The Research Of Plasma MiRNAs Biomarker For Multiple Myeloma

Multiple myeloma (MM) is a plasma cell malignancy characterized by the clonalaccumulation of monotypic plasma cells in the bone marrow. MM patients exhibit oneor more symptoms including hypercalcemia, bone destruction, anemia and renalfailure.Although morphologically similar, several subtypes of the disease have beenidentified at the genetic and molecular level. At the top hierarchical level, myelomacan be divided into hyperdiploid and non-hyperdiploid subtypes. Nearly half ofmyeloma casesare non-hyperdiploid, and mostly have recurrent IgH translocations,whereas hyperdiploid myeloma is characterized by multiple trisomies involving oddnumber chromosomes. A number of secondary genetic lesions associated with diseaseprogression and survival have been identified including RAS mutation, MYCdysregulation, deletions of chr13and chr17, chromosome1abnormalities and nuclearfactor-kappa B (NF-κB) activation. Despite advances in molecular genetics, themechanism behind MM initiation and progression remains to be elucidated and thedisease remains incurable largely.The study found that microRNAs (miRNAs) belong to ncRNA not only involvein embryonic development and cell differentiation but also play a key role intumorigenesis. Some researchers claimed number of miRNA genes is as high as1000.Each miRNA molecules have hundreds of different conservative and nonconservative target genes, it is estimated that50%of the genes are modulated bymiRNA. Almost all discovered miRNA are highly conserved in the neighboringspecies, suggesting a conservative on its functions. miRNAs have been shown toregulate several developmental and physiological processes including embryonicdevelopment, apoptosis, cell division, differentiation, death, hematopoiesis andcancer.Many experiments confirmed that miRNA dyregulted in tumor,which indicatethat miRNAs act as tumor suppressors and oncogenes. The miRNA expressionprofiling studies had found tissue-specific expression miRNA. The expressionspectrum of miRNA in different tissues and disease showed the apparent tissuespecificity. Furthermore, miRNA profiles have been associated with different tumourtypes, such as liver cancer, lung cancer, colon cancer, ovarian cancer, leukemia andother malignancies. Studies show potential for microRNAs to be the biomarks oftumors. miRNA released from tumor cells into the body fluids can be used as diagnosticmarkers in a series of cancer. Recent studies reported that miRNA molecules can bereleased into the blood, breast milk and breast ductal fluid from the breast cancer cells,the miRNA molecules in these body fluids can be used as alternative molecules forcancer molecular markers. To confirm whether circulating miRNA as a tumor-specific,the study analyzed a variety of cancers (melanoma, breast cancer, colon cancer,prostate cancer and kidney cancer) in patients with preoperative range ofcancer-related miRNA levels. The results showed that miR-195in breast cancerpatients with increased specificity, unlike other cancer patients and healthy people. Inaddition, compared to control populations in the circulation of patients with breastcancer the level of miR-195and let-7a were significantly decreased. These findingssuggested that circulating miRNA had the potential to become a biomarker for earlydiagnosis of breast cancer. Circulating miRNA had represent potentially informativebiomarkers for disease diagnosis and prognosis, especially for early-stage cancer.In recent studies,miRNAs were involved in the pathogenesis of MM. MiRNAmicroarray detection of miRNA-29b expression levels decreased significantly inmyeloma cells of MM patients and human myeloma cell lines (HMCLs). Overexpressed miR-29b in HMCLs could induce apoptosis and increase the activity ofcaspase-3. Further studies showed miRNA-29b induced apoptosis, improve the caspaseactivity and antagonism of IL-6promote tumor effect in HMCLs by targeting Mcl-1(myeloid-cell-leukemia1). This study suggested that miR-29b played as an importanttumor suppressor in MM. the expression of MiR-106b~25cluster, miR-181a/b andmiR-19a/b elevated in the MM bone marrow plasma cells compared with the healthycontrols. Moreover miR-106b-25cluster, miR-32, miR-181a/b and miR-19a/btargeted SOCS1(IL-6inhibitor) and p300-CBP,(p53pathway components).Xenograftstudies using human MM cell linestreated with miR-19a and b, and miR-181a and