基因沉默(eiRNA->HBV)技术治疗HBV！

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Ercole Biotech And Novosom Complete First Phase of Gene Research Trial 05-07-2004

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RESEARCH TRIANGLE PARK, N.C. and HALLE, GERMANY -- Ercole Biotech, Inc., and novosom AG announced the successful completion of the first phase of their research collaboration. The research involves the intracellular targeted delivery of Ercole's alternative splicing oligonucleotides by using novosom's SMARTICLES delivery system.

The first phase results confirmed the effectiveness of SMARTICLES in targeting Ercole's splice switching drug to specific organs while reducing the total quantity of oligonucleotide required to produce the desired effect.

"We are pleased to be working with an industry leader such as Ercole in the delivery of their proprietary oligonucleotides designed to modulate alternative splicing. This technology has the potential to be a therapeutic breakthrough for a number of diseases," says Dr. Steffen Panzner, CEO of novosom AG.

"It is a major accomplishment to demonstrate the functionality of novosom's SMARTICLES in delivering Ercole's proprietary oligonucleotides directly into the cells of targeted tissue. These studies further confirm that Smarticles offer substantial potential for the delivery of antisense molecules."

Alternative splicing is a natural process in which a single gene can encode multiple related, yet distinct, protein products. Most mammalian genes are interrupted by pieces of DNA, called introns, which are selectively removed from RNA message produced from a gene through the process of RNA splicing.

Cells may remove and splice together different segments of RNA from the same gene, resulting in alternatively spliced gene products. In many cases, the alternatively spliced products can have very different biological effects, with one form contributing to human disease and another form preventing human disease.

Novosom's SMARTICLES enable the targeted delivery of oligonucleotides, plasmids and other materials directly into living cells. Smarticles represent a novel class of liposomes that are fully charge-reversible, which offers a number of advantages in delivering certain new classes of therapeutic agents. Smarticles are stable in blood and distribute in the same manner as traditional anionic liposomes. In stark contrast to anionic liposomes, Smarticles?become positively charged during endocytosis and thus provide a mechanism for endosome release and intracellular delivery.

The research collaboration between Ercole and novosom is now continuing into its second phase.

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ABOUT ERCOLE BIOTECH, INC.: Ercole BioTech, Inc. is a biopharmaceutical company developing a new class of drugs that modulate alternative splicing, a natural gene regulatory mechanism. Ercole's Splice Switching Drugs are applicable to major human diseases including cancer, inflammatory diseases, and cardiovascular disorders. Ercole was founded in 2002 with technology licensed from The University of North Carolina at Chapel Hill, discovered by Dr. Ryszard Kole, an expert on alternative splicing and human disease. Additional information about Ercole BioTech Inc. is available at www.ercolebiotech.com.

ABOUT NOVOSOM AG: Novosom is a drug delivery company that has developed a unique platform technology for the effective and efficient intracellular delivery of oligonucleotides, plasmids and other materials into living cells. Novosom's patented SMARTICLES?are capable of encapsulating large quantities of cargo, remain stable under physiological conditions, and then effectively deliver their payload to targeted delivery sites. Novosom was founded in 1999 by Dr. Steffen Panzner, an expert on the transport of protein molecules through the cell membrane. Novosom collaborates with a number of leading pharmaceutical companies to develop highly effective delivery vehicles for siRNA or antisense. Novosom is also active in the development of sustained release formulations for therapeutic proteins. Additional information about novosom is available at www.novosom.com.

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SMARTICLES® define a novel class of liposomes: fully charge-reversible particles. They are negatively charged under physiological conditions, but as the pH drops down to 5 or 4 during endocytosis, the vector surface becomes neutral and eventually positively charged. This unique property guarantees stable and aggregate-free travel within the bloodstream, but the acidification from endocytosis switches the charge of SMARTICLES®, leading to mem-brane fusion and the escape of the cargo from the endosome. Delivering a cargo via SMARTICLES®: The vehicle attaches to the cell membrane and gets internalized. Ongoing acidification results in charge reversal, and membrane fusion occurs. Cargo is directly released into the cytosol.

