Junrong Ma

MicroRNA-10b regulates tumorigenesis in neurofibromatosis type 1

MicroRNAs (miRNAs) are frequently deregulated in human tumors, and play important roles in tumor development and progression. The pathological roles of miRNAs in neurofibromatosis type 1 (NF1) tumorigenesis are largely unknown. We demonstrated that miR‐10b was up‐regulated in primary Schwann cells isolated from NF1 neurofibromas and in cell lines and tumor tissues from malignant peripheral nerve sheath tumors (MPNSTs). Intriguingly, a significantly high level of miR‐10b correlated with low neurofibromin expression was found in a neuroectodermal cell line: Ewing’s sarcoma SK‐ES‐1 cells. Antisense inhibiting miR‐10b in NF1 MPNST cells reduced cell proliferation, migration and invasion. Furthermore, we showed that NF1 mRNA was the target for miR‐10b. Overexpression of miR‐10b in 293T cells suppressed neurofibromin expression and activated RAS signaling. Antisense inhibition of miR‐10b restored neurofibromin expression in SK‐ES‐1 cells, and decreased RAS signaling independent of neurofibromin in NF1 MPNST cells. These results suggest that miR‐10b may play an important role in NF1 tumorigenesis through targeting neurofibromin and RAS signaling. (Cancer Sci 2010)

MiR-335 Inhibits Small Cell Lung Cancer Bone Metastases via IGF-IR and RANKL Pathways

Small cell lung cancer (SCLC) is a rapidly progressing, incurable cancer that frequently spreads to bone. New insights are needed to identify therapeutic targets to prevent or retard SCLC metastatic progression. Human SCLC SBC-5 cells in mouse xenograft models home to skeletal and nonskeletal sites, whereas human SCLC SBC-3 cells only pervade nonskeletal sites. Because microRNAs (miRNA) often act as tumor regulators, we investigated their role in preclinical models of SCLC. miRNA expression profiling revealed selective and reduced expression of miRNA (miR)-335 and miR-29a in SBC-5 cells, compared with SBC-3 cells. In SBC-5 cells, miR-335 expression correlated with bone osteolytic lesions, whereas miR-29a expression did not. Overexpression of miR-335 in SBC-5 cells significantly reduced cell migration, invasion, proliferation, colony formation, and osteoclast induction in vitro. Importantly, in miR-335 overexpressing SBC-5 cell xenografts (n = 10), there were minimal osteolytic lesions in the majority of mice and none in three mice. Expression of RANK ligand (RANKL) and insulin-like growth factor-I receptor (IGF-IR), key mediators of bone metastases, were elevated in SBC-5 as compared with SBC-3 cells. Mechanistically, overexpression of miR-335 in SBC-5 cells reduced RANKL and IGF-IR expression. In conclusion, loss of miR-335 promoted SCLC metastatic skeletal lesions via deregulation of IGF-IR and RANKL pathways and was associated with metastatic osteolytic skeletal lesions. Implications: These preclinical findings establish a need to pursue the role of miR-335 in human SCLC with metastatic skeletal disease. Mol Cancer Res; 12(1); 101–10. ©2013 AACR.

MicroRNA-204 critically regulates carcinogenesis in malignant peripheral nerve sheath tumors

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive soft tissue sarcomas accounting for 3%-10% of all soft tissue sarcomas. Neurofibromatosis type 1 (NF1) is the most important known risk factor. MPNSTs are often diagnosed at an advanced stage when distant metastases have developed. Although surgical resection remains the main treatment for MPNSTs, complete surgical resection is rarely possible. The prognosis for patients with MPNSTs is poor. There is an urgent need for improved therapies. To this end, we investigated whether microRNA (miR), specifically miR-204, might be implicated in MPNSTs because it is located at a cancer-associated genomic region exhibiting high frequency of loss of heterozygosity in tumors. We show that miR-204 expression is downregulated in NF1 and non-NF1 MPNST tumor tissues and in tumor cell lines. Restoring miR-204 expression in MPNST cell lines STS26T (non-NF1), ST88-14 (NF1), and T265p21 (NF1) significantly reduces cellular proliferation, migration, and invasion in vitro. Restoring miR-204 expression in STS26T decreases tumor growth and malignant progression in vivo. We also report that miR-204 inhibits Ras signaling and expression of high mobility group gene A2. These findings support the hypothesis that miR-204 plays critical roles in MPNST development and tumor progression. miR-204 may represent a novel biomarker for diagnosis and a candidate target with which to develop effective therapies for MPNSTs.

Hyperactivation of mTOR critically regulates abnormal osteoclastogenesis in neurofibromatosis type 1

Individuals with nerofibromatosis Type 1 (NF1) frequently suffer a spectrum of bone pathologies, such as abnormal skeletal development (scoliosis, congenital bowing, and congenital pseudoarthroses, etc), lower bone mineral density with increased fracture risk. These skeletal problems may result, in part, from abnormal osteoclastogenesis. Enhanced RAS/PI3K activity has been reported to contribute to abnormal osteoclastogenesis in Nf1 heterozygous (Nf1+/−) mice. However, the specific downstream pathways linked to NF1 abnormal osteoclastogenesis have not been defined. Our aim was to determine whether mammalian target of rapamycin (mTOR) was a key effector responsible for abnormal osteoclastogenesis in NF1. Primary osteoclast‐like cells (OCLs) were cultured from Nf1 wild‐type (Nf1+/+) and Nf1+/− mice. Compared to Nf1+/+ controls, there were 20% more OCLs induced from Nf1+/− mice. Nf1+/− OCLs were larger and contained more nuclei. Hyperactive mTOR signaling was detected in Nf1+/− OCLs. Inhibition of mTOR signaling by rapamycin in Nf1+/− OCLs abrogated abnormalities in cellular size and number. Moreover, we found that hyperactive mTOR signaling induced abnormal osteoclastogenesis major through hyper‐proliferation. Our research suggests that neurofibromin directly regulates osteoclastogenesis through mTOR signaling pathway. Inhibiting mTOR may represent a viable strategy to treat NF1 bone diseases.

Double suicide genes driven by kinase domain insert containing receptor promoter selectively kill human lung cancer cells

BackgroundTo investigate the selective killing efficacy of the double suicide genes driven by KDR promoter.Materials and methodsA double suicide gene system with the KDR promoter, pcDNA3-KDRp-CDglyTK, was constructed and transfected into lung cancer cell lines L9981 and NL9980, and human hepatocellular carcinoma cell line HepG2. The efficiency and specificity of the double suicide gene system were assayed by in vitro cellular proliferation and apoptosis, as well as in vivo xenograft studies.ResultsThe transgenic CD and TK genes were only expressed in L9981 and NL9980 but not in HepG2 cells. Pre-treating transfected cells with 5-Fc and GCV significantly reduced proliferation, enhanced apoptosis in L9981 and NL9980 but not in HepG2 cells. The tumor formed by L9981 and NL9980 cells with the double suicide gene system was much smaller in vivo.ConclusionTumor targeted expression of CDglyTK gene driven by KDR promotor represents a novel strategy for effective gene therapy of tumor with intrinsic KDR.