Methods and matrices: approaches to identifying miRNAs for Nasopharyngeal carcinoma

Plieskatt, Jordan; Rinaldi, Gabriel; Feng, Yanjung; Levine, Paul H.; Easley, Samantha; Martinez, Elizabeth Ferreira; Hashmi, Salman; Sadeghi, N.; Brindley, Paul J.; Bethony, Jeffrey M.; Mulvenna, Jason

doi:10.1186/1479-5876-12-3

Cited by 33 publications

(56 citation statements)

References 58 publications

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“…To address this problem, we extracted RNA using the miRNeasy serum/blood plasma kit (Qiagen), which uses phenol and chloroform, and as a positive control, spiked plasma with C. elegans miR-39 mimic (Qiagen) [16]. The yield of RNA derived from plasma in both the QIAamp and miRNAeasy methods was 8-12 ng/μl, and was of similar quality to that we have recovered from sera [17], although RNA from plasma exhibited lower 260/230 ratios, (<0.7) with the miRNeasy kit. However, when RNA isolated using miRNeasy was subjected to qPCR, the C t values did not improve, including for control C. elegans miR-39 which was employed to spike plasma (Figure 1A and B).…”

Section: Resultsmentioning

confidence: 99%

Circumventing qPCR inhibition to amplify miRNAs in plasma

et al. 2014

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BackgroundCirculating microRNAs (c-miRNAs) have be identified in saliva, urine and blood, which has led to increasing interest in their development as biomarkers for diverse diseases including cancers. One of the key advantages of c-miRNAs over other biomarkers is the ability to be amplified and quantified by quantitative PCR (qPCR). However, at phlebotomy when whole blood is dispensed into heparinized tubes, residual levels of the anti-coagulant lithium heparin may remain in the plasma and hence with RNA isolated from the plasma. This can confound the detection of c-miRNAs by qPCR because it inhibits reverse transcriptase (RT). Here we present a procedure, modified from earlier techniques, to detect c-miRNAs in plasma that improves sensitivity and streamlines performance.FindingsTreatment of total RNA isolated from human blood plasma with Bacteroides heparinase I during reverse transcription at 37°C for one hour improved sensitivity and performance of the qPCR. This is in comparison to no treatment or treatment of the RNA prior to RT, which is the current suggested method and exposes plasma to Flavobacterium heparinum heparinase I for up to 2 hours before RT. This modest alteration improved qPCR performance and resulted in lowered threshold cycles (Ct) for detection of the target sequence, candidate c-miRNA biomarkers, and controls. It also reduced the expense and number of processing steps, shortening the duration of the assay and minimizing exposure of RNA to elevated temperatures.ConclusionIncorporating Bacteroides heparinase I treatment into conventional RT protocols targeting c-miRNA in plasma can be expected to expedite the discovery of biomarkers.

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Section: Resultsmentioning

confidence: 99%

Circumventing qPCR inhibition to amplify miRNAs in plasma

et al. 2014

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“…Multiple reports confirmed the abundant expression of the BART miRNAs in NPC tissues in the absence of BHRF1 miRNAs. A recent study revealed that all 44 BART miRNAs were expressed in NPC tumours, at broadly similar levels (Plieskatt et al, 2014). Interestingly, approximately one-sixth of the total miRNA content in NPC cells attributed to BART miRNAs, suggesting that BART miRNAs provided a considerable portion of the total regulatory influence of both viral and cellular genes in NPC tumour cells (Lo et al, 2012).…”

Section: Bart Mirnamentioning

confidence: 98%

“…Number of Studies miRNAs Up 2 miR-15b (Chen et al, 2009;Plieskatt et al, 2014), miR-18a (Chen et al, 2009;Li et al, 2011), miR-205 (Chen et al, 2009Plieskatt et al, 2014), miR-25 (Chen et al, 2009;Plieskatt et al, 2014) 1 miR-106a (Chen et al, 2009), miR-1268a (Plieskatt et al, 2014), miR1268b (Plieskatt et al, 2014), miR-1303 (Plieskatt et al, 2014), miR-1304 (Plieskatt et al, 2014), miR-1305 (Plieskatt et al, 2014), miR-138 (Chen et al, 2009), miR-142-3p (Chen et al, 2009, miR-151 (Sengupta et al, 2008), miR-155 (Chen et al, 2009), miR-17 (Chen et al, 2009), miR-184 (Plieskatt et al, 2014), miR-192 (Sengupta et al, 2008), miR-196b (Chen et al, 2009, miR-21 (Plieskatt et al, 2014), miR-27a (Plieskatt et al, 2014), miR-378 , miR-4677 (Plieskatt et al, 2014), miR-4791 (Plieskatt et al, 2014), miR548n (Plieskatt et al, 2014), miR-6510 (Plieskatt et al, 2014), miR-92a (Plieskatt et al, 2014), miR-944 (Plieskatt et al,2014) Down 3 miR-34b (Chen et al, 2009;Li et al, 2011;Sengupta et al, 2008), miR-100 (Chen et al, 2009;Li et al, 2011;Plieskatt et al, 2014), miR-152 (Chen et al, 2009;Li et al, 2011;Plieskatt et al, 2014…”

