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1. 发明公开

US20230295627A1 Novel Replicase Cycling Reaction (RCR) and the Related SamRNA Designs Thereof 审中-公开
公开(公告)号：US20230295627A1
公开(公告)日：2023-09-21
申请号：US18156231
申请日：2023-01-18
申请人： Shi-Lung Lin , Sam Lin , Chun-Hung Lin
发明人： Shi-Lung Lin , Sam Lin , Chun-Hung Lin
IPC分类号： C12N15/113 , C12N9/12 , C12Q1/686
CPC分类号： C12N15/113 , C12N9/127 , C12Q1/686 , C12N2310/141 , C12Y207/07048
摘要： This invention generally relates to a novel composition of RNA/mRNA medicines as well as vaccines produced by using replicase- and/or RNA-dependent RNA polymerase (RdRp)-mediated RNA cycling reaction (RCR). The present invention is useful for developing a variety of self-amplifying RNA/mRNA (samRNA) medicines and vaccines containing at least a replicase/RdRp-binding site in the 5′- or 3′-end, or both, of any desired RNA molecule, including but not limited to antisense RNA (aRNA), small interferring RNA (siRNA), short hairpin RNA (shRNA), microRNA (miRNA)/miRNA precursor, long non-coding RNA (lnRNA) and mRNA. These RNA molecules can be either in single-stranded or in double-stranded, or mixed, conformation. The samRNA so obtained is useful not only for producing RNA-based vaccines and/or medicines but also for generating the mRNA-associated proteins, peptides, and/or antibodies under a proper in-vitro or in-cell translation condition. The replicase/RdRp-binding sites used in samRNA are derived or modified from coronaviral (e.g. COVID-19) and/or hepatitis C viral (HCV) RNA-dependent RNA polymerases (RdRp) in either single-stranded or double-stranded compositions.

2. 发明授权

US09453219B2 Cosmetic designs and products using intronic RNA 有权
标题翻译：使用内含子RNA的化妆品设计和产品
公开(公告)号：US09453219B2
公开(公告)日：2016-09-27
申请号：US12003662
申请日：2007-12-28
申请人： Shi-Lung Lin , David TS Wu
发明人： Shi-Lung Lin , David TS Wu
IPC分类号： C12N15/113 , C12N15/11
CPC分类号： C12N15/111 , C12N15/113 , C12N2310/111 , C12N2310/141 , C12N2320/30 , C12N2320/50
摘要： The present invention relates to a method and composition for generating a non-naturally occurring intron and its components capable of being processed into small hairpin RNA (shRNA) and/or microRNA (miRNA) molecules by skin cells and thus inducing specific gene silencing effects on skin pigment-related genes and/or aging-causing genes in the cells. The gene silencing effects so obtained are not only useful for lightening and whitening skin colors but also useful for suppressing unwanted aging gene activities in skins.
摘要翻译：本发明涉及一种用于产生非天然存在的内含子及其组分的方法和组合物，其能够被皮肤细胞加工成小发夹RNA（shRNA）和/或微小RNA（miRNA）分子，从而诱导特定的基因沉默效应皮肤色素相关基因和/或衰老基因。如此获得的基因沉默效应不仅可用于减轻和增白皮肤颜色，还可用于抑制皮肤中不需要的老化基因活性。

