Augmentation of in-vitro HIV replication in peripheral blood mononuclear cells of AIDS and ARC patients by tumour necrosis factor.
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AIDS-Related Complex↗
Biomedical subjects
G Soma
Publications and source records attributed to G Soma.
Immunochemotherapy for murine bladder tumor with a new human recombinant tumor necrosis factor (rTNF-S), VP-16 and hyperthermia.
Effects of a new recombinant human tumor necrosis factor (rTNF-S) on the growth of murine bladder tumor (MBT-2) were studied. The growth of MBT-2 was significantly inhibited by intratumor injection of rTNF-S. Moreover etoposide (VP-16) and hyperthermia at relatively low temperature enhanced the antitumor effect of rTNF-S. Since TNF is considered to have species preference, these observations may be relevant to treatments for human bladder tumor with human TNF.
Animals↗
Enhancement of HIV replication and giant cell formation by tumor necrosis factor.
Human T cell lines Molt-4, Jurkat, and TLOm-1 were infected by HIV-1 in the presence of various concentrations of r-TNF. The infectivity of HIV-1 was monitored by an indirect immunofluorescence method using anti-HIV-1-positive human serum. We found that r-TNF enhanced the replication of HIV-1. HIV-1-induced giant cell formation between HIV-1 chronically infected Molt-4 cells and HIV-1-uninfected Molt-4 cells was accelerated by r-TNF. The median tissue culture infectious doses (TCID50) of HIV-1 determined in the presence of TNF revealed that TNF apparently accelerated the time of the appearance of CPE but did not affect final titer of HIV-1.
Cells, Cultured↗
Cytocidal effect of tumor necrosis factor on cells chronically infected with human immunodeficiency virus (HIV): enhancement of HIV replication.
Tumor necrosis factor (TNF), a monokine initially described as a tumoricidal agent, facilitated the replication of human immunodeficiency virus (HIV) in vitro. The viability of human T-cell line MOLT-4/HIV, chronically infected with HIV, was affected by the addition of a low dose (10 ng/ml) of recombinant TNF-alpha (rTNF-alpha), while uninfected MOLT-4 cells were resistant to treatment with rTNF-alpha at concentrations up to 1,000 ng/ml. A marked increase in the level of HIV-specific RNA was detected in MOLT-4/HIV cells as early as 1 h after exposure to rTNF-alpha and reached almost maximum level within 6 h. Production of HIV particles from MOLT-4/HIV was also increased at 6 h after treatment with rTNF-alpha. Nearly identical phenomena were observed in CCRF-CEM/HIV, Jurkat/HIV, and H9/HIV cells, although the sensitivity of these cell lines to rTNF-alpha varied. A human T-lymphotropic virus type 1-infected cell line, MT-4, was insensitive to treatment with rTNF-alpha.
Antibodies, Monoclonal↗
Immunohistochemical evidence for the production of tumor necrosis factor by murine MH134 tumor cells as well as monocytes in tumor lesions after systemic administration of a polyalcoholized mannoglucan from Microellobosporia grisea.
The endogenous production of tumor necrosis factor (TNF) by tumor tissues was examined. MH134 tumor cells as well as monocytes in MH134 tumor tissues produced TNF after systemic administration of a polyalcoholized mannoglucan (MGA) from Microellobrosporia grisea as shown by the indirect immunofluorescence technique. MH134 tumor cells also produced TNF when stimulated with lipopolysaccharide in vitro. These results suggest that regression of MH134 hepatoma can be ascribed to TNF produced not only by monocytes but also by tumor cells themselves in the tumor tissue.
Animals↗
Secretion of tumor necrosis factor during fetal and neonatal development of the mouse: ontogenic inflammation.
Inflammation plays an important role in homeostasis of the body. We therefore can assume that an inflammatory state occurs during ontogenesis of animals. To address this problem, we examined the ability of tumor necrosis factor (TNF), one of the inflammatory mediators, to be secreted by mouse cells during development. We cultured cells prepared from various parts of fetuses (10-19 days of gestation) and postnatal brains by collagenase digestion and assayed the secreted TNF activity by the L-929 cytotoxicity test. We found TNF activity by fetal cells without any stimulation. The spontaneous secretion of TNF was relatively high at around 13-15 days of gestation. The secretion was enhanced by lipopolysaccharide (LPS), showing that fetal cells are in an activated state for TNF secretion. These TNF activities were neutralized completely by rabbit anti-murine TNF antibody. Spontaneous and LPS-enhanced secretion by postnatal brain cells reached a peak around 7 days after birth, and thereafter declined rapidly. This time course was well correlated to the increase in the weight of brain. The producing cells were negative in macrophage marker surface antigen, and heterogeneous in relation to adherence and phagocytic activity, showing that TNF is secreted by various types of cells in the fetal body. These results suggest the presence of an inflammation-like state during ontogenesis. We consider that this "ontogenic inflammation" may be the prototype of inflammation, which can regulate homeostasis of the adult body.
