University of Washington

Oncogen

Other

Fred Hutchinson Cancer Center

Oncogen Limited Partnership

Timothy M Rose

Kurt Strand

Marnix L Bosch

George J Todaro

Anthony F Purchio

Gregory D Plowman

Karl E Hellstrom

Ingegerd E Hellstrom

Joseph P Brown

Shiu-Lok Hu

Charles D Estin

Sridhar Pennathur

Peter S Linsley

David W Leung

Hans Marquardt

Mohammed Shoyab

Joyce M Zarling

Marcia B Hanson

Najma Malik

J Ken Rose

A Gregory Bruce

Hybrid cytokines containing four helical regions, each of which is derived from a corresponding &agr;-helical region in leukemia inhibitory factor (LIF), granulocyte colony stimulating factor (G-CSF), interleukin-6 (IL-6) or oncostatin-M (OSM) are disclosed. These hybrid cytokines may further contain linking regions also derived from corresponding linking regions in these factors. The hybrid cytokines offer a unique spectrum of activities useful in treating conditions for which the native cytokines are useful or in treating conditions characterized by an excess of the native cytokines.

Hybrid cytokines

Expression cassettes for enhanced expression and production of a polypeptide of interest in prokaryotic cells are provided. The expression cassettes provide for production of the polypeptide of interest so that such polypeptide can either be secreted from the host cell in an active conformation or conveniently processed and renatured to a functional state. Preferably, the polypeptide of interest is expressed as a fusion protein, particularly fused to a leader sequence from a highly expressed bacterial or bacteriophage gene. The polypeptide of interest may subsequently be cleaved from the leader sequence and refolded, or used as a fusion protein.

Expression systems for preparation of polypeptides in prokaryotic cells

Therapeutic hybrid cytokines, having a size ranging from about 10 to about 30 kDa, comprise portions of cytokines: leukemia inhibitory factor (LIF), granulocyte-colony stimulating factor (G-CSF), interleukin-6 (IL-6), interleukin-11 (IL-11), oncostatin-M (OSM), and ciliaryneurotrophic factor (CNTF). Hybrid cytokines comprise three or four &agr;-helical sequences selected from &agr;-helical sequences of IL-6, G-CSF, LIF, IL-11, CNTF and OSM and linking sequences of 5-40 amino acids in length, selected from the linking sequences of IL-6, G-CSF, LIF, IL-11, CNTF and OSM or other, desirable linking sequences. In the hybrid cytokines, at least one &agr;-helical sequence is derived from a different cytokine than at least one other &agr;-helical sequence; or, at least one linking sequence of a cytokine differentiates the hybrid cytokine from a corresponding cytokine. In hybrid cytokines having three &agr;-helical sequences selected from the group of cytokines listed, a possible fourth sequence may not correspond to any &agr;-helical sequence in any of the cytokines. Hybrid cytokines, radically different from any corresponding cytokines, have unexpected advantages in selective biological mechanisms over a much broader range of biological activities than exhibited by cytokines.

This invention relates to polynucleotides encoding Glycoprotein B from the RFHV/KSHV subfamily of gamma herpes viruses, three members of which are characterized in detail. DNA extracts were obtained from Macaque nemestrina and Macaque mulatta monkeys affected with retroperitoneal fibromatosis (RF), and human AIDS patients affected with Kaposi's sarcoma (KS). The extracts were amplified using consensus-degenerate oligonucleotide probes designed from known protein and DNA sequences of gamma herpes viruses. The nucleotide sequences of a 319 base pair fragment are about 76% identical between RFHV1 and KSHV, and about 60-63% identical with the closest related gamma herpes viruses outside the RFHV/KSHV subfamily. Protein sequences encoded within these fragments are are about 91% identical between RFHV1 and KSHV, and &lt;.about.65% identical to that of other gamma herpes viruses. The full-length KSHV Glycoprotein B sequence comprises a transmembrane domain near the N-terminus, and a plurality of potentially antigenic sites in the extracellular domain. Materials and methods are provided to characterize Glycoprotein B encoding regions of members of the RFHV/KSHV subfamily, including but not limited to RFHV1, RFHV2, and KSHV Peptides, polynucleotides, and antibodies of this invention can be used for diagnosing infection, and for eliciting an immune response against Glycoprotein B.

