CN107502609A - A kind of miRNA of targeting cJun signal paths and its preparation method and application - Google Patents

A kind of miRNA of targeting cJun signal paths and its preparation method and application Download PDF

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CN107502609A
CN107502609A CN201710807381.8A CN201710807381A CN107502609A CN 107502609 A CN107502609 A CN 107502609A CN 201710807381 A CN201710807381 A CN 201710807381A CN 107502609 A CN107502609 A CN 107502609A
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mirna
cjun
disease
mir
slow virus
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钱政江
李翔
杨海洋
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Shenzhen Institute of Advanced Technology of CAS
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Shenzhen Institute of Advanced Technology of CAS
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Abstract

The present invention relates to a kind of miRNA of targeting cJun signal paths and its preparation method and application, microRNA miR 4632 and its application more particularly to a kind of targeted inhibition cJun genes, the nucleotides sequence of the miRNA are classified as SEQ ID NO.1 or have the nucleotide sequence of at least 90% homogeneity with it.MiRNA of the present invention has endogenous, selectively targeted suppression cJun (cellular proto-oncogene) signal path, can prepare the medicine for suppressing the signal path abnormal activation diseases caused due to cJun mediations.

Description

A kind of miRNA of targeting cJun signal paths and its preparation method and application
Technical field
The present invention relates to biomedical sector, more particularly to a kind of miRNA of targeting cJun signal paths and its preparation side Method and application, and in particular to a kind of microRNA-miR-4632 of targeted inhibition cJun genes and its application, more particularly to Hsa-miR-4632 is being prepared by the application in the medicine of the signal path abnormal activation diseases caused of cJun mediations.
Background technology
Cellular proto-oncogene cJun is transcription factor important in core, while is also nuclear transcriptional activation albumen l The forming member of (activation protein, AP-1) family；CJun joins by single transcription factor or in the form of AP-1 With very polygenic transcriptional control, and various biological functions are played with this.Cell is receiving cell factor, growth signals, purple After the various outer signals such as outside line or environmental contaminants stimulate, swash via EGFR-TK associated receptor and cJun amino terminals The transmission of the signaling molecules such as enzyme JNK, so as to induce cJun activation, DNA fragmentation special in nucleus is ultimately applied to, adjusted Target gene is transcribed.CJun signal paths participate in the functions such as regulating cell propagation, differentiation, cycle and apoptosis.
Research shows that cJun mediates the abnormal activation of signal path, with myocardial hypertrophy, cerebral ischemia re-pouring injured, mental and physical efforts The occurrence and development phase of the cardiovascular and cerebrovascular diseases such as exhaustion, myocardial infarction, ischemia reperfusion, hypertension and atherosclerosis Close.In addition, the signal path continuous activation also with enterogastric diseases, diseases associated with inflammation, prostate, the nervous system disease and pernicious Tumour (such as cancer of the esophagus, oophoroma) forms closely related.Therefore, cJun signal paths great potential is as treatment relevant disease Medicine novel targets.
However, current exploitation is relatively fewer for the specific inhibitor of cJun signal paths.CN 104080469A are disclosed The purposes of new jnk inhibitor molecule and its by treating the purposes in the method for handling human or animal body, its main pin For cJun upstream signaling molecule JNK, the shortcomings of biological stability is poor, availability is relatively low, preparation technology is cumbersome.
Other also include peptide for the specific inhibitor of cJun signal paths and intend peptide inhibitor, the suppression of natural products class Preparation and the micromolecular inhibitor by computer medicine virtual screening scientific discovery.(1) peptide and intend peptide inhibitor be with The amino acid residue sequence of correlation molecule Phosphorylated products is the phosphoeptide of stencil design on JNK signal paths, JNK can be blocked to believe Number cJun is transferred to so as to reach inhibition, but such inhibitor easily metabolic inactivation, and bioavilability is relatively low in vivo； (2) natural products class inhibitor mainly includes the native chemical products such as terpene and flavonoids, has to JNK signal paths and suppresses to make Natural activity molecule, so as to suppress the transmission of cJun signals, but obtain such bioactive compound complex process, flow It is cumbersome, and can not be specifically cJun signal paths are suppressed；(3) by computer medicine virtual screening technology, can obtain A series of there must be the micromolecular compound of inhibitory activity to JNK signal paths, but the micromolecular compound synthesis much screened It is more difficult, and be difficult to keep high biostatic activity.
