Skip to main content
  • AACR Publications
    • Blood Cancer Discovery
    • Cancer Discovery
    • Cancer Epidemiology, Biomarkers & Prevention
    • Cancer Immunology Research
    • Cancer Prevention Research
    • Cancer Research
    • Clinical Cancer Research
    • Molecular Cancer Research
    • Molecular Cancer Therapeutics

AACR logo

  • Register
  • Log in
  • My Cart
Advertisement

Main menu

  • Home
  • About
    • The Journal
    • AACR Journals
    • Subscriptions
    • Permissions and Reprints
    • Reviewing
  • Articles
    • OnlineFirst
    • Current Issue
    • Past Issues
    • Meeting Abstracts
    • Cancer Immunology Essentials
    • Collections
      • COVID-19 & Cancer Resource Center
      • "Best of" Collection
      • Editors' Picks
  • For Authors
    • Information for Authors
    • Author Services
    • Best of: Author Profiles
    • Submit
  • Alerts
    • Table of Contents
    • Editors' Picks
    • OnlineFirst
    • Citation
    • Author/Keyword
    • RSS Feeds
    • My Alert Summary & Preferences
  • News
    • Cancer Discovery News
  • COVID-19
  • Webinars
  • Search More

    Advanced Search

  • AACR Publications
    • Blood Cancer Discovery
    • Cancer Discovery
    • Cancer Epidemiology, Biomarkers & Prevention
    • Cancer Immunology Research
    • Cancer Prevention Research
    • Cancer Research
    • Clinical Cancer Research
    • Molecular Cancer Research
    • Molecular Cancer Therapeutics

User menu

  • Register
  • Log in
  • My Cart

Search

  • Advanced search
Cancer Immunology Research
Cancer Immunology Research
  • Home
  • About
    • The Journal
    • AACR Journals
    • Subscriptions
    • Permissions and Reprints
    • Reviewing
  • Articles
    • OnlineFirst
    • Current Issue
    • Past Issues
    • Meeting Abstracts
    • Cancer Immunology Essentials
    • Collections
      • COVID-19 & Cancer Resource Center
      • "Best of" Collection
      • Editors' Picks
  • For Authors
    • Information for Authors
    • Author Services
    • Best of: Author Profiles
    • Submit
  • Alerts
    • Table of Contents
    • Editors' Picks
    • OnlineFirst
    • Citation
    • Author/Keyword
    • RSS Feeds
    • My Alert Summary & Preferences
  • News
    • Cancer Discovery News
  • COVID-19
  • Webinars
  • Search More

    Advanced Search

Research Articles

IgA-Mediated Killing of Tumor Cells by Neutrophils Is Enhanced by CD47–SIRPα Checkpoint Inhibition

Louise W. Treffers, Toine ten Broeke, Thies Rösner, J.H. Marco Jansen, Michel van Houdt, Steffen Kahle, Karin Schornagel, Paul J.J.H. Verkuijlen, Jan M. Prins, Katka Franke, Taco W. Kuijpers, Timo K. van den Berg, Thomas Valerius, Jeanette H.W. Leusen and Hanke L. Matlung
Louise W. Treffers
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Toine ten Broeke
2Immunotherapy Laboratory, Center for Translational Immunology, University Medical Center Utrecht, Utrecht, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Thies Rösner
3Section for Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, Christian-Albrechts-University, Kiel, Germany.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
J.H. Marco Jansen
2Immunotherapy Laboratory, Center for Translational Immunology, University Medical Center Utrecht, Utrecht, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Michel van Houdt
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Steffen Kahle
3Section for Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, Christian-Albrechts-University, Kiel, Germany.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Karin Schornagel
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Paul J.J.H. Verkuijlen
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jan M. Prins
4Department of Internal Medicine, Division of Infectious Diseases, Academic Medical Center, University of Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Katka Franke
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Taco W. Kuijpers
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
5Emma Children's Hospital, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Timo K. van den Berg
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
6Department of Molecular Cell Biology and Immunology, Amsterdam UMC, Vrije Universiteit Amsterdam, Amsterdam Infection and Immunity Institute, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Thomas Valerius
3Section for Stem Cell Transplantation and Immunotherapy, Department of Internal Medicine II, Christian-Albrechts-University, Kiel, Germany.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Thomas Valerius
Jeanette H.W. Leusen
2Immunotherapy Laboratory, Center for Translational Immunology, University Medical Center Utrecht, Utrecht, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Hanke L. Matlung
1Sanquin Research, and Landsteiner Laboratory, Amsterdam UMC, University of Amsterdam, Amsterdam, the Netherlands.
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • For correspondence: h.matlung@sanquin.nl
DOI: 10.1158/2326-6066.CIR-19-0144 Published January 2020
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

