Long noncoding RNA MAPKAPK5‐AS1 promotes colorectal cancer proliferation by partly silencing p21 expression

Colorectal cancer (CRC) is the third most common malignancy in the world, and long noncoding RNA (lncRNA) plays a critical role in carcinogenesis. Here, we report a novel lncRNA, MAPKAPK5‐AS1, that acts as a critical oncogene in CRC. In addition, we attempted to explore the functions of MAPKAPK5‐AS1 on tumor progression in vitro and in vivo. Quantitative RT‐PCR was used to examine the expression of MAPKAPK5‐AS1 in CRC tissues and cells. Expression of MAPKAPK5‐AS1 was significantly upregulated in 50 CRC tissues, and increased expression of MAPKAPK5‐AS1 was found to be associated with greater tumor size and advanced pathological stage in CRC patients. Knockdown of MAPKAPK5‐AS1 significantly inhibited proliferation and caused apoptosis in CRC cells. We also found that p21 is a target of MAPKAPK5‐AS1. In addition, we are the first to report that MAPKAPK5‐AS1 plays a carcinogenic role in CRC. MAPKAPK5‐AS1 is a novel prognostic biomarker and a potential therapeutic candidate for CRC cancer.     

MET is a potential target for use in combination therapy with EGFR inhibition in triple‐negative/basal‐like breast cancer

MET, a cell surface receptor for hepatocyte growth factor, is involved in the development of triple‐negative/basal‐like breast cancer (TNBC/BLBC). However, its utility as a therapeutic target in this subtype of breast cancer is poorly understood. To evaluate MET fully as a potential therapeutic target for TNBC/BLBC, we investigated the relationship between MET expression and clinical outcomes of patients with breast cancer and the functional effect of MET inhibition. Using automated immunohistochemistry (Ventana), we analyzed MET expression in 924 breast cancer patients with relevant clinicopathologic parameters. BLBC showed the strongest relationship with MET expression (57.5%, p < 0.001). High expression of MET in breast cancer resulted in poor overall survival (p = 0.001) and disease‐free survival (DFS, p = 0.010). MET expression was relatively high in TNBC cell lines, and the silencing of MET via small interfering RNA reduced cell proliferation and migration. We observed reduced TNBC cell viability after treatment with the MET inhibitor PHA‐665752. In the most drug‐resistant cell line, MDA‐MB‐468, which showed elevated epidermal growth factor receptor (EGFR) expression, silencing of EGFR resulted in increased sensitivity to PHA‐665752 treatment. We confirmed that PHA‐665752 synergizes with the EGFR inhibitor erlotinib to decrease the viability of MDA‐MB‐468 cells. TNBC patients coexpressing MET and EGFR showed significantly worse DFS than that in patients expressing EGFR alone (p = 0.021). Our findings strongly suggest that MET may be a therapeutic target in TNBC and that the combined therapy targeting MET and EGFR may be beneficial for the treatment of TNBC/BLBC patients.     

KIFC1 is a novel potential therapeutic target for breast cancer

Kinesin-like protein KIFC1, a normally nonessential kinesin motor, plays a critical role in centrosome clustering in cancer cells and is essential for the survival of cancer cells. Herein, we reported that KIFC1 expression is up-regulated in breast cancer, particularly in estrogen receptor negative, progesterone receptor negative and triple negative breast cancer, and is not associated with epidermal growth factor receptor 2 status. In addition, KIFC1 is highly expressed in all 8 tested human breast cancer cell lines, but is absent in normal human mammary epithelial cells and weakly expressed in 2 human lung fibroblast lines. Moreover, KIFC1 silencing significantly reduced breast cancer cell viability. Finally, we found that PJ34, a potent small molecule inhibitor of poly(ADP-ribose) polymerase, suppressed KIFC1 expression and induced multipolar spindle formation in breast cancer cells, and inhibited cell viability and colony formation within the same concentration range, suggesting that KIFC1 suppression by PJ34 contributes to its anti-breast cancer activity. Together, these results suggest that KIFC1 is a novel promising therapeutic target for breast cancer.     