Therefore, prior art has yet to be improved and developed, and need to research and develop the more preferable cJun kinase signal pathway of effect and suppress Agent.
The content of the invention
In view of the shortcomings of the prior art, the invention provides a kind of miRNA of targeting cJun signal paths and its preparation side Method and application, the miRNA have endogenous, selectively targeted suppression cJun (cellular proto-oncogene) signal path, can made The standby medicine for suppressing the signal path abnormal activation diseases caused due to cJun mediations.
To use following technical scheme up to this purpose, the present invention：
In a first aspect, the present invention provides a kind of miRNA of targeted inhibition cJun signal paths, the nucleotides of the miRNA Sequence is SEQ ID NO.1 or has the nucleotide sequence of at least 90% homogeneity with it.
Nucleotide sequence shown in the SEQ ID NO.1 is as follows：5’-UGCCGCCCUCUCGCUGCUCUAG-3’.
According to the present invention, the nucleotides sequence of the miRNA is classified as the nucleotide sequence shown in SEQ ID NO.1.
In the present invention, by being overexpressed miRNA of the present invention, i.e. miR-4632 in human pulmonary artery smooth muscle cells, it can press down Activation or expression of the PDGF-BB (PDGFBB) processed to cJun signal paths；Tested, sent out by cell function MiRNA of the present invention is now overexpressed, i.e. miR-4632 can suppress PDGF-BB (PDGFBB) activation cJun signals Path induction, the abnormality proliferation of human pulmonary artery smooth muscle cells.
In the present invention, the preparation method of the miRNA includes the method for the synthesis of mature sequence full genome and clonal expression, institute The method for stating clonal expression is the ordinary skill in the art, and those skilled in the art can be entered using suitable method as needed Row structure, is not particularly limited herein.
In a specific embodiment, the construction method of the miRNA comprises the following steps：
(1) primer is designed, using human gene group DNA as template, enters performing PCR amplification；
(2) fragment by PCR amplifications is inserted between the XhoI and EcoRI of plv4/EGFP slow virus carriers；
(3) by the slow virus carrier transfectional cell of structure, miRNA is expressed.
The nucleotide sequence of the primer is as shown in SEQ ID NO.2-3；
Nucleotide sequence shown in the SEQ ID NO.2 is as follows：5’-CCGCTCGAGGACGAGCAGGACTGCGGA- 3’；
Nucleotide sequence shown in the SEQ ID NO.3 is as follows：5’-CCGGAATTCCAAGGACCTGAGCCCCAC- 3’.
Second aspect, the present invention provide miRNA as described in relation to the first aspect and are used to diagnosing, prevent, treat or in advance preparing The medicine of disease or the purposes of kit are assessed afterwards.
According to the present invention, the disease is disease caused by the signal path abnormal activation of cJun mediations.
According to the present invention, the disease is cardiovascular and cerebrovascular disease, enterogastric diseases, diseases associated with inflammation, leukaemia, forefront In gland, the nervous system disease, tumour or pulmonary hypertension any one or at least two combination.
In the present invention, the cardiovascular and cerebrovascular disease includes myocardial hypertrophy, cerebral ischemia re-pouring injured, heart failure, cardiac muscle The cardiovascular and cerebrovascular diseases such as infarct, ischemia reperfusion, hypertension and atherosclerosis；The tumour includes cancer of the esophagus or ovum The malignant tumours such as nest cancer.
" cerebral ischemia re-pouring " in the present invention refers to that ligaturing mouse coronary artery left anterior descending branch causes one section of myocardial ischemia After time, unclamping ligature makes cardiac muscle reach Reperfu- sion.
The third aspect, the present invention provide a kind of slow virus carrier, and the slow virus carrier includes as described in relation to the first aspect MiRNA nucleotide sequence.
Fourth aspect, the present invention provide a kind of recombinant slow virus, by comprising the slow virus carrier as described in the third aspect and The recombinant slow virus that packaging helper plasmid cotransfection mammalian cell obtains；
Preferably, the mammalian cell is HEK293T cells.
5th aspect, the present invention provide a kind of pharmaceutical composition, and the composition is included as described in relation to the first aspect miRNA。
According to the present invention, the effective dose of the miRNA is 10-200mg/kg, such as can be 10mg/kg, 20mg/ kg、30mg/kg、40mg/kg、50mg/kg、60mg/kg、70mg/kg、80mg/kg、90mg/kg、100mg/kg、120mg/kg、 130mg/kg, 150mg/kg, 160mg/kg, 180mg/kg or 200mg/kg.
