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研究生:林宜君
研究生(外文):Lin, Yi-Chun
論文名稱:長效型混成桿狀病毒結合CRISPRi系統並應用於調控肝細胞癌(HCC)中之miR-221/222表現量
論文名稱(外文):Inhibition of miR-221/222 Expression in Hepatocellular Carcinoma via Baculovirus-Mediated CRISPRi System
指導教授:胡育誠胡育誠引用關係
指導教授(外文):Hu, Yu-Chen
口試委員:江啟勳陳冠宇
口試委員(外文):Chiang, Chi-ShiunChen, Kuan-Yu
口試日期:2017-07-21
學位類別:碩士
校院名稱:國立清華大學
系所名稱:化學工程學系所
學門:工程學門
學類:化學工程學類
論文種類:學術論文
論文出版年:2017
畢業學年度:105
語文別:中文
論文頁數:86
中文關鍵詞:CRISPRi肝細胞癌微小核醣核酸
外文關鍵詞:CRISPRiHCCmicroRNAmiR-221/222
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  在醫學技術不斷進步下,癌症治療已有很大的進展,但是肝細胞癌 (hepatocellular carcinoma, HCC)仍是一個非常普遍且致命的癌症,因此開發有效且安全的治療方式為癌症治療一大課題。MicroRNA (微核醣核酸)為長度約20-24個鹼基的短片段RNA,其主要功能為可與mRNA的5’UTR或3’UTR的部分核苷酸配對,造成目標mRNA降解,或影響其轉譯。自microRNA被發現後,科學家致力於研究其與疾病的關聯,而其中尤以癌症居首,透過調控miRNAs作為癌症治療的策略也不斷地推陳出新。而近年來新興的CRISPRi (CRISPR interference)基因調控系統,擁有僅需設計sgRNA (single guide RNA)來辨識目標基因即可達到抑制基因或miRNA表現的功效,不僅設計便捷,且能同時調控多個目標基因。因此我們認為開發一套CRISPRi系統來調控miRNAs並應用於癌症基因治療深具潛力。
  我們初步證實,與正常肝組織細胞相比,肝癌細胞Mahlavu中之miR-221及miR-222過量表現 (Mahlavu為一具有高轉移性之肝癌細胞株),而其抑癌基因PTEN之mRNA表現量則偏低,根據文獻報導指出,肝癌細胞中過量表現的miR-221及miR-222,會促使肝癌的腫瘤生長及轉移,而過量的miR-221也會抑制其目標基因PTEN的表現。本實驗即希望建立以Sleeping Beauty (SB)轉位系統為基礎之長效型混成桿狀病毒(Baculovirus, BV)載體,結合CRISPRi調控系統,抑制miR-221及miR-222的上游啟動子與轉錄起始點,使miR-221及miR-222的轉錄受到干擾,下調miR-221及miR-222的表現量進而達到促進抑癌基因的表現量。我們建構了同時表現dCas9及三組sgRNA之桿狀病毒並藉由轉導送入Mahlavu細胞中,以qRT-PCR分析miR-221/222之表現量發現,在多次獨立的轉導實驗中,我們所建立之桿狀病毒無法穩定的對miR-221/222造成抑制,因此我們將CRISPRi系統針對dCas9進核及sgRNA結構加以優化,期望能夠達到穩定且良好的抑制效果,完成新的混成桿狀病毒後,我們以qRT-PCR分析轉導Mahlavu後內源性miR-221/222之表現量,發現仍然無法達到抑制效果。於本研究後續討論中,我們會討論造成抑制效果差的原因,以及對於未來與本研究相關的改進方向。
In spite of the development of new therapies to cancers, hepatocellular carcinoma (HCC) is still one of the most malignant tumors because of high operative mortality, recurrence rate and lack of survival benefit, thus entailing the need to develop new and strong therapy. Small non-coding RNAs consisting of 21-25 nucleotides called microRNAs (miRNAs) induce mRNA degradation or suppress mRNA translation by binding to the 3’-untranslated region of target mRNAs. Deregulation of miRNAs facilitates cancer development through the upregulation of oncogenes and the silencing of tumor suppressor genes, which plays pivotal regulatory roles in cancers, and tumor suppressor miRNAs can modulate gene regulatory networks to inhibit tumor cell proliferation, invasion and metastasis, hence holding great promise as a therapeutics strategy for HCC. For gene suppression, a novel RNA-guided CRISPR interference (CRISPRi) technology was recently well-developed, which only requires co-expression of dead Cas9 (dCas9) and single guide RNA (sgRNA) that target specific genomic sequences without permanently altering genomic DNA.
