Ran Sun

MicroRNA-34a suppresses cell proliferation by targeting LMTK3 in human breast cancer mcf-7 cell line.

Breast cancer remains the leading cause of cancer mortality in females, and about 70% of the primary breast cancer patients are diagnosed ERα-positive, which is the most common type of breast cancer. MicroRNA-34a (miR-34a) has been shown to be a master regulator of tumor suppression in many types of cancers including breast cancer. However, the role of miR-34a in ERα-positive breast cancer has not been elucidated. Here, we find that in MCF-7, which is an ERα-positive breast cancer cell line, miR-34a is remarkably downregulated after E2 treatment. Overexpression of miR-34a by lentivirus suppresses cell proliferation, S phase ratio, and tumor formation in an E2-dependent manner in vitro. According to the mRNA sequence, lemur tyrosine kinase 3 (LMTK3), which is an important regulator of estrogen receptor alpha (ERα), is a predicted target of miR-34a. This is confirmed by dual luciferase reporter assay and the decrease of LMTK3 mRNA and protein levels after overexpression of miR-34a. Moreover, miR-34a overexpression decreases AKT signaling pathway and increases ERα phosphorylation status. Taken together, these results suggest that miR-34a inhibits breast cancer proliferation by targeting LMTK3 and might be used as an anti-ERα agent in breast cancer therapy.

A human microsatellite DNA-mimicking oligodeoxynucleotide with CCT repeats negatively regulates TLR7/9-mediated innate immune responses via selected TLR pathways

n Abstractn n A human microsatellite DNA-mimicking ODN (MS ODN) composed of CCT repeats, designated as SAT05f, has been studied for its capacity of negatively regulating innate immunity induced by TLR7/TLR9 agonists in vitro and in mice. The result showed that SAT05f could down-regulate TLR7/9-dependent IFN-α production in cultured human PBMC stimulated by inactivated Flu virus PR8 or HSV-1 or CpG ODN or imiquimod, protect d-GalN-treated mice from lethal shock induced by TLR9 agonist, not by TLR3/4 agonist. In addition, SAT05f significantly inhibit IFN-α production from purified human plasmacytoid cells (pDCs) stimulated by CpG ODN. Interestingly, SAT05f could up-regulate CD80, CD86, and HLA-DR on the pDCs in vitro, implying that SAT05f-mediated inhibition on IFN-α production could be related to the activation of pDCs. The data suggest that SAT05f could be developed as a candidate medicament for the treatment of TLR7/9 activation-associated diseases by inhibiting TLR7/9 signaling pathways.n n

High expression of miR-105-1 positively correlates with clinical prognosis of hepatocellular carcinoma by targeting oncogene NCOA1

Increasing evidence supports that microRNA (miRNA) plays a significant functional role in cancer progression by directly regulating respective targets. In this study, the expression levels of miR-105-1 and its target gene were analyzed using genes microarray and hierarchical clustering analysis followed by validation with quantitative RT-PCR in hepatocellular carcinoma (HCC) and normal liver tissues. We examined the expression of nuclear receptor coactivator 1 (NCOA1), the potential target gene of miR-105-1, following the transfection of miR-105-1 mimics or inhibitors. Our results showed that miR-105-1 was downregulated in HCC tissues when compared with normal liver tissues and patients with lower miR-105-1 expression had shorter overall survival (OS) and progression free survival (PFS). Moreover, NCOA1 was confirmed to be a direct target of miR-105-1. Furthermore, concomitant high expression of NCOA1 and low expression of miR-105-1 correlated with a shorter median OS and PFS in HCC patients. In conclusion, our results provide the first evidence that NCOA1 is a direct target of miR-105-1 suggesting that NCOA1 and miR-105-1 may have potential prognostic value and may be useful as tumor biomarkers for the diagnosis of HCC patients.

