WO2005097205A2 - Dna virus microrna and methods for inhibiting same - Google Patents
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- WO2005097205A2 WO2005097205A2 PCT/US2005/011232 US2005011232W WO2005097205A2 WO 2005097205 A2 WO2005097205 A2 WO 2005097205A2 US 2005011232 W US2005011232 W US 2005011232W WO 2005097205 A2 WO2005097205 A2 WO 2005097205A2
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Definitions
- the invention relates to an isolated nucleic acid molecule comprising the sequence of a DNA virus microRNA.
- FIG. 3 Schematic representation of miR-BART2-guided cleavage of BALF5 mRNA. Lytic genes are shown as black boxes and genes for which the expression has not been characterized are indicated in gray (GenBank entry V01555). The miR-B ART2 sequence is aligned relative to the nucleotide sequence and the processing site of the BALF5 mRNA. The prediction position of BALF5 mRNA cleavage coincides with the mapped terminus of the 3.7 kb processed form.
- KSHV miRNAs are differentially regulated upon induction of the lytic cycle.
- Northern blots for KSHV miR-Kla, miR-K6 and miR-K7 made from total RNA isolated from a KSHV negative (BJAB) cell line and from BCBLl cells at 24h, 48h and 72h after TPA treatment.
- the DNA virus infects mammalian cells.
- mammals include laboratory animals, such as dogs and cats, farm animals, such as cows, horses and sheeps, laboratory animals, such as rats, mice and rabbits, and primates, such as monkeys and humans.
- the DNA virus can be a single stranded or double stranded DNA virus.
- the DNA virus microRNA molecule comprises a maximum number of fifty moieties, preferably a maximum of forty, more preferably a maximum of thirty, even more preferably a maximum of twenty-five, and most preferably a maximum of twenty-three moieties.
- a suitable range of minimum and maximum numbers of moieties may be obtained by combining any of the above minima with any of the above maxima.
- Ci to C 4 alkoxy - Ci to C 4 alkyl group Another suitable example of a substituent at the 2' position of a modified ribonucleotide moiety is a Ci to C 4 alkoxy - Ci to C 4 alkyl group.
- the to C 4 alkoxy (alkyloxy) and Ci to C alkyl group may comprise any of the alkyl groups described above.
- the preferred Ci to C 4 alkoxy - to C alkyl group is methoxyethyl. See structure 4 in figure 1.
- Non-naturally occurring amino acids also include derivatives of naturally occurring amino acids.
- the derivative of a naturally occurring amino acid may, for example, include the addition or one or more chemical groups to the naturally occurring amino acid.
- the amino acids can be identical or different from one another.
- Bases are attached to the amino acid unit by molecular linkages. Examples of linkages are methylene carbonyl, ethylene carbonyl and ethyl linkages. (Nielsen et al, Peptide Nucleic Acids-Protocols and Applications, Horizon Scientific Press, pages 1-19; Nielsen et al., Science 254: 1497-1500.)
- linkages are methylene carbonyl, ethylene carbonyl and ethyl linkages.
- the PNA has many desirable properties, including high melting temperatures (Tm), high base-pairing specificity with nucleic acid and an uncharged molecular backbone. Additionally, the PNA does not confer RNase H sensitivity on the target RNA, and generally has good metabolic stability.
- up to ten percent, and preferably up to five percent of the contiguous bases can be additions, deletions, mismatches or combinations thereof.
- Additions refer to the insertion in the contiguous sequence of any moiety described above comprising any one of the bases described above.
- Deletions refer to the removal of any moiety present in the contiguous sequence.
- Mismatches refer to the substitution of one of the moieties comprising a base in the contiguous sequence with any of the above described moieties comprising a different base.
- no more than fifty percent, and preferably no more than thirty percent, of the contiguous moieties contain deoxyribonucleotide backbone units.
- Table E and F show the number of contiguous bases and the maximum number of deoxyribonucleotide backbone units.
- the moiety corresponding to position 11 in a naturally occurring DNA virus microR A sequence can be an addition, deletion or mismatch.
- the invention provides an isolated microRNP comprising any of the isolated nucleic acid sequences described above or analogs of the DNA virus microRNAs described above.