3. 发明授权

US09394538B2 Development of universal cancer drugs and vaccines 有权
标题翻译：开发普遍的癌症药物和疫苗
公开(公告)号：US09394538B2
公开(公告)日：2016-07-19
申请号：US12792413
申请日：2010-06-02
申请人： Shi-Lung Lin , David T S Wu
发明人： Shi-Lung Lin , David T S Wu
IPC分类号： C12N15/11 , C12Q1/68 , C12N5/02 , C07H21/04 , C12N15/113 , C12N15/63
CPC分类号： C12N15/113 , C12N15/63 , C12N15/635 , C12N2310/141 , C12N2330/51
摘要： This invention generally relates to a design and method for developing novel anti-tumor/cancer drugs, vaccines and therapies, using microRNA (miRNA) and its shRNA homologues/derivatives. More particularly, the present invention relates to the use of a nucleic acid composition capable of expressing mir-302-like gene silencing effectors upon delivery into human cells and then silencing mir-302-targeted cell cycle regulators and oncogenes, resulting in an inhibitory effect on tumor/cancer cell growth and metastasis. Mir-302 is the most predominant miRNA found in human embryonic stem (hES) and induced pluripotent stem (iPS) cells, yet its function is unclear. The present invention establishes that in humans mir-302 concurrently suppressed both cyclin-E-CDK2 and cyclin-D-CDK4/6 pathways and eventually blocked over 70% of the G1-S transition. Simultaneously, mir-302 also silences BMI-1, a cancer stem cell marker, and subsequently promotes the tumor suppressor functions of p16Ink4a and p14/p19Arf in inhibiting CDK4/6-mediated cell proliferation. Therefore, the present invention for the first time reveals the tumor suppressor function of mir-302 in humans. This novel finding advances the design and method for developing new cancer drugs, vaccines and therapies directed against multiple kinds of human tumors and cancers, in particular including, but not limited, malignant skin, prostate, breast and liver cancers as well as various tumors.
摘要翻译：本发明一般涉及使用微小RNA（miRNA）及其shRNA同源物/衍生物开发新的抗肿瘤/癌症药物，疫苗和疗法的设计和方法。更具体地说，本发明涉及在递送至人细胞后能够表达mir-302样基因沉默效应物，然后使mir-302靶向的细胞周期调节因子和致癌基因沉默的核酸组合物的用途，从而产生抑制作用对肿瘤/癌细胞的生长和转移。 Mir-302是人类胚胎干细胞（hES）和诱导多能干（iPS）细胞中最主要的miRNA，但其功能尚不清楚。本发明确定人mir-302同时抑制细胞周期蛋白E-CDK2和细胞周期蛋白-D-CDK4 / 6途径，并最终阻断70％以上的G1-S转换。同时，mir-302还沉默BMI-1，一种癌症干细胞标记物，随后促进p16Ink4a和p14 / p19Arf抑制CDK4 / 6介导的细胞增殖的肿瘤抑制功能。因此，本发明首次揭示了人类mir-302的肿瘤抑制功能。该新发现推进了针对多种人类肿瘤和癌症开发新的癌症药物，疫苗和疗法的设计和方法，特别是包括但不限于恶性皮肤，前列腺癌，乳腺癌和肝癌以及各种肿瘤。

4. 发明申请

US20150209377A1 Use of Novel Monosaccharide-like Glycylated Sugar Alcohol Compositions for Designing and Developing Anti-Diabetic Drugs 有权
标题翻译：使用新型单糖样糖基化糖醇组合物设计和开发抗糖尿病药物
公开(公告)号：US20150209377A1
公开(公告)日：2015-07-30
申请号：US14585978
申请日：2014-12-30
申请人： Shi-Lung Lin , Samantha Chang-Lin , Yi-Wen Lin , Donald Chang
发明人： Shi-Lung Lin , Samantha Chang-Lin , Yi-Wen Lin , Donald Chang
IPC分类号： A61K31/7012 , A61K31/221
CPC分类号： A61K31/221 , A61K9/0053 , A61K9/08 , A61K31/7012 , A61K31/7024 , C12N15/111 , C12N15/113 , C12N2310/141 , C12N2320/32 , C12N2330/50 , C12N2330/51
摘要： This invention is related to a novel sugar-like chemical composition and its use for diabetes therapy. Particularly, the present invention teaches the use of monosaccharide-like glycylated sugar alcohol compounds to block or reduce sugar absorption in diabetes patients, so as to prevent the risk of hyperglycemia symptoms. Glycylation of sugar alcohols is a totally novel reaction that has never been reported before. Therefore, the novelty of the present invention is that for the first time glycylated sugar alcohols not only was found but also was found to be useful for treating Diabetes mellitus. In addition, the present invention teaches a method for producing these glycylated sugar alcohols. In sum, the present invention includes not only a kind of novel sugar-like chemical compositions and its use for treating diabetes but also a state-of-the-art protocol and methodology for producing such a novel composition via glycylation of sugars and sugar alcohols.
摘要翻译：本发明涉及新型糖类化学组合物及其用于糖尿病治疗的用途。特别地，本发明教导了使用单糖样的糖化糖醇化合物来阻止或减少糖尿病患者的糖吸收，以防止高血糖症状的风险。糖醇的糖基化是一种完全新颖的反应，从未在之前报道过。因此，本发明的新颖性不仅首次发现了糖化糖醇，而且被发现可用于治疗糖尿病。此外，本发明教导了一种制备这些糖基化糖醇的方法。总之，本发明不仅包括一种新型的糖类化学组合物及其用于治疗糖尿病的用途，而且还包括通过糖和糖醇的糖基化制备这种新型组合物的最新方案和方法。

