IGF-I is a pleiotropic hormone reported to affect linear growth, glucose metabolism, organ homeostasis, and the immune and neurologic systems. In contrast to IGF-II, IGF-I is expressed at low levels embryonically and has been thought to be more important for postnatal growth and development. To investigate the role of IGF-I in normal development we generated mice with an inactive IGF-I gene by homologous recombination in ES cells. Heterozygous mice are healthy and fertile, but they are 10-20% smaller than wild-type littermates and have lower than normal levels of IGF-I. The size reduction is attributable to a decrease in organs and muscle and bone mass. However, all tissues appear histologically normal. At birth homozygous mutant mice (IGF-I -/-) are <60% body weight of wild type. Greater than 95% of IGF-I -/-pups die perinatally. Histopathology is characterized by underdevelopment of muscle tissue. Lungs of late embryonic and neonates also appeared less organized with ill-defined alveolae. IGF-I appears to be essential for correct embryonic development in mice.

DNA clones encoding the complete 2,351 amino acid sequence for human factor VIII have been isolated and used to produce biologically active factor VIII in cultured mammalian cells. The recombinant protein corrects the clotting time of plasma from haemophiliacs and has many of the biochemical and immunological characteristics of serum-derived factor VIII.

The primary structure of factor VIII consists of 2332 amino acids that exhibit 3 distinct structural domains, including a triplicated region (A domains), a unique region of 909 amino acids (B domain), and a carboxy-terminal duplicated region (C domains), that are arranged in the order A1-A2-B-A3-C1-C2. The B domain (residues 741-1648) of factor VIII is lost when factor VIII is activated by thrombin, which proteolytically processes factor VIII to active subunits of Mr 50,000 (domain A1), 43,000 (domain A2), and 73,000 (domains A3-C1-C2). To determine if the B domain is required for factor VIII coagulant activity, a variant was constructed by using recombinant DNA techniques in which residues 797-1562 were eliminated. This shortened the B domain from 909 to 142 amino acids. This variant factor VIIIdes-797-1652 was expressed in mammalian cells and was found to be functional. The factor VIIIdes-797-1562 protein was purified and shown to be processed by thrombin in the same manner as full-length factor VIII. The factor VIIIdes-797-1562 variant also bound to von Willebrand factor (vWF) immobilized on Sepharose. These results indicate that most of the highly glycosylated B domain of factor VIII is not required for the expression of factor VIII coagulant activity and its interaction with vWF.

Purpose: This study examined the effectiveness of early and prolonged mu4D5 (the murine form of trastuzumab/ Herceptin) treatment in transgenic mice that overexpress human HER2 (huHER2), under the murine mammary tumor virus promoter, as a model of huHER2-overexpressing breast cancer.Experimental Design: Mice were randomly assigned to one of three treatment groups and received i.p. injections from 17 weeks of age until either 52 weeks of age or morbidity. Fourteen mice received 100 mg/kg mu4D5, 14 mice received 100 mg/kg antiherpes simplex virus glycoprotein D control antibody, and 11 mice received a diluent control.Results: High levels of huHER2 expression were detectable in mammary glands of young virgin founder mice. Mammary adenocarcinomas were frequently found in female founders and progeny at an average age of 28 weeks, with some progressing to metastatic disease. The incidence of mammary tumors was significantly reduced, and tumor growth inhibition was observed in mice receiving mu4D5 compared with control mice. In addition, Harderian gland neoplasms, highly associated with overexpression of hu-HER2 in this transgenic line, were entirely absent in the mu4D5 treatment group, indicating down-regulation of hu-HER2 in vivo activity.Conclusions: Early intervention with mu4D5 was of benefit in our transgenic mice at high risk for developing huHER2-overexpressing breast cancer. This study suggests a potential benefit of early treatment with Herceptin in HER2-positive primary breast cancer.

Growth hormone (GH) is an important regulator of postnatal growth, acting on a wide variety of target tissues. Here, we show that local production of GH in osteoblasts is able to stimulate bone growth directly without significant systemic effects. Mice were made transgenic by microinjection of an osteocalcin-human GH (osteocalcin-hGH) gene construct in which approximately 1,800 bp of the rat osteocalcin promoter was fused to the hGH gene. Five lines of transgenic mice, each with measurable amounts of serum hGH (ranging from 1 to 1,000 ng/ml), were analyzed. Northern (RNA) blot hybridization showed that the hGH transcript was detectable only in the bone. Further characterization of hGH mRNA distribution by in situ hybridization revealed that in neonates the most intense signal was found in periosteal osteoblasts and in odontoblasts, while in adults, trabecular and endosteal osteoblasts were favored. In one transgenic line (992-1), hGH was expressed at a much lower level and had minimal systemic effects; however, the local concentrations of hGH in bone were sufficient to stimulate bone growth in these animals.