Louisiana State University and Agricultural & Mechanical College

Transgenrx, Inc.

Other

Helix Biomedix Inc.

Frederick M Enright

Philip H Elzer

Jesse M Jaynes

Richard K Cooper

William Hansel

Patricia A Melrose

Lane D Foil

Samuel M McCann

Wen H Yu

Kenneth L Koonce

Robert P Hammer

Mark L McLaughlin

William C Fioretti

Thomas Scott Yokum

Gerhardt G Schurig

Kenneth L White

John H Wyckoff, Iii

Alexander J Winter

Samuel M Mccann

Amphipathic lytic peptides are ideally suited to use in a ligand/cytotoxin combination to specifically inhibit cells that are driven by or are dependent upon a specific ligand interaction; for example, to induce sterility or long-term contraception, or to attack tumor cells, or to selectively lyse virally-infected cells, or to attack lymphocytes responsible for autoimmune diseases. The peptides act directly on cell membranes, and need not be internalized. Administering a combination of gonadotropin-releasing hormone (GnRH) (or a GnRH agonist) and a membrane-active lytic peptide produces long-term contraception or sterilization in animals in vivo. Administering in vivo a combination of a ligand and a membrane-active lytic peptide kills cells with a receptor for the ligand. The compounds are relatively small, and are not antigenic. Lysis of gonadotropes has been observed to be very rapid (on the order of ten minutes.) Lysis of tumor cells is rapid. The two components&#x2014;the ligand and the lytic peptide&#x2014;may optionally be administered as a fusion peptide, or they may be administered separately, with the ligand administered slightly before the lytic peptide, to activate cells with receptors for the ligand, and thereby make those cells susceptible to lysis by the lytic peptide. The compounds may be used in gene therapy to treat malignant or non-malignant tumors, and other diseases caused by clones or populations of 'normal' host cells bearing specific receptors (such as lymphocytes), because genes encoding a lytic peptide or encoding a lytic peptide/peptide hormone fusion may readily be inserted into hematopoietic stem cells or myeloid precursor cells.

Ligand/lytic peptide methods of use

Methods and compositions are presented for the administration of transposon-based vectors to an animal or human to provide gene therapy to the animal or human.

Gene therapy using transposon-based vectors

Methods and compositions are presented for the administration of transposon based vectors to an animal or human to provide gene therapy to the animal or human.

Genes encoding hormone and lytic peptides

'Minimalist' antimicrobial peptides are disclosed based on 50 to 80% &#x3b1;,&#x3b1;-dialkylated amino acids. The peptides are short, cationic, amphipathic, and possess a high helix propensity. Polar &#x3b1;,&#x3b1;-dialkylated amino acids are also disclosed. These peptides are easy and inexpensive to synthesize via solid-phase techniques. The peptides exhibit in vitro anti-bacterial properties at concentrations that are not lethal to normal mammalian cells. The peptides exhibit in vivo bioactivity against intracellular pathogens.

Short amphipathic peptides with activity against bacteria and intracellular pathogens

Amphipathic lytic peptides are ideally suited to use in a ligand/cytotoxin combination to induce sterility or long-term contraception in mammals. The peptides act directly on cell membranes, and need not be internalized. Administering a combination of gonadotropin-releasing hormone (GnRH) (or a GnRH agonist) and a membrane-active lytic peptide produces long-term contraception or sterilization in mammals in vivo. The compounds are relatively small, and are not antigenic. Lysis of gonadotropes has been observed to be very rapid (on the order of ten minutes.) The two components&mdash;the ligand and the lytic peptide&mdash;may optionally be administered as a fusion peptide, or they may be administered separately, with the ligand administered slightly before the lytic peptide, to activate cells with receptors for the ligand, and thereby make those cells susceptible to lysis by the lytic peptide.

Compositions and methods for contraception in or sterilization of mammals

Amphipathic lytic peptides are ideally suited to use in a ligand/cytotoxin combination to specifically inhibit cells that are driven by or are dependent upon a specific ligand interaction; for example, to induce sterility or long-term contraception, or to attack tumor cells, or to selectively lyse virally-infected cells, or to attack lymphocytes responsible for autoimmune diseases. The peptides act directly on cell membranes, and need not be internalized. Administering a combination of gonadotropin-releasing hormone (GnRH) (or a GnRH agonist) and a membrane-active lytic peptide produces long-term contraception or sterilization in animals in vivo. Administering in vivo a combination of a ligand and a membrane-active lytic peptide kills cells with a receptor for the ligand. The compounds are relatively small, and are not antigenic. Lysis of gonadotropes has been observed to be very rapid (on the order of ten minutes.) Lysis of tumor cells is rapid. The two components&mdash;the ligand and the lytic peptide&mdash;may optionally be administered as a fusion peptide, or they may be administered separately, with the ligand administered slightly before the lytic peptide, to activate cells with receptors for the ligand, and thereby make those cells susceptible to lysis by the lytic peptide. The compounds may be used in gene therapy to treat malignant or non-malignant tumors, and other diseases caused by clones or populations of &ldquo;normal&rdquo; host cells bearing specific receptors (such as lymphocytes), because genes encoding a lytic peptide or encoding a lytic peptide/peptide hormone fusion may readily be inserted into hematopoietic stem cells or myeloid precursor cells.