CYP24A1 is a potential biomarker for the progression and prognosis of human colorectal cancer

Our study aims to fully evaluate clinicopathological and prognostic values of CYP24A1 in colorectal cancer (CRC) patients. Tissue microarrays of formalin-fixed and paraffin-embedded tumor samples and matched adjacent nontumor colorectal tissues from 99 CRC patients were studied for CYP24A1 protein expression by immunohistochemistry. Messenger RNA expression of CYP24A1 was further evaluated by quantitative real-time polymerase chain reaction in 12 pairs of fresh frozen CRC samples. CYP24A1 expression was significantly higher in CRC tissues compared to corresponding noncancerous tissues. The expression of CYP24A1 protein in CRC was correlated with the depth of tumor invasion (P = .000), lymph node metastasis (P = .030), venous permeation (P = .016), and overall survival (P = .008). A Kaplan-Meier analysis of the CRC patients with high CYP24A1 expression showed significantly reduced overall survival and disease-free survival compared to the patients with low expression (P = 0.026 and .009). A prognostic significance of CYP24A1 was also found in the subgroup of venous permeation condition classification. A multivariate Cox regression analysis showed that CYP24A1 expression was an independent prognostic factor for CRC recurrence (P = .032). In conclusion, CYP24A1 expression is closely associated with CRC progression, and it might be a novel prognostic biomarker for CRC.

Effect of Duration of Ex Vivo Ischemia Time and Storage Period on RNA Quality in Biobanked Human Renal Cell Carcinoma Tissue

AbstractBackgroundnRNA degradation is a major problem in tissue banking, and the effects of the ex vivo ischemia time, storage time, and transport conditions on RNA integrity and gene expression have not been well understood.MethodsnA total of 100 fresh-frozen clear cell carcinoma tissues and matched normal tissues during five storage periods (≤6, 7–12, 13–18, 19–24, and 25–30xa0months) were chosen to detect RNA quality. At surgery, fresh kidney cancer tissues from five patients were cut into pieces and snap frozen. Additional fresh tissue pieces were (1) left at room temperature, (2) kept on ice, or (3) placed in normal saline before being snap frozen after 0.5, 1, 2, or 4xa0h. RNA integrity was determined by microchip electrophoresis, and gene expression was analyzed by real-time polymerase chain reaction.Results Altogether, 82xa0% of kidney cancer specimens banked using a standardized protocol yielded RNA with an RNA integrity number of ≥7 (7.77xa0±xa00.95, 7.87xa0±xa00.37, 7.15xa0±xa01.46, 8.10xa0±xa00.64, and 7.11xa0±xa01.08 during the five storage periods, respectively). RNA remained intact after 4xa0h on ice, whereas degradation was found in tissues left at room temperature or in saline. Expression of genes in certain functional pathways changed during storage under three conditions.ConclusionsMore than 80xa0% of the banked kidney cancer biospecimens collected following a standardized protocol yielded high-quality RNA. Fresh human cancer tissue samples should be transported on ice before biobanking to avoid a major reduction in RNA quality. The presented data should be considered in attempts to further standardize tissue biospecimen collection and banking.

Inhibition of a C‐rich oligodeoxynucleotide on activation of immune cells in vitro and enhancement of antibody response in mice

To explore the possibility that human mitochondrial genomic DNA‐mimicking oligodeoxynucleotides could regulate the immune response, a series of mitochondrial DNA‐based oligodeoxynucleotides (MTODNs) were designed and studied to determine their immunoregulatory effects on immune cells activated by toll‐like receptor (TLR) stimulation. The results showed that a C‐rich MTODN, designated MT01, was able to inhibit the proliferation of human peripheral blood mononuclear cells (PBMCs) induced by cytosine–phosphate–guanosine (CpG) oligodeoxynucleotides (ODNs) and the production of type I interferon (IFN) from human PBMCs stimulated by TLR agonists, including inactivated influenza virus, imiquimod, inactivated herpes simplex virus‐1 (HSV‐1) and CpG ODNs. In addition, MT01 inhibited the CpG ODN‐enhanced antibody response and this inhibition could be related to the antagonism of TLR9‐activation pathways in B cells. Notably, unlike the G‐rich suppressive ODNs reported, MT01 is composed of ACCCCCTCT repeats. These data imply that MT01 represents a novel class of immunosuppressive ODNs that could be candidate biologicals with therapeutic use in TLR activation‐associated diseases.

Human microsatellite DNA mimicking oligodeoxynucleotides down-regulate TLR9-dependent and -independent activation of human immune cells.