The chemistry behind SMARTICLES® is simple: a well defined mixture of an- ionic and cationic headgroups is present in the same lipid membrane. Histi- dine, a natural occuring amino acid, is a typical example for such a head group. Once injected, SMARTICLES® are distributed in the body in the same way as classic liposomes. Tiny openings in the capillaries are a necessary prere- quisite for the escape from the bloodstream. Thus liver and spleen along with sites of inflammation and tumors are primary targets for the SMARTICLES®. See also "routes and rules" (PDF) Novosom believes that its technology offers significant advantages as com- pared to all other delivery methods and is the only functional delivery method available for the in vivo delivery of nucleotides to targeted cells. COMPARSION OF COMMERCIALLY AVAILABLE DELIVERY SYSTEMSHigh payload of nucleic acidsDelivery to cytosolNo aggregation in serumLow cost, ease of preparationLiposomes--++Lipofection++-+Polymers (e.g.PEI)++-+Viral Vectors+++-SMARTICLES®++++ Unlike cationic liposomes or polymers, SMARTICLES® perform well in the living organism. While cationic liposomes are known to efficiently incorporate and transfer cargo into cultured cells, they lose their functionality in vivo. They aggregate in blood or serum and this aggregation interferes with the delivery of their cargo. See also "the liposome dilemma" (PDF) Potential applications In sharp contrast to classic liposomes, SMARTICLES® provide a mechanism for endosome escape and thus deliver directly into the cytosol.

If you got interested, contact our Business Development or Novosom directly, see under contact. > topIn vitro transfectionCase study: in vitro transfection Novosom´s novel lipid based SMARTICLES® were shown to enable the targeted, therapeutic delivery of siRNA and antisense molecules, allowing pharmaceutical researchers to turn these research tools into effective drugs. Target validation in vitro uses a number of commercially available transfec- tants. However, none of them enables the direct preclinical testing. Animal trials, as a first step to evaluate functionality, already require different delivery systems. Novosom’s SMARTICLES® transfection technology solves these problems. Our in vitro transfection approach requires no additives or complex proce- dures. Adherent or suspension cell lines are cultured as usual, then drug loaded SMARTICLES® are added and the mixture is centrifuged under gentle conditions. The result - excellent in vitro transfection with a delivery system that is ready for use in preclinical trials!Cy3-Bcl2 SMARTICLES®free Cy3-Bcl2* ng Cy3- labelled antisense against Bcl240 fold enhanced uptake: Hela cells were centrifuged (1h, 150 x g, 37°C) together with the indicated amounts of Cy3-labeled antisense molecules, either as free drug or encapsulated in SMARTICLES® formulation.Novosom´s "reverse engineering strategy" now enables SMARTICLES®, designed for in vivo delivery of nucleic acids, to be used in vitro without further additives. > topInflammation bannedAntisense delivery Inflammatory bowel diseases, including Crohn’s disease and ulcerative colitis, are characterized by chronic inflammation of the gastrointestinal tract. Although their etiology remains unknown, the inappropriate and sustained activation of the mucosal immune system seems crucial to the development of these diseases. The pivotal role of CD 154 / CD 40 interactions in the regulation of immune responses has been firmly established and blockage of the CD 154 / CD 40 interactions may expert a potent therapeutic effect in Crohn´s disease. Our SMARTICLES® technology offers an effective delivery system for oligo- nucleotides, including the CD´s oligonucleotides. We have encapsulated a special CD 40 antisense oligonucleotide into SMATICLES® - novel liposo- mal vectors for in vivo transfection. The SMARTICLES® formulation effectively targeted and treated inflammation in a rat colitis model after intracolonic in-jection. Colitis was induced in rats using a single intracolonic application of TNBS. Subsequently, SMARTICLES® encapsulated CD 40 oligonucleotides were administered by intracolonic injection. Seven days after induction of colitis, the colonic inflammation and damage were assessed by macroscopic evaluation. There was a significant reduction in the most severe types of damage in the group treated with the SMARTICLES® encapsulated CD 40 oligonucleotide. Histological analysis of CD 40 antisense oligonucleotides effects in TNBS-induced colitis:

