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CD16 is indispensable for antibody-dependent cellular cytotoxicity by human monocytes

View Article: PubMed Central - PubMed

ABSTRACT

Antibody-dependent cellular cytotoxicity (ADCC) is exerted by immune cells expressing surface Fcγ receptors (FcγRs) against cells coated with antibody, such as virus-infected or transformed cells. CD16, the FcγRIIIA, is essential for ADCC by NK cells, and is also expressed by a subset of human blood monocytes. We found that human CD16− expressing monocytes have a broad spectrum of ADCC capacities and can kill cancer cell lines, primary leukemic cells and hepatitis B virus-infected cells in the presence of specific antibodies. Engagement of CD16 on monocytes by antibody bound to target cells activated β2-integrins and induced TNFα secretion. In turn, this induced TNFR expression on the target cells, making them susceptible to TNFα-mediated cell death. Treatment with TLR agonists, DAMPs or cytokines, such as IFNγ, further enhanced ADCC. Monocytes lacking CD16 did not exert ADCC but acquired this property after CD16 expression was induced by either cytokine stimulation or transient transfection. Notably, CD16+ monocytes from patients with leukemia also exerted potent ADCC. Hence, CD16+ monocytes are important effectors of ADCC, suggesting further developments of this property in the context of cellular therapies for cancer and infectious diseases.

No MeSH data available.


Related in: MedlinePlus

Cell-cell contact mediated through β2-integrins is needed for ADCC to take place.(A) CD16+ monocytes were co-cultured with SKBR3 at E:T ratio of 10:1. The x-axis depicts the conditions at which tumour targets were used. SKBR3 cells labelled with BATDA are depicted with black diamond inside and are either uncoated or trast-coated (antibody symbol). Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to trast-coated SKBR3 and based on One-way ANOVA (****p ≤ 0.0001), ns = not significant. (B) ADCC by CD16+ monocytes was assessed using time-lapsed imaging (see also Supplementary Movie). Interaction of CD16+ monocyte (black arrow) and trast-coated SKBR3 (dotted) was tracked at 20 secs intervals for up to 2 hrs and static images at various times as indicated are shown. Brightfield images were visualised under a FV-1000 confocal system with an inverted Olympus IX81 microscope at 200x magnification using Fluoview software FV10-ASW 2.0. Data shown is a representative of 3 independent experiments where >10 different cells were followed in each experiment. (C) CD16+ monocytes were either untreated or pre-treated with blocking antibodies for the indicated integrins prior to co-culturing with either uncoated (white bar) or trast-coated SKBR3 (grey bar) at E:T ratio of 10:1. Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to untreated trast-coated SKBR3 based on One-way ANOVA (****p ≤ 0.0001), ns = not significant.
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f4: Cell-cell contact mediated through β2-integrins is needed for ADCC to take place.(A) CD16+ monocytes were co-cultured with SKBR3 at E:T ratio of 10:1. The x-axis depicts the conditions at which tumour targets were used. SKBR3 cells labelled with BATDA are depicted with black diamond inside and are either uncoated or trast-coated (antibody symbol). Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to trast-coated SKBR3 and based on One-way ANOVA (****p ≤ 0.0001), ns = not significant. (B) ADCC by CD16+ monocytes was assessed using time-lapsed imaging (see also Supplementary Movie). Interaction of CD16+ monocyte (black arrow) and trast-coated SKBR3 (dotted) was tracked at 20 secs intervals for up to 2 hrs and static images at various times as indicated are shown. Brightfield images were visualised under a FV-1000 confocal system with an inverted Olympus IX81 microscope at 200x magnification using Fluoview software FV10-ASW 2.0. Data shown is a representative of 3 independent experiments where >10 different cells were followed in each experiment. (C) CD16+ monocytes were either untreated or pre-treated with blocking antibodies for the indicated integrins prior to co-culturing with either uncoated (white bar) or trast-coated SKBR3 (grey bar) at E:T ratio of 10:1. Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to untreated trast-coated SKBR3 based on One-way ANOVA (****p ≤ 0.0001), ns = not significant.

Mentions: To determine whether ADCC by CD16+ monocytes required direct contact with target cells, we tested ADCC against mixtures of trastuzumab-coated and uncoated SKBR3 cells labeled with BATDA. We detected release of the BATDA label when the antibody-coated cells were the ones pre-labeled, and not when the non-coated cells were carrying the BATDA (Fig. 4A). Thus, cells lacking antibody coating were not lysed by CD16+ monocytes, even when in the same culture as antibody-coated cells that were being actively lysed.


