Mucosal challenge of Macaca nemestrina with simian immunodeficiency virus (SIV) following SIV nucleocapsid mutant DNA vaccination.

A simian immunodeficiency virus (SIV)(Mne) DNA clone was constructed that produces viruses containing a four amino acid deletion in the second zinc finger of the nucleocapsid (NC) domain of the Gag polyprotein. Viruses produced from this clone, although non-infectious both in vitro and in vivo, complete a majority of the steps in a single retroviral infection cycle. Eight pig-tailed macaques (Macaca nemestrina) were inoculated intramuscularly and subcutaneously three times over the course of 24 weeks with the NC mutant expressing DNA. These macaques, and four controls, were then challenged mucosally (intrarectally) with the homologous virus (SIV Mne CL E11S) and monitored for evidence of infection and clinical disease. Prior to challenge, a measurable humoral immune response was noted in four of eight immunized macaques. After challenge, all 12 macaques became infected, although four immunized animals greatly restricted their viral replication, and one immunized animal that controlled replication remains antibody negative. No disease has been evidence during the 46-week period of monitoring after challenge.

Detection of serum antibodies in Ethiopian baboons that cross-react with SIV, HTLV-I, and type D retroviral antigens.

Baboons (Papio cynocephalus) imported from Ethiopia were screened for antibodies to various primate retroviruses by immunoblotting. Antibodies that cross-reacted with SIV/Mne or with type D viral antigens were detected in approximately one-third of these animals. In addition, 20% of these baboons had antibodies that cross-reacted with HTLV-I viral antigens. These data suggest that wild-caught baboons are infected with retroviruses only partially related to known primate viral isolates.

Characterization of clones of HIV-1 infected HuT 78 cells defective in gag gene processing and of SIV clones producing large amounts of envelope glycoprotein.

Single-cell clones of HIV-1 (FRE-3) or SIV/Mne infected HuT 78 cells were obtained by plating dilutions of virally infected HuT 78 cells on a monolayer of sheep choroid plexus cells in 96-well microtiter plates. Several of these clones produce HIV-1 virus mutants that accumulate the gag precursor polyprotein and lack a functional protease. These protease-deficient viruses are non-infectious and consist of aberrant "immature" virus particles as determined by electron microscopy. Several SIV mutants are also described that produce large amounts of either the envelope glycoprotein gp120 or the nucleic acid binding gag protein. These mutants are useful for the purification of these retroviral proteins, in developing assays of protease inhibitors, and in preparing SIV envelope protein vaccines.

Molecular characterization and comparison of simian immunodeficiency virus isolates from macaques, mangabeys, and African green monkeys.

Simian immunodeficiency virus (SIV)/Mne has been inoculated into three species of macaques and into baboons. Virus was isolated from all the macaques who subsequently died at 15 to 120 weeks (mean 80 weeks) with various manifestations of immune deficiency. Individual animals varied in their viral antibody profile as a function of time after infection. Independent SIV isolates obtained from African green monkeys and magabeys were compared to SIV/Mne for their ability to replicate in lymphocytes and macrophages and with respect to the immunological relatedness of their viral proteins. Antibodies present in human immunodeficiency virus-2 (HIV-2)-infected individuals were readily detected by the virus produced by a single-cell clone of SIV/Mne.

Inoculation of baboons and macaques with simian immunodeficiency virus/Mne, a primate lentivirus closely related to human immunodeficiency virus type 2.

