scholarly journals Recombinant immunotoxins containing anti-Tac(Fv) and derivatives of Pseudomonas exotoxin produce complete regression in mice of an interleukin-2 receptor-expressing human carcinoma

Blood ◽  
1994 ◽  
Vol 83 (2) ◽  
pp. 426-434 ◽  
Author(s):  
RJ Kreitman ◽  
P Bailon ◽  
VK Chaudhary ◽  
DJ FitzGerald ◽  
I Pastan
Keyword(s):  
Interleukin 2 ◽  
Target Cells ◽  
Complete Regression ◽  
Single Chain ◽  
Pseudomonas Exotoxin ◽  
Gene Encoding ◽  
Human Carcinoma ◽  
Interleukin 2 Receptor ◽  
Variable Domains

Anti-Tac(Fv)-PE40 is a recombinant single-chain immunotoxin composed of the variable domains of the monoclonal antibody anti-Tac, which binds to the p55 subunit of the interleukin-2 receptor (IL-2R), and a truncated form of Pseudomonas exotoxin (PE), which does not bind to the PE receptor (Chaudhary et al, Nature 339:394, 1989). Whereas its cytotoxic activity toward autoimmune and malignant target cells has been established, its efficacy in vivo remains unknown. To establish an animal model, we produced ATAC-4 cells by transfecting the gene encoding the low-affinity IL-2R (p55) into A431 epidermoid carcinoma cells. ATAC-4 cells contained low-affinity IL-2Rs (2 x 10(5)/cell) and formed tumors in nude mice. In tissue culture, protein synthesis in ATAC-4 cells was inhibited 50% (IC50) at 0.06 ng/mL (0.9 pmol/L) of anti-Tac(Fv)-PE40. IC50s for the derivatives anti-Tac(Fv)-PE38, which is missing PE amino acids 365–380, and anti-Tac(Fv)-PE38KDEL, which contains the same deletion plus the KDEL carboxyl terminus, were 0.04 and 0.025 ng/mL, respectively. All the agents produced complete tumor regressions in ATAC-4 tumor-bearing mice and anti-Tac(Fv)-PE38KDEL had significant antitumor activity at 1% of the LD50. The dose limiting toxicity of anti-Tac(Fv)-PE38KDEL was from hemorrhagic liver necrosis, which was observed at approximately 55% of the LD50.

scholarly journals Recombinant immunotoxins containing anti-Tac(Fv) and derivatives of Pseudomonas exotoxin produce complete regression in mice of an interleukin-2 receptor-expressing human carcinoma

Blood ◽  
1994 ◽  
Vol 83 (2) ◽  
pp. 426-434 ◽  
Author(s):  
RJ Kreitman ◽  
P Bailon ◽  
VK Chaudhary ◽  
DJ FitzGerald ◽  
I Pastan
Keyword(s):  
Interleukin 2 ◽  
Target Cells ◽  
Complete Regression ◽  
Single Chain ◽  
Pseudomonas Exotoxin ◽  
Gene Encoding ◽  
Human Carcinoma ◽  
Interleukin 2 Receptor ◽  
Variable Domains

Abstract Anti-Tac(Fv)-PE40 is a recombinant single-chain immunotoxin composed of the variable domains of the monoclonal antibody anti-Tac, which binds to the p55 subunit of the interleukin-2 receptor (IL-2R), and a truncated form of Pseudomonas exotoxin (PE), which does not bind to the PE receptor (Chaudhary et al, Nature 339:394, 1989). Whereas its cytotoxic activity toward autoimmune and malignant target cells has been established, its efficacy in vivo remains unknown. To establish an animal model, we produced ATAC-4 cells by transfecting the gene encoding the low-affinity IL-2R (p55) into A431 epidermoid carcinoma cells. ATAC-4 cells contained low-affinity IL-2Rs (2 x 10(5)/cell) and formed tumors in nude mice. In tissue culture, protein synthesis in ATAC-4 cells was inhibited 50% (IC50) at 0.06 ng/mL (0.9 pmol/L) of anti-Tac(Fv)-PE40. IC50s for the derivatives anti-Tac(Fv)-PE38, which is missing PE amino acids 365–380, and anti-Tac(Fv)-PE38KDEL, which contains the same deletion plus the KDEL carboxyl terminus, were 0.04 and 0.025 ng/mL, respectively. All the agents produced complete tumor regressions in ATAC-4 tumor-bearing mice and anti-Tac(Fv)-PE38KDEL had significant antitumor activity at 1% of the LD50. The dose limiting toxicity of anti-Tac(Fv)-PE38KDEL was from hemorrhagic liver necrosis, which was observed at approximately 55% of the LD50.

