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1.
Clin Pharmacol Ther ; 65(6): 615-29, 1999 Jun.
Article in English | MEDLINE | ID: mdl-10391667

ABSTRACT

BACKGROUND: Interleukin-12 (IL-12) is a cytokine that promotes type-1 helper T-cell responses and may have therapeutic utility in the treatment of cancer, asthma, and a variety of infectious diseases. METHODS: In a phase I trial, recombinant human IL-12 (rHuIL-12) was administered subcutaneously once a week at a fixed dose of 0.1 to 1.0 microg/kg to 24 patients with renal cell carcinoma. A similar study was later performed in mice to evaluate the mechanism of down-regulation of pharmacokinetic-pharmacodynamic response observed in patients with cancer. RESULTS: Adverse events, serum IL-12 levels, and serum levels of interferon-gamma (IFN-gamma) and interleukin-10 (IL-10) produced in response to IL- 12 were all maximum in the week after the first dose of rHuIL-12 and decreased after long-term administration. Similar to these results, repetitive subcutaneous administration of recombinant mouse IL-12 (rMoIL-12) to normal mice led to down-regulation of serum levels of IL-12 and IFN-gamma measured 5 hours after rMoIL-12 injection. Down-regulation of IL-12 serum levels was inversely correlated with the up-regulation of IL-12 receptor expression and may be the result of increased clearance of rMoIL-12 from serum by binding to lymphoid cells expressing increased amounts of IL-12 receptor. The down-regulation of serum IFN-gamma levels correlated with decreased IFN-gamma messenger ribonucleic acid expression and may result from feedback inhibition of IL-12 signaling or from a more specific inhibition of IFN-gamma synthesis. CONCLUSION: Administration of rHuIL-12 in fixed weekly doses resulted in decreased serum levels of IL-12 and of IFN-gamma, a secondary cytokine believed to be critical to response of IL-12. A better understanding of the complex regulation of the pharmacokinetic-pharmacodynamic response to IL-12 should facilitate the development of more effective dosing regimens for its use in the clinic.


Subject(s)
Adjuvants, Immunologic/pharmacology , Carcinoma, Renal Cell/drug therapy , Gene Expression Regulation, Neoplastic , Interleukin-12/pharmacology , Kidney Neoplasms/drug therapy , Adjuvants, Immunologic/administration & dosage , Adjuvants, Immunologic/adverse effects , Adjuvants, Immunologic/pharmacokinetics , Adult , Aged , Animals , Carcinoma, Renal Cell/blood , Down-Regulation , Drug Administration Schedule , Female , Humans , Interferon-gamma/blood , Interferon-gamma/genetics , Interleukin-12/administration & dosage , Interleukin-12/adverse effects , Interleukin-12/blood , Interleukin-12/pharmacokinetics , Kidney Neoplasms/blood , Male , Mice , Mice, Inbred C57BL , Middle Aged , RNA, Messenger/analysis , Recombinant Proteins/pharmacology , beta 2-Microglobulin/metabolism
2.
Ann N Y Acad Sci ; 795: 1-12, 1996 Oct 31.
Article in English | MEDLINE | ID: mdl-8958912

ABSTRACT

Mo(p40)2 is a potent IL-12 antagonist that interacts strongly with the beta 1 subunit of the IL-12R to block binding of moIL-12 to the high-affinity mouse IL-12R. Mo(p40)2, alone or in synergy with the 2B5 mAb specific for the moIL-12 heterodimer, blocked IL-12-induced responses in vitro, Mo(p40)2 was thus used alone or with 2B5 mAb to examine the role of IL-12 in vivo, Mo(p40)2 caused a dose-dependent inhibition of both the rise in serum IFN-gamma levels in mice injected with endotoxin and the Th1-like response to immunization with KLH. Treatment with mo(p40)2 plus 2B5 anti-moIL-12 mAb also suppressed DTH responses to methylated bovine serum albumin but not specific allogeneic CTL responses in vivo. In each of these models, responses seen in mice treated with mo(p40)2 +/- 2B5 anti-moIL-12 mAb were similar to those observed in IL-12 knockout mice. Thus, mo(p40)2 can act as a potent IL-12 antagonist in vivo, as well as in vitro, and is currently being used to investigate the role of IL-12 in the pathogenesis of some Th1-associated autoimmune disorders in mice.


