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1. 发明申请

US20090023186A1 USE OF VALPROIC ACID FOR ENHANCING PRODUCTION OF RECOMBINANT PROTEINS IN MAMMALIAN CELLS 审中-公开
标题翻译：用于增强在马马里亚细胞中重组蛋白生产的有效性
公开(公告)号：US20090023186A1
公开(公告)日：2009-01-22
申请号：US11781252
申请日：2007-07-22
申请人： Markus Hildinger , Gaurav Backliwal , Florian Wurm
发明人： Markus Hildinger , Gaurav Backliwal , Florian Wurm
IPC分类号： C12P21/00
CPC分类号： C12P21/02 , C07K16/00
摘要： Culturing cells for the commercial production of proteins for diagnosis and therapy is a costly and time consuming process. The equipment required is expensive, and production cost are high. In order to provide commercially viable processes it is desirable to use cell lines which produce large quantities of product with each production run. However, most cells do not produce large quantities of desired product per se either because they do not produce a large quantity of product per unit of time (specific productivity) or because they do not survive long enough in the culture medium (time). Here, we identified that addition of a valproic acid compound to the culture medium increases overall (batch) yield and titer. More importantly, compared to the widely used sodium butyrate, batch yields using a valproic acid compound as a medium additive are significantly higher.
摘要翻译：用于商业生产蛋白质用于诊断和治疗的培养细胞是昂贵且耗时的过程。所需设备价格昂贵，生产成本高。为了提供商业可行的方法，期望使用在每次生产运行时产生大量产品的细胞系。然而，大多数细胞本身并不产生大量的所需产品，因为它们每单位时间（比生产率）不产生大量的产品，或者因为它们在培养基（时间）内不能足够长的时间生存。在这里，我们确定向培养基中加入丙戊酸化合物提高了总体（批）产率和滴度。更重要的是，与广泛使用的丁酸钠相比，使用丙戊酸化合物作为中等添加剂的批产量显着更高。

2. 发明申请

US20080145893A1 Method for producing a recombinant protein at high specific productivity, high batch yield and high volumetric yield by means of transient transfection 审中-公开
标题翻译：通过瞬时转染以高比生产率，高批量产量和高体积产量生产重组蛋白的方法
公开(公告)号：US20080145893A1
公开(公告)日：2008-06-19
申请号：US11309732
申请日：2006-09-17
申请人： Markus Hildinger , Sarah Wulhfard , Gaurav Backliwal , David Hacker , Maria De Jesus , Florian Maria Wurm
发明人： Markus Hildinger , Sarah Wulhfard , Gaurav Backliwal , David Hacker , Maria De Jesus , Florian Maria Wurm
IPC分类号： C12P21/06 , C12N15/09 , C12N5/06 , C07K14/50
CPC分类号： C12N15/85 , C12N15/67 , C12N2500/36 , C12N2500/40 , C12N2500/90 , C12N2500/92 , C12N2501/113 , C12N2510/02
摘要： Recombinant proteins are of great commercial interest. Yet, most current production methods in mammalian cells involve the time- and labor-consuming step of creating stable cell lines. Production methods based on transient gene expression are advantageous in terms of speed and versatility, yet, thus far, those methods have not shown the specific productivity, batch yield and volumetric yield to be an economic alternative to stable cell lines. The inventors improved on the methodology of transient transfection and achieved commercially relevant yields in terms of specific productivity (exceeding 35 pg per cell per day), batch yield (exceeding 700 mg/l) and volumetric yield.
摘要翻译：重组蛋白具有很大的商业利益。然而，哺乳动物细胞中目前的大多数生产方法涉及创建稳定细胞系的耗时和耗时的步骤。基于瞬时基因表达的生产方法在速度和多功能性方面是有利的，但是到目前为止，这些方法尚未显示比生产力，分批产量和体积产量作为稳定细胞系的经济替代物。本发明人改进了瞬时转染的方法，并且在比生产率（每个细胞每天超过35pg），批产量（超过700mg / l）和体积产量方面获得了商业上相关的产量。