According to the present invention, described pharmaceutical composition also includes any in pharmaceutically acceptable virus, carrier or auxiliary material It is a kind of or at least two combination.
In the present invention, including pharmaceutically acceptable virus, carrier or auxiliary material be selected from chitosan, cholesterol, liposome or Nano particle etc..
6th aspect, the present invention provide a kind of kit for being used for diagnosis and/or prognosis evaluation disease, the kit bag Include miRNA as described in relation to the first aspect and/or the probe or primer for specific detection miRNA.
According to the present invention, the nucleotide sequence of the probe or primer is as shown in SEQ ID NO.2-3；
Nucleotide sequence shown in the SEQ ID NO.2 is as follows：5’-CCGCTCGAGGACGAGCAGGACTGCGGA- 3’；
Nucleotide sequence shown in the SEQ ID NO.3 is as follows：5’-CCGGAATTCCAAGGACCTGAGCCCCAC- 3’.
Compared with prior art, the present invention has the advantages that：
(1) present invention discover that small miRNA is using cJun as target spot, the signal is suppressed by suppressing the expression of cJun molecules The activity of path, it can be used to treat as a kind of new cJun kinase inhibitors and be drawn by cJun kinase signal pathway abnormal activations The disease risen；
(2) miRNA of the invention is human endogenous's property miRNA, smaller to the toxicity of human body, can be preferably by human body profit With, can by be injected intravenously import human body in, privileged site is transported by blood circulation, so as to reach therapeutic purposes；
(3) miRNA of the invention can be used to treat cardiovascular disease as a kind of new cJun kinase signal pathway inhibitor Disease, enterogastric diseases, diseases associated with inflammation, prostate, the nervous system disease and malignant tumour etc..
Brief description of the drawings
Figure 1A is miR-4632 and cJ Jun 3 '-UTR binding sites；
Figure 1B is the influence for being overexpressed miR-4632 to cJun-UTR or cJun-UTR-mut uciferase activities, wherein, CJun-UTR-mut is the mutational vector that 3 '-UTR are built based on binding site；
Fig. 1 C influence to be overexpressed miR-4632 and human pulmonary artery smooth muscle cells cJun total proteins being expressed, wherein, Mimic Ctrl is the miR-4632 analogs of chemical synthesis；
After Fig. 1 D is are overexpressed miR-4632 to PDGFBB stimulation human pulmonary artery smooth muscle cells, phosphorylation activation state CJun protein expressions influence, wherein, Mimic Ctrl are that the structure of chemical synthesis is similar but insignificant sequence, Mimic The unused PDGF processing of Ctrl-1, Mimic Ctrl-2 are with PDGF processing；
Fig. 2A is after EdU detects PDGFBB stimulation human pulmonary artery smooth muscle cells, to be overexpressed miR-4632 cell proliferations The Representative fluorescence picture of influence, wherein, Mimic Ctrl are that the structure of chemical synthesis is similar but insignificant sequence, Mimic The unused PDGF processing of Ctrl-1, Mimic Ctrl-2 are with PDGF processing；Fig. 2 B are that EdU detections PDGFBB stimulates people's pulmonary artery smooth After myocyte, the statistical result that miR-4632 cell proliferations influence is overexpressed, wherein, Mimic Ctrl are the knot of chemical synthesis Structure is similar but insignificant sequence, the unused PDGF processing of Mimic Ctrl-1, and Mimic Ctrl-2 are with PDGF processing；
Fig. 2 C are after PDGFBB stimulates human pulmonary artery smooth muscle cells, are overexpressed shadows of the miR-4632 to PCNA protein expressions Ring, wherein, Mimic Ctrl are that the structure of chemical synthesis is similar but insignificant sequence, at the unused PDGF of Mimic Ctrl-1 Reason, Mimic Ctrl-2 are with PDGF processing.
Embodiment
Further to illustrate the technological means and its effect of the invention taken, below in conjunction with accompanying drawing and by specific real Mode is applied to further illustrate technical scheme, but the present invention is not limited in scope of embodiments.
Unless stated otherwise, the various raw materials employed in following examples derive from commercially available, and used method is equal For conventional technical means.