Subject(s)
Interleukin-12/antagonists & inhibitors , Animals , Antibodies, Monoclonal/immunology , CHO Cells , Cricetinae , Cytotoxicity, Immunologic , Hypersensitivity, Delayed/immunology , Interferon-gamma/biosynthesis , Interleukin-12/chemistry , Lymph Nodes/cytology , Lymph Nodes/immunology , Lymphocyte Activation/drug effects , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Molecular Weight , Receptors, Interleukin/metabolism , Receptors, Interleukin-12 , Recombinant Proteins , Structure-Activity Relationship , Th1 Cells/immunology
3.
Immunity ; 4(5): 471-81, 1996 May.
Article in English | MEDLINE | ID: mdl-8630732

ABSTRACT

IL-12 is a cytokine that can exert regulatory effects on T and NK cells and promote Th1 responses. To delineate further the physiologic role of IL-12 in immunity, mice deficient for this cytokine were generated. IL-12-deficient mice were impaired but not completely lacking in the ability to produce IFN gamma following endotoxin administration and to mount a Th1 response in vivo, as measured by antigen-induced IFN gamma secretion by immune lymph node cells in vitro. In contrast, secretion of IL-4 was enhanced, while proliferation and secretion of IL-2 and IL-10 were normal following antigen stimulation. DTH responses were significantly reduced in IL-12-deficient mice, but no defect in allogeneic CTL responses was observed. These results indicate that IL-12 plays an essential role in regulating IFN gamma production and in facilitating normal DTH responses. However, other phenomena associated with Th1 responses and cell-mediated immunity, i.e., IL-2 secretion and CTL generation, were not compromised in the absence of IL-12.


Subject(s)
Interferon-gamma/biosynthesis , Interleukin-12/deficiency , Interleukin-12/genetics , Lymphokines/biosynthesis , Mice, Mutant Strains/immunology , Animals , Base Sequence , Genetic Vectors/immunology , Killer Cells, Natural/immunology , Lymphocyte Activation/genetics , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Mutant Strains/growth & development , Mice, Mutant Strains/metabolism , Molecular Sequence Data , Th1 Cells/immunology , Th1 Cells/metabolism
4.
Eur J Immunol ; 26(2): 345-50, 1996 Feb.
Article in English | MEDLINE | ID: mdl-8617302

ABSTRACT

We previously described the cloning of a cDNA encoding an interleukin-12 receptor (IL-12R) subunit, designated beta, that bound IL-12 with low affinity when expressed in COS cells. We now report that a pair of monoclonal antibodies (mAb), 2B10 and 2.4E6, directed against different epitopes on the IL-12R beta chain, when used in combination, strongly inhibited IL-12-induced proliferation of activated T cells, IL-12-induced secretion of interferon-gamma by resting peripheral blood mononuclear cells (PBMC), and IL-12-mediated lymphokine-activated killer cell activation. The mAb had no effect on lymphoblast proliferation induced by IL-2, -4, or -7. Thus, the IL-12R beta chain appears to be an essential component of the functional IL-12R on both T and natural killer (NK) cells. We previously observed that high affinity IL-12R were expressed on activated T and NK cells, but not B cells. Studies using flow cytometry and reverse transcription-polymerase chain reaction analysis showed that IL-12R beta chain was expressed on several human T, NK, and (surprisingly) B cell lines, but not on non-lymphohematopoietic cell lines. The Kit225/K6 (T cell) and SKW6.4 (B cell) lines were found to express the greatest amounts of IL-12R beta chain (800-2500 sites/cell); however, Kit225/K6 but not SKW6.4 cells bound IL-12. Similar to SKW6.4 B cells, activated tonsillar B lymphocytes expressed IL-12R beta chain but, consistent with previous results, did not display detectable IL-12 binding. Likewise, up to 72% of resting PBMC from normal volunteer donors expressed IL-12R beta, but did not bind measurable amounts of IL-12. These results indicate that expression of IL-12R beta is essential, but not sufficient, for expression of functional IL-12R. We speculate that expression of functional, high-affinity IL-12R may require the presence of a second subunit that is more restricted in its expression than IL-12R beta.