We found that the miR-221&222 are overexpressed in Mahlavu cells compared with normal liver sample, and the tumor suppressor gene, PTEN, is relatively low expressed in hepatocarcinoma cells, especially Mahlavu cells. We intend to develop a baculovirus-mediated, CRISPRi-based gene regulation approach to specificly inhibit miR-221/222 expression. We constructed 2 baculovirus vectors exploiting the Sleeping Beauty system (SB) for sustained transgene expression: Bac-dCas9K-gRNAabc expressed dCas9 protein and three sgRNAs that target upstream of miR-221/222 cluster, Bac-dCas9K-gRNAØ expressed dCas9 protein and sgRNA that doesn’t have targeting site. By co-transducing Mahlavu cells with another SB transposase-expressing Bac-SB/W, we found that Bac-dCas9K-gRNAabc couldn’t stably attenuate miR-221/222 levels. To solve this problem, we design another 2 baculovirus which contain 2 and 3 nucleus localization signals (NLS), and we also optimize the sgRNA scaffold sequence to enhance the affinity between dCas9 protein and sgRNA. As the result, we still couldn’t stably inhibit the highly expressed oncomiR.
摘要 I
Abstract II
致謝 III
目錄 IV
圖目錄 VII
表目錄 IX
第一章 文獻回顧 1
1-1 桿狀病毒表現系統 1
1-1-1 桿狀病毒表現系統之發展 1
1-1-2 桿狀病毒/哺乳動物細胞表現系統 1
1-2 RNAi系統 4
1-2-1 microRNAs (miRNAs) 4
1-2-2 miRNA cluster於疾病中之影響 5
1-2-3 以miRNAs為基礎的治療方法 5
1-3 CRISPR基因編輯與調控系統 6
1-3-1 CRISPR/Cas9基因剪輯系統 6
1-3-2 CRISPRi基因調控系統 8
1-3-3 sgRNA標的位置及scaffold之設計 9
1-3-4 核定位訊號(NLS, Nucleus localization signal) 10
1-4 RNAi與CRISPRi比較 11
1-5 肝細胞癌(Hepatocellular Carcinoma, HCC) 12
1-5-1 背景 12
1-5-2 miRNAs在肝臟疾病的作用 13
1-5-3 CRISPR基因編輯與調控系統結合RNAi在肝臟疾病的作用 14
1-6 長期表現系統Sleeping Beauty (SB) 14
1-7 研究動機與目的 16
第二章 實驗材料與方法 25
2-1 細胞培養 25
2-1-1 昆蟲細胞之培養 25
2-1-2 哺乳動物細胞之培養 25
2-2 質體建構之方法 26
2-2-1 PCR反應 26
2-2-2 DNA引子黏合反應(primer annealing) 26
2-2-3 限制酶(DNA restriction enzyme)反應及接合酶(DNA ligase)反應 27
2-3 重組桿狀病毒建構與製備 27
2-3-1 重組桿狀病毒之建構 27
2-3-1-1 pTA-TypeC及pTA-TypeD之建構 27
2-3-1-2 pTA-sgRNA-2.0之建構 27
2-3-1-3 sgRNA之設計與建構方法 28
2-3-1-4 含有dCas9-KRAB表現匣之質體建構 30
2-3-1-5 pBac-IR/DR及pBac-WPRE之建構 31
2-3-1-6 重組桿狀病毒donor plasmid之建構 32
2-3-1-7 pBac-SB/W之建構 33
2-3-2 重組桿狀病毒製備與放大 33
2-3-2-1 重組表現載體之轉置(transposition)反應(Bac-to-Bac system) 33
2-3-2-2 重組Bacmid之分離及轉染昆蟲細胞 33
2-3-2-3 重組桿狀病毒之放大 34
2-3-3 超高速離心濃縮桿狀病毒 35
2-3-4 重組桿狀病毒之感染效價 35
2-4 桿狀病毒之轉導策略 35
2-5 測試CRISPRi系統質體之建構 36
2-6 實驗分析及方法 37
2-6-1 即時偵測同步定量聚合酶連鎖反應分析(qRT-PCR analysis) 37
2-6-2 西方墨點法(Western blot analysis) 40
2-6-3 統計學分析 41
第三章 實驗結果 61
3-1 小鼠肝癌細胞之測試 61
3-1-1 miR-214及 miR-199a於小鼠肝癌細胞之內源性表現量 61
3-1-2 PTEN蛋白質於小鼠肝癌細胞之內源性表現量 61
3-2 人類肝癌細胞之測試 62
3-2-1 不同miRNA於人類肝癌細胞內源性表現量 62
3-2-2 PTEN mRNA於Mahlavu細胞內源性表現量 62
3-3 驗證CRISPRi系統於Mahlavu中之調控效果 62
3-3-1 qRT-PCR測定miR-221、miR-222表現量 62
3-3-2 確認CRISPRi於Mahlavu細胞中標的位置之序列 63
3-3-3 利用螢光系統測試桿狀病毒轉導細胞後之dCas9功能 64