The miRNA miR-4632 of embodiment 1 preparation
MiR-4632 mature sequence is that 5 '-UGCCGCCCUCUCGCUGCUCUAG-3 ' (22bp, SEQ ID NO.1) can It is made in the following manner：
1) chemical synthesis
MiR-4632 analog (miR-4632mimic) has the synthesis of Guangzhou Rui Bo bio tech ltd, is double-strand RNA, endogenous sexal maturity miR-4632 high level expressions can be simulated；Standard purification, in -20 DEG C of preservations.Dissolved with the water without RNase Be configured to 20M storing liquid, be placed in -80 DEG C it is standby, be diluted to required concentration during use.
2) miR-4632 of viral vector mediation expression
(1) in miRBase (http://www.mirbase.org/) obtain in database miR-4632 mature sequence and Precursor sequence (pre-miRNA) information.Two about 300bp of mature sequence flanking sequence is found in human genome database, According to flanking sequence design primer, and XhoI and EcoRI restriction enzyme site and protection base are added respectively, design primer：
- CCGCTCGAGGACGAGC the AGGACTGCGGA-3 ' of sense primer 5 ' (SEQ ID NO.2)；
The sequence of anti-sense primer is 5 '-CCGGAATTCCAAGGACCTGAGCCC CAC-3 ' (SEQ ID NO.3)；
(2) using human gene group DNA as template, performing PCR is entered by the primer of design and expanded, PCR reaction conditions are：95 DEG C of changes Property 2 minutes, 95 DEG C 30 seconds, 55 DEG C 30 seconds, 72 DEG C 40 seconds carry out 35 circulation, 72 DEG C extend 3 minutes；
(3) after the PCR primer Ago-Gel of amplification is called, with glue reclaim kit (E.Z.N.A Gel Extraction Kit, Omgega) fragment recovery is carried out, double digestion is carried out with restriction enzyme XhoI and EcoRI (NEB), PCR fragment after digestion is illustrated after being reclaimed by DNA purification kits (E.Z.N.A Cycle-Pure Kit, Omgega) As Insert Fragment, insert in PLV4/EGFP slow virus carriers；Carrier is with restriction enzyme XhoI and EcoRI NEB) carry out Reclaimed, purified as carrier segments after double digestion；
(4) Insert Fragment is connected 2h with 16 DEG C using T4 ligases (Promega) with carrier segments, then produced connection Thing is converted to E. coli competent STBL3, and 37 DEG C are incubated overnight the well-grown single bacterium colony of rear picking corresponding containing carrier Expanded in the LB culture mediums of antibiotic, sequence verification is carried out after extracting plasmid；
(5) DNA vector made from can be transfected into cell, transient expression miRNA, i.e. miR-4632.
Obtained slow virus carrier is packed available for slow virus, and step is as described below：
Cell line needed for (1 ') slow virus packaging is HEK293T cell lines, and required plasmid includes Lentiviral PLVX-miR-4632 and slow virus packaging plasmid.Cell transfecting uses conventional calcium phosphate transfection method, with 2M CaCl2With plasmid Mix, 2 × HBS solution is then added dropwise and jog mixes, is finally slowly uniformly added into mixed solution in cell culture fluid. Transfection changes nutrient solution after 12 hours；
(2 ') after transfecting 48-72 hours, the cell culture fluid containing virion is collected, 4000rpm room temperatures centrifuge 10 points Clock, collect supernatant and dispense be placed in -80 DEG C it is standby；
(3 ') packaged slow virus can be used for the selected cell of infection, and being obtained after being screened by antibiotic puromycin can The stable cell for being overexpressed miR-4632.