Subject(s)
Interleukin-12/metabolism , Leukocytes, Mononuclear/chemistry , Receptors, Interleukin/metabolism , Receptors, Interleukin/physiology , Antibodies, Monoclonal/pharmacology , B-Lymphocytes/immunology , Base Sequence , Binding Sites/immunology , Binding, Competitive/immunology , Cell Line , Cell Separation , Humans , Interleukin-12/chemistry , Leukocytes, Mononuclear/immunology , Lymphocyte Activation , Molecular Sequence Data , Palatine Tonsil/cytology , Receptors, Interleukin/immunology , Receptors, Interleukin-12
5.
J Interferon Cytokine Res ; 15(4): 377-83, 1995 Apr.
Article in English | MEDLINE | ID: mdl-7627813

ABSTRACT

Although IL-12 has been reported to synergize with c-kit ligand (KL) in promoting hematopoietic stem cell proliferation in vitro, administration of recombinant mouse IL-12 (rIL-12) to normal mice caused a dose- and time-dependent anemia, leukopenia, and thrombocytopenia in vivo. Decreased numbers of bone marrow cells were recovered from the tibiae of IL-12-treated mice, and histologic examination of the marrow revealed a loss of mature neutrophils and red blood cell precursors. However, simultaneously with the suppression of hematopoiesis in the bone marrow, the IL-12-treated mice developed splenomegaly, which was largely caused by a marked enhancement of splenic extramedullary hematopoiesis of the erythroid, myeloid, and megakaryocytic lineages. These histologic observations were confirmed by colony-forming cell assays in which administration of IL-12 was shown to cause a time-dependent decrease in bone marrow CFU-GM, CFU-E, and BFU-E hematopoietic colony-forming cells while causing an increase in splenic CFU-GM and BFU-E colony-forming cells. All these effects were reversible upon cessation of IL-12 treatment. The observation that in IL-12-treated mice hematopoiesis was suppressed in the marrow but enhanced in the spleen suggests that myelosuppression was not caused by a direct effect of IL-12 on hematopoietic progenitors. It seems likely that myelosuppression was caused instead by an IL-12-induced alteration in the local environment of the marrow.


Subject(s)
Bone Marrow/drug effects , Hematopoiesis, Extramedullary/drug effects , Hematopoiesis/drug effects , Interleukin-12/pharmacology , Spleen/drug effects , Anemia/chemically induced , Animals , Dose-Response Relationship, Drug , Leukopenia/chemically induced , Mice , Mice, Inbred C57BL , Recombinant Proteins/pharmacology , Reference Values , Thrombocytopenia/chemically induced
6.
J Biol Chem ; 270(11): 5864-71, 1995 Mar 17.
Article in English | MEDLINE | ID: mdl-7890716

ABSTRACT

Mouse IL-12 acts on both mouse and human cells; human IL-12 acts only on human cells. This species specificity is determined by the p35 subunit of the IL-12 heterodimer. Since mouse and human p35 sequences are 60% identical, the determinants for the species specificity most likely residue in the nonhomologous sequences of mouse p35. To identify the regions on the p35 subunit interacting with the mouse IL-12 receptor, we constructed a series of chimeric mouse-human p35 molecules by replacing mouse sequences with the nonhomologous human counterparts. An IL-12 heterodimer containing a mouse-human p35 chimera with five residues changed in three discontinuous sites had drastically reduced (750-3000-fold) bioactivities on mouse cells. However, the competitive binding activity of the same mutant IL-12 heterodimer on mouse cells was only reduced 30-fold relative to wild-type IL-12. These findings therefore suggest that 1) the mouse p35 subunit participates in both receptor binding and signaling, 2) the mutations introduced into p35 affect signaling to a much greater extent than receptor binding, and 3) the five residues identified on p35 are required for interacting with the mouse, but not with the human IL-12 receptor and as such contribute extensively to the observed species specificity of IL-12.