In vitro and in vivo effects of BT563, an anti-interleukin-2 receptor monoclonal antibody

1994 ◽  
Vol 7 (s1) ◽  
pp. 556-558 ◽  
Author(s):  
T. VanGelder ◽  
C. R. Daane ◽  
L. M. B. Vaessen ◽  
C. J. Hesse ◽  
W. Weimar ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Interleukin 2 ◽  
Interleukin 2 Receptor ◽  
In Vivo Effects ◽  
Receptor Monoclonal Antibody

scholarly journals Regulation of the growth and functions of cloned murine large granular lymphocyte lines by resident macrophages.

1985 ◽  
Vol 162 (4) ◽  
pp. 1161-1181 ◽  
Author(s):  
N Minato ◽  
T Amagai ◽  
J Yodoi ◽  
T Diamanstein ◽  
S Kano
Keyword(s):  
Bone Marrow Cells ◽  
Interleukin 2 ◽  
Functional Expression ◽  
Suppressive Effect ◽  
Leukemic Cells ◽  
Target Cells ◽  
Significant Heterogeneity

Using cloned lines with the morphology of large granular lymphocytes (LGL) from BALB/c mice, we studied the exact requirements for proliferation and their functional characteristics, as well as their regulation. Although these cloned LGL lines were interleukin 2 (IL-2) dependent for growth, experiments using human recombinant IL-2 (rIL-2), known to be active on murine cells, indicated that IL-2 was a necessary but not sufficient factor. Coexistance of normal macrophages in addition to rIL-2 was found to support continuous proliferation of cloned LGL in vitro. This role of macrophages could be replaced by partially purified IL-1 derived from macrophage-conditioned medium. An IL-2 binding assay using 125I-rIL-2 suggested that the role of normal macrophages was to selectively induce and/or maintain high affinity IL-2 receptors (IL-2R) (Kd, 0.2-0.5 nM) without affecting low affinity ones (Kd, 10-30 nM). Functional studies indicated that most of the LGL clones killed various combinations of representative groups of natural killer (NK)-susceptible target cells, including leukemic cells (YAC-1, RL male 1), virus-infected cells (HeLa-measles, HeLa-herpes simplex virus), and normal bone marrow cells (BMC), whereas none of them affected any of NK-resistant target cells, including uninfected HeLa cells. Some of these clones also suppressed in vitro hematopoiesis. Such characteristic cytotoxic spectra, as well as serological phenotypes (Thy-1+, Lyt-1-2-, asialo GM1-positive, T200+, TdT-, Fc receptor-positive) indicated that these LGL clones exactly represent endogenous NK cells, rather than a variety of anomalous killer cells generated in various culture conditions. Although there was significant heterogeneity of cytotoxic spectrum among LGL clones, no clonotypic distribution of specificities was observed. Normal macrophages were found to modulate the functional expression of LGL clones. They augmented the cytotoxic potential of the clones against leukemic and virus-infected targets, but suppressed intrinsic reactivity against normal BMC. Similarly, LGL clones maintained with macrophages showed much less suppressive effect on in vitro hematopoiesis. The present observations on the interaction of cloned LGL and normal macrophages provide a basic explanation for the mechanisms by which the immediate responsiveness to IL-2 of the NK effector system, without exogenous stimulation, and the functional selectivity toward abnormal rather than normal cells, are actively maintained in vivo.