- HCMV ACUCUCACGGGAAGGCUAQUUA CUACAAACUAGCAUUCUGGUGA UCUUUCCAGGUGUCUUCAACGA AGCCUGGAUCUCACCGUCACUU GCGGUGAAGAAGGGGAGGACGA AACGCUCUCGUCAGGCUUGUCA GACAUCGUCACACCUAUCAUAA CGGUCCGAGCCACUGAGCGGUU UCAUCCACCUGAACAGACCGCU CGCGGGAGCUCUCCAAGUGGAU CGCCCACGGUCCGGGCACAAUC
- the anti-DNA virus microRNA molecule can be modified as described above for DNA virus microRNA molecules.
- the contiguous moieties in the anti-DNA virus microRNA molecule are complementary to the corresponding DNA virus microRNA molecule.
- the degree of complementarity of the anti-DNA virus microRNA molecules are subject to the restrictions described above for analogs of DNA virus microRNA molecules, including the restriction relating to wobble base pairs, as well as those relating to additions, deletions and mismatches.
- the anti-DNA virus microRNA molecule contains at least one modified moiety in the at least ten contiguous bases and/or comprises a chemical cap.
- the moiety in the anti-DNA virus microRNA molecule at the position corresponding to position 11 of a naturally occurring DNA virus microRNA is non- complementary.
- the moiety in the anti-DNA virus microRNA molecule corresponding to position 11 of a naturally occurring DNA virus microRNA can be rendered non-complementary by any means described above, including by the introduction of an addition, deletion or mismatch, as described above.
- the nucleic acid molecule, DNA virus microRNA molecule or anti-DNA virus microRNA molecule is preferably isolated, which means that it is essentially free of other nucleic acids. Essentially free from other nucleic acids means that the nucleic acid molecule, DNA virus microRNA molecule, anti-DNA virus microRNA molecule, or microRNP molecule, such as those described above, is at least about 90%, preferably at least about 95% and, more preferably at least about 98% free of other nucleic acids.
- the molecule is essentially pure, which means that the molecule is free not only of other nucleic acids, but also of other materials used in the synthesis and isolation of the molecule. Materials used in synthesis include, for example, enzymes. Materials used in isolation include, for example, gels, such as SDS-PAGE.
- the molecule is at least about 90% free, preferably at least about 95% free and, more preferably at least about 98% free of other nucleic acids and such other materials.
- DNA virus microRNA molecules and anti-DNA virus microRNA molecules of the present invention have numerous in vitro, ex vivo, and in vivo applications.
- the microRNA molecules and/or anti-microRNA molecules of the present invention can be introduced into a cell to study the function of the microRNA.
- Any DNA viral microRNA molecule and/or anti-DNA viral microRNA molecule mentioned above can be introduced into a cell for studying their function.
- a microRNA in a cell is inhibited with a suitable anti-microRNA molecule.
- the activity of a microRNA molecule in a cell can be enhanced by introducing into the cell one or more additional microRNA molecules.
- the function of the microRNA can be inferred by observing changes associated with inhibition and/or enhanced activity of the microRNA in the cell.
- the invention relates to a method for inhibiting microRNP activity in a cell.
- the method for inhibiting microRNP activity in a cell comprises introducing into the cell a single-stranded anti-DNA virus microRNA molecule.
- the microRNP comprises a DNA virus microRNA molecule.
- the microRNA molecule comprises a sequence of bases complementary to the sequence of bases in a single stranded anti-DNA virus microRNA molecule.
- any anti-DNA virus microRNA molecule can be used in the method for inhibiting microRNP activity in a cell, as long as the anti-DNA virus microRNA is complementary, subject to the restrictions described above, to the DNA virus microRNA present in the microRNP.
- the anti-DNA virus microRNA molecules of the present invention are capable of inhibiting microRNP activity by binding to the DNA virus microRNA in the microRNP in a host cell.
- MicroRNP activity refers to the cleavage or the repression of translation of the target sequence.
- the target sequence may be any sequence which is partially or perfectly complementary to the sequence of bases in a DNA virus microRNA.
- the target sequence can be, for example, a viral or host messenger RNA.
- a DNA virus can produce a microRNA which is complementary to a host derived target sequence that is beneficial to the host cell for defending against the viral infection.