Ligand/lytic peptide compositions and methods of use

&ldquo;Minimalist&rdquo; antimicrobial peptides are disclosed based on 50 to 80% &agr;,&agr;-dialkylated amino acids. The peptides are short, cationic, amphipathic, and possess a high helix propensity. Polar &agr;,&agr;-dialkylated amino acids are also disclosed. These peptides are easy and inexpensive to synthesize via solid-phase techniques. The peptides exhibit in vitro anti-bacterial properties at concentrations that are not lethal to normal mammalian cells. The peptides exhibit in vivo bioactivity against intracellular pathogens.

A novel class of antimicrobial agents for animal species including cecropins, attacins, lysozymes, phage derived polypeptides, such as those transcribed from gene 13 of phage 22, an S protein from lambda phage, and an E protein from phage PhiXl74, as well as, synthetically derived polypeptides of similar nature. The antimicrobial agents can be used to treat microbial infections and as components of medicinal compositions. The genes encoding for such antimicrobial agents can be used to transform animal cells, especially embryonic cells. The transformed animals including such antimicrobial cells are also included.

Therapeutic antimicrobial polypeptides, their use and methods for preparation

Vaccines against facultative intracellular pathogens are disclosed. A host is vaccinated with non-viable but metabolically active agents. The non-viable agents produce immunogenic components that elicit protective host immune responses, with minimal likelihood of host infection by the vaccine agent. Living agents, either attenuated or virulent, are exposed to a dose of gamma irradiation (or other strong mutagen) that is sufficient to limit or prevent the replication of the agents within the host, but that is insufficient to stop the metabolic activities of the agent. In vitro exposure of a microbial agent to the damaging effects of gamma irradiation or of another strong mutagen induces certain stress responses in the infectious agent. These stress responses are similar to the stress responses that the virulent agent would produce within the tissues of the host. The stress responses include the production of antigens that stimulate appropriate host immune responses when the irradiated agent is used in a vaccine. Examples of facultative intracellular pathogens for which non-viable vaccine agents may be made in accordance with the present invention include various bacterial pathogens (e.g., Brucella sp.,,, Mycobacterium sp.,,, Salmonella sp.,,); various mycotic pathogens (e.g., Blastomyces, Histoplasma, Cocidioides); and various protozoal pathogens (e.g., Leishmania, Trypanosomas).

Method for protecting a mammalian host against infection by Brucella

Novel means have been discovered for increasing the resistance of an animal host (including humans) to diseases caused by intracellular bacteria, protozoa, and viruses. The infection treated may, for example, be equine infectious anemia, or infection by the human immunodeficiency virus. Novel means have also been found for treating tumors. Augmentation of the host's defenses against infectious diseases or tumors is achieved by 'arming' the host's cells with an exogenous gene encoding a natural or synthetic lytic peptide. For example, the transfection of hematopoietic stem cells and embryonic cells will produce animals with enhanced disease resistance; and transfection of TIL (tumor infiltrating lymphocytes) cells or other cells can be used in the treatment of tumors. Genes coding for a cecropin or other native or synthetic lytic peptide can be transferred and stably expressed in mammalian, bony fish, other vertebrate, and other animal cells. The transformed cells have the ability to produce and secrete a broad spectrum chemotherapeutic agent that has a systemic effect on certain pathogens, particularly pathogens that might otherwise evade or overcome host defenses. Novel means have also been discovered for transforming a eukaryotic cell with a gene under the control of an exogenous promoter that is responsive to an inducer of an acute-phase peptide or protein. For example, a moth promoter for the cecropin B gene has been found to function appropriately in a mammalian cell.

Transformed eukaryotic cells

Novel means have been discovered for increasing the resistance of an animal host (including humans) to diseases caused by intracellular bacteria, protozoa, and viruses. The infection treated may, for example, be equine infectious anemia, or infection by the human immunodeficiency virus. Novel means have also been found for treating tumors Augmentation of the host's defenses against infectious diseases or tumors is achieved by 'arming' the host's cells with an exogenous gene encoding a natural or synthetic lytic peptide. For example, the transfection of hematopoietic stem cells and embryonic cells will produce animals with enhanced disease resistance; and transfection of TIL (tumor infiltrating lymphocytes) cells or other cells can be used in the treatment of tumors. Genes coding for a cecropin or other native or synthetic lytic peptide can be transferred and stably expressed in mammalian, bony fish, other vertebrate, and other animal cells. The transformed cells have the ability to produce and secrete a broad spectrum chemotherapeutic agent that has a systemic effect on certain pathogens, particularly pathogens that might otherwise evade or overcome host defenses.

Transgenic fish capable of expressing exogenous lytic peptides

Novel means have been discovered for increasing the resistance of a mammalian host (including humans) to diseases caused by intracellular bacteria, protozoa, and viruses. Novel means have also been found for treating tumors. Augmentation of the host's defenses against infectious diseases or tumors is achieved by 'arming' the host's cells with a gene encoding the lytic peptide cecropin B. The host's own leukocytes, other cells involved in resistance to infection, or other cells are transformed, and expression of the gene is induced when needed to combat pathogens. Transfection of hematopoietic stem cells with the cecropin B gene will enhance disease resistance in mammals; and transfection of TIL (tumor infiltrating lymphocytes) cells or other cells can be used in the treatment of tumors. The transformed cells have the ability to produce and secrete a broad spectrum chemotherapeutic agent which has a systemic effect on certain pathogens, particularly pathogens that might otherwise evade or overcome host defenses. The peptide's expression is preferably induced only in areas of infection, where it will most effectively augment the host's defense systems. Expression of the exogenous gene does not damage a healthy recipient cell.

Growing community of inventors

Baton Rouge, LA, United States of America

Frederick M Enright