University of California

Other

Michael S German

Ira D Goldfine

Stephen S Rothman

Lori Sussel

Hiroshi Inoue

Francis C Szoka, Jr

Graeme I Bell

John L Rubenstein

Dennis J Hartigan-O'Connor

Hiroto Furuta

M Alan Permutt

Roger A Pedersen

Maike Sander

Juanito J Meneses

Joseph Lin

The present invention features polypeptides having activity of human neurogenin3 (hNgn3), and nucleic acid encoding such polypeptide. The invention also features use of islet transcription factors such as hNgn3 to facilitate production of pancreatic islet cells from progenitor cells, and to facilitate insulin delivery by production of islet cells so produced.

Polynucleotides comprising Neurogenin3 promoter and bHLH encoding domains

The present invention relates to the production of islet cells and insulin in a subject by providing for expression of an islet transcription factors in the pancreas of the subject, by for example, introduction of nucleic acid encoding the transcription factor neurogenin3 or a factor that induces neuorgenin3 expression. The present invention also relates to methods for using a islet transcription factor gene and the islet transcription factor polypeptide to alter cellular differentiation in culture or in vivo to produce new &#x3b2;-cells to treat patients with diabetes mellitus.

Production of pancreatic islet cells and delivery of insulin

The present invention provides methods of delivering a secreted protein into the bloodstream of a mammal. A nucleic acid molecule encoding the protein is introduced into the gastrointestinal tract of the mammal, and the nucleic acid molecule enters an intestinal epithelial cell, where the protein is produced and secreted into the bloodstream of the mammal.

Delivery of therapeutic gene products by intestinal cell expression

The present invention features a human neurogenin3 (Ngn3) polypeptide and nucleotide sequences encoding Ngn3 polypeptides. In a particular aspect, the polynucleotide is the nucleotide sequence of SEQ ID NO:1. In addition, the invention features isolated nucleic acid sequence comprising an Ngn3 promoter, as well as a polynucleotide sequences that hybridize under stringent conditions to SEQ ID NO:1. In related aspects the invention features expression vectors and host cells comprising polynucleotides that encode a human Ngn3 polypeptide. The present invention also relates to antibodies that bind specifically to a human Ngn3 polypeptide, methods for producing human Ngn3 polypeptides, methods for identifying &bgr;-cell precursor cells expressing Ngn3, methods for using the Ngn3 gene and the Ngn3 polypeptide to alter cellular differentiation in culture or in vivo to produce new &bgr;-cells to treat patients with diabetes mellitus, and identification of individuals at risk for diabetes by detecting alteration in Ngn3 coding and regulatory sequences and Ngn3 expression levels.

Human neurogenin 3-encoding nucleotide sequences

Secretory gland cells, particularly pancreatic and salivary gland cells, are genetically altered to operatively incorporate a gene which expresses a protein which has a desired therapeutic effect on a mammalian subject. The expressed protein is secreted directly into the gastrointestinal tract and/or blood stream to obtain therapeutic blood levels of the protein thereby treating the patient in need of the protein. The transformed secretory gland cells provide long term therapeutic cures for diseases associated with a deficiency in a particular protein or which are amenable to treatment by overexpression of a protein.

Gene therapy by secretory gland expression

The invention features methods for delivering a polypeptide to the bloodstream of a subject by introduction of a nucleic acid construct into secretory gland cells(e.g., cells of salivary gland, pancreas, or liver). In general, the method involves introduction of a nucleic acid construct into a secretory gland duct, which introduction results in expression of a gene product encoded by the introduced construct and delivery of the gene product into the bloodstream of the subject.

Delivery of polynucleotides by secretory gland expression

The present invention features a human Nkx-6.1 polypeptide and nucleotide sequences encoding Nkx-6.1 polypeptides. In a particular aspect, the polynucleotide is the nucleotide sequence of SEQ ID NO:1. In addition, the invention features polynucleotide sequences that hybridize under stringent conditions to SEQ ID NO:1. In related aspects the invention features expression vectors and host cells comprising polynucleotides that encode a human Nkx-6.1 polypeptide. The present invention also relates to antibodies that bind specifically to a human Nkx-6.1 polypeptide, and methods for producing human Nkx-6.1 polypeptides.

Human NKX-6.1 polypeptide-encoding nucleotide sequences

Intestinal epithelial cells of a mammalian subject are genetically altered to operatively incorporate a gene which expresses a protein which has a desired effect. The method of the invention comprises administration of a formulation containing DNA to the gastrointestinal tract, preferably by an oral route. The expressed recombinant protein is secreted directly into the bloodstream. Of particular interest is the use of the method of the invention to provide for short term delivery of gene products to the bloodstream.

Delivery of gene products by intestinal cell expression

Gene delivery by secretory gland expression

The present invention features a human Nkx-2.2 polypeptide and nucleotide sequences encoding Nkx-2.2 polypeptides. In a particular aspect, the polynucleotide is the nucleotide sequence of SEQ ID NO:1. In addition, the invention features polynucleotide sequences that hybridize under stringent conditions to SEQ ID NO:1. In related aspects the invention features expression vectors and host cells comprising polynucleotides that encode a human Nkx-2.2 polypeptide. The present invention also relates to antibodies that bind specifically to a human Nkx-2.2 polypeptide, and methods for producing human Nkx-2.2 polypeptides.

Human Nkx-2.2 polypeptide-encoding nucleotide sequences

Intestinal epithelial cells of a mammalian subject are genetically altered to operatively incorporate a gene which expresses a protein which has a desired therapeutic effect. Intestinal cell transformation is accomplished by administration of a formulation composed primarily of naked DNA, and is preferably administered orally. Oral or other intragastrointestinal routes of administration provide a simple method of administration, while the use of naked nucleic acid avoids the complications associated with use of viral vectors to accomplish gene therapy. The expressed protein is secreted directly into the gastrointestinal tract and/or blood stream to obtain therapeutic blood levels of the protein thereby treating the patient in need of the protein. The transformed intestinal epithelial cells provide short or long term therapeutic cures for diseases associated with a deficiency in a particular protein or which are amenable to treatment by overexpression of a protein.

Systemic gene therapy by intestinal cell transformation

The present invention features mouse models for Nkx-2.2 gene function and for Nkx-6.1 gene function, wherein the transgenic mouse is characterized by having a defect in Nkx-2.2 gene function or a defect in Nkx-6.1 gene function (where, because Nkx-2.2 acts upstream of Nkx-6.1, a defect in Nkx-2.2 gene function affects Nkx-6.1 gene function) and by having a decreased number of insulin-producing cells relative to a normal mouse. Where the transgenic mouse contains a defect in Nkx-2.2 gene function, the mouse is further characterized by a decreased number of serotonin-producing cells relative to a normal mouse. The transgenic mice may be either homozygous or heterozygous for the Nkx-2.2 or Nkx-6.1 defect.

NKX-2.2 and NKX-6.1 transgenic mouse models for diabetes, depression,

Secretory gland cells, particularly pancreatic, hepatic, and salivary gland cells, are genetically altered to operatively incorporate a gene which expresses a protein which has a desired therapeutic effect on a mammalian subject. The expressed protein is secreted directly into the bloodstream to obtain therapeutic levels of the protein thereby treating the patient in need of the protein. The transformed secretory gland cells provide long term or short term therapies for diseases associated with a deficiency in a particular protein or which are amenable to treatment by overexpression of a protein.

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San Francisco, CA, United States of America

Michael S German