Glycoprotein B of the RFHV/KSHV subfamily of herpes viruses

This invention provides isolated polynucleotides encoding DNA polymerases of three members of a subfamily of gamma herpes viruses. Two were obtained from macaque monkeys affected with retroperitoneal fibromatosis, the other from human AIDS patients affected with Kaposi's sarcoma. A 454-base pair fragment encoding a region near the active site of the DNA polymerase is 69-83% identical amongst the three viruses, but only 54-68% identical with other known gamma herpes sequences and &lt;55% identical with alpha and beta herpes sequences. Also provided are polynucleotides encoding DNA polymerase from related viruses in the RFHV/KSHV subfamily. Polynucleotides prepared according to the sequence data can be used as reagents to detect and characterize related sequences. Such reagents may be used to detect members of the RFHV/KSHV subfamily, including but not limited to RFHV, RFHV2, and KSHV. Corresponding polypeptides and peptide fragments may be obtained by expressing the polynucleotide or by chemical synthesis. They may be used for detecting specific antibody potentially present in the serum of infected subjects. They may also be used for designing or screening pharmaceutical compounds that limit viral replication by inhibiting DNA polymerase activity.

DNA polymerase of gamma herpes viruses associated with Kaposi's sarcoma

Novel compositions comprising Oncostatin M and congeners thereof, as well as methods for their preparation and methods for their use are provided. The compositions may be prepared by isolation from natural sources, or by recombinant means in either prokaryotic or eukaryotic host cells. In addition, the DNA and polypeptide sequences for Oncostatin M are disclosed. The compositions find use in modulating growth of cells, in particular inhibition of tumor cell proliferation, and stimulation of normal cell growth, especially cells involved in hematopoiesis. Cell growth inhibition compositions may additionally include an adjunctive agent comprising at least one of a transforming growth factor, tumor necrosis factor, or an interferon. Receptors having high affinity for Oncostatin M may additionally be used to screen polypeptides for Oncostatin M-like activity. Methods for use of antibodies to the compositions and probes specific for Oncostatin M mRNA as a means for detecting tumor cells are also provided.

Oncostatin M and novel compositions having anti-neoplastic activity

Peptides or proteins related to a melanoma associated antigen are described. These are produced in large quantities via recombinant DNA techniques and/or by chemical synthetic methods. The peptides or proteins can be used as immunogens in vaccine formulations which can induce an immune response that selectively destroys melanoma cells in a vaccinated individual. Where the peptides or proteins are expressed by a recombinant virus, inactivated or live virus vaccine formulations may be prepared.

Recombinant viruses encoding the human melanoma-associated antigen

Vaccines against melanoma

A continuous concrete encased conduit laying apparatus is attached to the rear of a tractor in a cantilevered fashion and can be raised and lowered into the ground by the tractor. The apparatus has two sections which are pivotally attached to each other. The forward section is the gouger, and it has a vertical bar having a sharp leading ripper edge. At the base of the bar are attached a wedge and a cone-shaped expander for creating a cavity in the ground as the apparatus is being pulled through the ground. The rear section which is the conduit layer, has a vertical guide tube, a concrete chute attached behind it, and a second expander forming the base of the layer. There is a hopper for holding the fresh concrete at the top of the chute. A quantity of conduit is fed through the guide tube as the apparatus is in operation. The cavity created by the two expanders is filled with fresh concrete as the conduit is laid in the cavity. The second expander can be round or rectangular in cross-section for holding several conduits, and for creating various cross-section encasements. The method includes the steps of laying the conduit and encasing it with concrete in the cavity.

Continuous conduit laying apparatus

Growing community of inventors

Seattle, WA, United States of America

Timothy M Rose