Ho Yong Park

Identification of Developmental Endothelial Locus-1 on Circulating Extracellular Vesicles as a Novel Biomarker for Early Breast Cancer Detection

Purpose: Currently, there are no molecular biomarkers for the early detection of breast cancer. This study focused on identifying surface proteins found on circulating extracellular vesicles (EVs) for detecting early-stage breast cancer. Experimental Design: Circulating EVs, isolated from the plasma of 10 patients with breast cancer (stages I and II) and 5 healthy controls, were analyzed using LC-MS/MS. Developmental endothelial locus-1 protein (Del-1) was selected as a candidate biomarker. Two different ELISAs were used to measure Del-1 in plasma samples from healthy controls (n = 81), patients with breast cancer (n = 269), breast cancer patients after surgical resection (n = 50), patients with benign breast tumors (n = 64), and patients with noncancerous diseases (n = 98) in two cohorts. Results: Plasma Del-1 levels were significantly higher (P < 0.0001) in patients with breast cancer than in all controls and returned to almost normal after tumor removal. The diagnostic accuracy of Del-1 was AUC, 0.961 [95% confidence interval (CI), 0.924–0.983], sensitivity of 94.70%, and specificity of 86.36% in test cohort and 0.968 (0.933–0.988), 92.31%, and 86.62% in validation cohort for early-stage breast cancer by one type of ELISA. Furthermore, Del-1 maintained diagnostic accuracy for patients with early-stage breast cancer using the other type of ELISA [0.946 (0.905–0.972), 90.90%, and 77.14% in the test cohort; 0.943 (0.900–0.971), 89.23%, and 80.99% in the validation cohort]. Conclusions: Del-1 on circulating EVs is a promising marker to improve identification of patients with early-stage breast cancer and distinguish breast cancer from benign breast tumors and noncancerous diseases. Clin Cancer Res; 22(7); 1757–66. ©2015 AACR.

Biological function of long noncoding RNA snaR in HER2-positive breast cancer cells

Purpose: Long noncoding RNA, snaR (small NF90-associated RNA), has been reported to be upregulated in various cancer cell lines. We evaluated the additional role of snaR in HER2-positive breast cancer cell lines. Methods: We explored changes of expression of snaR among the selected long noncoding RNAs which have a potential in cancer proliferation or progression. The proliferation, migration, and invasion of HER2-positive breast cancer cells (SK-BR3) were evaluated by snaR with RNA interruption in 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide, wound-healing assay, and Transwell assay. Results: The expression of snaR was remarkably upregulated in SK-BR3 cell lines together with ANRIL, while the SFMBT2 was downregulated in SK-BR3 cell lines. Although Nespas, 7SK, PSF inhibiting RNA, mascRNA, Hoxa11as, NRON, AK023948, MER11C, p53 mRNA, CAR Intergenic 10, HUC 1 and 2, ZFAS1, SCA8, and SNHG5 were also upregulated and UCA1 was downregulated, the differences were not dominent. Based on the expression result, we explored the functional role of snaR in HER2-positive breast cancer. Downregulation of snaR with small interfering RNA was identified to significanlty inhibit migration as well as proliferation of SK-BR3 cells. Conclusion: In this study, snaR was identified as upregulated and to play a role in cancer progression of HER2-positive breast cancer cells. These results suggest snaR as a potential biomarker for HER2-positive breast cancer.

AQP5 Variants Affect Tumoral Expression of AQP5 and Survival in Patients with Early Breast Cancer

