AR) expression in TNBCs has been noted in up to 50% of tumors suggesting it has clinical relevance. The non-canonical NF-kB pathway is also upregulated in this patient group and it has been reported that that there is crosstalk between these pathways. Therefore the aim of this study was to examine the expression of IKKα and AR in breast cancer tissue samples, to assess if combining these markers increased prognostic power. Methods Immunohistochemistry was performed on tissue microarray of 410 patients to assess proteins level of IKK alpha and AR. Protein expression levels were assessed using the weighted histoscore (WHS) method. The median was employed as the cut off for IKK alpha and 1% as cut off for AR. Expression was analyzed for associations with cancer-specific survival (CSS) and recurrence-free survival (RFS). Results In a cohort of 370 breast cancers nether AR nor IKK alpha alone or combined were associated with CSS or RFS. Stratifying patients by ER status did not impact CSS or RFS. However, in TNBC patients (n=82) high expression of AR was associated with shorter CSS (HR 2.55 95 CI 1.61-5.59, p=0.013). To assess if combining AR and IKK alpha increased prognostic power, AR and IKK alpha were combined into a single score: 0= low expression of both or high expression of one and 1= high expression of both. In the full cohort or when stratified by ER status the score was not associated with CSS or RFS, however in TNBC the combined score potentiated the effect observed with AR alone, (HR 1.68 95 CI 1.20-2.33, p=0.001). Patient CSS was stratified from 11.5 years to 4.6 years and was independently associated with CSS when compared with common clinicopathological factors (HR 1.56 95CI 1.11-2.21, p=0.011). In addition, the combined score was associated with decrease radiotherapy use (p=0.032), increased recurrence rate (p=0.014), decreased cytotoxic T cells (p=0.007), B cells (p=0.043) and macrophages (p=0.037). Conclusions A combined AR and IKK alpha score is an independent prognostic classification for patients with TNBC. Patients with high expression of both AR and IKK alpha a significantly reduced survival and were immune cell cold. This study suggests that this patient group will not benefit from immunotherapy but dual targeting with anti-androgens and IKK alpha selective inhibitors could offer a novel therapeutic strategy for this patient group. Citation Format: Roseweir AK, Khongthong P, Dickson K, Bennett L, Edwards J. Dual targeting of androgen receptor and IKK alpha is a potential therapeutic strategy for triple negative breast cancer [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-10-13.