Engineering therapeutic antibodies for improved function

2002 ◽  
Vol 30 (4) ◽  
pp. 487-490 ◽  
Author(s):  
L. G. Presta ◽  
R. L. Shields ◽  
A. K. Namenuk ◽  
K. Hong ◽  
Y. G. Meng
Keyword(s):  
Binding Sites ◽  
Mononuclear Cells ◽  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Fcγ Receptor ◽  
Peripheral Blood Mononuclear ◽  
Adcc Assay ◽  
Human Igg1 ◽  
Improved Function

The binding sites on human IgG1 for human Fcγ receptor (FcγR) I, FcγRIIa, FcγRIIb, FcγRIIIa and neonatal FcR have been mapped. A common set of IgG1 residues is involved in binding to all FcγRs, while FcγRII and FcγRIII utilize distinct sites outside this common set. In addition to residues which abrogated binding to the FcγR, several positions were found which improved binding only to specific FcγRs or simultaneously improved binding to one type of FcγR and reduced binding to another type. Selected IgG1 variants with improved binding to FcγRIIIa were then tested in an in vitro antibody-dependent cellular cytotoxicity (ADCC) assay and showed an enhancement in ADCC when either peripheral blood mononuclear cells or natural killer cells were used.

scholarly journals GM-CSF enhances 3F8 monoclonal antibody-dependent cellular cytotoxicity against human melanoma and neuroblastoma

Blood ◽  
1989 ◽  
Vol 73 (7) ◽  
pp. 1936-1941
Author(s):  
BH Kushner ◽  
NK Cheung
Keyword(s):  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Gm Csf ◽  
Tumor Associated Antigen ◽  
Human Granulocytes ◽  
Adcc Assay ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

3F8 is a murine monoclonal IgG3 antibody specific for the tumor- associated antigen ganglioside GD2. Previous in vitro studies suggest that tumor regressions observed in a phase I clinical trial of 3F8 may be attributable to complement activation by 3F8 and to 3F8-dependent cellular cytotoxicity (ADCC) with lymphocytes. We now describe 3F8- mediated ADCC of GD2-positive tumor targets (melanoma and neuroblastoma) with human granulocytes and report that recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) enhanced this phenomenon. Cytotoxicity required binding of 3F8 to the low-affinity Fc receptor type III (CD16) on the granulocytes and was poor with tumor-binding monoclonal antibodies of other immunoglobulin (ie, non-IgG3) subclasses. GM-CSF (2 to 20 ng/mL) increased ADCC by 93% to 267% at limiting dilutions of 3F8 (1 microgram/mL). With most GD2- positive cell lines tested, this effect translated into a tenfold or greater augmentation in 3F8 efficiency at mediating ADCC. Comparable enhancement occurred whether GM-CSF was present in the ADCC assay or granulocytes were incubated with GM-CSF and washed before the assay. Nonoxidative mechanisms may be important for ADCC since 3F8 mediated ADCC with granulocytes from two children with chronic granulomatous disease; this cytotoxicity was also enhanced by GM-CSF. Since GM-CSF induces a neutrophilia in patients, our data suggest that this cytokine may have the potential of amplifying 3F8 antitumor activity in patients by increasing effector cell numbers and by priming granulocytes for greater cytotoxicity.

scholarly journals GM-CSF enhances 3F8 monoclonal antibody-dependent cellular cytotoxicity against human melanoma and neuroblastoma

