The purpose of this study was to understand the characteristics of PDEF protein expression in breast and prostate cancer progression. A polyclonal antibody specific to PDEF was raised and reacted with tissue microarrays consisting of benign breast, in situ ductal, invasive ductal and invasive lobular breast carcinomas. The antibody was also reacted with tissue microarrays including benign prostate, prostate intra-epithelial neoplasias and prostate carcinomas. Increased expression of PDEF was identified in 18%, 50%, 46% and 51% of benign breast tissues, intraductal, invasive ductal and invasive lobular carcinomas, respectively. Importantly, in matched samples of benign breast versus tumor, 90% showed higher expression of PDEF in the tumor tissue. Moreover, in invasive breast carcinomas, increased PDEF expression tended to correlate with Her2/neu over expression. Increased expression of PDEF was also found in 27%, 33% and 40% of benign prostate tissues, PIN samples and prostate adenocarcinomas, respectively. Again, in matching samples of cancer versus benign and cancer versus PIN, 68% and 70% respectively showed increased expression in the malignant tissue. Moreover, PDEF was found to be more highly expressed in tumors with intermediate or high Gleason score compared to low grade tumors (P<0.01). Additionally, R1881 treatment induced PDEF expression in the LNCaP prostate tumor cell line, suggesting regulation of PDEF by androgens in vivo. Together, these results for the first time show frequent increased expression of PDEF protein in breast and prostate tumors and support a role for PDEF in breast and prostate cancer progression.

Summary This phase I dose-escalation trial evaluates the feasibility of single-dose preoperative partial breast irradiation delivered with external beam techniques in early stage breast cancer patients. No acute dose-limiting toxicity was observed at 15, 18, or 21Gy. Paired pre- and post-radiation imaging and tumor biopsies offer unique insight into the biology of breast cancer radiation response. Purpose Women with biologically favorable early stage breast cancer are increasingly treated with accelerated partial breast radiation (PBI). However, treatment-related morbidities have been linked to the large post-operative treatment volumes required for external beam PBI. Relative to external beam delivery, alternative PBI techniques require equipment that is not universally available. To address these issues, we designed a phase I trial utilizing widely available technology to 1) evaluate the safety of a single radiation treatment delivered preoperatively to the small-volume, intact breast tumor and 2) identify imaging and genomic markers of radiation response. Methods Women 55 or older with clinically node negative, ER+ and/or PR+, HER2-, T1 invasive carcinomas or low-intermediate grade in situ disease ≤2cm were enrolled (n=32). Intensity-modulated radiotherapy was used to deliver 15 Gy (n=8), 18 Gy (n=8), or 21Gy (n=16) to the tumor with a 1.5cm margin. Lumpectomy was performed within 10 days. Paired pre- and post-radiation MRI images and patient tumor samples were analyzed. Results No dose-limiting toxicity was observed. At a median follow-up of 23 months, there have been no recurrences. Physician-rated cosmetic outcomes were good/excellent and chronic toxicities were grade 1-2 (fibrosis, hyperpigmentation) in patients receiving preoperative radiation only. Evidence of dose-dependent changes in vascular permeability, cell density, and expression of genes regulating immunity and cell death were seen in response to radiation. Conclusions Preoperative single-dose radiotherapy to intact breast tumors is well-tolerated. Radiation response is marked by early indicators of cell death in this biologically favorable patient cohort. This study represents a first-step towards a novel PBI approach. Preoperative radiation should be tested in future clinical trials as it has the potential to challenge the current treatment paradigm and provide a path forward to identify radiation response biomarkers.