3-4 CRISPRi系統之優化與測試其效果 64
3-4-1 CRISPRi系統之優化 64
3-4-2 螢光系統測試優化之CRISPRi抑制效果 65
3-4-3 優化後之CRISPRi系統於Mahlavu細胞中的調控效果 65
3-5 結論 65
第四章 討論與未來展望 76
4-1 與本研究相似之文獻比較 76
4-2 CRISPRi系統無法抑制Mahlavu中內源性miR-221/222表現量 77
4-3 未來展望 78
第五章 參考文獻 80
Airenne KJ, Hu YC, Kost TA, Smith RH, Kotin RM, Ono C, Matsuura Y, Wang S, Yla-Herttuala S. 2013. Baculovirus: an insect-derived vector for diverse gene transfer applications. Mol Ther 21: 739-749.
Aronovich EL, Bell JB, Khan SA, Belur LR, Gunther R, Koniar B, Schachern PA, Parker JB, Carlson CS, Whitley CB, McIvor RS, Gupta P, Hackett PB. 2009. Systemic correction of storage disease in MPS I NOD/SCID mice using the sleeping beauty transposon system. Mol Ther 17: 1136-1144.
Bak XY, Yang J, Wang S. 2010. Baculovirus-transduced bone marrow mesenchymal stem cells for systemic cancer therapy. Cancer Gene Ther 17: 721-729.
Balakrishnan B, Jayandharan GR. 2014. Basic biology of adeno-associated virus (AAV) vectors used in gene therapy. Curr Gene Ther 14: 86-100.
Baldo A, van den Akker E, Bergmans HE, Lim F, Pauwels K. 2013. General considerations on the biosafety of virus-derived vectors used in gene therapy and vaccination. Curr Gene Ther 13: 385-394.
Bao L, Yan Y, Xu C, Ji W, Shen S, Xu G, Zeng Y, Sun B, Qian H, Chen L, Wu M, Su C, Chen J. 2013. MicroRNA-21 suppresses PTEN and hSulf-1 expression and promotes hepatocellular carcinoma progression through AKT/ERK pathways. Cancer Lett 337: 226-236.
Bikard D, Jiang W, Samai P, Hochschild A, Zhang F, Marraffini LA. 2013. Programmable repression and activation of bacterial gene expression using an engineered CRISPR-Cas system. Nucleic Acids Res 41: 7429-7437.
Boettcher M, McManus MT. 2015. Choosing the Right Tool for the Job: RNAi, TALEN, or CRISPR. Mol Cell 58: 575-585.
Callegari E, Gramantieri L, Domenicali M, D'Abundo L, Sabbioni S, Negrini M. 2015. MicroRNAs in liver cancer: a model for investigating pathogenesis and novel therapeutic approaches. Cell Death Differ 22: 46-57.
Chalberg TW, Portlock JL, Olivares EC, Thyagarajan B, Kirby PJ, Hillman RT, Hoelters J, Calos MP. 2006. Integration specificity of phage phiC31 integrase in the human genome. J Mol Biol 357: 28-48.
Chang H, Yi B, Ma R, Zhang X, Zhao H, Xi Y. 2016. CRISPR/cas9, a novel genomic tool to knock down microRNA in vitro and in vivo. Sci Rep 6: 22312.
Chen B, Gilbert LA, Cimini BA, Schnitzbauer J, Zhang W, Li GW, Park J, Blackburn EH, Weissman JS, Qi LS, Huang B. 2013. Dynamic imaging of genomic loci in living human cells by an optimized CRISPR/Cas system. Cell 155: 1479-1491.
Chen CL, Luo WY, Lo WH, Lin KJ, Sung LY, Shih YS, Chang YH, Hu YC. 2011. Development of hybrid baculovirus vectors for artificial MicroRNA delivery and prolonged gene suppression. Biotechnol Bioeng 108: 2958-2967.