Article Figures & Data

Figures

  • Additional Files
  • Figure 1.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 1.

    Downregulation of CD47 expression promoted IgA-mediated ADCC by neutrophils. A, Representative histograms (n = 2) showing the expression of CD47 and HER2/neu or EGFR in SKBR3 and A431 cells (top) and their respective CD47KD or CD47KO variants (bottom). B and C, Genetic disruption of CD47–SIRPα interactions led to increased ADCC of tumor cells (SKBR3 and A431) by freshly isolated neutrophils using increasing concentrations of the indicated IgG1 (black lines) or their IgA2 variants (gray lines). Data shown are means ± SEM pooled from two to three experiments with 2 to 3 donor samples per experiment with n = 2–5 (B, SKBR3) and n = 3–4 (C, A431) individual donors. Statistics shown in B and C are for the highest antibody concentrations or highest E:T ratio using two-way ANOVA with Tukey correction for multiple tests. ***, P < 0.001. Cmab, cetuximab; Tmab, trastuzumab.

  • Figure 2.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 2.

    Different approaches to inhibit CD47–SIRPα interactions enhanced IgA-mediated ADCC. ADCC of cancer cells by freshly isolated neutrophils without (white background) and with inhibition of CD47–SIRPα interactions (gray background) by either using cell lines with no (A431-CD47KO) or reduced (SKBR3-CD47KD) expression of CD47 (A), inhibition of CD47 with B6H12 F(ab′)2 antibodies (B), or inhibition of SIRPα using antibody 12C4 (C). Used are either anti-HER2-IgA2 antibody against SKBR3 or anti-EGFR-IgA2 antibody (either at 5 μg/mL) against A431 cells, respectively. Data shown are means ± SEM pooled from two to three experiments with 2 to 3 individual donor samples per experiment with n = 8 (A), n = 7–8 (B), and n = 6–8 (C). Statistics were performed by a paired t test (**, P < 0.01; ***, P < 0.001).

  • Figure 3.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 3.

    IgA-mediated trogocytosis of cancer cells by neutrophils. A–F, Representative live-cell imaging stills at different time points of a neutrophil binding to an anti-HER2-IgA2–opsonized cancer cell and trogocytosing pieces of fluorescent cancer cell membrane. Scale bar, 10 μm. G, Trogocytosis of SKBR3 cells opsonized with increasing concentrations of trastuzumab or anti-HER2-IgA2. Tmab, trastuzumab. ADCC (H) and trogocytosis (I) of anti-EGFR-IgA2–opsonized A431 cancer cells by freshly isolated neutrophils isolated from healthy controls or FHL5 patients (for patient characteristics, see patients B and C described in ref. 32) combined without (white background) or with CD47–SIRPα inhibition by knockout of CD47 in the target cells (gray background). ADCC (J) and trogocytosis (K) of anti-HER2-IgA2–opsonized SKBR3 cells by freshly isolated neutrophils isolated from healthy controls or CGD patients (for patient characteristics, see Materials and Methods), combined without (white background) or with CD47–SIRPα inhibition by knockdown of CD47 in the target cells (gray background). Data shown are means ± SEM pooled from two separate experiments with two replicates of 2 individual FHL5 patient samples and 11 total healthy controls (H and I), three separate experiments with 3 (K) or 4 (J) individual CGD patient samples and 4-day controls (J and K), or pooled from two experiments with 1 to 2 donor samples per experiment with n = 4 (G) individual donors. Statistics shown in G are for the highest antibody concentrations using two-way ANOVA with Tukey correction for multiple tests. No statistics could be performed on FHL5 patient data in H and I due to low patient count (n = 2). Statistics shown in J and K using one-way ANOVA with Sidak correction for multiple tests (ns, nonsignificant; *, P < 0.05; **, P < 0.01; ***, P < 0.001). MFI, mean fluorescence intensity.