5. 发明授权

US08895525B2 Preventing hyaluronan-mediated tumorigenetic mechanisms using intronic RNAs 有权
标题翻译：使用内含子RNA预防透明质酸介导的肿瘤发生机制
公开(公告)号：US08895525B2
公开(公告)日：2014-11-25
申请号：US12740334
申请日：2008-10-29
申请人： Shao-Yao Ying , Shi-Lung Lin
发明人： Shao-Yao Ying , Shi-Lung Lin
IPC分类号： A61K48/00 , C07H21/02 , C07H21/04
CPC分类号： C12N15/1135 , C12N2310/141 , C12Q1/6886 , C12Q2600/112 , C12Q2600/178
摘要： Patterns of microRNA (miRNA) expression are correlated to the degrees of tumor cell differentiation in human prostate cancer. MiRNAs can complementarily bind to either oncogenes or tumor suppressor genes, resulting in targeted gene silencing and thus changes of cellular tumorigenecity. Using miRNA microarray analysis, 8 down-regulated and 3 up-regulated known miRNAs in androgen-independent human prostate cancer cell lines, such as LNCaP C4-2B and PC3, compared to those androgen-dependent cell lines, such as LNCaP and PC3-AR9 were consistently detected. Fluorescent in-situ hybridization assays in human prostate cancer tissue arrays containing sixty patients at different stages also showed the same miRNA expression patterns in hormone-refractory prostate carcinomas (HRPC) compared to androgen-sensitive non-cancerous prostate epithelium. In-vitro tumorigenecity assays using one of the identified miRNAs, mir-146a, were performed to provide validation of its function in prostate cancer. Gain-of-function transfection of mir-146a markedly suppressed its targeted ROCK1 gene expression in androgen-independent PC3 cells, consequently resulting in reduced cancer cell proliferation, invasion and metastasis to human bone marrow endothelial cell monolayers. Since ROCK1 is the key kinase for activating hyaluronan-mediated HRPC transformation in vivo and in PC3 cells, mir-146a should function as a tumor-suppressor gene in modulating the ROCK1-associated tumorigenecity.
摘要翻译： microRNA（miRNA）表达的模式与人类前列腺癌的肿瘤细胞分化程度相关。 MiRNA可以互补结合癌基因或肿瘤抑制基因，导致靶向基因沉默，从而导致细胞致瘤性的变化。与雄激素依赖性细胞系（如LNCaP和PC3）相比，使用miRNA微阵列分析，8个下调和3个上调已知的与雄激素非依赖性前列腺癌细胞系（如LNCaP C4-2B和PC3）的miRNA相比，一直检测到AR9。与雄激素敏感的非癌前列腺上皮相比，含有60位不同阶段患者的人类前列腺癌组织阵列中的荧光原位杂交测定也显示与激素难治性前列腺癌（HRPC）相同的miRNA表达模式。使用鉴定的miRNA之一的mir-146a进行体外肿瘤发生测定，以提供其在前列腺癌中的功能的验证。 mir-146a功能的功能转染显着抑制雄激素依赖性PC3细胞中靶向的ROCK1基因表达，从而导致癌细胞增殖，侵袭和转移到人类骨髓内皮细胞单层。由于ROCK1是在体内和PC3细胞中激活透明质酸介导的HRPC转化的关键激酶，因此mir-146a应该作为调节ROCK1相关致瘤性的肿瘤抑制基因起作用。