Chen CL, Wu JC, Chen GY, Yuan PH, Tseng YW, Li KC, Hwang SM, Hu YC. 2015. Baculovirus-mediated miRNA regulation to suppress hepatocellular carcinoma tumorigenicity and metastasis. Mol Ther 23: 79-88.
Chen CY, Lin CY, Chen GY, Hu YC. 2011. Baculovirus as a gene delivery vector: recent understandings of molecular alterations in transduced cells and latest applications. Biotechnol Adv 29: 618-631.
Chen CY, Wu HH, Chen CP, Chern SR, Hwang SM, Huang SF, Lo WH, Chen GY, Hu YC. 2011. Biosafety assessment of human mesenchymal stem cells engineered by hybrid baculovirus vectors. Mol Pharm 1505-1514.
Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, Jiang W, Marraffini LA, Zhang F. 2013. Multiplex genome engineering using CRISPR/Cas systems. Science 339: 819-823.
Copeland MF, Politz MC, Pfleger BF. 2014. Application of TALEs, CRISPR/Cas and sRNAs as trans-acting regulators in prokaryotes. Curr Opin Biotechnol 29: 46-54.
Deltcheva E, Chylinski K, Sharma CM, Gonzales K, Chao Y, Pirzada ZA, Eckert MR, Vogel J, Charpentier E. 2011. CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III. Nature 471: 602-607.
Di Leva G, Gasparini P, Piovan C, Ngankeu A, Garofalo M, Taccioli C, Iorio MV, Li M, Volinia S, Alder H, Nakamura T, Nuovo G, Liu Y, Nephew KP, Croce CM. 2010. MicroRNA cluster 221-222 and estrogen receptor alpha interactions in breast cancer. J Natl Cancer Inst 102: 706-721.
Dismuke DJ, Tenenbaum L, Samulski RJ. 2013. Biosafety of recombinant adeno-associated virus vectors. Curr Gene Ther 13: 434-452.
Dominguez AA, Lim WA, Qi LS. 2016. Beyond editing: repurposing CRISPR-Cas9 for precision genome regulation and interrogation. Nat Rev Mol Cell Biol 17: 5-15.
Ebert MS, Sharp PA. 2010. MicroRNA sponges: progress and possibilities. RNA 16: 2043-2050.
Ehrhardt A, Engler JA, Xu H, Cherry AM, Kay MA. 2006. Molecular analysis of chromosomal rearrangements in mammalian cells after phiC31-mediated integration. Hum Gene Ther 17: 1077-1094.
Fan QW, Weiss WA. 2010. Targeting the RTK-PI3K-mTOR axis in malignant glioma: overcoming resistance. Curr Top Microbiol Immunol 347: 279-296.
Fulco CP, Munschauer M, Anyoha R, Munson G, Grossman SR, Perez EM, Kane M, Cleary B, Lander ES, Engreitz JM. 2016. Systematic mapping of functional enhancer-promoter connections with CRISPR interference. Science 354: 769-773.
Galardi S, Mercatelli N, Farace MG, Ciafre SA. 2011. NF-kB and c-Jun induce the expression of the oncogenic miR-221 and miR-222 in prostate carcinoma and glioblastoma cells. Nucleic Acids Res 39: 3892-3902.
Gao Y, Xiong X, Wong S, Charles EJ, Lim WA, Qi LS. 2016. Complex transcriptional modulation with orthogonal and inducible dCas9 regulators. Nat Methods 13: 1043-1049.
Garofalo M, Di Leva G, Romano G, Nuovo G, Suh SS, Ngankeu A, Taccioli C, Pichiorri F, Alder H, Secchiero P, Gasparini P, Gonelli A, Costinean S, Acunzo M, Condorelli G, Croce CM. 2009. miR-221&222 regulate TRAIL resistance and enhance tumorigenicity through PTEN and TIMP3 downregulation. Cancer Cell 16: 498-509.
Gilbert LA, Horlbeck MA, Adamson B, Villalta JE, Chen Y, Whitehead EH, Guimaraes C, Panning B, Ploegh HL, Bassik MC, Qi LS, Kampmann M, Weissman JS. 2014. Genome-Scale CRISPR-Mediated Control of Gene Repression and Activation. Cell 159: 647-661.
Gilbert LA, Larson MH, Morsut L, Liu Z, Brar GA, Torres SE, Stern-Ginossar N, Brandman O, Whitehead EH, Doudna JA, Lim WA, Weissman JS, Qi LS. 2013. CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes. Cell 154: 442-451.
Guilinger JP, Thompson DB, Liu DR. 2014. Fusion of catalytically inactive Cas9 to FokI nuclease improves the specificity of genome modification. Nat Biotechnol 32: 577-582.