MicroRNA‐493 is a prognostic factor in triple‐negative breast cancer

Breast cancer is one of the most common malignant diseases in women. Triple‐negative breast cancer (TNBC) shows higher aggressiveness and recurrence rates than other subtypes, and there are no effective targets or tailored treatments for TNBC patients. Thus, finding effective prognostic markers for TNBC could help clinicians in their ability to care for their patients. We used tissue microarrays (TMAs) to detect microRNA‐493 (miR‐493) expression in breast cancer samples. A miRCURY LNA detection probe specific for miR‐493 was used in in situ hybridization assays. Staining results were reviewed by two independent pathologists and classified as high or low expression of miR‐493. Kaplan–Meier survival plots and multivariate Cox analysis were carried out to clarify the relationship between miR‐493 and survival. In the Kaplan–Meier analysis, patients with high miR‐493 expression had better disease‐free survival than patients with low miR‐493 expression. After adjusting for common clinicopathological factors in breast cancer, the expression level of miR‐493 was still a significant prognostic factor in breast cancer. Further subtype analysis revealed that miR‐493 expression levels were only significantly prognostic in TNBC patients. These results were validated in the Molecular Taxonomy of Breast Cancer International Consortium database for overall survival. We proved the prognostic role of miR‐493 in TNBC by using one of the largest breast cancer TMAs available and validated it in a large public RNA sequencing database.     

Subtyping of triple‐negative breast cancer: Implications for therapy

Triple‐negative breast cancer (TNBC) is a heterogeneous disease; gene expression analyses recently identified 6 distinct TNBC subtypes, each of which displays a unique biology. Exploring novel approaches for the treatment of these subtypes is critical, especially because the median survival for women with metastatic TNBC is less than 12 months, and virtually all women with metastatic TNBC ultimately will die of their disease despite systemic therapy. To date, not a single targeted therapy has been approved for the treatment of TNBC, and cytotoxic chemotherapy remains the standard treatment. In this review, the authors discuss recent developments in subtyping TNBC and the current and upcoming therapeutic strategies being explored in an attempt to target TNBC. Cancer 2015;121:8–16 . © 2014 American Cancer Society.     

Role of inhibitor of yes‐associated protein 1 in triple‐negative breast cancer with taxol‐based chemoresistance

Triple‐negative breast cancer (TNBC) is highly clinically aggressive and taxol‐based chemoresistance remains a big TNBC therapeutic problem to be solved. Verteporfin, a small molecular yes‐associated protein 1 (YAP1) inhibitor, is little known as an antitumor drug for TNBC. Our data showed that YAP1 expression was associated with early relapse in tissue samples of patients with TNBC taxol chemoresistance (P < .001). Verteporfin reduced migration and enhanced apoptosis or autophagy of a taxol‐resistant MDA‐MB‐231 cell line in vitro. Knockdown of YAP1 increased epithelial‐mesenchymal transition response in a taxol‐resistant TNBC cell line. In an in vivo experiment, we found that verteporfin was able to shrink tumor weight and volume and decreased Ki67 expression in a taxol‐resistant mouse model. Our results provide evidence that verteporfin could be a chemosensitizer for TNBC patients with taxol‐based treatment.     

c‐Src inhibitor selectively inhibits triple‐negative breast cancer overexpressed Vimentin in vitro and in vivo