Chimeric Fv-ζ or Fv-ε receptors are not sufficient to induce activation or cytokine production in peripheral T cells

Blood ◽  
2000 ◽  
Vol 96 (5) ◽  
pp. 1999-2001 ◽  
Author(s):  
Thomas Brocker
Keyword(s):  
T Cells ◽  
Cytokine Production ◽  
Interleukin 2 ◽  
Tumor Rejection ◽  
Adoptive T Cell Therapy ◽  
Chimeric Antibody ◽  
Single Chain ◽  
T Cell Therapy ◽  
Chimeric Receptors

In current clinical trials, chimeric antibody-like receptors fused to signaling domains derived from TCR-ζ or Fc(ε)RIγ-chain are tested for their ability to lyse tumor cells in vivo. In this study, the function of primary T cells expressing such receptors has been investigated in transgenic mice. These receptors cannot induce proliferation of resting T cells or trigger the production of optimal amounts of cytokines. It is further demonstrated that an initial low presence of cytokine message and protein is disappearing rather fast, whereas the triggering of endogenous TCR/CD3 in the same cells leads to normal prolonged cytokine production. The direct clinical relevance of these findings is further underlined by the increased in vivo tumor rejection by T cells expressing chimeric receptors in presence of exogenous interleukin-2. Therefore, adoptive T-cell therapy using primary T cells transfected with single chain receptors might benefit substantially from the accompanying administration of cytokines.

scholarly journals Binding of HMG-I(Y) Imparts Architectural Specificity to a Positioned Nucleosome on the Promoter of the Human Interleukin-2 Receptor α Gene

2000 ◽  
Vol 20 (13) ◽  
pp. 4666-4679 ◽  
Author(s):  
R. Reeves ◽  
W. J. Leonard ◽  
M. S. Nissen
Keyword(s):  
T Cell ◽  
Transcriptional Activation ◽  
Interleukin 2 ◽  
Proximal Promoter ◽  
Cell Mediated Immunity ◽  
Core Particle ◽  
The Core ◽  
Interleukin 2 Receptor

ABSTRACT Transcriptional induction of the interleukin-2 receptor alpha-chain (IL-2Rα) gene is a key event regulating T-cell-mediated immunity in mammals. In vivo, the T-cell-restricted protein Elf-1 and the general architectural transcription factor HMG-I(Y) cooperate in transcriptional regulation of the human IL-2Rα gene by binding to a specific positive regulatory region (PRRII) in its proximal promoter. Employing chromatin reconstitution analyses, we demonstrate that the binding sites for both HMG-I(Y) and Elf-1 in the PRRII element are incorporated into a strongly positioned nucleosome in vitro. A variety of analytical techniques was used to determine that a stable core particle is positioned over most of the PRRII element and that this nucleosome exhibits only a limited amount of lateral translational mobility. Regardless of its translational setting, the in vitro position of the nucleosome is such that DNA recognition sequences for both HMG-I(Y) and Elf-1 are located on the surface of the core particle. Restriction nuclease accessibility analyses indicate that a similarly positioned nucleosome also exists on the PRRII element in unstimulated lymphocytes when the IL-2Rα gene is silent and suggest that this core particle is remodeled following transcriptional activation of the gene in vivo. In vitro experiments employing the chemical cleavage reagent 1,10-phenanthroline copper (II) covalently attached to its C-terminal end demonstrate that HMG-I(Y) protein binds to the positioned PRRII nucleosome in a direction-specific manner, thus imparting a distinct architectural configuration to the core particle. Together, these findings suggest a role for the HMG-I(Y) protein in assisting the remodeling of a critically positioned nucleosome on the PRRII promoter element during IL-2Rα transcriptional activation in lymphocytes in vivo.

scholarly journals Neutralizationof Enteric Coronaviruses with Escherichia coli CellsExpressing Single-Chain Fv-AutotransporterFusions

2003 ◽  
Vol 77 (24) ◽  
pp. 13396-13398 ◽  
Author(s):  
Esteban Veiga ◽  
Víctor de Lorenzo ◽  
Luis Angel Fernández
Keyword(s):  
Escherichia Coli ◽  
Infectious Agent ◽  
Target Cells ◽  
Single Chain ◽  
E Coli ◽  
Single Chain Antibodies ◽  
Single Chain Fv ◽  
Transmissible Gastroenteritis ◽  
Iga Protease

ABSTRACT We report here that fusions of single-chain antibodies (scFvs) to the autotransporter β domain of the IgA protease of Neisseria gonorrhoeae are instrumental in locating virus-neutralizing activity on the cell surface of Escherichia coli. E. coli cells displaying scFvs against the transmissible gastroenteritis coronavirus on their surface blocked in vivo the access of the infectious agent to cultured epithelial cells. This result raises prospects for antiviral strategies aimed at hindering the entry into target cells by bacteria that naturally colonize the same intestinal niches.