- the DNA virus microRNA which is packaged in a microRNP, will inhibit the beneficial effect of the target sequence. Accordingly, the introduction of the anti-DNA virus microRNA molecule inhibits the RNP activity, and thereby reduces harm from the virus.
- a host cell can defend against a viral infection by transcribing a gene which is harmful to the virus.
- the gene may induce the cell to undergo apoptosis, and therefore the gene is harmful to the virus.
- a DNA virus microRNA complementary to the target sequence transcribed by the host cell is beneficial to the virus, because the DNA virus micro RNA (in a microRNP) will inhibit the ability of the host cell to undergo apoptosis. Accordingly, the introduction of DNA virus microRNA molecules promotes survival of the cell, thereby enhancing the infection.
- the cell can be any cell capable of being infected with a particular DNA virus.
- Particular cells infected by a particular DNA virus are well known to those skilled in the art. For example, it is well known to those in the art that EBV preferentially infects B lymphocytes.
- microRNA molecules or anti-microRNA molecules can be introduced into a cell by any method known to those skilled in the art.
- the molecules can be injected directly into a cell, such as by microinjection.
- the molecules can be contacted with a cell, preferably aided by a delivery system.
- Useful delivery systems include, for example, liposomes and charged lipids.
- Liposomes typically encapsulate oligonucleotide molecules within their aqueous center. Charged lipids generally form lipid-oligonucleotide molecule complexes as a result of opposing charges. These liposomes-oligonucleotide molecule complexes or lipid-oligonucleotide molecule complexes are usually internalized in cells by endocytosis.
- the liposomes or charged lipids generally comprise helper lipids which disrupt the endosomal membrane and release the oligonucleotide molecules.
- RNA molecules or an anti-microRNA into a cell include use of delivery vehicles, such as dendrimers, biodegradable polymers, polymers of amino acids, polymers of sugars, and oligonucleotide-binding nanoparticles.
- delivery vehicles such as dendrimers, biodegradable polymers, polymers of amino acids, polymers of sugars, and oligonucleotide-binding nanoparticles.
- pluoronic gel as a depot reservoir can be used to deliver the anti-microRNA oligonucleotide molecules over a prolonged period. The above methods are described in, for example, Hughes et al., Drug Discovery Today 6, 303-315 (2001); Liang et al. Eur. J. Biochem.
- Targeting of a microRNA molecule or an anti-microRNA molecule to a particular cell can be performed by any method known to those skilled in the art.
- the microRNA molecule or anti-microRNA molecule can be conjugated to an antibody or ligand specifically recognized by receptors on the cell.
- the antibody can be against the cell receptor CD 19, CD20, CD21, CD23 or a ligand to these receptors.
- the invention provides a method for treating a DNA virus infection is a mammal in need thereof.
- the method comprises introducing into the mammal an anti-DNA virus microRNA molecule.
- the anti-DNA virus microRNA molecules can be introduced into the mammal by any method known to those in the art.
- the above described methods for introducing the anti-DNA molecules into a cell can also be used for introducing the molecules into a mammal.
- EBV negative BL-41 and EBV positive BL41/95 cells were described previously (Torsteinsdottir et al., Int. J. Cancer 1989, 43:273) and were maintained in RPMI 1640 (Gibco) supplemented with 10% FBS.
- BL41/95 but not BL-41 contained EBV, as confirmed by Western blot analysis using antibodies against EBNA-1.
- EBV miRNA expression For analysis of EBV miRNA expression, we also cultured Hodgkin's lymphoma (HD) cells L540 and HD-MY-Z (EBV negative) and RPMI 6666 (EBV positive) and the Burkitt' s lymphoma (BL) cells Ramos (EBV negative), Ous and Mutu (EBV positive), and EBV positive Marmoset B95-8 cells that produce infectious B95-8 viral particles. These cell lines were also maintained in RPMI 1640 (Gibco) supplemented with 10% FBS. The KSHV positive BCBLl cell line was described previously (Renne et al. Nat. Med. 1996, 2:342-346) and was maintained in RPMI 1640 (Gibco) supplemented with 10% FBS.