  • Figure 4.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 4.

    Inhibition of CD47–SIRPα did not result in alternative FcαR-mediated ADCC. Neutrophil-mediated trogocytosis (A) and ADCC (B) of SKBR3 (white background) and SKBR3-CD47KD (gray background) cells using IgA2-Her2 antibody with inhibition of Syk, PI3K, myosin light chain kinase, CD11b/CD18 integrins and calcium mobilization. Data shown are means ± SEM pooled from two experiments with 2 donor samples per experiment with n = 4 individual donors. Statistics are shown using one-way ANOVA with Sidak correction for multiple tests (*, P < 0.05; ****, P < 0.001).

  • Figure 5.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 5.

    Interference with CD47–SIRPα interactions enhanced tumor eradication in a xenogeneic long-term in vivo model. A, Schematic overview of the in vivo xenograft model and the injection scheme. Long-term in vivo tumor growth comparing the maternal A431-SCR cell line (B) with one with no expression of CD47 (A431-CD47KO; C). Treatment was started when mice had palpable tumors (day 6) with either PBS (light gray line), a single intravenous injection of 50 μg cetuximab (black line), or an intravenous injection of 250 μg anti-EGFR-IgA2 followed by four intraperitoneal injections of 250 μg (dark gray line) to compensate for the shorter half-life of IgA compared with IgG. Tumor outgrowth was measured with calipers, and volume was calculated as length × width × height. Data shown are means ± SEM from one experiment with 8 mice per group in B and C. Statistics performed are for day 17 using two-way ANOVA with Tukey correction for multiple tests (ns, nonsignificant; *, P < 0.05). Cmab, cetuximab.

  • Figure 6.
    • Download figure
    • Open in new tab
    • Download powerpoint
    Figure 6.

    Inhibition of SIRPα effectively triggered syngeneic tumor cell killing by neutrophils. A, Expression of CD47 and HER2 or EGFR in Ba/F3 cells transfected with human HER2 or EGFR, respectively. B, ADCC of trastuzumab or anti-HER2-IgA2 (either at 5 μg/mL)–opsonized Ba/F3-HER2 cells by mouse neutrophils isolated from wild-type (NTg) or FcαR-transgenic (FcαR-Tg) mice combined without (white background) or with inhibition of SIRPα (MY-1; gray background). C, Same as B but with cetuximab or anti-EGFR-IgA2–opsonized Ba/F3-EGFR cells. D, In vivo inhibition of SIRPα (MY-1) was combined with trastuzumab or anti-HER2-IgA2 treatment in a short intraperitoneal model. The ratio of Ba/F3-HER2 and Ba/F3 cells in wild-type mice [PBS, isotype + Tmab, anti-SIRPα (MY-1) + Tmab] or FcαR-transgenic mice [isotype + anti-HER2-IgA2, anti-SIRPα (MY-1) + anti-HER2-IgA2] in the peritoneal washes was determined by flow cytometry. E, Number of granulocytes (Ly6G+/CD11b+) present in the peritoneal cavity at the end of the experiment in D. F, Overview of all cells present in the peritoneal cavity during the experiment shown in D. G, Inhibition of SIRPα (MY-1) is combined with anti-HER2-IgA2 and neutrophil depletion (Ly6G) in an in vivo mouse experiment, showing the ratio of Ba/F3-HER2 and Ba/F3 cells in wild-type mice [isotype, anti-SIRPα (MY-1), anti-SIRPα (MY-1) + anti-HER2-IgA2 (NTg)] or FcαR-transgenic mice [isotype + anti-HER2-IgA2, anti-SIRPα (MY-1) + anti-HER2-IgA2, anti-SIRPα (MY-1) + anti-HER2-IgA2 + Ly6G]. Data shown for B and C are means ± SEM pooled from two experiments with 2 mice per experiment with a total of n = 4 individual mice. Data shown for D–F are means ± SEM from one experiment with 6 mice per group with A representative of n = 2, B n = 3–6, C n = 6, and D n = 6 individual mice. Statistics shown for B and C are calculated by one-way ANOVA with Dunnett correction for multiple tests. Statistics shown for D and G are calculated by paired one-way ANOVA with Sidak correction for multiple tests. Statistics performed for E are calculated by one-way ANOVA with Sidak correction for multiple tests (ns, nonsignificant; *, P < 0.05; **, P < 0.01; ***, P < 0.001). Cmab, cetuximab; Tmab, trastuzumab.