Hildebrandt-Eriksen ES, Aarup V, Persson R, Hansen HF, Munk ME, Orum H. 2012. A locked nucleic acid oligonucleotide targeting microRNA 122 is well-tolerated in cynomolgus monkeys. Nucleic Acid Ther 22: 152-161.
Ho TT, Zhou N, Huang J, Koirala P, Xu M, Fung R, Wu F, Mo YY. 2015. Targeting non-coding RNAs with the CRISPR/Cas9 system in human cell lines. Nucleic Acids Res 43: e17.
Horvath P, Barrangou R. 2010. CRISPR/Cas, the immune system of bacteria and archaea. Science 327: 167-170.
Housden BE, Valvezan AJ, Kelley C, Sopko R, Hu Y, Roesel C, Lin S, Buckner M, Tao R, Yilmazel B, Mohr SE, Manning BD, Perrimon N. 2015. Identification of potential drug targets for tuberous sclerosis complex by synthetic screens combining CRISPR-based knockouts with RNAi. Sci Signal 8: rs9.
Huang S, He X. 2011. The role of microRNAs in liver cancer progression. Br J Cancer 104: 235-240.
Huo W, Zhao G, Yin J, Ouyang X, Wang Y, Yang C, Wang B, Dong P, Wang Z, Watari H, Chaum E, Pfeffer LM, Yue J. 2017. Lentiviral CRISPR/Cas9 vector mediated miR-21 gene editing inhibits the epithelial to mesenchymal transition in ovarian cancer cells. J Cancer 8: 57-64.
Hwang BY, Schaffer DV. 2013. Engineering a serum-resistant and thermostable vesicular stomatitis virus G glycoprotein for pseudotyping retroviral and lentiviral vectors. Gene Ther 20: 807-815.
Hwang WY, Fu Y, Reyon D, Maeder ML, Tsai SQ, Sander JD, Peterson RT, Yeh JR, Joung JK. 2013. Efficient genome editing in zebrafish using a CRISPR-Cas system. Nat Biotechnol 31: 227-229.
Ivics Z, Izsvak Z. 2011. Nonviral gene delivery with the sleeping beauty transposon system. Hum Gene Ther 22: 1043-1051.
Izsvak Z, Ivics Z, Plasterk RH. 2000. Sleeping Beauty, a wide host-range transposon vector for genetic transformation in vertebrates. J Mol Biol 302: 93-102.
Janssen HL, Reesink HW, Lawitz EJ, Zeuzem S, Rodriguez-Torres M, Patel K, van der Meer AJ, Patick AK, Chen A, Zhou Y, Persson R, King BD, Kauppinen S, Levin AA, Hodges MR. 2013. Treatment of HCV infection by targeting microRNA. N Engl J Med 368: 1685-1694.
Jiang Y, Chen B, Duan C, Sun B, Yang J, Yang S. 2015. Multigene editing in the Escherichia coli genome via the CRISPR-Cas9 system. Appl Environ Microbiol 81: 2506-2514.
Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E. 2012. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337: 816-821.
Kalderon D, Roberts BL, Richardson WD, Smith AE. 1984. A short amino acid sequence able to specify nuclear location. Cell 39: 499-509.
Kataoka C, Kaname Y, Taguwa S, Abe T, Fukuhara T, Tani H, Moriishi K, Matsuura Y. 2012. Baculovirus GP64-mediated entry into mammalian cells. J Virol 86: 2610-2620.
Kawaguchi T, Ohkawa K, Imanaka K, Tamai C, Kawada N, Ikezawa K, Uehara H, Itou Y, Nakanishi K, Katayama K. 2012. Lipiodol accumulation and transarterial chemoembolization efficacy for HCC patients. Hepatogastroenterology 59: 219-223.
Kost TA, Condreay JP. 2002. Recombinant baculoviruses as mammalian cell gene-delivery vectors. Trends Biotechnol 20: 173-180.
Kren BT, Unger GM, Sjeklocha L, Trossen AA, Korman V, Diethelm-Okita BM, Reding MT, Steer CJ. 2009. Nanocapsule-delivered Sleeping Beauty mediates therapeutic Factor VIII expression in liver sinusoidal endothelial cells of hemophilia A mice. J Clin Invest 119: 2086-2099.
Lanford RE, Hildebrandt-Eriksen ES, Petri A, Persson R, Lindow M, Munk ME, Kauppinen S, Orum H. 2010. Therapeutic silencing of microRNA-122 in primates with chronic hepatitis C virus infection. Science 327: 198-201.