3. 发明申请

US20050019927A1 DECREASING GENE EXPRESSION IN A MAMMALIAN SUBJECT IN VIVO VIA AAV-MEDIATED RNAi EXPRESSION CASSETTE TRANSFER 审中-公开
标题翻译：通过AAV介导的RNAi表达载体转移，在VIVO中降低MAMMALIAN受体中的基因表达
公开(公告)号：US20050019927A1
公开(公告)日：2005-01-27
申请号：US10604340
申请日：2003-07-13
申请人： Markus Hildinger , Alberto Auricchio
发明人： Markus Hildinger , Alberto Auricchio
IPC分类号： A61K38/00 , A61K48/00 , C12N5/02 , C12N15/113 , C12N15/864 , C12N15/86
CPC分类号： C12N15/113 , A61K38/00 , A61K48/00 , C12N15/86 , C12N2310/111 , C12N2310/14 , C12N2750/14143 , C12N2840/20
摘要： Decreasing the expression of genes in a mammalian subject has multiple applications ranging from cancer therapy to anti-infective therapy or treatment of autosomal dominant genetic disorders. Yet, there is still a lack of efficient technologies to achieve that goal in mammalian subjects in vivo. The present invention relates to methods for decreasing gene expression by administering to a mammalian subject a recombinant adeno-associated viral vector in vivo with said vector comprising an RNA interference (RNAi) expression cassette whose RNA expression products directly or indirectly lead to a decrease in expression of the corresponding RNAi target gene. Upon successful transduction with the recombinant adeno-associated viral vector, the RNA expression products of the RNAi expression cassette will decrease the cellular concentration of the mRNA transcripts of the RNAi target gene, thus resulting in decreased concentration of the protein encoded by the RNAi target gene.
摘要翻译：减少哺乳动物受试者中基因的表达具有多种应用，从癌症治疗到抗感染治疗或治疗常染色体显性遗传疾病。然而，在哺乳动物体内仍然缺乏有效的技术来实现这一目标。本发明涉及降低基因表达的方法，其通过向哺乳动物受试者施用重组腺相关病毒载体，所述载体包含RNA干扰（RNAi）表达盒，其RNA表达产物直接或间接导致表达降低的相应RNAi靶基因。在用重组腺相关病毒载体成功转导后，RNAi表达盒的RNA表达产物将降低RNAi靶基因的mRNA转录物的细胞浓度，从而导致由RNAi靶基因编码的蛋白质的浓度降低。

4. 发明授权

US07943374B2 Super-size adeno-associated viral vector harboring a recombinant genome larger than 5.7 kb 失效
标题翻译：超大尺寸腺相关病毒载体携带大于5.7kb的重组基因
公开(公告)号：US07943374B2
公开(公告)日：2011-05-17
申请号：US11161890
申请日：2005-08-21
申请人： Markus Hildinger
发明人： Markus Hildinger
IPC分类号： C12N15/63 , C12N15/85
CPC分类号： C12N15/86 , A61K48/00 , C12N2750/14143
摘要： Prior art teaches an effective packaging capacity for adeno-associated virus and adeno-associated viral vectors of 4.1 kb to 4.9 kb as well as a packaging limit of 5.2 kb to 5.6 kb. However, the inventor discovered that this packaging limit as well as that effective packaging capacity does not apply to all AAV serotypes: Whereas it is true that efficient packaging of AAV serotype 2 is limited to less than 5 kb, the inventor discovered that one can efficiently package more than 6 kb of genetic information into AAV capsids of other AAV serotypes, particularly into capsids of AAV serotype 5 and—to a lesser extent—into capsids of AAV serotype 7. This discovery will be useful in the context of gene therapy where large transgenes will have to be transferred such as the ABCA4 coding sequence, the Factor VIII coding sequence, the B-deleted Factor VIII coding sequence or minidystrophin genes.
摘要翻译：现有技术教导了4.1kb至4.9kb的腺相关病毒和腺相关病毒载体的有效包装能力以及5.2kb至5.6kb的包装限制。然而，发明人发现，这种包装限制以及有效的包装能力并不适用于所有AAV血清型：鉴于AAV血清型2的有效包装确实被限制在小于5kb，发明人发现可以有效地将超过6kb的遗传信息包装到其他AAV血清型的AAV衣壳中，特别是AAV血清型5的衣壳中，并且在较小程度上包装到AAV血清型7的衣壳中。该发现将在基因治疗的背景下有用，其中大的必须转移转基因，如ABCA4编码序列，因子VIII编码序列，缺失B因子VIII编码序列或微小营养不良蛋白基因。

5. 发明申请

US20100311116A1 FAST GENERATION OF HIGH EXPRESSION STABLE CELL LINES EXPRESSING RECOMBINANT PROTEINS UNDER MINIMAL AND SHORT-TERM SELECTIVE PRESSURE 审中-公开
标题翻译：在最小和短期选择压力下快速表达表达重组蛋白的高表达稳定细胞系
公开(公告)号：US20100311116A1
公开(公告)日：2010-12-09
申请号：US12478558
申请日：2009-06-04
申请人： Florian M. Wurm , Markus Hildinger , Maria De Jesus , Mattia Matasci , David Hacker
发明人： Florian M. Wurm , Markus Hildinger , Maria De Jesus , Mattia Matasci , David Hacker
IPC分类号： C12N15/87 , C40B50/06 , C12P21/00
CPC分类号： C12P21/02 , C12N9/22 , C12N15/85 , C12N2800/40 , C12N2800/90
摘要： The present invention provides a novel method for the fast generation of high expression stable cell lines for the production of recombinant proteins with high efficacy of stable integration while using low selective pressure for only a short period of time. The method uses transiently expressed piggybac transposase to mediate stable integration of a transgene of interest flanked by the PB transposon termini.
摘要翻译：本发明提供了一种用于快速产生高表达稳定细胞系的新方法，用于生产具有稳定整合的高效力的重组蛋白质，同时在短时间内使用低选择压力。该方法使用瞬时表达的piggybac转座酶介导侧翼为PB转座子末端的感兴趣的转基因的稳定整合。