- Primary human foreskin fibroblasts were cultured in MEM (GIBCO) supplemented with 10% FCS, 10 U/ml moronal, and 10 ⁇ g/ml neomycin sulphate. Cells at 90% confluency were infected with HCMV strain VR1814 at 5 PFU/cell and harvested when a strong cytopathic effect was visible, usually at about 4-5 days post-infection.
- RNA preparation, cloning procedure and Northern blot analysis were performed as described previously (Lagos-Quintana et al., Curr. Biol. 2002, 12:735). RNA size fractionation and cloning procedure have also been described. Northern blot analysis was performed as described (Lagos-Quintana et al, Curr. Biol. 2002, 12:735) loading 30 ⁇ g or 15 ⁇ g of total RNA per lane and using 5' P-radiolableled oligodeoxynucleotides complementary to the miRNA sequence.
- Bacterial colonies were picked into 96 well plates filled with 20 ⁇ l sterile water per well, then diluted 1 : 1 into a second 96 well plate containing 10 ⁇ l PCR cocktail (2 ⁇ l lOx Sigma JumpStart PCR buffer, 2 ⁇ l 2 mM deoxynucleoside triphosphate mixture, 0.4 ⁇ l each 10 ⁇ M M13 universal and reverse primers, 0.35 ⁇ l 1 U/ ⁇ l JumpStart REDAccuTaq DNA polymerase (Sigma), and 4.85 ⁇ l water.
- the PCR cycling program consisted of 1'30" at 94 °C.
- miRanda (Enright et al., Genome Biol, 2003, 5:RI, 1) was used to identify miRNA binding sequences in the 3' UTR sequences.
- the thresholds used for this scan were S:90 and ,G: -17 kcal/mol. Targets that were in the 90th percentile of the raw alignment scores were selected as candidate miRNA targets.
- GenBank http://www.ncbi.nih.gov/Genbank index.html
- a dataset of human tRNA sequences http://rna.wustl.edu/GtRDB/Hs/Hs-seqs.html
- a dataset of human and mouse sn/snoR A sequences http://mbcr.bcm.tmc.edu/smallRNA/Database
- microRNAs http://www.sanger.ac.uk/Software/Rfam/microRNA/
- predictions of microRNAs 35
- the repeat element annotation of the HG16 human genome assembly http://genome.cse.ucsc.edu).
- EBV microRNAs originated from 5 different dsRNA precursors that are clustered in two regions of the EBV genome ( Figures 2A and B).
- the EBV microRNAs were all readily detectable by Northern blotting, including the approximately 60-nt fold-back precursor for 3 of the 5 microRNAs (Figure 2C).
- the first microRNA cluster is located within the mRNA of the BHFR1 gene encoding a distant Bcl-2 homolog, and we refer to these three microRNAs as miR-BHRFl-1 to miR-BHRFl-3.
- miRBHFRl-1 is located in the 5' UTR and miR-BHFRl-2 and -3 are positioned in the 3' UTR of the BHRFl mRNA. Structurally similar microRNA gene organization has been observed for some mammalian microRNAs that flank open reading frames in expressed sequence tags.
- EBV microRNAs cluster in intronic regions of the BART gene, and we refer to them as miR-BARTl and miR-BART2. Since microRNAs function in RNA silencing pathways either by targeting mRNAs for degradation or by repressing translation, we identified new viral regulators of host and/or viral gene expression.
- EBV latently infected cells can be found in three different latent stages (I to III, Figure 2 A) that are characterized by the expression of various subsets of the latent genes: six nuclear antigens (EBNAs 1, 2, 3 A, B, C, and EBNA-LP), three latent membrane proteins (LMPs 1, 2A and 2B), two non-coding RNAs (EBERs 1 and 2) and transcripts from the BamHI A region (BARTs/CSTs) whose coding capacity is still controversial.
- EBNAs 1, 2, 3 A, B, C, and EBNA-LP three latent membrane proteins
- LMPs 1, 2A and 2B three latent membrane proteins
- EBERs 1 and 2 two non-coding RNAs
- BARTs/CSTs transcripts from the BamHI A region
- RNAs from a latent-stage-III EBV cell line that expresses all latent genes.
- EBV microRNAs were detected in all latent stages consistent with the reported expression of BART during every stage of EBV infection.