CD16 is indispensable for antibody-dependent cellular cytotoxicity by human monocytes
Cell-cell contact mediated through β2-integrins is needed for ADCC to take place.(A) CD16+ monocytes were co-cultured with SKBR3 at E:T ratio of 10:1. The x-axis depicts the conditions at which tumour targets were used. SKBR3 cells labelled with BATDA are depicted with black diamond inside and are either uncoated or trast-coated (antibody symbol). Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to trast-coated SKBR3 and based on One-way ANOVA (****p ≤ 0.0001), ns = not significant. (B) ADCC by CD16+ monocytes was assessed using time-lapsed imaging (see also Supplementary Movie). Interaction of CD16+ monocyte (black arrow) and trast-coated SKBR3 (dotted) was tracked at 20 secs intervals for up to 2 hrs and static images at various times as indicated are shown. Brightfield images were visualised under a FV-1000 confocal system with an inverted Olympus IX81 microscope at 200x magnification using Fluoview software FV10-ASW 2.0. Data shown is a representative of 3 independent experiments where >10 different cells were followed in each experiment. (C) CD16+ monocytes were either untreated or pre-treated with blocking antibodies for the indicated integrins prior to co-culturing with either uncoated (white bar) or trast-coated SKBR3 (grey bar) at E:T ratio of 10:1. Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to untreated trast-coated SKBR3 based on One-way ANOVA (****p ≤ 0.0001), ns = not significant.
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f4: Cell-cell contact mediated through β2-integrins is needed for ADCC to take place.(A) CD16+ monocytes were co-cultured with SKBR3 at E:T ratio of 10:1. The x-axis depicts the conditions at which tumour targets were used. SKBR3 cells labelled with BATDA are depicted with black diamond inside and are either uncoated or trast-coated (antibody symbol). Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to trast-coated SKBR3 and based on One-way ANOVA (****p ≤ 0.0001), ns = not significant. (B) ADCC by CD16+ monocytes was assessed using time-lapsed imaging (see also Supplementary Movie). Interaction of CD16+ monocyte (black arrow) and trast-coated SKBR3 (dotted) was tracked at 20 secs intervals for up to 2 hrs and static images at various times as indicated are shown. Brightfield images were visualised under a FV-1000 confocal system with an inverted Olympus IX81 microscope at 200x magnification using Fluoview software FV10-ASW 2.0. Data shown is a representative of 3 independent experiments where >10 different cells were followed in each experiment. (C) CD16+ monocytes were either untreated or pre-treated with blocking antibodies for the indicated integrins prior to co-culturing with either uncoated (white bar) or trast-coated SKBR3 (grey bar) at E:T ratio of 10:1. Data plotted are mean ± SD, n = 3. ****p ≤ 0.0001 with respect to untreated trast-coated SKBR3 based on One-way ANOVA (****p ≤ 0.0001), ns = not significant.
Mentions: To determine whether ADCC by CD16+ monocytes required direct contact with target cells, we tested ADCC against mixtures of trastuzumab-coated and uncoated SKBR3 cells labeled with BATDA. We detected release of the BATDA label when the antibody-coated cells were the ones pre-labeled, and not when the non-coated cells were carrying the BATDA (Fig. 4A). Thus, cells lacking antibody coating were not lysed by CD16+ monocytes, even when in the same culture as antibody-coated cells that were being actively lysed.

View Article: PubMed Central - PubMed

ABSTRACT

Antibody-dependent cellular cytotoxicity (ADCC) is exerted by immune cells expressing surface Fcγ receptors (FcγRs) against cells coated with antibody, such as virus-infected or transformed cells. CD16, the FcγRIIIA, is essential for ADCC by NK cells, and is also expressed by a subset of human blood monocytes. We found that human CD16− expressing monocytes have a broad spectrum of ADCC capacities and can kill cancer cell lines, primary leukemic cells and hepatitis B virus-infected cells in the presence of specific antibodies. Engagement of CD16 on monocytes by antibody bound to target cells activated β2-integrins and induced TNFα secretion. In turn, this induced TNFR expression on the target cells, making them susceptible to TNFα-mediated cell death. Treatment with TLR agonists, DAMPs or cytokines, such as IFNγ, further enhanced ADCC. Monocytes lacking CD16 did not exert ADCC but acquired this property after CD16 expression was induced by either cytokine stimulation or transient transfection. Notably, CD16+ monocytes from patients with leukemia also exerted potent ADCC. Hence, CD16+ monocytes are important effectors of ADCC, suggesting further developments of this property in the context of cellular therapies for cancer and infectious diseases.

No MeSH data available.


Related in: MedlinePlus