Background: Our previous study showed the association of AQP5 upregulation with cancer proliferation and migration in breast cancer cell lines and with unfavorable prognosis in patients with early breast cancer (EBC). In the current study, we analyzed the association of AQP5 variants or their haplotypes with AQP5 expression and their prognostic impact for survival in patients with EBC. Methods: Three AQP5 polymorphisms (rs74091166, rs3736309, and rs1964676) were selected based on the SNP database and genotyped using the Sequenom MassARRAY in 374 out of 447 patients with EBC in whom AQP5 expression had been investigated in our previous study. Results: The allele frequencies of the selected variants in the current study were similar to those from Asian data previously reported. In a univariate analysis, both rs74091166 and rs1964676 were statistically associated with survival as a dominant model of minor allele. Moreover, a multivariate survival analysis revealed that the CC genotype of rs1964676 is an independent prognostic marker of survival in EBC patients, regardless of stage, tumor subtype, and adjuvant treatment [hazard ratio = 0.399, 0.384, and 0.205; p = 0.021, 0.027, and 0.016 for disease-free survival (DFS), distant DFS, and disease-specific survival, respectively]. In particular, the CT/TT genotype of rs1964676 showed an association with strong expression of AQP5 (58.6 vs. 26.0%; p = 0.001), without any associations with clinical or pathological characteristics including tumor subtype, stage, or histologic grade. Conclusion: The current study suggests AQP5 rs1964676 as a new potential prognostic marker in patients with EBC involved in AQP5 expression.

Del-1 Expression as a Potential Biomarker in Triple-Negative Early Breast Cancer

Objective: A differential diagnostic role for plasma Del-1 was proposed for early breast cancer (EBC) in our previous study. We examined tumoral Del-1 expression and analyzed its prognostic impact among patients with EBC. Methods: Del-1 mRNA expression was assessed in breast epithelial and cancer cells. Meanwhile, the tumoral expression of Del-1 was determined based on tissue microarrays and immunohistochemistry results from 440 patients. Results: While a high Del-1 mRNA expression was found in all the breast cancer cell lines, the expression was significantly higher in MDA-MB-231. Tumoral expression of Del-1 was also significantly associated with a negative expression of estrogen receptor or progesterone receptor, and low expression of Ki-67, particularly in the case of triple-negative breast cancer (TNBC) (p < 0.036). Furthermore, a correlation was found between Del-1 expression and an aggressive histological grade, nuclear mitosis, and polymorphism, suggesting a possible role in tumor progression. In the survival analysis, a worse distant disease-free survival trend was noted for the group overexpressing Del-1. Conclusion: While all the investigated breast cancer cell lines exhibited Del-1 expression, the expression rate and intensity were specifically prominent in TNBC. In addition, based on its relationship to an unfavorable histology and worse survival trend, Del-1 could act as a molecular target in TNBC patients.

Abstract 2781: Exosomal Del-1 as a potent diagnostic marker for breast cancer: A prospective cohort study

Background: We previously demonstrated a diagnostic role of exosomal del-1 with two separated groups of breast cancer patients. In the current study, therefore, we aimed to confirm the diagnostic role in a prospective study with breast cancer by measuring plasma exosomal del-1 before and after surgery. Patients and methods: To identify the optimal time of sampling after surgery, serial blood at day 1, 3, 5, and 7 after surgery was collected from 22 patients with early breast cancer. Thereafter, One hundred fifteen patients with breast cancer who underwent curative surgery were enrolled in the prospective cohort study to compare difference in plasma exosomal del-1 measured by ELISA at the time of diagnosis and post-surgery. Results: Among all 22 patients for optimal sampling time after surgery, exosomal del-1 was higher than 0.5 at the time of diagnosis and then normalized at POD1. Among 115 patients for the confirmatory set, 109 (94.8%) patients showed a normalization of del-1 lower than 0.5 after surgery and 10 patients showed del-1> 0.4 . Median f/u duration of 22 months, 9 patients experienced relapse (4 locoregional and 5 distant), where 3 out of 6 in high group (>0.5), and 2 out of 4 in borderline group (0.4-0.5), and 4 out of 105 in normalized group ( Conclusion: In a prospective cohort study, we confirmed that exosomal del-1 has a potent diagnostic role in breast cancer. Furthermore, del-1 was also identified to dramatically decrease after curative surgery. Our current findings suggest its potential prognostic role as well as diagnostic role in breast cancer patients. Citation Format: Soo Jung Lee, Jeeyeon Lee, Yee Soo Chae, Jin Hyang Jung, Ho Yong park, Moon-Chang Baek, Pyong-Gon Moon. Exosomal Del-1 as a potent diagnostic marker for breast cancer: A prospective cohort study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2781. doi:10.1158/1538-7445.AM2017-2781

Abstract 2797: MiR-137 and MiR-496 target Del-1 and affect triple negative breast cancer progression