- HD Hodgkin' s lymphoma
- BL Burkitt' s lymphoma
- BART microRNAs were detected in all latent stages consistent with the reported expression of BART during every stage of EBV infection.
- BHRFl microRNAs The expression pattern of BHRFl microRNAs is dependent on the EBV latency stage. While cell lines in stage II and III expressed BHRFl microRNAs ( Figure 2D, lanes 5-6), only one of the two stage I cell lines expressed BHRFl microRNAs ( Figure 2D, lanes 7, 8). Latency I cell lines are thought to express only EBNA 1, the EBERs and the BARTs.
- BHRFl protein is only detected in lytic stage, latent stage EBV transcripts encompassing the BHRFl region were observed previously. It is likely that the microRNAs BHRFl -1 to 3 are also expressed during lytic stage along with the BHRFl protein. The high- level transcription of BHRFl during the lytic cycle may exceed the cellular microRNA processing capacity and unprocessed transcripts could then be translated.
- EBV microRNAs targets To identify targets for EBV microRNAs, we used a computational method recently developed for prediction of Drosophila microRNAs targets (Enright et al, Genome Biol, 2003, 5:RI, 1). A set of approximately 20,000 non-redundant human 3' UTRs and the genome sequence of EBV were searched for potential microRNA binding sites. The top scoring hits for which a gene function annotation was available, are listed in Table 3. The majority of predicted host cell targets have more than one binding site for the viral microRNA, and approximately 50% of these are additionally targeted by one or several host cell microRNAs. Multiple microRNA binding sites are believed to act synergistically and increase targeting efficiency in a cooperative nonlinear fashion. Table 3. Predicted host cell target mRNAs of EBV microRNA.
- the gene name is indicated as recommended by HUGO, and the gene function annotation was extracted from Ensemble.
- the number of predicted microRNA binding sites in the 3 ' UTR of the targ gene (NS) and a percentile score ranking the target site predictions (%-ile) are indicated. If human microRNAs are also predicted to bind to a putative EBV microRNA regulated target, it is indicated in the last column.
- the predicated human microRNA binding site are also conserved in the orthologous mRNAs in mouse.
- BHRFl -3 NSEPl Y Box Binding protein 1 94 miR-95, miR-216, miR-136 BHRFl-3, TGIF 5'-TG-3' Interacting factor, Homeobox protein TGIF 1,1 97 miR-194 BART2 97
- BHRFl-3 PRF1 Perform 1 precursor 1 99 Signal transduction BART1 CXCL12 Stromal cell derived factor 1 precursor, Pre-B growth 100 miR-106, miR-135 Stimulating factor miR-197 BART2 GAB2 GRB2- Associated Binding Protein 2 4 100 miR- 155 BART2 TNFRSF 1 A Tumor Necrosis Factor Receptor Superfamily member 1 A 2 99 BHRFl -2 PIK3R1 Phosphatidylinositol 3-kinase regulatory Alpha Subunit 1 92 let-7b BHRFl -2, B7RP-1 B7 homolog, ICOS ligand precursor 1,3 97, miR-155 BART2 99
- EBV microRNA targets are prominent regulators of cell proliferation and apoptosis, which are presumably important for growth control of the infected cells.
- microRNA modulation of cell proliferation also provides new leads for studying the association of EBV with several cancerous malignancies.
- Another important group of EBV microRNA targets are B-cell specific chemokines and cytokines, which are important for leukocyte activation and/or chemotaxis. Down-regulation of these genes presumably contributes to escape of EBV-infected B cells from activated cytotoxic T cells.
- Additional targets include transcriptional regulators and components of signal transduction pathways that are critical for maintaining or switching between EBV lytic and latent stages.
- miR-BART2 is capable of targeting the virally encoded DNA polymerase BALF5 for degradation ( Figure 3).
- miR-BART2 is transcribed anti- sense to the BALF5 transcript and is therefore perfectly complementary to the BALF5 3' UTR and able to subject this mRNA for degradation.
- the clustered miRBHRFl-2 and -3 are complementary to the transcript encoding the lytic gene BFLF2 ( Figure 2A), whose function is currently unknown.