Oncogene c‐Src has been found to be a potential target for the treatment of triple‐negative breast cancer (TNBC). However, the therapeutic effects of the c‐Src inhibitor on TNBC patients are controversial compared to those on cell lines. The molecular mechanisms of the inhibitory effects of the c‐Src inhibitor on TNBC remain unclear. Herein, we showed that a specific c‐Src inhibitor, PP2, was effective in inhibiting phosphorylation of c‐Src in 4 cell lines: T‐47D, SK‐BR‐3, SUM1315MO2, and MDA‐MB‐231, regardless of hormone receptors and human epidermal growth factor receptor 2 (HER2) expression levels. Giving PP2 preferentially reduced the S phase of cell cycles and inhibited colony formation in SUM1315MO2 and MDA‐MB‐231, but not in SK‐BR‐3 and T‐47D cells. Furthermore, PP2 effectively blocked cell migration/invasion and epithelial‐mesenchymal transition (EMT) in TNBC cell lines, SUM1315MO2 and MDA‐MB‐231. An EMT biomarker, vimentin, was highly expressed in 2 TNBC cell lines when they were compared with SK‐BR‐3 and T‐47D cells. Further depletion of vimentin by shRNA remarkably attenuated the inhibitory effects of the c‐Src inhibitor on TNBC cells in vitro and in vivo, indicating a crucial action of vimentin to affect the function of c‐Src in TNBC. This study provides an important rationale for the clinic to precisely select TNBC patients who would benefit from c‐Src inhibitor treatment. This finding suggests that traditional markers for TNBC are not sufficient to precisely define this aggressive type of cancer. Vimentin is identified as an important biomarker to enable categorization of TNBC.     

BRCA1/2‐negative hereditary triple‐negative breast cancers exhibit BRCAness

BRCA1/2‐associated breast cancers are sensitive to poly(ADPribose) polymerase (PARP) inhibitors and platinum compounds mainly due to their deficiency in DNA repair via homologous recombination (HR). However, approximately only 15% of triple‐negative breast cancers (TNBCs) are BRCA1/2‐associated. TNBCs that exhibit BRCAness (a phenotype reflecting impaired HR in BRCA1/2‐negative tumors) are also regarded sensitive to PARP inhibitors and platinum compounds. Thus, we hypothesized that hereditary BRCA1/2‐negative TNBCs may exhibit BRCAness. To find a subset of hereditary BRCA1/2‐negative TNBCs among 360 TNBCs, we first identified a group of 41 hereditary TNBCs by analyzing the family histories of the patients. Next, we tested this group for the presence of germline BRCA1/2 mutations, and finally, we compared the expression levels of 120 genes involved in HR and five other major mechanisms of DNA damage repair between BRCA1/2‐associated and BRCA1/2‐negative subgroups of hereditary TNBCs using real‐time PCR arrays. Approximately 73% of the hereditary TNBCs were BRCA1/2‐associated and 27% were BRCA1/2‐negative. The expression levels of the analyzed genes showed no significant differences between these two subgroups indicating the BRCAness of the BRCA1/2‐negative hereditary TNBCs and thereby distinguishing a novel subset of TNBCs as a potential target for PARP inhibitors or platinum‐based therapy. The results show the significance of family history in selecting patients with TNBC for therapies directed at incompetent DNA repair (e.g., PARP inhibitors and/or platinum‐based therapies) and indicate that a relatively simple strategy for broadening the target group for these modes of treatment is to identify patients with hereditary TNBCs.     

Cripto-1 as a novel therapeutic target for triple negative breast cancer

Triple-negative breast cancer (TNBC) presents the poorest prognosis among the breast cancer subtypes and no current standard therapy. Here, we performed an in-depth molecular analysis of a mouse model that establishes spontaneous lung metastasis from JygMC(A) cells. These primary tumors resembled the triple-negative breast cancer (TNBC) both phenotypically and molecularly. Morphologically, primary tumors presented both epithelial and spindle-like cells but displayed only adenocarcinoma-like features in lung parenchyma. The use of laser-capture microdissection combined with Nanostring mRNA and microRNA analysis revealed overexpression of either epithelial and miRNA-200 family or mesenchymal markers in adenocarcinoma and mesenchymal regions, respectively. Cripto-1, an embryonic stem cell marker, was present in spindle-like areas and its promoter showed activity in primary tumors. Cripto-1 knockout by the CRISPR-Cas9 system inhibited tumor growth and pulmonary metastasis. Our findings show characterization of a novel mouse model that mimics the TNBC and reveal Cripto-1 as a TNBC target hence may offer alternative treatment strategies for TNBC.     

