Dna Vec Research Inc.

Other

Id Pharma Co., Ltd.

Akihiro Iida

Mamoru Hasegawa

Yasuji Ueda

Makoto Inoue

Masanori Kobayashi

Yoshikazu Yonemitsu

Hiroshi Ban

Takahiro Hirata

Tsuyoshi Tokusumi

Hiroaki Kinoh

Toshihiro Nakajima

Yoshiyuki Nagai

Masanori Miyazaki

Katsuo Sueishi

Makoto Asakawa

Tsugumine Shu

Yasuhiro Ikeda

Toshiaki Tabata

Kenji Nakamaru

Hiroyuki Sakakibara

Kaio Kitazato

Hidekazu Kuma

Fumino Tokito

Yumiko Tokusumi

The present invention provides methods for improving the efficiency of inducing pluripotent stem cells, as well as vectors and compositions for use therein. In the induction of pluripotent stem cells which contains the step of introducing a vector that contains the KLF gene, OCT gene, and SOX gene in this order, the efficiency of pluripotent stem cell induction was successfully increased significantly by further introducing a vector that contains the KLF gene but not the OCT gene and the SOX gene. The methods of the present invention have an excellent feature in that they allow efficient induction of pluripotent stem cells under a temperature condition closer to the physiological environment, and prompt vector removal after the pluripotent stem cell induction. The present invention enables more efficient induction of pluripotent stem cells.

Method for improving efficiency of inducing pluripotent stem cell

The present invention provides methods for producing a minus-strand RNA viral vector, which comprise using a promoter comprising a cytomegalovirus enhancer and a chicken &#x3b2;-actin promoter, to induce the transcription of the genome RNA of a minus-strand RNA viral vector and the expression of minus-strand RNA viral proteins that form a ribonucleoprotein with the genome RNA. The methods of the present invention enable high efficiency production of highly safe minus-strand RNA viral vectors. The methods of the present invention are particularly useful for producing minus-strand RNA viral vectors that are deficient in envelope-constituting protein genes.

Methods for producing minus-strand RNA viral vectors using hybrid promoter comprising cytomegalovirus enhancer and chicken &#x3b2;-actin promoter

The present invention provides novel methods for treating diseases associated with apoptotic degeneration in ocular tissue cells by effective administration of pigment epithelium derived factor (PEDF). The present inventors studied PEDF as a means to prevent ganglion cell death, the final pathology of glaucoma. The present invention is particularly focused on SIV vectors for effective methods for delivering PEDF, and constructed an SIV-PEDF vector. When the SIV-PEDF vector was administered subretinally to an ischemia reperfusion model and NMDA-induced model, a significant suppression effect on ganglion cell death was observed. The present inventors therefore discovered that the SIV-PEDF vector is an effective pharmaceutical agent for treating diseases associated with apoptotic degeneration in ocular tissue cells, such as glaucoma.

Therapeutic agents for diseases associated with apoptotic degeneration in ocular tissue cells that use SIV-PEDF vectors

The present invention provides cell fusogenic vectors having replicative ability, whose protease-dependent tropism has been modified. M gene-deficient viral vectors encoding modified F proteins, in which the cleavage site of the F protein of paramyxovirus is modified to be cleaved by different proteases, were produced. In cells transfected with these vectors, the genomic RNA present in the vectors is replicated, and cell fusogenic infection spreads to neighboring cells depending on the presence of other proteases; however, no viral particles are released. The vectors of this invention, encoding the F proteins which are cleaved by proteases whose activity is enhanced in cancer, show cancer growth suppressive effect in vivo.

Vectors with modified protease-dependent tropism

The present invention provides a retroviral vector containing a membrane protein having a hemagglutinin activity. The present inventors constructed a retroviral vector pseudotyped by the membrane protein having a hemagglutinin activity. This viral vector showed gene transfer at a high efficiency into host cells. In particular, it was established that genes can be transferred thereby at a high efficiency into cells into which genes can hardly be transferred by the conventional techniques, for example, blood cells and hematopoietic cells including hematopoietic stem cells, and mucous cells including mucosa epithelial cells. The viral vector of the present invention is highly useful as a vector for gene therapy.

Pseudotype retroviral vectors containing membrane proteins having hemagglutinin activity

The present invention provides cell fusogenic vectors having replicative ability, whose protease-dependent tropism has been modified. M gene-deficient viral vectors encoding modified F proteins, in which the cleavage site of the F protein ofis modified to be cleaved by different proteases, were produced. In cells transfected with these vectors, the genomic RNA present in the vectors is replicated, and cell fusogenic infection spreads to neighboring cells depending on the presence of other proteases; however, no viral particles are released. The vectors of this invention, encoding the F proteins which are cleaved by proteases whose activity is enhanced in cancer, show cancer growth suppressive effect in vivo.

F gene-deficient virus virions are successfully recovered by using an F gene-deficient Sendai virus genomic cDNA. Further, F gene-deficient infectious viral particles are successfully constructed by using F-expressing cells as helper cells. Also, F gene and HN gene-deficient virus virions are successfully recovered by using a virus genomic cDNA deficient in both F gene and HN gene. Further, F gene and HN gene-deficient infectious viral particles are successfully produced by using F- and HN-expressing cells as helper cells. A virus deficient in F gene and HN gene and having F protein is constructed by using F-expressing cells as helper cells. In addition, M gene-deficient infectious virus particles were produced using helper cells expressing M protein. From cells infected with M gene-deficient viruses, release of virus-like particles was inhibited. Further, a VSV-G pseudo type virus is successfully constructed by using VSV-G-expressing cells. Techniques for constructing these deficient viruses contribute to the development of vectors of Paramyxoviridae usable in gene therapy.

Envelope gene-deficient Paramyxovirus vector

A paramyxovirus vector capable of Extransfecting foreign genes and having a replication capacity, is provided. A Sendai virus vector comprising a foreign gene can be constructed by inserting a foreign gene between the viral genes of the Sendai virus genome. These Sendai viruses have a replication capacity and express the foreign gene in transfected cells. The expression level of the foreign gene is higher towards the 3&prime; end of negative strand RNA, and especially, a high level of expression is obtained when the foreign gene is inserted before the NP gene, and between P gene and M gene. Conversely, the expression decreases towards 5&prime; end of negative strand RNA, and especially, a relatively low level of expression is obtained when the when the foreign gene is inserted between HN gene and L gene, and between F gene and HN gene. Thus, the vector of the invention enables the regulation of the expression level of a foreign gene. The vector is useful for gene therapy due to its safety, high gene transfer efficiency, and capacity to express a foreign gene at a high level.

Paramyxovirus vectors used for transfer of foreign genes

Growing community of inventors

Ibaraki, Japan

Akihiro Iida