Blood ◽  
1989 ◽  
Vol 73 (7) ◽  
pp. 1936-1941 ◽  
Author(s):  
BH Kushner ◽  
NK Cheung
Keyword(s):  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Gm Csf ◽  
Tumor Associated Antigen ◽  
Human Granulocytes ◽  
Adcc Assay ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract 3F8 is a murine monoclonal IgG3 antibody specific for the tumor- associated antigen ganglioside GD2. Previous in vitro studies suggest that tumor regressions observed in a phase I clinical trial of 3F8 may be attributable to complement activation by 3F8 and to 3F8-dependent cellular cytotoxicity (ADCC) with lymphocytes. We now describe 3F8- mediated ADCC of GD2-positive tumor targets (melanoma and neuroblastoma) with human granulocytes and report that recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) enhanced this phenomenon. Cytotoxicity required binding of 3F8 to the low-affinity Fc receptor type III (CD16) on the granulocytes and was poor with tumor-binding monoclonal antibodies of other immunoglobulin (ie, non-IgG3) subclasses. GM-CSF (2 to 20 ng/mL) increased ADCC by 93% to 267% at limiting dilutions of 3F8 (1 microgram/mL). With most GD2- positive cell lines tested, this effect translated into a tenfold or greater augmentation in 3F8 efficiency at mediating ADCC. Comparable enhancement occurred whether GM-CSF was present in the ADCC assay or granulocytes were incubated with GM-CSF and washed before the assay. Nonoxidative mechanisms may be important for ADCC since 3F8 mediated ADCC with granulocytes from two children with chronic granulomatous disease; this cytotoxicity was also enhanced by GM-CSF. Since GM-CSF induces a neutrophilia in patients, our data suggest that this cytokine may have the potential of amplifying 3F8 antitumor activity in patients by increasing effector cell numbers and by priming granulocytes for greater cytotoxicity.

scholarly journals Perforin gene expression in granular lymphocyte proliferative disorders

Blood ◽  
1990 ◽  
Vol 75 (3) ◽  
pp. 704-708 ◽  
Author(s):  
K Oshimi ◽  
Y Shinkai ◽  
K Okumura ◽  
Y Oshimi ◽  
H Mizoguchi
Keyword(s):  
Northern Blot Analysis ◽  
Mononuclear Cells ◽  
Cytolytic Activity ◽  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Peripheral Blood Mononuclear ◽  
Histocompatibility Complex ◽  
Low Levels ◽  
Granular Lymphocytes ◽  
Rule Out

By Northern blot analysis using a cDNA clone of the perforin gene, we studied the levels of perforin mRNA in peripheral blood mononuclear cells from 11 cases of granular lymphocyte-proliferative disorders (GLPDs). The granular lymphocytes studied were characterized by morphologic, immunophenotypic, and immunogenotypic analyses. Cytolytic functions of the lymphocytes assayed included nonmajor histocompatibility complex-requiring cytotoxicity, anti-CD3-redirected cytotoxicity, antibody-dependent cellular cytotoxicity, and lectin- dependent cellular cytotoxicity. The results showed that in lymphocytes with strong cytolytic functions high levels of perforin mRNA existed, whereas in lymphocytes with weak or undetectable levels of cytolytic functions, low levels of perforin mRNA existed. Because the levels of perforin mRNA correlated with those of cytolytic functions, perforin is probably a mediator in cytolytic functions of granular lymphocytes in patients with GLPDs. When the lymphocytes were cultured for 1 day, however, the levels of cytolytic activity were increased, and those of perforin mRNA were decreased. Therefore, we cannot rule out the possibility that factors other than perforin protein are involved in the cytolytic functions of granular lymphocytes.

scholarly journals Perforin gene expression in granular lymphocyte proliferative disorders

Blood ◽  
1990 ◽  
Vol 75 (3) ◽  
pp. 704-708 ◽  
Author(s):  
K Oshimi ◽  
Y Shinkai ◽  
K Okumura ◽  
Y Oshimi ◽  
H Mizoguchi
Keyword(s):  
Northern Blot Analysis ◽  
Mononuclear Cells ◽  
Cytolytic Activity ◽  
Cellular Cytotoxicity ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Peripheral Blood Mononuclear ◽  
Histocompatibility Complex ◽  
Low Levels ◽  
Granular Lymphocytes ◽  
Rule Out

Abstract By Northern blot analysis using a cDNA clone of the perforin gene, we studied the levels of perforin mRNA in peripheral blood mononuclear cells from 11 cases of granular lymphocyte-proliferative disorders (GLPDs). The granular lymphocytes studied were characterized by morphologic, immunophenotypic, and immunogenotypic analyses. Cytolytic functions of the lymphocytes assayed included nonmajor histocompatibility complex-requiring cytotoxicity, anti-CD3-redirected cytotoxicity, antibody-dependent cellular cytotoxicity, and lectin- dependent cellular cytotoxicity. The results showed that in lymphocytes with strong cytolytic functions high levels of perforin mRNA existed, whereas in lymphocytes with weak or undetectable levels of cytolytic functions, low levels of perforin mRNA existed. Because the levels of perforin mRNA correlated with those of cytolytic functions, perforin is probably a mediator in cytolytic functions of granular lymphocytes in patients with GLPDs. When the lymphocytes were cultured for 1 day, however, the levels of cytolytic activity were increased, and those of perforin mRNA were decreased. Therefore, we cannot rule out the possibility that factors other than perforin protein are involved in the cytolytic functions of granular lymphocytes.