Locoregional recurrence of triple‐negative breast cancer after breast‐conserving surgery and radiation

BACKGROUND:

The results of radiation on the local control of triple receptor‐negative breast cancer (negative estrogen [ER], progesterone [PR], and HER‐2/neu receptors) was studied.

METHODS:

Conservative surgery and radiation were used in 753 patients with T1‐T2 breast cancer. Three groups were defined by receptor status: Group 1: ER or PR (+); Group 2: ER and PR (?) but HER‐2 (+); and Group 3: triple‐negative (TN). Factors analyzed were age, menopausal status, race, stage, tumor size, lymph node status, presentation, grade, extensive in situ disease, margins, and systemic therapy. The primary endpoint was 5‐year locoregional recurrence (LRR) isolated or total with distant metastases.

RESULTS:

ER‐ and PR‐negative patients were statistically significantly more likely to be black, have T2 disease, have tumors detectable on both mammography and physical examination, have grade 3 tumors, and receive chemotherapy. There were no significant differences noted with regard to ER? and PR? patients by HER‐2 status. There was a significant difference noted in rates of first distant metastases (3%, 12%, and 7% for Groups 1, 2, and 3, respectively; P = .009). However, the isolated 5‐year LRR was not significantly different (2.3%, 4.6%, and 3.2%, respectively; P = .36) between the 3 groups.

CONCLUSIONS:

Patients with TN breast cancer do not appear to be at a significantly increased risk for isolated LRR at 5 years and therefore remain appropriate candidates for breast conservation. Cancer 2009. © 2009 American Cancer Society.     

Treatment outcomes and clinicopathologic characteristics of triple‐negative breast cancer patients who received platinum‐containing chemotherapy

The aim of this study was to evaluate the role of platinum‐containing chemotherapy for metastatic triple‐negative breast cancer (TNBC) patients in terms of the response rate (RR) and progression‐free survival. A second aim was to characterize the clinical behavior at the time of relapse of TNBC. We retrospectively analyzed the clinical outcomes of patients with metastatic breast cancer who received taxane–platinum chemotherapy as the first‐ or second‐line treatment, focusing on the TN phenotype. In total, 257 patients with metastatic breast cancer received platinum‐containing chemotherapy at Samsung Medical Center from 1999 to 2006. Of these patients, 106 patients with available data on estrogen (ER), progesterone (PgR) and human epidermal growth factor receptor‐2 (HER2) receptor status received taxane–platinum regimen as the first‐ or second‐line treatment. The overall RR of patients with TNBC was 39%. This rate did not differ significantly from those of patients with other phenotypes. The time to death after chemotherapy (19 vs. 50 months, p = 0.037) and overall survival (OS) (21 vs. 56 months, p = 0.030) differed significantly between patients with TNBC and non‐TNBC. TNBC showed a unique locoregional infiltration pattern at relapse, which might reflect its aggressive clinical behavior. Despite the similar response to platinum‐containing chemotherapy, patients with TNBC had a shorter OS than patients with non‐TNBC. The implication of TN phenotype as poor prognostic factor is uncertain, because it needs to be defined whether poor outcome is related to the rapid growing characteristics of tumor itself or the resistance to drug therapy. Further prospective studies are warranted. © 2008 Wiley‐Liss, Inc.     

LINC01605 promotes aerobic glycolysis through lactate dehydrogenase A in triple‐negative breast cancer