Subject(s)
Interleukin-12/chemistry , Interleukin-12/pharmacology , Lymphocyte Activation/drug effects , Amino Acid Sequence , Animals , Binding, Competitive , Cell Line , Chlorocebus aethiops , Humans , Interleukin-12/biosynthesis , Kinetics , Macromolecular Substances , Mice , Molecular Sequence Data , Mutagenesis, Site-Directed , Point Mutation , Protein Structure, Tertiary , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/chemistry , Recombinant Fusion Proteins/pharmacology , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/pharmacology , Sequence Homology, Amino Acid , Transfection
7.
Int Immunol ; 6(1): 157-67, 1994 Jan.
Article in English | MEDLINE | ID: mdl-7908534

ABSTRACT

IL-12 is a heterodimeric cytokine that has been shown to enhance natural killer (NK) and cytotoxic T lymphocyte (CTL) responses, and to induce IFN-gamma production in vitro. In this study, we have examined the effects in vivo of administering purified murine rIL-12 to normal mice. Daily injections of rIL-12 i.p. (1 ng to 10 micrograms/day) caused dose-dependent enhancement of NK cell lytic activity in the spleens and livers of treated mice. Histologic examination of the livers of IL-12-treated mice revealed focal mononuclear cell infiltrates, and flow cytometry studies indicated that the livers of IL-12-treated mice contained increased numbers of NK cells, CD8+ T cells, and monocytes. Liver and splenic lymphoid cells from IL-12-treated mice, unlike liver and splenic lymphoid cells from control mice, spontaneously secreted IFN-gamma in vitro, suggesting that they had been induced by IL-12 to produce IFN-gamma in vivo. Consistent with this, IFN-gamma could be detected in the serum of IL-12-treated mice. In mice which had been immunized by footpad injection of allogeneic splenocytes, daily administration of rIL-12 i.p. was shown to enhance the specific CTL response in the draining lymph nodes. Thus, these studies demonstrate that IL-12 can enhance NK and CTL activity and induce IFN-gamma production in vivo, as well as in vitro, and suggest possible mechanisms by which IL-12 may exert therapeutic effects in the treatment of some tumors and infectious diseases.


Subject(s)
Interferon-gamma/biosynthesis , Interleukins/pharmacology , T-Lymphocytes, Cytotoxic/drug effects , Animals , Cytotoxicity Tests, Immunologic , Immunophenotyping , Interferon-gamma/drug effects , Interleukin-12 , Killer Cells, Natural/drug effects , Liver/drug effects , Lung/drug effects , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Recombinant Proteins/pharmacology , Spleen/drug effects , Tumor Cells, Cultured
8.
Rev. chil. pediatr ; 57(6): 536-41, nov.-dic. 1986. ilus
Article in Spanish | LILACS | ID: lil-40103

ABSTRACT

Se presenta un paciente evaluado en el PRN por presentar sexo ambiguo cuyo estudio hormonal fue normal pero la evaluación citogenética demostró la existencia de una translocación (Y;14) en mosaico: 45,X / 45,X, t (Y;14), posible de explicar por una translocación de media cromátide. Durante una hernioplastía se demostró la existencia de derivados mullerianos y dos gónadas de impresión equívoca que, histológicamente, correspondieron a testículo embrionario y a gónada veteada. Con el diagnóstico de disgenesia gonadal mixta del testículo fue ulteriormente extirpado. Se enfatiza la importancia de los estudios citogenéticos, hormonales e histopatológicos en la evaluación diagnóstica de pacientes con sexo ambiguo


Subject(s)
Infant, Newborn , Humans , Male , Gonadal Dysgenesis/diagnosis , Translocation, Genetic , Disorders of Sex Development , Mosaicism
9.
Rev. chil. pediatr ; 57(2): 164-70, mar.-abr. 1986. ilus, tab
Article in Spanish | LILACS | ID: lil-39802

ABSTRACT

En los últimos años se han delineado tres entidades que involucran trisomía del cromosoma 22 y que comparten ciertas manifestaciones clínicas: el síndrome de ojo de gato, la trisomía parcial y la trisomía total 22, cuya existencia en recién nacidos aún está en controversia. Se describe un caso de trisomía 22 total, por translocación (15;22) "de novo", que falleció en el período de recién nacido con malformaciones pulmonares no descritas, y dos pacientes con una trisomía 22 parcial, (47, XX y XY, + der(22), t(11;22) (q25;q13) mat) debida a una segregación 3:1 en sus madres, quienes son portadoras de una translocación balanceada (11;22). Se discuten los hallazgos clínicos y citogenéticos de los probandos y sus familiares, así como los riesgos de recurrencia de estas trisomías


Subject(s)
Infant, Newborn , Infant , Child, Preschool , Humans , Male , Female , Chromosomes, Human, 21-22 and Y , Trisomy
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