6. 发明申请

US20060276391A1 Use of compounds that interfere with the Hedgehog signaling pathway for the manufacture of a medicament for preventing, inhibiting, and/or reversing ocular diseases related with ocular neovascularization 失效
标题翻译：使用干扰Hedgehog信号传导途径的化合物来制造用于预防，抑制和/或逆转与眼睛新生血管形成相关的眼部疾病的药物
公开(公告)号：US20060276391A1
公开(公告)日：2006-12-07
申请号：US11002882
申请日：2004-12-03
申请人： Alberto Auricchio , Markus Hildinger , Enrico Surace
发明人： Alberto Auricchio , Markus Hildinger , Enrico Surace
IPC分类号： A61K38/17 , A61K48/00 , A61K31/58 , A61K31/56
CPC分类号： A61K38/1709 , A61K31/00 , A61K31/4355 , A61K31/445 , A61K31/56 , A61K31/58 , C07K16/18 , C12N15/113 , C12N15/86 , C12N2310/14 , C12N2750/14143
摘要： The present invention concerns the use of compounds that interfere with the hedgehog signaling pathway for the manufacture of a medicament for preventing, inhibiting, and/or reversing ocular diseases related with ocular neovascularization. Partcularly, the above-mentioned diseases are (wet) age-related macular degeneration, (proliferative) diabetic retinopathy, neovascular glaucoma, retinal vein occlusion, or retinopathy of prematurity (ROP).
摘要翻译：本发明涉及干扰刺猬蛋白信号传导途径的化合物在制备用于预防，抑制和/或逆转与眼睛新生血管形成相关的眼部疾病的药物中的用途。特别地，上述疾病是（湿性）年龄相关性黄斑变性，（增殖性）糖尿病性视网膜病变，新生血管性青光眼，视网膜静脉阻塞或早产儿视网膜病变（ROP）。

7. 发明授权

US06548301B2 Retroviral gene transfer vectors 有权
标题翻译：逆转录病毒基因转移载体
公开(公告)号：US06548301B2
公开(公告)日：2003-04-15
申请号：US09306417
申请日：1999-05-06
申请人： Wolfram Ostertag , Christopher Baum , Markus Hildinger
发明人： Wolfram Ostertag , Christopher Baum , Markus Hildinger
IPC分类号： C12N1585
CPC分类号： C12N15/86 , C12N2740/13043 , C12N2840/20 , C12N2840/44
摘要： The invention relates to novel retroviral gene transfer vectors, preferably expression vectors, which, because of a reduced content of viral genes, are distinguished by a higher safety standard and an expression of non-viral nucleotide sequences in higher amounts.
摘要翻译：本发明涉及新型的逆转录病毒基因转移载体，优选表达载体，其由于病毒基因的含量降低而被更高的安全性标准和更高量的非病毒核苷酸序列的表达区分。

8. 发明授权

US07517870B2 Use of compounds that interfere with the hedgehog signaling pathway for the manufacture of a medicament for preventing, inhibiting, and/or reversing ocular diseases related with ocular neovascularization 失效
标题翻译：使用干扰刺猬蛋白信号传导途径的化合物来制造用于预防，抑制和/或逆转与眼睛新生血管形成相关的眼部疾病的药物
公开(公告)号：US07517870B2
公开(公告)日：2009-04-14
申请号：US11002882
申请日：2004-12-03
申请人： Alberto Auricchio , Markus Hildinger , Enrico Maria Surace
发明人： Alberto Auricchio , Markus Hildinger , Enrico Maria Surace
IPC分类号： A61K38/00 , A61K38/17
CPC分类号： A61K38/1709 , A61K31/00 , A61K31/4355 , A61K31/445 , A61K31/56 , A61K31/58 , C07K16/18 , C12N15/113 , C12N15/86 , C12N2310/14 , C12N2750/14143
摘要： The present invention concerns the use of compounds that interfere with the hedgehog signaling pathway for the manufacture of a medicament for preventing, inhibiting, and/or reversing ocular diseases related with ocular neovascularization. Particularly, the above-mentioned diseases are (wet) age-related macular degeneration, (proliferative) diabetic retinopathy, neovascular glaucoma, retinal vein occlusion, or retinopathy of prematurity (ROP).
摘要翻译：本发明涉及干扰刺猬蛋白信号传导途径的化合物在制备用于预防，抑制和/或逆转与眼睛新生血管形成相关的眼部疾病的药物中的用途。特别地，上述疾病是（湿性）年龄相关性黄斑变性，（增殖性）糖尿病性视网膜病变，新生血管性青光眼，视网膜静脉阻塞或早产儿视网膜病变（ROP）。