scholarly journals Engineered ACE2-Fc counters murine lethal SARS-CoV-2 infection through direct neutralization and Fc-effector activities

2021 ◽  
Author(s):  
Yaozong Chen ◽  
Lulu Sun ◽  
Irfan Ullah ◽  
Guillaume Beaudoin-Bussieres ◽  
Sai Priya Anand ◽  
...  
Keyword(s):  
Cellular Cytotoxicity ◽  
Therapeutic Activity ◽  
Converting Enzyme ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Angiotensin Converting Enzyme 2 ◽  
Effector Functions ◽  
Therapeutic Setting ◽  
Human Igg1 ◽  
Complement Deposition

Soluble Angiotensin-Converting Enzyme 2 (ACE2) constitutes an attractive antiviral receptor decoy that targets the vulnerable site on SARS-CoV-2 spike (S). Here, using structure-guided approaches, we developed divalent ACE2 molecules by grafting the extracellular ACE2-domain onto a human IgG1 or IgG3 (ACE2-Fc). These ACE2-Fcs harbored structurally validated mutations that enhance S binding and remove enzymatic activity. The lead variant bound tightly to S, mediated in vitro neutralization of SARS-CoV-2 variants of concern (VOCs) with sub-nanomolar IC50 and was capable of robust Fc-effector functions, including antibody-dependent cellular cytotoxicity, phagocytosis and complement deposition. When tested in a stringent K18-hACE2 mouse model, the lead variant prevented or delayed lethal SARS-CoV-2 infection in a prophylactic or therapeutic setting utilizing the combined effect of neutralization and Fc-effector functions. These data confirm the utility of ACE2-Fcs as valuable agents in preventing and eliminating SARS-CoV-2 infection and demonstrate that ACE2-Fc therapeutic activity require Fc-effector functions.

Preliminary Report of In vitro and In vivo Effectiveness of Dornase Alfa on SARS-CoV-2 Infection (Preprint)

2020 ◽  
Author(s):  
Hacer Kuzu Okur ◽  
Koray Yalcin ◽  
Cihan Tastan ◽  
Sevda Demir ◽  
Bulut Yurtsever ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Respiratory Tract ◽  
Mononuclear Cells ◽  
Multiple Organ ◽  
Peripheral Blood Mononuclear ◽  
Dornase Alfa ◽  
Tract Infections ◽  
Adult Peripheral Blood

UNSTRUCTURED Dornase alfa, the recombinant form of the human DNase I enzyme, breaks down neutrophil extracellular traps (NET) that include a vast amount of DNA fragments, histones, microbicidal proteins and oxidant enzymes released from necrotic neutrophils in the highly viscous mucus of cystic fibrosis patients. Dornase alfa has been used for decades in patients with cystic fibrosis to reduce the viscoelasticity of respiratory tract secretions, to decrease the severity of respiratory tract infections, and to improve lung function. Previous studies have linked abnormal NET formations to lung diseases, especially to acute respiratory distress syndrome (ARDS). Coronavirus disease 2019 (COVID-19) pandemic affected more than two million people over the world, resulting in unprecedented health, social and economic crises. The COVID-19, viral pneumonia that progresses to ARDS and even multiple organ failure, is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). High blood neutrophil levels are an early indicator of SARS-CoV-2 infection and predict severe respiratory diseases. A similar mucus structure is detected in COVID-19 patients due to the accumulation of excessive NET in the lungs. Here, we show our preliminary results with dornase alfa that may have an in-vitro anti-viral effect against SARS-CoV-2 infection in a bovine kidney cell line, MDBK without drug toxicity on healthy adult peripheral blood mononuclear cells. In this preliminary study, we also showed that dornase alfa can promote clearance of NET formation in both an in-vitro and three COVID-19 cases who showed clinical improvement in radiological analysis (2-of-3 cases), oxygen saturation (SpO2), respiratory rate, disappearing of dyspnea and coughing.

Sign in / Sign up

Export Citation Format

Share Document