6. 发明授权

US6130040A Method for generating directly covalent bonding between interstrand nucleotides 失效
标题翻译：在星际核苷酸之间直接共价连接的方法
公开(公告)号：US6130040A
公开(公告)日：2000-10-10
申请号：US4070
申请日：1998-01-08
申请人： Shi-Lung Lin
发明人： Shi-Lung Lin
IPC分类号： C07H21/00 , C12Q1/68 , C07H19/00 , C12P19/34
CPC分类号： C07H21/00
摘要： The present invention provides a fast, simple and direct covalent bond formation between two strands of nucleotide sequences. Non-modified first strand nucleotide sequences are hybridized with second strand nucleotide sequences, of which certain specific base structure(s) is modified by chemical reagents in order to generate covalent bonding with the first strand. While the hybridization of these two strand nucleotide sequences generates double-stranded hybrid duplexes between their homologues, covalent bond formation occurs in the region of modified base-pairs. Since neither a polymerase chain restriction nor a restriction enzyme digestion can be performed with the covalently bonded hybrid duplexes, the present invention can be used to subtract common sequences during subtractive hybridization, to inhibit nonspecific contamination during subcloning and to increase binding stability of antisense probes during in situ hybridization as well as gene therapy.
摘要翻译：本发明提供了两条核苷酸序列之间的快速，简单和直接的共价键形成。未修饰的第一链核苷酸序列与第二链核苷酸序列杂交，其中某些特异性碱基结构被化学试剂修饰以产生与第一链的共价键。虽然这两条链核苷酸序列的杂交在其同源物之间产生双链杂交双链体，但共价键形成发生在修饰的碱基对的区域。由于不能用共价键合的杂交双链体进行聚合酶链限制和限制性内切酶消化，本发明可用于在减法杂交期间减去常见序列，以抑制亚克隆过程中的非特异性污染，并增加反义探针的结合稳定性原位杂交以及基因治疗。

7. 发明授权

US08372969B2 RNA interference methods using DNA-RNA duplex constructs 有权
标题翻译：使用DNA-RNA双链体构建的RNA干扰方法
公开(公告)号：US08372969B2
公开(公告)日：2013-02-12
申请号：US12911654
申请日：2010-10-25
申请人： Shao-Yao Ying , Shi-Lung Lin
发明人： Shao-Yao Ying , Shi-Lung Lin
IPC分类号： C07H21/04
CPC分类号： C12N15/1096
摘要： The present invention provides novel compositions and methods for suppressing the function or activity of a targeted gene through a novel intracellular piRNA-mediated RNAi mechanism, using RNA-DNA duplex constructs. The invention further provides novel methods and compositions for generating or producing RNA-DNA duplex agents, whose quantity is high enough to be used for the invention's gene silencing transfection and possibly in therapeutics applications. This improved RNA-polymerase chain reaction (RNA-PCR) method utilizes thermocycling steps of promoter-linked DNA or RNA template synthesis, in vitro transcription and then reverse transcription to bring up the amount of RNA-DNA duplexes up to two thousand folds within one round of the above procedure for using in D-RNAi-directed gene silencing.
摘要翻译：本发明提供了使用RNA-DNA双链构建体通过新的细胞内piRNA介导的RNAi机制抑制靶基因的功能或活性的新型组合物和方法。本发明还提供用于产生或产生RNA-DNA双链体的新方法和组合物，其量足够高以用于本发明的基因沉默转染并且可能在治疗应用中。这种改进的RNA聚合酶链反应（RNA-PCR）方法利用启动子连接的DNA或RNA模板合成的热循环步骤，体外转录，然后逆转录，将RNA-DNA双链体的量在一个以上用于D-RNAi定向基因沉默的方法。

8. 发明申请

US20100249212A1 Gene Silencing Using mRNA-cDNA Hybrids 有权
标题翻译：基因沉默使用mRNA-cDNA杂交
公开(公告)号：US20100249212A1
公开(公告)日：2010-09-30
申请号：US12629845
申请日：2009-12-02
申请人： Shi-Lung Lin , Cheng-Ming Chuong , Randall B. Widelitz
发明人： Shi-Lung Lin , Cheng-Ming Chuong , Randall B. Widelitz
IPC分类号： A61K31/7088 , C07H21/04 , A61P31/12
CPC分类号： C12N15/1096
摘要： The present invention provides novel compositions and methods for suppressing the expression of a targeted gene using mRNA-cDNA duplexes. The invention further provides novel methods and compositions for generating amplified mRNA-cDNA hybrids, whose quantity is high enough to be used for the invention's gene silencing transfection. This improved RNA-polymerase chain reaction method uses thermocycling steps of promoter-linked double-stranded cDNA or RNA synthesis, in vitro transcription and then reverse transcription to amplify the amount of mRNA-cDNA hybrids up to two thousand folds within one round of the above procedure.
摘要翻译：本发明提供使用mRNA-cDNA双链体抑制靶基因表达的新型组合物和方法。本发明还提供了用于产生扩增的mRNA-cDNA杂交体的新方法和组合物，其量足够高以用于本发明的基因沉默转染。这种改进的RNA聚合酶链反应方法使用启动子连接的双链cDNA或RNA合成的热循环步骤，体外转录，然后逆转录以在上述一轮之内扩增两千倍的mRNA-cDNA杂交体的量程序。