scholarly journals Endogenous IL-2 in Cancer Cells: A Marker of Cellular Proliferation

1998 ◽  
Vol 46 (5) ◽  
pp. 603-611 ◽  
Author(s):  
Torsten E. Reichert ◽  
Simon Watkins ◽  
Joanna Stanson ◽  
Jonas T. Johnson ◽  
Theresa L. Whiteside
Keyword(s):  
Cell Cycle ◽  
Tumor Cells ◽  
Cell Lines ◽  
Cellular Proliferation ◽  
Histological Grade ◽  
Interleukin 2 ◽  
Ki 67 ◽  
Human Carcinoma

We have previously demonstrated that interleukin-2 (IL-2) receptors, IL-2 protein, and mRNA for IL-2 are present in human carcinomas in vitro and in vivo. Carcinoma cells synchronized in the G2/M-phase of the cell cycle express significantly more intracytoplasmic IL-2 as well as IL-2R-β and -γ than tumor cells in the G0/G1-phase. Here we evaluated immunohistologically the cell cycle-dependent distribution of the proliferation-associated Ki-67 antigen and expression of the cytokine IL-2 in four different carcinoma cell lines. In addition, 34 tissue samples from patients with squamous cell carcinomas of the head and neck were simultaneously analyzed for Ki-67 and IL-2 expression and the data were correlated to the histological grade of the tumors. All tumor cell lines were shown to express IL-2 in the Golgi complex. The strongest IL-2 expression was seen in tumor cells undergoing mitosis, identified by double staining with the antibody to Ki-67. In the tumor tissue, the highest level of co-expression of IL-2 and Ki-67 was observed in poorly differentiated carcinomas, with a labeling index (LI) of 67.2% for IL-2 and 68.8% for Ki-67. Well-differentiated carcinomas showed a significantly lower expression of both proteins (LI 35.0% for IL-2 and 26.5% for Ki-67). The correlation between the labeling indices was statistically significant ( r = 0.747; p<0.001). These results demonstrate that IL-2 expression in human carcinoma tissues is strongly associated with cell proliferation and significantly correlates with the histological tumor grade.

scholarly journals Homeostatic action of interleukin-4 on endogenous and recombinant interleukin-2-induced activated killer cell function

Blood ◽  
1991 ◽  
Vol 77 (6) ◽  
pp. 1283-1289
Author(s):  
C Bello-Fernandez ◽  
C Bird ◽  
HE Heslop ◽  
DJ Gottlieb ◽  
JE Reittie ◽  
...  
Keyword(s):  
Tumor Necrosis Factor ◽  
Marrow Transplantation ◽  
Tumor Necrosis ◽  
Cell Function ◽  
Interleukin 2 ◽  
Gamma Interferon ◽  
Target Cells ◽  
Recombinant Interleukin 2 ◽  
Necrosis Factor

Cytokine-secreting, major histocompatibility complex-unrestricted activated killer (AK) cells are toxic to a wide range of virus-infected or malignant target cells and may be generated endogenously, eg, after bone marrow transplantation, or by infusion of cytokines such as recombinant interleukin-2 (rIL-2). Although AK cells secrete cytokines such as gamma-interferon and tumor necrosis factor, which are themselves able to recruit fresh cytokine-secreting AK cells, activation in both settings is short-lived, implying the existence of homeostatic regulatory mechanisms. We now demonstrate one mechanism by which rapid homeostasis is achieved. We show that IL-4 is produced in patients with both endogenously and exogenously generated AK cells. The cytokine was detected in serum after marrow transplantation, and IL-4 transcripts appeared in circulating lymphocytes during rIL-2 infusion. Although IL-4 inhibited the induction phase of AK cell function, it had no significant inhibitory effect on the ability of AK cells from these individuals to respond to restimulation. Nonetheless, neutralization of the IL-4 induced during cell activation doubled the half-life of AK function, once activating stimuli were removed, from 18 to 44 hours and produced a 2-log increase in AK cell secretion of tumor necrosis factor and gamma-interferon. These data suggest that IL-4 induced in vivo during lymphocyte activation abbreviates AK cell responses once the triggering stimuli have been removed. Neutralization of endogenous IL-4 in vivo by appropriate monoclonal antibodies might prolong the duration of AK function.

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