Breast cancer is the most prevalent cancer diagnosed in women and the major malignancy that threatens women health, thus we explored the role of long noncoding RNA LINC01605 in triple‐negative breast cancer (TNBC). We collected tissue samples from TNBC patients and cultured breast cancer cells to detect LINC01605 levels by RT‐PCR. We then constructed LINC01605 knockdown and LINC01605 overexpressed TNBC cell lines, cell proliferation was measured by CCK‐8 and colony formation assays, cell migration and invasion were measured by Transwell assay, and aerobic glycolysis of cells was detected. Furthermore, a downstream target gene was found, and its role was confirmed by mouse allogeneic tumor formation. It discovered that LINC01605 expression was significantly increased in TNBC patients, and its high expression predicted a low survival prognosis for TNBC patients. Stable knockdown of LINC01605 remarkably inhibited cell proliferation, migration, and invasion, as well as aerobic glycolysis by inhibiting lactate dehydrogenase A in TNBC cell lines. Notably, knockdown of LINC01605 suppressed in vivo tumor formation and migration in TNBC transplanted mice. In conclusion, targeting long noncoding RNA LINC01605 might serve as a therapeutic candidate strategy to treat patients with TNBC.     

HB‐EGF orchestrates the complex signals involved in triple‐negative and trastuzumab‐resistant breast cancer

A number of therapeutic strategies targeting epidermal growth factor receptor (EGFR) have not always led to success in the present state of breast cancer therapy. Notably, there is currently no way to treat trastuzumab‐resistant and triple‐negative breast cancer (TNBC). Here, we found that heparin‐binding epidermal growth factor‐like growth factor (HB‐EGF), a member of the EGFR ligands, was predominantly expressed in breast cancer and that treatment with crossreacting material 197 (CRM197), a specific inhibitor of HB‐EGF, blocked ERK as well as AKT activation via complexes of EGFR and unknown growth factor receptors in TNBC or through complexes of EGFR and human epidermal growth factor receptor‐2 in trastuzumab‐resistant breast cancer, caused significant cell apoptosis and inhibited tumor growth. Accordingly, we can provide a novel concept that a certain EGFR ligand is recognized as a rational target against breast cancer. In addition, it is plausible that CRM197 could be an effective anticancer agent for molecularly targeted therapies.     

Differential response of triple‐negative breast cancer to a docetaxel and carboplatin‐based neoadjuvant treatment

BACKGROUND:

In this study, the authors evaluated whether a pathologic complete response (pCR) or a clinical complete response (cCR) to neoadjuvant treatment in patients with locally advanced breast cancer differed among the 3 subtypes of breast cancer: triple‐negative breast cancer (TNBC), human epidermal growth factor receptor 2 (HER2)‐positive breast cancer, and hormone receptor‐positive/HER2‐negative breast cancer. Whether a cCR or a pCR was correlated with fewer recurrences and better survival also was investigated.

METHODS:

Patients with stage II/III breast cancer received 4 cycles of neoadjuvant docetaxel and carboplatin (TC) every 3 weeks. Patients with HER2‐positive tumors were randomized to receive either additional weekly trastuzumab preoperatively or TC alone. Postoperatively, all patients received 4 cycles of TC, and all HER2‐positive patients received a total of 52 weeks of trastuzumab. The recurrence‐free survival (RFS) and overall survival (OS) rates at 2 years were reported.

RESULTS:

Seventy‐four patients were enrolled, including 11 patients with TNBC, 30 patients with HER2‐positive tumors, and 33 patients with hormone receptor‐positive/HER2‐negative tumor. The cCR rates were 45.4%, 50% and 40.6% in TNBC, HER2‐positive, and hormone receptor‐positive/HER2‐negative groups, respectively. The pCR rate for the entire group was 26.8%, and patients with TNBC had the best response (54.6%) followed by patients with HER2‐positive tumors (24.1%) and patients with hormone receptor‐positive/HER2‐negative tumors (19.4%; P = .0126). The pCR rate for patients with HER2‐positive tumors improved from 7% to 40% if trastuzumab was added (P = .08). Infiltrating ductal cancer, TNBC, negative estrogen receptor and/or progesterone receptor status, tumor classification predicted a pCR (P ≤ .05). Multivariate analysis using a logistic regression test indicated that tumor type was an independent predictor. The RFS rate for patients who did versus patients who did not achieve a pCR was 93.8% versus 78.4% at 2 years, respectively, and 83.3% versus 58% at 3 years, respectively (P = .1227); whereas, for patients who did versus patients who did not achieve a cCR, the RFS rate was 80.9% versus 83.9%, respectively, at 2 years and 65% versus 64.3%, respectively, at 3 years (P = .999).