In the descending colon of control animals (B), the mucosa was usually lost and the bowel wall thickened because of the prominent infiltration of leukocytes and moderate fibrosis. The bowel wall structure of the animals which were administered with the SMARTICLES®-formulation (C) displayed the appearance of healthy control rats (A). The microscopic scoring also revealed that treatment with the encapsulated CD 40 antisense (rAS3) resulted in a significant improvement. Encapsulated scrambled control oligonucleotides (rScr) were as ineffective as saline. Quantification of SMARTICLES® encapsulated CD 40 antisense oligonucleotides effects in TNBS induced colitis: Statistical analysis of microscopic scoring (n=7-8. * P< 0.01) Cont. - healthy animals, rAS3 - SMARTICLES® encapsulated CD 40 antisense, rScr - SMARTICLES® encapsulated scrambled control oligonucleotides

The histochemical investigations demonstrated a prominent infiltration of granulocytes in the affected colon of both untreated rats and with scrambled control oligonucleotides treated rats, especially in the most severely damaged regions. The mucosa and submucosa propria of animals treated with SMARTICLES® encapsulated CD 40 oligonucleotides show only few granulocytes. > topGene transferWith ease, without virus To cover the most obvious fields of application, we transferred fully intact, functional genes into animals. We here used the Green Fluorescent Protein (GFP), coded on a 7kb vector. Animals received a single intravenous dose of 50µg plasmid, either packed into SMARTICLES® or as naked DNA. Vector-dependent expression was found in liver, spleen, lungs and endothelia. A transient expression like this is already sufficient for therapeutic vaccina- tion. A repeated or more prolonged one will open the door towards gene therapy. A transient, but well targeted one is the key to cancer gene therapy. SMARTICLES® help to pave the way to future medicines. Expression of GFP in rat liver: Animals were given a single i.v. injection of a plasmid encapsulated in two different SMARTICLES® vectors. Nuclei were stained with DAPI (left) Green fluorescence develops after expression of GFP (right).

> topcmc dataChemical and physical characterization of SMARTICLES® particle size50.... 300nm zeta potentialpH-dependent viscositycomparable to water materialsphospholipids, proprietary lipids stability>24 h in serum shelf life>2 months @ room temperature tested circulation half-life3-4 h in organism Efficient loading - leakproof encapsulation SMARTICLES® efficiently encapsulate and protect the active ingredient. Once encapsulated, even very sensitive cargoes like siRNA become fully protected against serum components. SMARTICLES® are manufactured from charge-reversible lipids and novosom takes full advantage of this variable surface charge during the encapsulation procedure. Encapsulation of siRNA with different SMARTICLES® formulations: 10 out of 16 starting formulations encapsulated more than 20% of the starting material and were selected for further optimization.

All SMARTICLES® formulations for in vivo use were rigorously tested for serum stability. Leakage of fluorescent dyes provides very sensitive means of stability measurements. Once passed this test, a particular SMARTICLES® formulation was loaded with siRNA and experimentally challenged with full human serum. As monitored below, SMARTICLES® fully protect siRNA from degradation for more than 24 hours. Protection of siRNA in serum: siRNA was used in free form (left panel) or was encapsulated in SMARTICLES® (right panel) and incubated with human serum at 37°C for the indicated periods of time. Both strands of the oligonucleotide were fully protected against degradation even after 24 hours of incubation.

[此贴子已经被作者于2004-10-25 13:18:54编辑过]

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Contact novosom AG Weinbergweg 22 06120 Halle Germany Mail: [email protected] Tel.: +49 (0)345 55 59 836 Fax: +49 (0)345 55 59 846 CEO Dr. Steffen Panzner Tel.:+49 (0)345 55 59 845 Mail: [email protected] COO Elias Papatheodorou Tel.: +49 (0)345 55 59 646 Mail: [email protected] SMARTICLES® Dr. Gerold Endert Tel.:+49 (0) 345 55 59 639 Mail: [email protected]

良药尽欢颜

RNAi是我最关注的治疗方法了，也是攻克的最大希望，听说这两年在这方面出了很多新的研究和著述，请楼主及本坛高手注意搜集，让我们增加信心。

614

2008年都快过去了,有新的进展吗?

宁静温泉

I期临床已经展开。好象是明年底会有结果。

刘德华影子

成果漫漫积累的。

kangyuguaiguai

大篇大篇的英文，晕哦，看不懂。
哪位高手翻译出来，那就功德无量了。呵呵

614

论文是用来评职称用的，研究需要资金为基础的，老百姓是可以忽悠的．

dickens_yu

估计还要15年左右