Long non-coding RNAs: Fundamental regulators and emerging targets of cancer stem cells.

Cancer is one of the leading causes of death worldwide, primarily due to the dearth of efficient therapies that result in long-lasting remission. This is especially true in cases of metastatic cancer where drug resistance causes the disease to recur after treatment. One of the factors contributing to drug resistance, metastasis, and aggressiveness of the cancer is cancer stem cells (CSCs) or tumor-initiating cells. As a result, CSCs have emerged as a potential target for drug development. In the present review, we have examined and highlighted the lncRNAs with their regulatory functions specific to CSCs. Moreover, we have discussed the difficulties and various methods involved in identifying lncRNAs that can play a particular role in regulating and maintaining CSCs. Interestingly, this review only focuses on those lncRNAs with strong functional evidence for CSC specificity and the mechanistic role that allows them to be CSC regulators and be the focus of CSC-specific drug development.

LASP-1 interacts with ErbB2 in ovarian cancer cells.

LASP-1 was identified as a protein following mass spectrometric analysis of phosphoproteins consequent to signaling by ErbB2 in SKOV-3 cells. It has been previously identified as an oncogene and is located on chromosomal arm 17q 0.76 Mb centromeric to ErbB2. It is expressed in serous ovarian cancer cell lines as a 40 kDa protein. In SKOV-3 cells, it was phosphorylated and was inhibited by Lapatinib and CP7274714. LASP-1 co-immunoprecipitated with ErbB2 in SKOV-3 cells, suggesting a direct interaction. This interaction and phosphorylation were independent of the kinase activity of ErbB2. Moreover, the binding of LASP-1 to ErbB2 was independent of the tyrosine phosphorylation of LASP-1. LASP-1 was neither expressed on the surface epithelium of the normal ovary nor in the fallopian tube. It was expressed in 28% of ovarian tumours (n = 101) that did not significantly correlate with other clinical factors. In tumours from patients with invasive ductal carcinoma of the breast who had ErbB2 amplification (3+), LASP-1 was expressed in 3/20 (P < 0.001). Analysis of the expression of an independent dataset of ovarian and breast tumours from TCGA showed the significant co-occurrence of ErbB2 and LASP-1 (P < 0.01). These results suggest that LASP-1 and ErbB2 interaction could be important in the pathogenesis of ovarian cancer.

The hedgehog pathway regulates cancer stem cells in serous adenocarcinoma of the ovary.

PURPOSE: Signaling by cancer stem cells (CSCs) is known to occur at least in part through conserved developmental pathways. Here, the role of one of these pathways, i.e., the hedgehog pathway, was evaluated in high-grade serous ovarian carcinoma (HGSOC). METHODS AND RESULTS: We found that in HGSOC, hedgehog inhibitors (HHIs) GANT61, LDE225 and GDC0449 reduced or inhibited the formation of spheroids enriched in CSCs. Primary malignant cells (PMCs) in ascites from HGSOC patients cultured in the presence of HHIs showed significant reduction in CSCs. Sonic hedgehog (SHH) significantly increased the expression of ALDH1A1, which was inhibited by GANT61. In the presence of a SHH neutralizing antibody (5E1), a significant reduction in the number of spheroids was observed in HGSOC-derived cell lines. Further, the motility, migration and clonogenic growth of the cells were significantly reduced by HHIs. In the presence of GANT61, a reduction of cells from PMCs in the G0 phase of the cell cycle was observed. The magnitude of difference in expression of Gli1 in tumors from the same HGSOC patients at presentation and at interval debulking surgery was greater in patients who had a recurrence on follow up. GANT61 also significantly inhibited the growth of CSCs in nude mice. Finally, RNA sequencing of HGSOC cells treated with GANT61 showed a significantly reduced expression of CSC markers. CONCLUSIONS: Our results indicate that the hedgehog pathway plays an important role in maintaining the integrity of CSCs in HGSOC and could be a potential therapeutic target.

Expression of cancer stem cell markers CD24, EPHA1 and CD9 and their correlation with clinical outcome in epithelial ovarian tumours.