Animals↗
Production in Escherichia coli of human thymosin beta 4 as chimeric protein with human tumor necrosis factor.
We have produced thymosin beta 4 protein in Escherichia coli as a chimeric protein with tumor necrosis factor (TNF). The protein was abundantly expressed, was immunoreactive against both anti-thymosin beta 4 and anti-TNF antibodies, and retained cytotoxicity in a TNF assay using mouse L929 fibroblasts. This latter characteristic enabled the easy and simple purification of thymosin beta 4 merely by following the TNF activity. The chimeric protein was designed to have an Asp-Pro bridge between thymosin beta 4 and TNF which could be specifically cleaved under suitable acidic conditions to release the thymosin beta 4 from the chimeric protein. These results indicate that the expression system in E.coli of a chimeric protein composed of thymosin beta 4 and TNF is appropriate for obtaining an abundant amount of the beta 4 peptide, especially since its purification from tissues is usually difficult because of limited yield and obscurity of its biological activity.
Amino Acid Sequence↗
Exogenous and endogenous tumour necrosis factor therapy.
Although tumour necrosis factor (TNF) is known as a tumoricidal cytokine, it is also important in maintaining homoeostasis in living organisms by inducing an inflammation-like state. We consider that this monokine may also regulate ontogenesis in animals, because we found that a considerable amount of TNF is secreted by mouse embryonal cells at various stages of development. We named the inflammation-like state in which the TNF concentration was high 'ontogenic inflammation'. Induction of endogenous TNF in the adult body as in ontogenic inflammation should restore homoeostasis in patients suffering from chronic diseases including tumours. We have established several methods for inducing sufficient endogenous TNF to cause tumour regression in patients. For more efficient tumour therapy by ontogenic inflammation, the localization of TNF around tumour lesions was found to be critical. By using a newly constructed rTNF-S, which, like endogenous TNF, has a broader cytotoxic spectrum and fewer side effects than conventional rTNF, we developed a new therapy--exogenous/endogenous TNF therapy (EET therapy). With this therapy, exogenous and endogenous TNF can be targeted to and localized in tumours where they act synergistically. Our results suggest that ontogenic inflammation may be important, even in adults, for maintaining homoeostasis. The appropriate application of ontogenic inflammation should also be effective in treating other diseases, such as rheumatism.
Animals↗
Antitumor effect of systemic administration of novel recombinant tumor necrosis factor (rTNF-S) with less toxicity than conventional rTNF-alpha in vivo.
This study reported that acute toxicity (expressed as the LD50 value) of rTNF-SAM1 and rTNF-SAM2 which we constructed was considerably lower than that of rTNF-alpha (1). Following this finding, the antitumor effects of systemic administration of these novel recombinant tumor necrosis factors (TNF-S) (named rTNF-SAM1 and rTNF-SAM2) were examined in vivo on murine tumors (Meth A fibrosarcoma, MH134 hepatoma, and B16 melanoma). Both rTNF-SAM1 and rTNF-SAM2 could be administered systemically to tumor-bearing mice at doses up to 3 X 10(4) to 10(5) U. Growth of all tumors was significantly inhibited by systemic administration of rTNF-SAM1 or rTNF-SAM2 at a dose of greater than 3 X 10(4) U; at this dosage conventional rTNF-alpha could not be administered systemically to any of the mice strains due to its toxicity. These results confirm our previous finding that the rTNF-SAM group can be administered with safety at higher doses than rTNF-alpha, and revealed that these novel rTNF-S could be more promising antitumor drugs than rTNF-alpha in view of their lower toxicity compared with antitumor effect.
Animals↗
Primary structure of two mRNAs encoding putative salmon alpha-subunits of pituitary glycoprotein hormone.
1. By random sequence analyses, we isolated from the cDNA library of salmon pituitary glands two clones, the deduced amino acid sequences corresponding to the C-terminal region of which are almost the same as those of the alpha subunits of mammalian glycoprotein hormones. 2. Comparison of the nucleotide sequences and deduced amino acid sequences from these two clones with those of mammalian species revealed that the two newly-isolated cDNAs corresponded to mRNAs encoding the putative salmon pre-alpha subunit of glycoprotein hormones. 3. Homology in the nucleotide sequences of these two clones suggested that corresponding mRNAs may be encoded by separate genes which probably evolved from a common ancestral gene.