The miR-4632 targeted inhibition cJun molecules of embodiment 2 and its signal path
(1) cell culture：HEK293A cells (are purchased from ATCC, Manassas, VA), with the DMEN containing 10% hyclone Medium culture, the good cell of growth conditions is collected, with 6 × 104/ hole is inoculated in 24 orifice plates, 37 DEG C, 5%CO2Culture 24 Hour；
(2) human pulmonary artery smooth muscle cells is purchased from Lonza (Walkersville, MD), complete in the SMCM containing 5% serum Cultivated in full culture medium, collect the good cell of growth conditions, centrifugation technique, with 6 × 105It is inoculated in 60mm wares, 37 DEG C, 5%CO2Culture 24 hours；
(3) transfect：It is double glimmering with calcium phosphate method transfectional cell, cotransfection CCJUN 3 '-UTR when cell density is up to 70% Light element enzyme report carrier or 3 '-UTR Dual-Luciferase report carriers of mutation, miR-4632 over-express vectors pLVX-CMV- MiR-4632 or pLVX-CMV-control, transfection two days later, remove nutrient solution, PBS is twice afterwards with 50 1 × cracking of μ L Liquid cell lysis；
(4) the miR-4632 analogs (mimic) of chemical synthesis are used for the expression for being overexpressed or suppressing miR-4632, with Cel-miR-67 is purchased from the auspicious rich biology in Guangzhou as negative control.By cell kind in culture dish, when cell density reaches When 70%, transfected by Lipofectamine 2000 (Invitrogen), transfection changes culture medium after 6 hours, continues to cultivate It is used within 24 hours test；
Luciferase detects
(E1810, Promega) kit reference explanation book, which is detected, using Dual-Luciferase carries out uciferase activity inspection Survey, with firefly luciferase numerical value divided by internal reference renilla luciferase readings, you can the uciferase activity number corrected Value.By the uciferase activity numerical value of CCJUN 3 '-UTR carriers compared with control group.
The 3 '-UTR binding sites of the miR-4632 and cJun as shown in Figure 1A, fluorescence mycin testing result such as Figure 1B institutes Show, compared with the control, being overexpressed the CJUN uciferase activities of miR-4632 groups significantly reduces, and illustrates that miR-4632 may be with CJun 3 '-UTR can target combination；3 '-UTR mutational vector cJun-UTR-mut is built based on binding site, using prominent Variant vector carries out Dual-Luciferase Activity determination again, and compared with wild type UTR, the mutation of cJun binding sites causes fluorescence The recovery of plain enzymatic activity.
Protein hybridization detects
Total protein of cell is collected, polyacrylamide gel electrophoresis is carried out after protein quantification, is examined using cJun protein antibodies CJun expression is surveyed, as shown in Figure 1 C, compared with the control, human pulmonary artery smooth muscle cells endogenous can be suppressed by being overexpressed miR-4632 CJun expression；Phosphorylation cJun is detected using cJun phospho-ABs, (Ctrl-1 is the untreated expressions of PDGF compared with the control Cell function normal condition, Ctrl-2 are that PDGF processing represents cell function abnormality), as shown in figure iD, it is overexpressed miR- 4632 can significantly inhibit cJun phosphorylation.
The present embodiment result shows：By targetting, CJUN 3 '-UTR suppress CJUN expression to miR-4632 and CJUN swashs Enzyme signal path.
It is thin that the miR-4632 of embodiment 3 suppresses PDGF-BB (PDGFBB) induction people's arteria pulmonalis smooth muscle Born of the same parents breed.
CJun signal paths are activated to PDGF-BB (PDGFBB) to inquire into miR-4632, so as to cause The regulating and controlling effect of human pulmonary artery smooth muscle cells dysfunction：People's pulmonary artery is improved with expression miR-4632 slow-virus infection The level of miR-4632 in smooth muscle cell, it is true that the propagation detection of marker protein PCNA and EdU cell is then bred by cell The propagation for determining cell is horizontal.
EdU marks are carried out using EdU cell proliferation detecting kits (sharp rich biology), are carried out according to kit specification real Operation is tested, key step is as follows：1) cell transfecting：Inoculation 1 × 104Individual cell carries out transfection in fact in 48 orifice plates after cultivating 24h Test, carry out Nature enemy with serum free medium after second day culture 24h, then add PDGF-BB (PDGFBB) and 20 μM of EdU continue 4 hours；2) cell dyeing is taken pictures：Cell is fixed 30 points with 4% paraformaldehyde room temperature Clock, 0.5%Triton X-100 carry out film penetrating 10 minutes, PBS, and 150 1 × Apollo of μ L dyeing is then added per hole Also it is incubated 30 minutes, DNA 1 × Hochest (per the μ L of hole 150) dyeing 5 minutes, is taken pictures under fluorescence microscope.