- the down-regulation of lytic genes by viral microRNAs may contribute to establishment and maintenance of latent infection.
- KSHV Kaposi's sarcoma-associated herpesvirus
- GenBank http://www.ncbi.nih.gov/Genbank/index.html
- a dataset of human tRNA sequences http://rna.wustl.edu/GtRDB/Hs/Hs-seqs.html
- a dataset of human and mouse sn/snoRNA sequences http://mbcr.bcm.tmc.edu/smalIRNA/Database
- a database of miRNAs http://www.sanger.ac.ulJSoftware/Rfam/mirna
- predictions of miRNAs and the repeat element annotation of the HG16 human genome assembly from UCSC
- KSHV microRNAs originated from 10 different dsRNA precursors that are all clustered in the same region of the KSHV genome ( Figure 4A and 4B).
- the KSHV microRNAs were designated miR-Kl to miR-KlO.
- the cluster is located within the mRNA of the K12 gene encoding a protein named Kaposin, which possesses some oncogenic properties.
- miR-Kl is located within the coding sequence of K12.
- Previous reports suggest that the K12 coding sequence region is complex and encodes several proteins named Kaposin A, B, and C (see Figure 4A).
- MiR-Kla corresponds to the sequenced genome present in BCBLl cells.
- MiR-Klb appears to be derived from a sequence isolated from a primary effusion lymphoma (PEL) tumor.
- PEL primary effusion lymphoma
- MiR-K2 to miR-Kl 0 are located in the intronic region of a longer transcript encoding K12 whose promoter is located upstream of the ORF 72 (see Figure 4A).
- KSHV miRNAs mature and precursor sequences. In bold the mature form, underlined the non-functional star sequence that was cloned for miR-K2 and miR-K6.
- KSHV microRNA sequence (5' to 3') Hairpin precursor sequence (5' to 3 ') miRNA CUGGAGGCUUGGGGCGAUACCACCACU miR-Kla UAGUGUUGUCCCCCCGAGUGGC CGUUUGUCUGUUGGCGAUUAGUGUUG UCCCCCCGAGUGGCCAG CUGGAGGCUUGGGGCGAUACCACCACU miR-Klb UGGUGUUGUCCCCCCGAGUGGC CGUUUGUCUGUUGGCGAUUGGUGUUG UCCCCCCGAGUGGCCAG GGGUCUACCCAGCUGCGUAAACCCCGC miR-K2 * ACCCAGCUGCGUAAACCCCGCU CUGGGUAUACGCAGCUGCGUAA UGCGUAAACACAGCUGGGUAUACGCA GCUGCGUAAACCC CUUGUCCAGCAGCACCUAAUCCAUCG miR-K3 5p CCAGCAGCACCUAAUCCAUCGG 3pUGAUGGUUUUCGGGCUGUUGAG GCGGUCGGGCUGA
- HCMV is a ubiquitous member of the ⁇ -herpesvirus family. Although HCMV infection of healthy children and adults is normally asymptomatic, it remains a leading cause of birth defects and an important cause of mortality in immunocompromised individuals.
- RNAs were cloned from primary human foreskin fibroblasts lytically infected with HCMV clinical strain VR1814. We cloned 424 small RNAs deriving from the virus genome in HCMV infected cells. Of these, 171 sequences were cloned once, and were dispersed throughout the genome; the 253 remaining sequences were cloned multiple times and analysis of the genomic sequences flanking these suggested structures characteristic of miRNAs (Tables 7 and 8 and Figure 6).
- RNAs were located in the UL region of the genome, and five derived from the US region. Interestingly, five miRNAs, miR-UL3, miR-UL4, miR-US3, miR-US4 and miR-US5 are transcribed on the complementary strand to known open reading frames (ORFs) ( Figure 6A). These five miRNAs may be involved in the cleavage of the complementary transcripts, as previously described for EBV miR-BART2 and the DNA polymerase BALF5.
- UL114 is a homolog of the mammalian uracyl-DNA glycosylase and has been shown to be required for efficient viral DNA replication.
- U 150 is an ORF that is present in the clinical strains of HCMV, but not in the laboratory strains.