Amino Acid Sequence↗
Biological activities of novel recombinant tumor necrosis factor having N-terminal amino acid sequences derived from cytotoxic factors produced by THP-1 cells.
Eight species of novel recombinant tumor necrosis factor-S (rTNF-SAM group) were constructed in which N-terminal amino acid sequences were based on that of TNF-S from THP-1 cells with higher basicity than conventional rTNF-alpha. Two of this rTNF-SAM group, denoted as rTNF-SAM1 and rTNF-SAM2, showed more cytocidal activity on A549 lung carcinoma cells and G401 Wilm's tumor cells than did rTNF-alpha. In addition to these cell lines, rTNF-SAM1 revealed strong cytocidal activity on T24 bladder carcinoma cells, which are resistant to rTNF-alpha. Moreover, possible cachectin activity of rTNF-SAM2 seemed to be lower than that of conventional rTNF-alpha, suggesting that rTNF-SAM2 has less side effects. Actually, toxicity as expressed by LD50 value of rTNF-SAM2 as well as others of the rTNF-SAM group was significantly lower than that of conventional rTNF-alpha. Thus, newly constructed rTNF-SAM1 and rTNF-SAM2 should be more promising antitumor reagents for clinical use, since they were shown to be superior to conventional rTNF-alpha both in antitumor effect and in less side effects.
Amino Acid Sequence↗
TNF induces endogenous TNF in vivo: the basis of EET therapy as a combination of rTNF together with endogenous TNF.
Enough amounts of tumor necrosis factor (TNF) in mice serum for the therapy were observed by treatment with 100 units of recombinant human TNF-alpha (rHuTNF-alpha) followed by administration of OK-432 (a streptococcal preparation). Optimal time interval between rTNF and OK-432 to produce endogenous TNF was 3 h, and priming activity of rTNF persisted for at least 10 h. The same effect was observed using novel human recombinant TNF-SAM2 (rHuTNF-SAM2) developed by our group. Production of endogenous TNF using rTNF-alpha or rTNF-SAM2 as a priming reagent was almost equal among various mice strains. Induced TNF in mice serum was completely neutralized by anti-MuTNF antiserum, but not by anti-HuTNF monoclonal antibody. rMuTNF could also induce the priming state; however, the dose-response kinetics of the priming effect to produce endogenous TNF was different between rHuTNFs and rMuTNF-alpha, suggesting species specificity among rTNFs used. The therapeutic effect against Meth A and MH134 tumors in mice treated by rHuTNFs in combination with OK-432 was superior to that by single administration of either OK-432 or rHuTNFs or by successive administrations of OK-432. Especially, the antitumor effect against MH134 hepatoma was superior to that of any other treatment using known biological response modifiers so far experienced. These results suggest that such combination antitumor therapy as rTNF together with OK-432 should be applicable to cancer patients.
Animals↗
Improvement of cytotoxicity of tumor necrosis factor (TNF) by increase in basicity of its N-terminal region.
Two new recombinant TNFs (named rTNF-Scw1 and -Scw2) with higher basicity than conventional recombinant human TNF-alpha (rTNF-alpha) in the N-terminal region were constructed. Their sequences were constructed based on those of partially purified cytotoxic factors from the culture supernatant of acute monocytic leukemia cells THP-1, which unlike rTNF-alpha are cytotoxic to T24 bladder carcinoma cells in vitro. These new rTNF-Ss showed a broader cytotoxicity to tumor cells than rTNF-alpha. This increase in the basicity of the N-terminal region over that of conventional TNF significantly increased the cytotoxicity on tumor cells in vivo as well as in vitro.
Amino Acid Sequence↗
Endogenous production of tumor necrosis factor in normal mice and human cancer patients by interferons and other cytokines combined with biological response modifiers of bacterial origin.