BACKGROUND: There has been variability between laboratories in the identification of cancer stem cells (CSCs) markers for epithelial ovarian cancer (EOC). We have evaluated three new surface markers for EOC to identify CSCs precisely. METHODS: Three new putative CSCs specific surface markers CD9, CD24 and EPHA1 identified by a bioinformatics approach were evaluated in normal ovary, fallopian tube and ovarian tumours. RESULTS: The expression of CD9 alone was observed in normal ovarian surface epithelium and fallopian tube whereas CD24 and EPHA1 were not expressed (n= 5). CD24 was expressed in all tumours (N= 101) while CD9 and EPHA1 were expressed in 89 and 71 tumours, respectively. The statistical analysis showed significant correlation of the stage of the disease (p< 0.0001), type of surgery (p< 0.0001) and residual disease (p< 0.0001) with overall survival. Although expression of CD9, CD24 and EPHA1 was observed in the majority of tumours there was no significant correlation with outcome. In patients who underwent primary surgery, increased expression of CD24 significantly correlated with poor survival. The expression of CD24 was significantly reduced (p< 0.002) upon analysis of paired sections from patients prior to surgery and at interval debulking surgery (n= 16). CONCLUSION: These findings suggest that overexpression of these new markers may be useful in identifying and targeting ovarian CSCs and CD24 may be a putative CSCs marker in ovarian cancer.

Oncogenes in high grade serous adenocarcinoma of the ovary.

High grade serous ovarian cancer is characterized by relatively few mutations occurring at low frequency, except in TP53. However other genetic aberrations such as copy number variation alter numerous oncogenes and tumor suppressor genes. Oncogenes are positive regulators of tumorigenesis and play a critical role in cancer cell growth, proliferation, and survival. Accumulating evidence suggests that they are crucial for the development and the progression of high grade serous ovarian carcinoma (HGSOC). Though many oncogenes have been identified, no successful inhibitors targeting these molecules and their associated pathways are available. This review discusses oncogenes that have been identified recently in HGSOC using different screening strategies. All the genes discussed in this review have been functionally characterized both in vitro and in vivo and some of them are able to transform immortalized ovarian surface epithelial and fallopian tube cells upon overexpression. However, it is necessary to delineate the molecular pathways affected by these oncogenes for the development of therapeutic strategies.

Cancer stem cells contribute to angiogenesis and lymphangiogenesis in serous adenocarcinoma of the ovary.

The origin of blood and lymphatic vessels in high-grade serous adenocarcinoma of ovary (HGSOC) is uncertain. We evaluated the potential of cancer stem cells (CSCs) in HGSOC to contribute to their formation. Using spheroids as an in vitro model for CSCs, we have evaluated their role in primary malignant cells (PMCs) in ascites from previously untreated patients with HGSOC and cell lines. Spheroids from PMCs grown under specific conditions showed significantly higher expression of endothelial, pericyte and lymphatic endothelial markers. These endothelial and lymphatic cells formed tube-like structures, showed uptake of Dil-ac-LDL and expressed endothelial nitric oxide synthase confirming their endothelial phenotype. Electron microscopy demonstrated classical Weibel-Palade bodies in differentiated cells. Genetically, CSCs and the differentiated cells had a similar identity. Lineage tracking using green fluorescent protein transfected cancer cells in nude mice confirmed that spheroids grown in stem cell conditions can give rise to all three cells. Bevacizumab, a monoclonal antibody that targets vascular endothelial growth factor inhibited the differentiation of spheroids to endothelial cells in vitro. These results suggest that CSCs contribute to angiogenesis and lymphangiogenesis in serous adenocarcinoma of the ovary, which can be inhibited.

Therapeutic antibodies against cancer stem cells: a promising approach.

Monoclonal antibodies have been extensively used to treat malignancy along with routine chemotherapeutic drugs. Chemotherapy for metastatic cancer has not been successful in securing long-term remission of disease. This is in part due to the resistance of cancer cells to drugs. One aspect of the drug resistance is the inability of conventional drugs to eliminate cancer stem cells (CSCs) which often constitute less than 1-2% of the whole tumor. In some tumor types, it is possible to identify these cells using surface markers. Monoclonal antibodies targeting these CSCs are an attractive option for a new therapeutic approach. Although administering antibodies has not been effective, when combined with chemotherapy they have proved synergistic. This review highlights the potential of improving treatment efficacy using functional antibodies against CSCs, which could be combined with chemotherapy in the future.