bantagonists resulted in significant suppression of tumor growth in nude mice. MiR-25and miR-30d can binding with3'UTR of TP53, and reduces levels of p53protein andreducing the downstream gene expression of the p53pathway. In addition, miR-25andmiR-30d can affect apoptosis, cell cycle arrest. Inhibiting the expression of these twomolecules in the number of multiple myeloma cell lines could promote expression ofendogenous p53and induce apoptosis. This study suggests that miR-25and miR-30d inmyeloma cell lines can be negative regulation of TP53. Based on the above researches, we proposed a number of questions: wether themiRNA expression pattern can discriminate the multiple myeloma and normal controlWether microRNA expression signatures may help in the development of prognosticand diagnosis of multiple myeloma? To address these questions, we conducted thefollowing two-part study:1) determination of miRNA expression profiles in the plasmaof patients with multiple myeloma;2) to investigate association between miRNA andthe prognosis and clinical features.3)to investigate whether the miRNA associated withpathogensis of MM.Part I miRNA profile in plasma of multiple myeloma patientsmiRNAs are an important class of non-coding RNA involved in various biologicalprocesses of eukaryotic cells.The relationship between the abnormal miRNAexpression and disease caused widespread concern of the researchers.The main purpose of this part is to study plasma miRNA expression profiling inpatients with multiple myeloma. The application of high specificity of the TaqManmicroRNA microarray analysis of miRNA expression in plasma of patients withmultiple myeloma; application software analysis to analyze the differences betweenmultiple myeloma and normal controls; further detection of specific miRNA expressionusing real-time PCR analysis.Plasma miRNA expression spectrums of eight normal subjects and12patients withmultiple myeloma were analysed by SAM algorithm.there were135miRNAmolecules were detected in plasma, of which there are six miRNA (miR-148a, miR-20a,miR-99b, miR-221, of miR-181a and miR-625) were significantly upregulatedcompared with normal controls.Part II Examined the correlation between specific miRNA and karyotypes andsurvivalMM is a heterogeneous disease with some patients dying within a few weeks ofdiagnosis, whereas others live for longer than10years. It is increasingly evident thatthe genetic features of the tumor cells largely dictate the clinical heterogeneity of MM.However, it lakes t a simple and reliable indicators for prognosis of the disease.Manypreset studies found that serum miRNA is expected to become the new tumordiagnostic and prognostic molecular markers.Then, quantitative analysis of these miRNAs was performed in a large series of MMpatients to find that the levels of5miRNAs (miR-148a, miR-181a, miR-20a, miR-221 and miR-99b) were increased in plasma of patients. We correlated the expression of the5miRNAs with the patients' clinicopathologic data and found that several of them hadpotentially applications for prognosis the disease. High levels of miR-20a andmiR-148a were related to shorter relapse-free survival.This work has identified aberrantly circulating miRNA expression associated withthe genetic subtype and survival of MM. It suggests that plasma miRNAs havepotential as tumor biomarkers of clinical usage.Part III miR-148a plays the oncogene role in multiple myelomaResearched have shown that a number of miRNAs involved in the development ofMM. Moreover we found that the level of miR-148a increased in the plasma ofpatients.In addition, it's reported that the molecule also increased in tumor cells of MMpatients. However, the molecular mechanisms which involved in MM pathogenesishave needed to be studied.Downregulted the expression of miR-148a by transfected inhibitor to NCL-H929could decrease the activity of proliferation induce apoptosis.Identification of potentialmRNA targets for the differentially expressed miRNAs was performed usingTargetScan,miRANDA,and PicTar databases. Luciferase reporter gene assayconfirmed the miR-148a can be combined with the3'UTR of CDKN1B and PTEN. Theresults of western blot confirmed the miR-148a can inhibit the expression of targetgenesIn this study, we studied functions of key miRNAs in MM pathogenesis andmechanisms behind their deregulation.This research lay the foundation of moleculartargeting therapy for multiple myeloma.