Recent advances in the roles of exosomal microRNAs in neuroblastoma

Exosomal miRNAs (exo-miRs), universally found in biofluids, tissues, and/or conditioned medium of the cell cultures play a significant role in cell - cell communication, thus driving cancer progression and metastasis. Very few studies have explored the role of exo-miRs in the progression of children’s cancer - neuroblastoma. In this mini review, I briefly summarize the existing literature on the role of exo-miRs in the pathogenesis of neuroblastoma

Metastasis in neuroblastoma: the MYCN question

Oncogenic drivers like MYCN in neuroblastoma subsets continues to present a significant challenge owing to its strong correlation with high-risk metastatic disease and poor prognosis. However, only a limited number of MYCN-regulatory proteins associated with tumor initiation and progression have been elucidated. In this minireview, I summarize the recent progress in understanding the functional role of MYCN and its regulatory partners in neuroblastoma metastasis

Drug resistance and the functional role of microRNAs in neuroblastoma

Neuroblastoma is the most common solid tumor in infants. Most of the children diagnosed above one year of age, shows metastatic disease and poor prognosis. Chemotherapy is one of the principle mode of treatment. However, resistance to drug represent a major clinical obstacle for the effective treatment of cancer. MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression, and dysregulation of miRNAs is commonly observed in different cancer types. Recently, miRNAs are shown to modulate drug resistance; however, the role of miRNAs in drug resistant neuroblastoma is limited and poorly understood. This study is focused on the identification and the functional role of miRNAs in drug resistant neuroblastoma. In paper-I, we performed miRNA-profiling study to elucidate miRNAs that are expressed in the cell lines isolated from the same patients before (parental) and after chemotherapy (resistant). We observed differential expression of 42-miRNAs (34-downregulated and 8-upregulated) across parental and resistant cells. Interestingly, miR-376c-3p and miR-323a-3p were markedly downregulated in resistant cell lines and in a cohort of 226-primary neuroblastoma tumors. In Paper-II and -III, over-expression studies showed that miR-376c-3p and miR-323a-3p targets the oncogenes CCND1 and STAT3, respectively and induce G1-cell cycle arrest and apoptosis in neuroblastoma cells

Hsa-miR-323a-3p functions as a tumor suppressor and targets STAT3 in neuroblastoma cells

Background: Studies conducted in the last decades have revealed a role for the non-coding microRNAs (miRNAs) in cancer development and progression. Several miRNAs within the chromosome region 14q32, a region commonly deleted in cancers, are associated with poor clinical outcome in the childhood cancer neuroblastoma. We have previously identified miR-323a-3p from this region to be downregulated in chemotherapy treated neuroblastoma cells compared to pre-treatment cells from the same patients. Furthermore, in neuroblastoma tumors, this miRNA is downregulated in advanced stage 4 disease compared to stage 1–2. In this study, we attempt to delineate the unknown functional roles of miR-323a-3p in neuroblastoma. Methods: Synthetic miRNA mimics were used to overexpress miR-323a-3p in neuroblastoma cell lines. To investigate the functional roles of miR-323a-3p, cell viability assay, flow cytometry, reverse transcription-quantitative polymerase chain reaction, luciferase reporter assay and western blot were conducted on the neuroblastoma cell lines Kelly, SH-SY5Y and SK-N-BE(2)-C. Results: Ectopic expression of miR-323a-3p resulted in marked reduction of cell viability in Kelly, SH-SY5Y and SK-N-BE(2)-C by causing G1-cell cycle arrest in Kelly and SH-SY5Y and apoptosis in all the cell lines tested. Furthermore, mRNA and protein levels of signal transducer and activator of transcription 3 (STAT3) were reduced upon miR-323a-3p overexpression. A direct binding of the miR-323a-3p to the 3′UTR of STAT3 was experimentally validated by luciferase reporter assay, where miR-323a-3p reduced luminescent signal from full length STAT3 3′UTR luciferase reporter, but not from a reporter with mutation in the predicted seed sequence. Conclusions: miR-323a-3p inhibits growth of neuroblastoma cell lines through G1-cell cycle arrest and apoptosis, and the well-known oncogene STAT3 is a direct target of this miRNA

Hsa-miR-376c-3p targets cyclin D1 and induces G1-cell cycle arrest in neuroblastoma cells

High‑risk neuroblastoma is the most aggressive form of cancer in children. The estimated survival of children with high‑risk neuroblastoma is 40‑50% compared with low and intermediate risk neuroblastoma, which is >98 and 90‑95%, respectively. In addition, patients with high‑risk neuroblastoma often experience relapse following intensive treatments with standard chemotherapeutic drugs. Therefore alternative strategies are required to address this problem. MicroRNAs (miRNAs/miRs) are small, endogenously expressed non‑coding RNAs, which when deregulated have been demonstrated to serve significant roles in the tumorigenesis of a number of different types of cancer. Results from a previous deep sequencing study identified 22 downregulated miRNAs from the 14q32 miRNA cluster differentially expressed in neuroblastoma cell lines isolated from 6 patients at diagnosis and at relapse following intensive treatments. miR‑376c‑3p is one of the 22 miRNAs that was downregulated in the majority of the cell lines isolated from patients post treatment. The present study employed reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) to quantify the basic expression of miR‑376c‑3p in 6 neuroblastoma cell lines. The functional role of miR‑376c‑3p in the neuroblastoma cell lines was evaluated by alamar blue‑cell viability and propidium iodide‑flow cytometric assays. In addition, luciferase reporter assays, RT‑qPCR and western blotting were performed to identify and quantify the targets of miR‑376c‑3p in neuroblastoma cell lines. Ectopic expression of miR‑376c‑3p led to significant inhibition of cell viability and G1‑cell cycle arrest in multiple neuroblastoma cell lines by reducing the expression of cyclin D1, an oncogene critical for neuroblastoma pathogenesis. The results of the present study provide novel insights into the functional role of miR‑376c‑3p and suggest new approaches for the treatment of neuroblastoma