Section: Regulationmentioning

confidence: 99%

“…, miR-136(Plieskatt et al, 2014), miR-139(Plieskatt et al, 2014), miR-139-5p(Chen et al, 2009), miR-150, miR-151a(Plieskatt et al, 2014), miR-155), miR-182 (Plieskatt et al, 2014, miR-187(Chen et al, 2009), miR-199a (Plieskatt et al, 2014), miR-199b-5p (Chen et al, 2009), miR-200a (Chen et al, 2009), miR-200b (Chen et al, 2009), miR-204 (Chen et al, 2009Plieskatt et al, 2014), miR-21(Plieskatt et al, 2014), miR-212(Sengupta et al, 2008), miR-214(Plieskatt et al, 2014), miR-216(Sengupta et al, 2008), miR-217(Sengupta et al, 2008), miR-221, miR-26a(Plieskatt et al, 2014), miR-26b, miR-27b(Plieskatt et al, 2014), miR-29a, miR-29b, miR-300(Plieskatt et al, 2014), miR-3065(Plieskatt et al, 2014), miR-30a(Chen et al, 2009), miR-30d, miR-31(Chen et al, 2009), miR-335(Plieskatt et al, 2014), miR-342, miR-34c-3p(Chen et al, 2009), miR-34c-5p(Chen et al, 2009), miR-375, miR-376a(Plieskatt et al, 2014), miR-376c(Plieskatt et al, 2014), miR-376b-5p(Plieskatt et al, 2014), miR-425-5p, miR-4423(Plieskatt et al, 2014), miR-449, miR-449a(Chen et al, 2009), miR-450a(Plieskatt et al, 2014), miR-4792(Plieskatt et al, 2014), miR-488(Plieskatt et al, 2014), miR-504(Plieskatt et al, 2014), miR-532-3p(Chen et al, 2009), miR-542(Plieskatt et al, 2014), miR-556(Plieskatt et al, 2014), miR-574(Plieskatt et al, 2014), miR-576), miR-584 (Plieskatt et al, 2014, miR-585(Plieskatt et al, 2014), miR-625), miR-642 (Li et al, 2011), miR-768-3p (Li et al, 2011), miR-874 (Plieskatt et...…”

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confidence: 99%

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MicroRNAs serving as potential biomarkers and therapeutic targets in nasopharyngeal carcinoma: A critical review

Lee

Tan

Lam

et al. 2016

Critical Reviews in Oncology/Hematology

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Despite significant medical advancement, nasopharyngeal carcinoma (NPC) remains one of the most difficult cancers to detect and treat where it continues to prevail especially among the Asian population. miRNAs could act as tumour suppressor genes or oncogenes in NPC. They play important roles in the pathogenesis of NPC by regulating specific target genes which are involved in various cellular processes and pathways. In particular, studies on miRNAs related to the Epstein Barr virus (EBV)-encoded latent membrane protein one (LMP1) and EBVmiRNA- BART miRNA confirmed the link between EBV and NPC. Both miRNA and its target genes could potentially be exploited for prognostic and therapeutic strategies. They are also important in predicting the sensitivity of NPC to radiotherapy and chemotherapy. The detection of stable circulating miRNAs in plasma of NPC patients has raised the potential of miRNAs as novel diagnostic markers. To conclude, understanding the roles of miRNA in NPC will identify ways to improve the management of patients with NPC.

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“…Overexpression of miR-376a has been reported to attenuate the proliferation and migration of melanoma cells by targeting IGF1R, a tyrosine kinase receptor associated with tumorigenesis and metastasis [8]. MiR-376a also regulates proliferation, apoptosis, migration and invasion in metastatic prostate cancer and hepatocellular carcinoma [9][10], and functions as a tumor suppressor in nasopharyngeal carcinoma [11]. Nevertheless, few studies have focused on the function of miR-376a in lung cancer, especially NSCLC, despite its significance to human health.…”

Section: Introductionmentioning

confidence: 99%

MiR-376a inhibits non-small cell lung cancer cell progression by regulating Rab1A

Cao

Cheng

Zheng

et al. 2019

Preprint

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Post transcriptional gene regulation of microRNA-376a (miR-376a) plays a crucial role for tumorigenesis and cancer development. However, the potential role of miR-367a in non-small cell lung cancer (NSCLC) remains unclear. In this study, we investigated the crucial role of miR-376a in NSCLC by analyzing miR-376a expression as well as its target genes. Through overexpression strategies, we uncovered the molecular mechanisms underlying miR-376a-mediated tumorigenesis. Quantitative real-time PCR analysis demonstrated miR-376a levels to be significantly decreased in NSCLC cells compared with non-tumorigenic counterparts. Interestingly, miR-376a overexpression potentially repressed NSCLC cell proliferation, migration, and invasion, but increased apoptosis in A549 cells. Using bioinformatic approaches, we predicted that miR-376a targets Rab1A, and further luciferase fusion assay demonstrated Rab1A was a direct target of miR-376a and miR-376a inhibited cell proliferation by regulating the mRNA and protein levels of Rab1A in NSCLC cells. Overall, our findings uncover the miR-376a could suppress NSCLC cells progression via directly targeting Rab1A.

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Methods and matrices: approaches to identifying miRNAs for Nasopharyngeal carcinoma

Cited by 33 publications

References 58 publications

Circumventing qPCR inhibition to amplify miRNAs in plasma

Circumventing qPCR inhibition to amplify miRNAs in plasma

MicroRNAs serving as potential biomarkers and therapeutic targets in nasopharyngeal carcinoma: A critical review

MiR-376a inhibits non-small cell lung cancer cell progression by regulating Rab1A

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