Larson MH, Gilbert LA, Wang X, Lim WA, Weissman JS, Qi LS. 2013. CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nat Protoc 8: 2180-2196.
Larson MH, Gilbert LA, Wang XW, Lim WA, Weissman JS, Qi LS. 2013. CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nature Protocols 8: 2180-2196.
Li YP, Gottwein JM, Scheel TK, Jensen TB, Bukh J. 2011. MicroRNA-122 antagonism against hepatitis C virus genotypes 1-6 and reduced efficacy by host RNA insertion or mutations in the HCV 5' UTR. Proc Natl Acad Sci U S A 108: 4991-4996.
Liao YH, Chang YH, Sung LY, Li KC, Yeh CL, Yen TC, Hwang SM, Lin KJ, Hu YC. 2014. Osteogenic differentiation of adipose-derived stem cells and calvarial defect repair using baculovirus-mediated co-expression of BMP-2 and miR-148b. Biomaterials 35: 4901-4910.
Lin J, Teo S, Lam DH, Jeyaseelan K, Wang S. 2012. MicroRNA-10b pleiotropically regulates invasion, angiogenicity and apoptosis of tumor cells resembling mesenchymal subtype of glioblastoma multiforme. Cell Death Dis 3: e398.
Lindow M, Kauppinen S. 2012. Discovering the first microRNA-targeted drug. J Cell Biol 199: 407-412.
Liu W, Liu C, Zhu J, Shu P, Yin B, Gong Y, Qiang B, Yuan J, Peng X. 2012. MicroRNA-16 targets amyloid precursor protein to potentially modulate Alzheimer's-associated pathogenesis in SAMP8 mice. Neurobiol Aging 33: 522-534.
Lopez-Fraga M, Martinez T, Jimenez A. 2009. RNA interference technologies and therapeutics: from basic research to products. BioDrugs 23: 305-332.
Luo WY, Lin SY, Lo KW, Lu CH, Hung CL, Chen CY, Chang CC, Hu YC. 2013. Adaptive immune responses elicited by baculovirus and impacts on subsequent transgene expression in vivo. J Virol 87: 4965-4973.
Luo WY, Shih YS, Lo WH, Chen HR, Wang SC, Wang CH, Chien CH, Chiang CS, Chuang YJ, Hu YC. 2011. Baculovirus vectors for antiangiogenesis-based cancer gene therapy. Cancer Gene Ther 18: 637-645.
Luo Y, Xu X, An X, Sun X, Wang S, Zhu D. 2016. Targeted Inhibition of the miR-199a/214 Cluster by CRISPR Interference Augments the Tumor Tropism of Human Induced Pluripotent Stem Cell-Derived Neural Stem Cells under Hypoxic Condition. Stem Cells Int 2016: 3598542.
Mali P, Esvelt KM, Church GM. 2013. Cas9 as a versatile tool for engineering biology. Nat Methods 10: 957-963.
Mali P, Yang L, Esvelt KM, Aach J, Guell M, DiCarlo JE, Norville JE, Church GM. 2013. RNA-guided human genome engineering via Cas9. Science 339: 823-826.
Marquardt JU, Galle PR, Teufel A. 2012. Molecular diagnosis and therapy of hepatocellular carcinoma (HCC): an emerging field for advanced technologies. J Hepatol 56: 267-275.
Meding S, Nitsche U, Balluff B, Elsner M, Rauser S, Schone C, Nipp M, Maak M, Feith M, Ebert MP, Friess H, Langer R, Hofler H, Zitzelsberger H, Rosenberg R, Walch A. 2012. Tumor classification of six common cancer types based on proteomic profiling by MALDI imaging. J Proteome Res 11: 1996-2003.
Moldt B, Miskey C, Staunstrup NH, Gogol-Doring A, Bak RO, Sharma N, Mates L, Izsvak Z, Chen W, Ivics Z, Mikkelsen JG. 2011. Comparative genomic integration profiling of Sleeping Beauty transposons mobilized with high efficacy from integrase-defective lentiviral vectors in primary human cells. Mol Ther 19: 1499-1510.
Montini E, Held PK, Noll M, Morcinek N, Al-Dhalimy M, Finegold M, Yant SR, Kay MA, Grompe M. 2002. In vivo correction of murine tyrosinemia type I by DNA-mediated transposition. Mol Ther 6: 759-769.
Morgens DW, Deans RM, Li A, Bassik MC. 2016. Systematic comparison of CRISPR/Cas9 and RNAi screens for essential genes. Nat Biotechnol 34: 634-636.