A primate lymphotropic lentivirus was isolated on the human T-cell line HuT 78 after cocultivation of a lymph node from a pig-tailed macaque (Macaca nemestrina) that had died with malignant lymphoma. This isolate, originally designated M. nemestrina immunodeficiency virus (MnIV) and now classified as simian immunodeficiency virus (SIV/Mne), was inoculated intravenously into three juvenile rhesus monkeys (Macaca mulatta), three juvenile pig-tailed macaques (M. nemestrina), and two juvenile baboons (Papio cynocephalus). All six macaques became viremic by 3 weeks after inoculation, whereas neither of the baboons developed viremia. One pig-tailed macaque died at 15 weeks with suppurative peritonitis secondary to ulcerative, necrotizing colitis. Immunologic abnormalities included a marked decrease in CD4+ peripheral blood lymphocytes. Although five macaques mounted an antibody response to SIV/Mne, the animal that died at 15 weeks remained antibody negative. Three other macaques (two rhesus and one pig-tailed) died 66 to 87 weeks after inoculation after exhibiting progressive weight loss, anemia, and diarrhea. Histopathologic findings at necropsy included various manifestations of immune deficiency, nephropathy, subacute encephalitis, pancreatitis, adenocarcinoma, and lymphoid atrophy. SIV/Mne could be readily isolated from the spleens and lymph nodes of all necropsied macaques, and from the cerebrospinal fluid, brains, bone marrow, livers, and pancreas of some of the animals. SIV antigens were localized by avidin-biotin immunohistochemistry to pancreatic islet cells and to bone marrow endothelial cells. The data suggest that African baboons may be resistant to infection by SIV/Mne, whereas Asian macaques are susceptible to infection with this pathogenic primate lentivirus.

Two distinct tumor cell growth-inhibiting factors from a human rhabdomyosarcoma cell line.

Tumor cell growth-inhibiting factors (TIFs) have been shown to inhibit the growth of tumor cell lines in culture. TIF-1, the first TIF to be described, is a low-molecular-weight, acid- and heat-stable polypeptide with no antiviral activity. A second class of TIFs (TIF-2) has now been isolated from the conditioned media of a human rhabdomyosarcoma cell line and partially purified by polyacrylamide gel filtration, cation exchange, and reverse-phase high-pressure liquid chromatography. Partially purified preparations of TIF-2 inhibit the growth of a variety of human tumor cells in soft agar and monolayer cultures and are mitogenic for normal human and mouse cells. TIF-2 has no antiviral activity. The growth-inhibitory effects of TIF-2 are reversible when the affected cells are no longer exposed to the factor. Although both TIF-1 and TIF-2 are obtained from the same source, they can be distinguished by their molecular weight, heat lability, elution pattern from reverse-phase high-pressure liquid chromatography, and their effect on the growth of mink lung epithelial cells. The growth of a human tumor cell variant, selected for resistance to growth inhibition by TIF-1, is inhibited by TIF-2. TIFs may therefore be a family of related polypeptides which selectively inhibit the growth of tumor cells.

Isolation of tumor cell growth-inhibiting factors from a human rhabdomyosarcoma cell line.

Two types of growth-modulating factors were derived from the serum-free conditioned media of a human rhabdomyosarcoma cell line, A673. One type, Mr 18,000 to 22,000, competes for binding to epidermal growth factor receptors and enhances the growth of normal and tumor cells in soft agar. It has all of the biological properties ascribed to transforming growth factor type alpha (TGF alpha). A673 cells also produce factors which inhibit the growth of human tumor cells in soft agar and in monolayer cultures. These tumor cell growth-inhibiting factors (TIFs) are acid- and heat-stable peptides. The major TIF activities have molecular weights in the ranges of greater than 28,000, 18,000 to 22,000, 10,000 to 16,000, and 5,000 to 10,000 and do not possess the antiviral activity associated with interferon. Partially purified preparations of TIF-1 (Mr 10,000 to 16,000) inhibit the growth of all human tumor cell lines tested and stimulate the growth of normal human fibroblasts and epithelial cells in monolayer cultures. The growth of human lung carcinoma A549 cells in soft agar, which was enhanced by treatment with TGF alpha from A673-conditioned media, was inhibited by treatment with TIF-1 derived from the same media. The ratio of the two types of tumor cell-derived, growth-modulating factors (TIFs and TGF alpha), which are antagonistic in their biological effects, may determine the capacity of tumor cells for anchorage-independent growth.