To develop novel immunoregulatory oligodeoxynucleotides (ODNs), we have designed a series of ODNs based on the sequences in human microsatellite (MS) DNA. The ODNs, designated as human MS DNA mimicking ODNs (MS ODNs), have been studied for their inhibitory effects on human immune cells activated by TLR9-dependent and -independent stimulations. We find for the first time that MS08, a MS ODN composed entirely of TC dinucleotide (TC) repeats, inhibits CpG ODN (TLR9 ligand)-induced human PBMCs proliferation, CD80 and CD86 expression and production of interferon. In addition, MS08 also inhibits the proliferation of human PBMCs stimulated by PHA, PMA and alloantigens in a TLR9-independent manner. The inhibition correlates with competition of binding and uptake between MS08 and CpG ODN in human PBMCs. Structurally, TC, CT or CCT are revealed as essential suppressive motifs required for the inhibition. These findings suggest that TC repeat containing MS ODN could be of therapeutic use in pathologic situations due to excessive activation of immune cells.

Effects of oligodeoxynucleotide with CCT repeats on chronic graft versus host disease induced experimental lupus nephritis in mice

A synthesized single-stranded oligodeoxynucleotide (ODN), designed as SAT05f with the sequence of human microsatellite DNA, has been studied for its capacity of alleviating the lupus nephritis in the chronic graft versus host disease (cGVHD) induced lupus-prone mice. In cGVHD model mice, both of continuous and discontinuous treatment with SAT05f was effective on reducing anti-ssDNA antibody production, decreasing renal IgG deposition and delaying the onset of lupus nephritis. In addition, SAT05f could down-regulate TLR9 mRNA expression in splenocytes of cGVHD model mice. These results indicated that SAT05f could be developed as a new therapeutic agent for the treatment of lupus nephritis by inhibiting TLR9 signaling pathways.

An oligodeoxynucleotide capable of lessening acute lung inflammatory injury in mice infected by influenza virus.

Infection of influenza virus could induce acute lung inflammatory injury (ALII) that was at least partially caused by excessive innate immune responses. To study whether down-regulating Toll-like receptor (TLR)-mediated innate immune response could lessen influenza virus-induced ALII, a microsatellite DNA mimicking oligodeoxynucleotide (MS ODN), named as SAT05f capable of inhibiting TLR7/9-activation in vitro, was used to treat mice infected with FM1 virus. In parallel, two MS ODNs confirmed with less or no in vitro activities, named as MS19 and MS33, were used as controls. Unexpectedly, SAT05f failed to lessen ALII in the mice, whereas MS19 significantly inhibited the weight loss and displayed dramatic effect on lessening the ALII by reducing consolidation, hemorrhage, intra-alveolar edema and neutrophils infiltration in lungs of the mice. Meanwhile, MS19 could decrease the mortality of influenza virus infected mice and down-regulate TNF-α production in their lungs. The data suggest that MS19 might display its therapeutic role on ALII induced by influenza virus by reducing over-production of TNF-α.

Association of glutathione S-transferase M1, T1, and P1 polymorphisms with renal cell carcinoma: evidence from 11 studies.

The glutathione S-transferases (GSTs) are a gene superfamily of phase II metabolic enzymes that has attracted a considerable attention as a candidate gene for renal cell carcinoma (RCC) based on its enzyme function as a key factor in biotransformation pathways. In the past decade, a number of case–control studies were conducted to investigate the association of GST genetic polymorphisms and RCC risk. However, studies on the association between GST (GSTM1, GSTT1, and GSTP1) polymorphisms and RCC remain to be conflicting. To derive a more precise estimation of the relationship, a meta-analysis of 2,189 cases and 3,817 controls from 11 case–control studies was performed. Overall, the summarized odds ratio for RCC of the GSTM1 null and GSTT1 null polymorphisms was 1.02 (95xa0% confidence interval (CI) 0.91–1.15, Pu2009=u20090.70) and 1.28 (95xa0% CI 0.96–1.72, Pu2009=u20090.09), respectively. No significant results were observed in heterozygous and homozygous genotypes when compared with wild-type genotype for GSTP1 I105V polymorphism. However, the GSTM1–GSTT1 interaction analysis showed that the dual null genotype of GSTM1/GSTT1 was significantly associated with an increased RCC risk (odds ratio (OR)u2009=u20091.42, 95xa0% CI 1.14–1.76, Pu2009=u20090.001). In the stratified analyses by ethnicity, significant gene–disease association was obtained among Asians for GSTT1 and GSTP1 polymorphisms. In our meta-analysis, the associations between variations of GSTs and RCC may vary in different ethnic populations, and the interaction between unfavorable GST genotypes may exist.