O'Reilly M, Shipp A, Rosenthal E, Jambou R, Shih T, Montgomery M, Gargiulo L, Patterson A, Corrigan-Curay J. 2012. NIH oversight of human gene transfer research involving retroviral, lentiviral, and adeno-associated virus vectors and the role of the NIH recombinant DNA advisory committee. Methods Enzymol 507: 313-335.
Peck-Radosavljevic M. 2012. Back to basics: staging and prognosis in HCC for medical oncologist. J Hepatol 56: 488-489.
Pihlmann M, Askou AL, Aagaard L, Bruun GH, Svalgaard JD, Holm-Nielsen MH, Dagnaes-Hansen F, Bek T, Mikkelsen JG, Jensen TG, Corydon TJ. 2012. Adeno-associated virus-delivered polycistronic microRNA-clusters for knockdown of vascular endothelial growth factor in vivo. J Gene Med 14: 328-338.
Pineau P, Volinia S, McJunkin K, Marchio A, Battiston C, Terris B, Mazzaferro V, Lowe SW, Croce CM, Dejean A. 2010. miR-221 overexpression contributes to liver tumorigenesis. Proc Natl Acad Sci U S A 107: 264-269.
Qi LS, Larson MH, Gilbert LA, Doudna JA, Weissman JS, Arkin AP, Lim WA. 2013. Repurposing CRISPR as an RNA-guided platform for sequence-specific control of gene expression. Cell 152: 1173-1183.
Radzisheuskaya A, Shlyueva D, Muller I, Helin K. 2016. Optimizing sgRNA position markedly improves the efficiency of CRISPR/dCas9-mediated transcriptional repression. Nucleic Acids Res 44: e141.
Ramakrishnan V, Timm M, Haug JL, Kimlinger TK, Halling T, Wellik LE, Witzig TE, Rajkumar SV, Adjei AA, Kumar S. 2012. Sorafenib, a multikinase inhibitor, is effective in vitro against non-Hodgkin lymphoma and synergizes with the mTOR inhibitor rapamycin. Am J Hematol 87: 277-283.
Roggli E, Gattesco S, Caille D, Briet C, Boitard C, Meda P, Regazzi R. 2012. Changes in microRNA expression contribute to pancreatic beta-cell dysfunction in prediabetic NOD mice. Diabetes 61: 1742-1751.
Sakuma T, Nishikawa A, Kume S, Chayama K, Yamamoto T. 2014. Multiplex genome engineering in human cells using all-in-one CRISPR/Cas9 vector system. Sci Rep 4: 5400.
Scartozzi M, Faloppi L, Svegliati Baroni G, Loretelli C, Piscaglia F, Iavarone M, Toniutto P, Fava G, De Minicis S, Mandolesi A, Bianconi M, Giampieri R, Granito A, Facchetti F, Bitetto D, Marinelli S, Venerandi L, Vavassori S, Gemini S, D'Errico A, Colombo M, Bolondi L, Bearzi I, Benedetti A, Cascinu S. 2014. VEGF and VEGFR genotyping in the prediction of clinical outcome for HCC patients receiving sorafenib: The ALICE-1 study. Int J Cancer.
Schonrock N, Matamales M, Ittner LM, Gotz J. 2012. MicroRNA networks surrounding APP and amyloid-beta metabolism--implications for Alzheimer's disease. Exp Neurol 235: 447-454.
Senis E, Mockenhaupt S, Rupp D, Bauer T, Paramasivam N, Knapp B, Gronych J, Grosse S, Windisch MP, Schmidt F, Theis FJ, Eils R, Lichter P, Schlesner M, Bartenschlager R, Grimm D. 2016. TALEN/CRISPR-mediated engineering of a promoterless anti-viral RNAi hairpin into an endogenous miRNA locus. Nucleic Acids Res.
Singh H, Huls H, Kebriaei P, Cooper LJ. 2014. A new approach to gene therapy using Sleeping Beauty to genetically modify clinical-grade T cells to target CD19. Immunol Rev 257: 181-190.
Slaymaker IM, Gao L, Zetsche B, Scott DA, Yan WX, Zhang F. 2016. Rationally engineered Cas9 nucleases with improved specificity. Science 351: 84-88.
Song CW, Lee J, Lee SY. 2015. Genome engineering and gene expression control for bacterial strain development. Biotechnol J 10: 56-68.