9. 发明授权

US07662791B2 Gene silencing using mRNA-cDNA hybrids 失效
标题翻译：基因沉默使用mRNA-cDNA杂交
公开(公告)号：US07662791B2
公开(公告)日：2010-02-16
申请号：US09920342
申请日：2001-08-01
申请人： Shi-Lung Lin , Cheng-Ming Chuong , Randall B. Widelitz
发明人： Shi-Lung Lin , Cheng-Ming Chuong , Randall B. Widelitz
IPC分类号： A61K48/00
CPC分类号： C12N15/1096
摘要： The present invention provides novel compositions and methods for suppressing the expression of a targeted gene using mRNA-cDNA duplexes. The invention further provides novel methods and compositions for generating amplified mRNA-cDNA hybrids, whose quantity is high enough to be used for the invention's gene silencing transfection. This improved RNA-polymerase chain reaction method uses thermocycling steps of promoter-linked double-stranded cDNA or RNA synthesis, in vitro transcription and then reverse transcription to amplify the amount of mRNA-cDNA hybrids up to two thousand folds within one round of the above procedure.
摘要翻译：本发明提供使用mRNA-cDNA双链体抑制靶基因表达的新型组合物和方法。本发明还提供了用于产生扩增的mRNA-cDNA杂交体的新方法和组合物，其量足够高以用于本发明的基因沉默转染。这种改进的RNA聚合酶链反应方法使用启动子连接的双链cDNA或RNA合成的热循环步骤，体外转录，然后逆转录以在上述一轮之内扩增两千倍的mRNA-cDNA杂交体的量程序。

10. 发明授权

US5928872A Subtractive hybridization with covalently binding homology 失效
标题翻译：具有共价结合同源性的消减杂交
公开(公告)号：US5928872A
公开(公告)日：1999-07-27
申请号：US927859
申请日：1997-09-11
申请人： Shi-Lung Lin , Shao-Yao Ying
发明人： Shi-Lung Lin , Shao-Yao Ying
IPC分类号： C12N15/10 , C12Q1/68 , C12P19/34
CPC分类号： C12N15/1034 , C12Q1/6809
摘要： Excess amount of modified subtracter DNA from control cells is generated by carboxylating the base structures of its certain nucleotides with chemical agents in order to introduce covalent affinity between the modified subtracter and a non-modified tester DNA. Hybridization of the control subtracter and the experimental tester DNA is performed with a heat-melting and then cool-reassociation technique. While the desired different (heterologous) sequences remain in the form of hydrogen-binding, common (homologous) sequences of the hybridized DNA are covalently bonded to each other. Since the covalent bonding of the common sequences can not be broken during a polymerase chain reaction, resulting in no amplification of the common sequences but great amplification of the desired different sequences. The desired DNA sequences present after such covalent homologue subtraction and selective amplification represent those DNA sequences which only exist in the tester but not in the subtracter DNA library.
摘要翻译：通过用化学试剂羧化其某些核苷酸的碱基结构来产生来自对照细胞的过量的修饰的减毒DNA，以便在修饰的减毒剂和未修饰的测试DNA之间引入共价亲和力。控制减法器和实验检测器DNA的杂交用热熔化然后冷再结合技术进行。尽管期望的不同（异源）序列以氢结合的形式保留，杂交DNA的共同（同源）序列彼此共价键合。由于在聚合酶链反应期间共同序列的共价键不能被破坏，所以不能扩增常见序列，而是扩增所需的不同序列。在这种共价同系物减法和选择性扩增之后存在的所需DNA序列代表仅存在于测试者中但不在减毒菌DNA文库中的那些DNA序列。

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