As a result as shown in Fig. 2A, Fig. 2 B and Fig. 2 C, from Fig. 2A and Fig. 2 B, under PDGFBB stimulations, compared with the control (Ctrl-1, which is that PDGF is untreated, represents cell function normal condition, and Ctrl-2 is that PDGF processing represents cell function exception shape State), it is that human pulmonary artery smooth muscle cells propagation declines 32% respectively to be overexpressed miR-4632；From Fig. 2 C, miR- is overexpressed 4632, which also make one arteria pulmonalis smooth muscle proliferation marker albumen PCNA expression, lowers 31%；Illustrate that miR-4632 causes to growth factor Proliferation of Pulmonary Artery Smooth Muscle Cells there is obvious inhibitory action.
In summary, present invention discover that small miRNA is using cJun as target spot, suppressed by suppressing the expression of cJun molecules The activity of the signal path, it is extremely sharp by cJun kinase signal pathway that treatment can be used for as a kind of new cJun kinase inhibitors Caused disease living.
Applicant states that the present invention illustrates the method detailed of the present invention, but not office of the invention by above-described embodiment It is limited to above-mentioned method detailed, that is, does not mean that the present invention has to rely on above-mentioned method detailed and could implemented.Art Technical staff it will be clearly understood that any improvement in the present invention, equivalence replacement and auxiliary element to each raw material of product of the present invention Addition, selection of concrete mode etc., within the scope of all falling within protection scope of the present invention and disclosing.
Sequence table
<110>Shenzhen Institutes of Advanced Technology, Chinese Academy of Science
<120>A kind of miRNA of targeting cJun signal paths and its preparation method and application
<130> 2017
<141> 2017-09-08
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<213>Artificial synthesized sequence ()
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Claims (10)

1. a kind of miRNA of targeting cJun signal paths, it is characterised in that the nucleotides sequence of the miRNA is classified as SEQ ID NO.1 or the nucleotide sequence with it with least 90% homogeneity.
2. miRNA according to claim 1, it is characterised in that the nucleotides sequence of the miRNA is classified as SEQ ID NO.1 Shown nucleotide sequence.
3. miRNA according to claim 1 or 2 is used to diagnosing, prevents, treated or the medicine of prognosis evaluation disease preparing Or the purposes of kit.
4. purposes according to claim 3, it is characterised in that the disease is the signal path abnormal activation of cJun mediations Caused disease；
Preferably, the disease is cardiovascular and cerebrovascular disease, enterogastric diseases, diseases associated with inflammation, leukaemia, prostate, nerveous system Unite in disease, tumour or pulmonary hypertension any one or at least two combination.
5. a kind of slow virus carrier, it is characterised in that the slow virus carrier includes miRNA's as claimed in claim 1 or 2 Nucleotide sequence.
6. a kind of recombinant slow virus, it is characterised in that slow virus carrier as claimed in claim 5 and packaging auxiliary matter will be included The recombinant slow virus that grain cotransfection mammalian cell obtains；
Preferably, the mammalian cell is HEK293T cells.
7. a kind of pharmaceutical composition, it is characterised in that the composition includes miRNA as claimed in claim 1 or 2.
8. composition according to claim 7, it is characterised in that the effective dose of the miRNA is 10-200mg/kg；
Preferably, described pharmaceutical composition also include in pharmaceutically acceptable virus, carrier or auxiliary material any one or extremely Few two kinds combination.
9. a kind of kit for being used for diagnosis and/or prognosis evaluation disease, it is characterised in that the kit includes claim MiRNA described in 1 or 2 and/or the probe or primer for specific detection miRNA.
10. kit according to claim 9, it is characterised in that the nucleotide sequence such as SEQ of the probe or primer Shown in ID NO.2-3.
CN201710807381.8A 2017-09-08 2017-09-08 A kind of miRNA of targeting cJun signal paths and its preparation method and application Pending CN107502609A (en)

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PCT/CN2017/111054 WO2019047368A1 (en) 2017-09-08 2017-11-15 Mirna targeting cjun signaling pathway and preparation method therefor and use thereof

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AU2010210426B2 (en) * 2009-02-06 2015-06-11 Imago Pharmaceuticals, Inc. Inhibitors of Jun N-terminal kinase
AU2013324716B2 (en) * 2012-09-26 2019-06-13 Guandong Mijinjia Biotechnology Co., Ltd Oligomers with improved off-target profile
CA2956146A1 (en) * 2014-07-24 2016-01-28 Memorial Sloan Kettering Cancer Center Method and composition for targeted delivery of therapeutic agents

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