Sterling RK, Wright EC, Morgan TR, Seeff LB, Hoefs JC, Di Bisceglie AM, Dienstag JL, Lok AS. 2012. Frequency of elevated hepatocellular carcinoma (HCC) biomarkers in patients with advanced hepatitis C. Am J Gastroenterol 107: 64-74.
Suzuki T, Chang MO, Kitajima M, Takaku H. 2010. Baculovirus activates murine dendritic cells and induces non-specific NK cell and T cell immune responses. Cell Immunol 262: 35-43.
Trabalza A, Georgiadis C, Eleftheriadou I, Hislop JN, Ellison SM, Karavassilis ME, Mazarakis ND. 2013. Venezuelan equine encephalitis virus glycoprotein pseudotyping confers neurotropism to lentiviral vectors. Gene Ther 20: 723-732.
Unniyampurath U, Pilankatta R, Krishnan MN. 2016. RNA Interference in the Age of CRISPR: Will CRISPR Interfere with RNAi? Int J Mol Sci 17: 291.
van Rooij E, Marshall WS, Olson EN. 2008. Toward microRNA-based therapeutics for heart disease: the sense in antisense. Circ Res 103: 919-928.
van Rooij E, Purcell AL, Levin AA. 2012. Developing microRNA therapeutics. Circ Res 110: 496-507.
Villanueva A, Llovet JM. 2011. Targeted therapies for hepatocellular carcinoma. Gastroenterology 140: 1410-1426.
Vink CA, Gaspar HB, Gabriel R, Schmidt M, McIvor RS, Thrasher AJ, Qasim W. 2009. Sleeping beauty transposition from nonintegrating lentivirus. Mol Ther 17: 1197-1204.
Wei Z, Doria C, Liu Y. 2013. Targeted therapies in the treatment of advanced hepatocellular carcinoma. Clin Med Insights Oncol 7: 87-102.
Woltjen K, Michael IP, Mohseni P, Desai R, Mileikovsky M, Hamalainen R, Cowling R, Wang W, Liu P, Gertsenstein M, Kaji K, Sung HK, Nagy A. 2009. piggyBac transposition reprograms fibroblasts to induced pluripotent stem cells. Nature 458: 766-770.
Woo HY, Heo J. 2012. Sorafenib in liver cancer. Expert Opin Pharmacother 13: 1059-1067.
Wu C, Lin J, Hong M, Choudhury Y, Balani P, Leung D, Dang LH, Zhao Y, Zeng J, Wang S. 2009. Combinatorial control of suicide gene expression by tissue-specific promoter and microRNA regulation for cancer therapy. Mol Ther 17: 2058-2066.
Xia H, Ooi LL, Hui KM. 2013. MicroRNA-216a/217-induced epithelial-mesenchymal transition targets PTEN and SMAD7 to promote drug resistance and recurrence of liver cancer. Hepatology 58: 629-641.
Xu H, Xiao T, Chen CH, Li W, Meyer CA, Wu Q, Wu D, Cong L, Zhang F, Liu JS, Brown M, Liu XS. 2015. Sequence determinants of improved CRISPR sgRNA design. Genome Res 25: 1147-1157.
Yang MH, Chen CL, Chau GY, Chiou SH, Su CW, Chou TY, Peng WL, Wu JC. 2009. Comprehensive analysis of the independent effect of twist and snail in promoting metastasis of hepatocellular carcinoma. Hepatology 50: 1464-1474.
Yant SR, Meuse L, Chiu W, Ivics Z, Izsvak Z, Kay MA. 2000. Somatic integration and long-term transgene expression in normal and haemophilic mice using a DNA transposon system. Nat Genet 25: 35-41.
Yin Y, Cai X, Chen X, Liang H, Zhang Y, Li J, Wang Z, Chen X, Zhang W, Yokoyama S, Wang C, Li L, Li L, Hou D, Dong L, Xu T, Hiroi T, Yang F, Ji H, Zhang J, Zen K, Zhang CY. 2014. Tumor-secreted miR-214 induces regulatory T cells: a major link between immune evasion and tumor growth. Cell Res 24: 1164-1180.
Zhang W, Solanki M, Muther N, Ebel M, Wang J, Sun C, Izsvak Z, Ehrhardt A. 2013. Hybrid adeno-associated viral vectors utilizing transposase-mediated somatic integration for stable transgene expression in human cells. PLoS One 8: e76771.
Zhao Y, Dai Z, Liang Y, Yin M, Ma K, He M, Ouyang H, Teng CB. 2014. Sequence-specific inhibition of microRNA via CRISPR/CRISPRi system. Sci Rep 4: 3943.
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