Purpose: In a previous gene expression array study, we identified some 300 genes that were differentially expressed in human epidermal growth factor receptor tyrosine kinase 2 (HER2)p ositive versus HER2-negative breast cancer cells.We have now done validation experiments on a group of three cell membrane components that had previously not been implicated in breast cancer.We also studied the expression of three other cell membrane proteins known to play a role in mammary neoplasia. Experimental Design: By immunohistochemistry, we examined up to 130 archival breast carcinomas for Celsr2, E-cadherin, Kai1, and CD9 expression. The expression levels of NET-6 and TROP-2 were determined by quantitative reverse transcription-PCR in a subset of frozen tumors. We also studied fresh pellets and paraffin-embedded cell buttons of nine human breast cell lines. The relationship between the expression of all six membrane proteins and a variety of pathologic and biological variables, including estrogen receptor, HER2, and epidermal growth factor receptor status, was also examined. The NET-6 gene was transfected into a low-expressing cell line, and the effect on cellular morphology, growth, and invasion in vitro was recorded. Results: Celsr2 was down-regulated in one cell line and in 7% of breast cancers. E-cadherin, Kai1, and CD9 were down-regulated in 35%, 76%, and 79% of tumors, respectively, confirming the important role of these markers in human mammary neoplasia. In breast cancer cell lines and tissues, TROP-2 was generally expressed at low levels, although a few specimens showed relative overexpression. NET-6 levels were lower in HER2-negative breast carcinoma cells. In addition, NET-6 was markedly down-regulated in estrogen receptor^negative breast cancers, and expression was lowest in ''basal-like'' tumors. Ectopic expression of NET-6 in low-expressing MDA-MB-231cells altered cellular morphology, inhibited growth in vitro, and decreased invasion in a Boyden chamber assay. Conclusions: We have confirmed the expression of three new membrane markers that had previously not been implicated in human breast cancer, and one of them (NET-6) was correlated with HER2 and estrogen receptor status. NET-6 levels were decreased in estrogen receptor^negative and high-grade tumors, and ectopic expression of this gene had an inhibitory effect on proliferation and invasion. Thus, NET-6 may represent a novel breast cancer suppressor gene.

BackgroundMYCN amplification or N-Myc overexpression is found in approximately 40% NEPC and up to 20% CRPC patients. N-Myc has been demonstrated to drive disease progression and hormonal therapeutic resistance of NEPC/CRPC. Here, we aim to identify the molecular mechanisms underlying the N-Myc-driven therapeutic resistance and provide new therapeutic targets for those N-Myc overexpressed NEPC/CRPC.MethodsN-Myc overexpressing stable cell lines for LNCaP and C4–2 were generated by lentivirus infection. ADT-induced senescence was measured by SA-β-gal staining in LNCaP cells in vitro and in LNCaP xenograft tumors in vivo. Migration, cell proliferation and colony formation assays were used to measure the cellular response after overexpressing N-Myc or perturbing the miR-421/ATM pathway. CRISPR-Cas9 was used to knock out ATM in C4–2 cells and MTS cell viability assay was used to evaluate the drug sensitivity of N-Myc overexpressing C4–2 cells in response to Enzalutamide and ATM inhibitor Ku60019 respectively or in combination.ResultsN-Myc overexpression suppressed ATM expression through upregulating miR-421 in LNCaP cells. This suppression alleviated the ADT-induced senescence in vitro and in vivo. Surprisingly, N-Myc overexpression upregulated ATM expression in C4–2 cells and this upregulation promoted migration and invasion of prostate cancer cells. Further, the N-Myc-induced ATM upregulation in C4–2 cells rendered the cells resistance to Enzalutamide, and inhibition of ATM by CRISPR-Cas9 knockout or ATM inhibitor Ku60019 re-sensitized them to Enzalutamide.ConclusionsN-Myc differentially regulating miR-421/ATM pathway contributes to ADT resistance and Enzalutamide resistance development respectively. Combination treatment with ATM inhibitor re-sensitizes N-Myc overexpressed CRPC cells to Enzalutamide. Our findings would offer a potential combination therapeutic strategy using ATM kinase inhibitor and Enzalutamide for the treatment of a subset of mCRPC with N-Myc overexpression that accounts for up to 20% CRPC patients.Electronic supplementary materialThe online version of this article (10.1186/s12943-019-0941-2) contains supplementary material, which is available to authorized users.