Additional Files

  • Figures
  • Supplementary Data

    • Supplementary Figures - Supplementary Figures
    • Supplementary Data - Supplementary Materials and Methods
PreviousNext
Back to top
Cancer Immunology Research: 8 (1)
January 2020
Volume 8, Issue 1
  • Table of Contents
  • Table of Contents (PDF)
  • About the Cover
  • Editorial Board (PDF)

Sign up for alerts

View this article with LENS

Open full page PDF
Article Alerts
Sign In to Email Alerts with your Email Address
Email Article

Thank you for sharing this Cancer Immunology Research article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
IgA-Mediated Killing of Tumor Cells by Neutrophils Is Enhanced by CD47–SIRPα Checkpoint Inhibition
(Your Name) has forwarded a page to you from Cancer Immunology Research
(Your Name) thought you would be interested in this article in Cancer Immunology Research.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Citation Tools
IgA-Mediated Killing of Tumor Cells by Neutrophils Is Enhanced by CD47–SIRPα Checkpoint Inhibition
Louise W. Treffers, Toine ten Broeke, Thies Rösner, J.H. Marco Jansen, Michel van Houdt, Steffen Kahle, Karin Schornagel, Paul J.J.H. Verkuijlen, Jan M. Prins, Katka Franke, Taco W. Kuijpers, Timo K. van den Berg, Thomas Valerius, Jeanette H.W. Leusen and Hanke L. Matlung
Cancer Immunol Res January 1 2020 (8) (1) 120-130; DOI: 10.1158/2326-6066.CIR-19-0144

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Share
IgA-Mediated Killing of Tumor Cells by Neutrophils Is Enhanced by CD47–SIRPα Checkpoint Inhibition
Louise W. Treffers, Toine ten Broeke, Thies Rösner, J.H. Marco Jansen, Michel van Houdt, Steffen Kahle, Karin Schornagel, Paul J.J.H. Verkuijlen, Jan M. Prins, Katka Franke, Taco W. Kuijpers, Timo K. van den Berg, Thomas Valerius, Jeanette H.W. Leusen and Hanke L. Matlung
Cancer Immunol Res January 1 2020 (8) (1) 120-130; DOI: 10.1158/2326-6066.CIR-19-0144
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Jump to section

  • Article
    • Abstract
    • Introduction
    • Materials and Methods
    • Results
    • Discussion
    • Disclosure of Potential Conflicts of Interest
    • Authors' Contributions
    • Acknowledgments
    • Footnotes
    • References
  • Figures & Data
  • Info & Metrics
  • PDF
Advertisement

Related Articles

Cited By...

More in this TOC Section

  • Activin A Mediates Radiation-Induced Antitumor Immunity
  • Mechanisms to CD3-Bispecific Antibody Therapy
  • Improved Estimation of T-cell Receptor Repertoire Diversity
Show more Research Articles
  • Home
  • Alerts
  • Feedback
  • Privacy Policy
Facebook   Twitter   LinkedIn   YouTube   RSS

Articles

  • Online First
  • Current Issue
  • Past Issues
  • Cancer Immunology Essentials

Info for

  • Authors
  • Subscribers
  • Advertisers
  • Librarians

About Cancer Immunology Research

  • About the Journal
  • Editorial Board
  • Permissions
  • Submit a Manuscript
AACR logo

Copyright © 2021 by the American Association for Cancer Research.

Cancer Immunology Research
eISSN: 2326-6074
ISSN: 2326-6066

Advertisement