The priming effect of endogenous biological response modifiers (BRMs), interferons (IFNs), and interleukin-2 (IL2), and the triggering effect of BRMs of bacterial origin, OK-432 and Corynebacterium parvum, on endogenous production of the tumor necrosis factor (TNF) were investigated in mice. TNF activity in serum was measured by in vitro cytotoxicity assay with L-929 cells as a target. The i.v. injection of OK-432 (3KE per mouse) triggered TNF maximally (mean value: 30 U/ml) after 2 h, with a similar time course to that of triggering by lipopolysaccharide. The priming activities of IFNs and IL2 were examined in the system of TNF-triggering by OK-432. The i.v. injection of recombinant IFN-gamma (rIFN-gamma, 10(4) U per mouse) increased TNF production to 790 U/ml; this priming effect was observed just after its injection, was maximal after 2 to 6 h, and disappeared after 24 h. Other types of interferon, rIFN-alpha A/D(Bgl) (2 X 10(5) U per mouse), rIFN-beta (10(6) U per mouse), and natural IFN-alpha/beta (10(6) U per mouse) showed maximal priming activity 6 h after their injection (200 to 800 U/ml) but no effect just after their injection. Recombinant IL2 (10(6) U per mouse) had priming activity that showed a similar time course to that of interferons other than IFN-gamma (a maximal TNF production: 100 U/ml). The i.v. injection of C. parvum, like OK-432, triggered TNF production at doses of 0.06 and 0.3 mg per mouse 2 h after its injection and the triggered TNF activity was enhanced by rIFN-gamma. These findings suggest that combinations of the above endogenous BRMs as priming agents and OK-432 or C. parvum as a triggering agent could induce endogenous production of TNF even in human cancer patients. In fact, combined administration of rIFN-gamma and OK-432 produced TNF in human cancer patients. The advantage of this method for treatment of human cancer patients is discussed.
Animals↗
[Partial response of lung metastases from a renal cancer treated with endogenous tumor necrosis factor].
The strong tumor-selective cytocidal action of tumor necrosis factor (TNF) has been observed in vitro and in vivo. Sato et al. have reported that it was possible to induce a primed state of TNF triggering by injection of purified protein derivative (PPD) even a long time after BCG sensitization, suggesting that this treatment could be applied to human patients. In the present study, we achieved a partial response of a metastatic lesion in a patient with renal cancer by the induction of endogenous TNF by PPD and OK-432 (a streptococcal preparation). This study suggested the possible application of this therapy to also patients with malignant tumor which are highly resistant to any conventional antitumor therapy.
Biological Products↗
Detection of a countertranscript in promyelocytic leukemia cells HL60 during early differentiation by TPA.
We have isolated several cDNA clones corresponding to the mRNAs expressed during early phase of differentiation of promyelocytic leukemia cells HL60 by TPA. Two interrelated clones were examined, one pHH81 and the other pHH58 possessing inserts of 180 and 730 base pairs, respectively. Northern blot analyses of poly(A) RNAs from induced cells revealed that the clone pHH81 hybridized with 4.3kb RNA, while the clone pHH58 hybridized with 4.3kb and, in addition 0.7kb RNA. Sequence determination of those cDNA clones and extensive Northern blot analyses revealed that the inserts of these clones were derived from 4.3kb mRNAs. 0.7kb RNA was hybridized with only 5' upstream region of the clone pHH58, especially with the strand designed to detect anti-sense RNA. Thus we concluded that 0.7kb RNA is a countertranscript of 4.3kb RNA expressed during early differentiation of HL60 cells.
Base Sequence↗
Purification of rabbit tumor necrosis factor.
Rabbit tumor necrosis factor (TNF) was purified and shown by SDS-PAGE to be a single protein of 18 kDa. TNF in 355 ml rabbit serum was precipitated with ammonium sulfate, and purified by repeated DEAE-Sephadex and Sephacryl S-200 chromatographies, and the final fractionation on Blue-Sepharose 6B. By this procedure its yield was 22% and its specific activity was 2.4 X 10(7) U/mg protein. The sequence of the N-terminal 20 amino acids was determined.
Amino Acid Sequence↗
Tissue-specific regulation of a chicken delta-crystallin gene in mouse cells: involvement of the 5' end region.
A cloned delta-crystallin gene of the chicken is preferentially expressed in lens cells after introduction into various mouse tissues. The level of expression in the lens epithelium is 20 times higher than in fibroblasts. Taking advantage of this system, we attempted to define regulatory regions of the delta-crystallin gene using a variety of deletion and substitution mutants. The results indicate that tissue-specific regulation of the delta-crystallin gene is mediated by the 5' end region of the gene; sequences upstream from -93 are not required for expression and sequences downstream from +58 are not involved in tissue specificity. The high expression in lens cells requires 5' flanking sequences of 80-bp long from the cap site, whereas the low expression in fibroblasts requires an additional 12 bp upstream sequence. Expression of both types is lost in a mutant with only 51 bp of the 5' flanking sequence. Thus, fine deletion analysis demonstrated that expression in lens cells and expression in fibroblasts are distinct not only in level but in regulation.
Amino Acid Sequence↗