- miRNAs are either located in intergenic regions (miR-ULl, miR-USl and miR-US2) or in an intronic region (miR-UL2). It is interesting to note that miR-UL2 is located in the intron of UL36, which has been described as an inhibitor of apoptosis that suppresses caspase-8 activation.
- Table 7 Composition in percentage of small RNA cDNA libraries prepared from HCMV- infected human cell line according to sequence annotation (see Table 4). The annotation for viral sequences is based on the published genomic sequence of HCMV FIX-BAC isolate VR1814 (Genbank AC146907).
- HCMV miRNAs mature and precursor sequences. In bold the mature form, underlined the
- HCMV miRNA Mature sequence follows q' Hairpin precursor sequence CCUGUCUAACUAGCCUUC miR-ULl UAACUAGCCUUCCCGUGAGA 101 AACAUGUAUCUCACCAGA GG miR-ULl* UCACCAGAAUGCUAGUUUGUAG 11 CCACGUCGUUGAAGACAC miR-UL2 UCGUUGAAGACACCUGGAAAGA UCGCUCGGGCACGUUCUU GUGCGUGG GACAGCCUCCGGAUCACAU miR-UL3 AAGUGACGGUGAGAUCCAGGCU 22 GCCAGCCUAAGUGACGGU AGCAGGUGAGGUUGGGGC miR-UL4 UCGUCCUCCCCUUCUUCACCG GAUUGUGGCGAGAACGUC ACCGCC UGAACGCUUUCGUCGUGUGU miR-USl UGACAAGCCUGACGAGAGCGU UACAGACCAUGACAAGCC GGAGGCUUUCGCCACACCU miR-US2 UUAUGAUAGGU
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WO2009012263A2 (en) * | 2007-07-18 | 2009-01-22 | The Trustees Of Columbia University In The City Of New York | Tissue-specific micrornas and compositions and uses thereof |
EP2112235A1 (en) | 2008-04-24 | 2009-10-28 | Max-Planck-Gesellschaft zur Förderung der Wissenschaften e.V. | Compositions and methods for microRNA expression profiling of nasopharyngeal carcinoma |
US8513209B2 (en) | 2007-11-09 | 2013-08-20 | The Board Of Regents, The University Of Texas System | Micro-RNAS of the MIR-15 family modulate cardiomyocyte survival and cardiac repair |
US9163235B2 (en) | 2012-06-21 | 2015-10-20 | MiRagen Therapeutics, Inc. | Inhibitors of the miR-15 family of micro-RNAs |
US10556020B2 (en) | 2014-09-26 | 2020-02-11 | University Of Massachusetts | RNA-modulating agents |
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WO2009012263A2 (en) * | 2007-07-18 | 2009-01-22 | The Trustees Of Columbia University In The City Of New York | Tissue-specific micrornas and compositions and uses thereof |
WO2009012263A3 (en) * | 2007-07-18 | 2009-03-26 | Univ Columbia | Tissue-specific micrornas and compositions and uses thereof |
US8586726B2 (en) | 2007-07-18 | 2013-11-19 | The Trustees Of Columbia University In The City Of New York | Tissue-specific MicroRNAs and compositions and uses thereof |
US8513209B2 (en) | 2007-11-09 | 2013-08-20 | The Board Of Regents, The University Of Texas System | Micro-RNAS of the MIR-15 family modulate cardiomyocyte survival and cardiac repair |
US9078919B2 (en) | 2007-11-09 | 2015-07-14 | The Board Of Regents, The University Of Texas System | Micro-RNAs of the miR-15 family modulate cardiomyocyte survival and cardiac repair |
EP2112235A1 (en) | 2008-04-24 | 2009-10-28 | Max-Planck-Gesellschaft zur Förderung der Wissenschaften e.V. | Compositions and methods for microRNA expression profiling of nasopharyngeal carcinoma |
US9163235B2 (en) | 2012-06-21 | 2015-10-20 | MiRagen Therapeutics, Inc. | Inhibitors of the miR-15 family of micro-RNAs |
US10556020B2 (en) | 2014-09-26 | 2020-02-11 | University Of Massachusetts | RNA-modulating agents |
US11464873B2 (en) | 2014-09-26 | 2022-10-11 | University Of Massachusetts | RNA-modulating agents |
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