Background: Although Del-1 was recently proposed as a new biomarker for early breast cancer in our previous studies, the mechanisms of Del-1 expression are barely understood. In the current study, we selected two microRNAs (miR-137 and - 496), potentially affecting Del-1 expression in breast cancer and examined their impact on Del-1 expression in a variety of breast cancer cell lines to identify their potential role in Del-1 expression and thereby breast cancer development or progression. Methods: Del-1 mRNA and miR-137/- 496 levels were measured by qRT-PCR among breast epithelial (MCF10A) and cancer cells (MDA-MB-231, MCF7, SK-BR3 and T-47D). The effects of miR-137/- 496 on cell proliferation and invasion were detected using MTT, wound healing and Transwell assays. Furthermore, luciferase reporter assay was used to identify the direct regulation of Del-1 by miR-137 or - 496 in MDA-MB-231 cells. Plus, we analyzed the expressions of miR-137 or - 496 and Del-1 mRNA from 20 patients with triple negative early breast cancer. Results: miR-137 and - 496 levels were low in all breast cancer cell lines. As Del-1 mRNA expression was remarkably higher in MDA-MB-231 compared to the other breast cancer cell lines, further functional analyses were done with MDA-MB-231 representing triple negative breast cancer subtype. Both miR-137 and miR-496 were revealed to directly bind at the 3’-UTR of Del-1. Del-1 by Luciferase reporter assay and Del-1 expression was upregulated by inhibitors and reversed by both mimics of both miR-137 and miR-496. Furthermore, both miR-137 and miR-496 were also demonstrated to inhibit cell proliferation, migration and invasion of MDA-MB-231, suggesting that these miRNAs affect cancer progression via Del-1. MiR-137 and miR-496 were remarkably down-regulated in 7 out of 12 triple negative breast cancer tissues, in particular with high Ki67 and high histologic grade. Conclusion: Although Del-1 was recently introduced as a new biomarker for triple negative breast cancer, the mechanisms of Del-1 expression were barely identified. The current study firstly demonstrated that microRNA 137 and 496 are involved in Del-1 regulation by binding at Del-1 gene, affecting cancer progression by altering Del-1 expression. Citation Format: Jeeyeon Lee, Yee Soo Chae, Soo Jung Lee, Jin Hyang Jung, Ho Yong Park, Moon-Chang Baek. MiR-137 and MiR-496 target Del-1 and affect triple negative breast cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2797. doi:10.1158/1538-7445.AM2017-2797

Concordance of preoperative US-guided tattooing of axillary lymph nodes to sentinel lymph nodes and comparison of their pathologic results according to imaging modalities in breast cancer patients.

28 Background: We wanted to know the concordance of preoperative ultrasound (US)-guided tattooing of axillary lymph nodes (ALNs) to sentinel lymph node and to correlate MR and PET-CT findings with the final histologic results.nnnMETHODSnAxillary US examination was performed for all breast cancer patients before sentinel lymph node biopsy. The detected lymph nodes in US were classified as negative (group I, enlarged but image-benign) or positive (group II, image-suspicious) finding for metastases based on US, MRI and PET-CT findings. US-guided tattooing for ALNs was performed preoperatively by injection of 3cc of activated charcoal into the cortex of lymph node and the adjacent soft tissue. We evaluated their concordance to sentinel lymph node and correlated the histologic results of US tattooed LN according to each imaging modality.nnnRESULTSnForty ALNs were tattooed and sentinel nodes corresponded to tattooed nodes in all except one patient with a tattooed non-sentinel node. Ten in group I and 30 in group II on US, 18 in group I and 22 in group II on MR, and 19 in group I and 21 in group II on PET-CT. Eight cases had evidence of metastases in final histology, 2 (20.0%) in group I and 6 (20.0%) in group II on US, 4 (22.2%) in group I and 4 (18.1%) in group II on MR, and 6 (31.6%) in group I and 2 ( 9.5%) in group II on PET-CT.nnnCONCLUSIONSnUS-guided tattooing is a feasible method for marking ALNs. In addition, tattooed lymph nodes correlate well with sentinel nodes, which may obviate the need for additional localization for axillary staging.