CONCLUSIONS:

The current results indicated that the TC combination is promising for the treatment of TNBC. The addition of trastuzumab to TC improved the pCR rate significantly in patients with HER2‐positive breast cancer. Cancer 2010. © 2010 American Cancer Society.     

Long noncoding RNA actin filament‐associated protein 1 antisense RNA 1 promotes malignant phenotype through binding with lysine‐specific demethylase 1 and repressing HMG box‐containing protein 1 in non‐small‐cell lung cancer

The number of documented long noncoding RNAs (lncRNAs) has dramatically increased, and their biological functions and underlying mechanisms in pathological processes, especially cancer, remain to be elucidated. Actin filament‐associated protein 1 antisense RNA 1 (AFAP1‐AS1) is a 6810‐nt lncRNA located on chromosome 4p16.1 that was first reported to be upregulated in esophageal adenocarcinoma tissues and cell lines. Here we reported that AFAP1‐AS1, recruiting and binding to lysine‐specific demethylase 1 (LSD1), was generally overexpressed in human non‐small‐cell lung cancer (NSCLC) tissues using quantitative real‐time PCR. Higher AFAP1‐AS1 expression was significantly correlated with larger tumor size (P = .008), lymph node metastasis (P = .025), higher TNM stage (P = .024), and worse overall survival in NSCLC patients. In vitro experiments revealed that AFAP1‐AS1 downregulation inhibited cell migration and induced apoptosis; AFAP1‐AS1 knockdown also hindered tumorigenesis in vivo. Moreover, mechanistic investigations including RNA immunoprecipitation and ChIP assays validated that AFAP1‐AS1 repressed HMG box‐containing protein 1 (HBP1) expression by recruiting LSD1 to the HBP1 promoter regions in PC‐9 and H1975 cells. Furthermore, HBP1 functions as a tumor suppressor, and its ectopic expression hindered cell proliferation. Rescue assays determined that the oncogenic effect of AFAP1‐AS1 is partially dependent on the epigenetic silencing of HBP1. In conclusion, our results indicate that AFAP1‐AS1 is carcinogenic and that the AFAP1‐AS1/LSD1/HBP1 axis could constitute a new therapeutic direction for NSCLC.     

长链非编码 RNA AFAP1－AS1在乳腺癌中的表达及临床意义

目的：探讨长链非编码 RNA（long non －coding RNA,lncRNA）肌动蛋白纤维相关蛋白1－反义 RNA1（actinfilament －associated protein 1－antisense RNA1,AFAP1－AS1）在乳腺癌肿瘤组织中的表达情况及其临床意义。方法：采用荧光定量 PCR 方法检测76例乳腺癌组织及其对应癌旁组织中 AFAP1－AS1的表达情况。选取表达差异最大的5对组织,结合核浆分离的方法,检测 AFAP1－AS1在乳腺癌细胞中的表达位置。分析AFAP1－AS1表达与乳腺癌主要临床病理特征及患者生存曲线的相关性。通过 siRNA 干扰 AFAP1－AS1在乳腺癌细胞 SKBR －3、MCF －7中的表达,MTT 法检测其表达对乳腺癌细胞增殖的影响。结果：AFAP1－AS1在乳腺癌组织中的表达水平相较癌旁组织明显下调（P ＜0．05）,并且这一表达差异主要发生在细胞质。沉默AFAP1－AS1的表达会促进乳腺癌细胞增殖。同时,AFAP1－AS1在乳腺癌组织中的表达差异与临床分期和淋巴结转移有关（P ＜0．05）,且与患者生存率也明显相关（P ＜0．05）。结论：lncRNA AFAP1－AS1在乳腺癌组织中表达下调,影响乳腺癌细胞增殖,可能与乳腺癌的发生和发展有关,可能成为新的乳腺癌分子标志物。