9. 发明申请

US20090233334A1 CELL CULTIVATION AND PRODUCTION OF RECOMBINANT PROTEINS BY MEANS OF AN ORBITAL SHAKE BIOREACTOR SYSTEM WITH DISPOSABLE BAGS AT THE 1,500 LITER SCALE 审中-公开
标题翻译：细胞培养和生产重组蛋白质的方法是通过在1,500 LITER SCALE上的具有可拆卸袋的ORBITAL SHAKE生物反应器系统
公开(公告)号：US20090233334A1
公开(公告)日：2009-09-17
申请号：US12046374
申请日：2008-03-11
申请人： Markus Hildinger , Florian Wurm , Matthieu Stettler , Maria DeJesus
发明人： Markus Hildinger , Florian Wurm , Matthieu Stettler , Maria DeJesus
IPC分类号： C12P21/04 , C12N7/00 , C12N5/04 , C12N5/06 , C12N5/02 , C12N5/08 , C12N1/00
CPC分类号： C12P21/02 , C12M23/14 , C12M27/16 , C12M41/02
摘要： The present invention provides a novel method for culturing cells as well as a novel method for producing a recombinant protein by culturing cells at large scale (up to 1,500 L nominal volume and 750 L working volume), whereby an inflated bag provides a sterile, disposable cultivation chamber. The inflated bag is partially filled with liquid cultivation media and cells, and placed into a containment vessel. The containment vessel is positioned onto an orbitally shaken platform. The orbital shaking moves the containment vessel and thus the bag and induces thereby motion to the liquid contained therein (“shake mixing”). This motion (caused by orbital shaking) induces a dynamic force field that ensures cell suspension, bulk mixing, and oxygen transfer from the liquid surface to the respiring cells without damaging shear or foam generation.
摘要翻译：本发明提供了一种培养细胞的新方法以及通过大规模培养细胞（达1,500L标称体积和750L工作体积）来生产重组蛋白的新方法，由此充气袋提供无菌的，一次性的栽培室。充气袋部分地装有液体培养基和细胞，并放置在容纳容器中。安全壳位于轨道上摇摆的平台上。轨道振荡使得容纳容器和袋子移动并引起其运动到其中容纳的液体（“混合”）。这种运动（由轨道震荡引起）引起动态力场，确保细胞悬浮，本体混合和从液体表面到呼吸细胞的氧传递，而不损害剪切或泡沫产生。

10. 发明申请

US20090192115A1 USE OF COMPOUNDS THAT INTERFERE WITH THE HEDGEHOG SIGNALING PATHWAY FOR THE MANUFACTURE OF A MEDICAMENT FOR PREVENTING, INHIBITING, AND/OR REVERSING OCULAR DISEASES RELATED WITH OCULAR NEOVASCULARIZATION 审中-公开
标题翻译：使用与HEDGEHOG信号通道干扰用于预防，抑制和/或逆转与眼内新生血管相关的眼科疾病的药物的化合物
公开(公告)号：US20090192115A1
公开(公告)日：2009-07-30
申请号：US12348077
申请日：2009-01-02
申请人： Alberto Auricchio , Markus Hildinger , Enrico Maria Surace
发明人： Alberto Auricchio , Markus Hildinger , Enrico Maria Surace
IPC分类号： A61K31/711 , A61K31/7105
CPC分类号： A61K38/1709 , A61K31/00 , A61K31/4355 , A61K31/445 , A61K31/56 , A61K31/58 , C07K16/18 , C12N15/113 , C12N15/86 , C12N2310/14 , C12N2750/14143
摘要： The use of compounds that interfere with the hedgehog signaling pathway for the manufacture of a medicament for preventing, inhibiting, and/or reversing ocular diseases related with ocular neovascularization. Particularly, the above-mentioned diseases are (wet) age-related macular degeneration, (proliferative) diabetic retinopathy, neovascular glaucoma, retinal vein occlusion, or retinopathy of prematurity (ROP).
摘要翻译：使用干扰刺猬蛋白信号传导途径的化合物来制造用于预防，抑制和/或逆转与眼睛新生血管形成相关的眼部疾病的药物。特别地，上述疾病是（湿性）年龄相关性黄斑变性，（增殖性）糖尿病性视网膜病变，新生血管性青光眼，视网膜静脉阻塞或早产儿视网膜病变（ROP）。

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