In vitro VLA-4 blockade results in an impaired NK cell-mediated immune surveillance against melanoma

Ilaria Gandoglia, Federico Ivaldi, Paolo Carrega, Eric Armentani, Guido Ferlazzo, Gianluigi Mancardi, Nicole Kerlero de Rosbo, Antonio Uccelli, Alice Laroni

Research output: Contribution to journalArticle

Abstract

Natalizumab (NTZ) is a monoclonal antibody targeting the α4β1 integrin (CD49d/CD29), very late antigen-4 (VLA-4), which is approved for treatment of relapsing-remitting multiple sclerosis (RR-MS). A possible association between NTZ treatment and a higher risk of melanoma is under debate. Natural Killer (NK) cells, which express VLA-4, represent an innate barrier limiting spreading of melanoma under steady state conditions. Indeed, because of their expression of activating receptors, they are very efficient in recognizing and killing melanoma cells without the need of a previous priming. For this reason, we aimed at assessing whether NK-cell functions might be impaired by sustained exposure to NTZ. To investigate this possibility we isolated NK cells from healthy donors and tested their cytotoxic and migratory functions against primary melanoma cells derived from subcutaneous and lymph node metastases. Flow cytometry analysis demonstrated expression of CD49d on both freshly isolated NK cells and activated NK cells. Moreover, VLA-4 and its receptor, vascular cell adhesion protein-1 (VCAM-1) were similarly expressed on freshly isolated NK cells. However, upon a short exposure to NTZ, expression of VLA-4 on NK cells decreased. Analysis of NK receptor expression upon exposure of NK cells from three healthy donors to NTZ indicated that DNAM-1 and NKp46 are apparently decreased, while NKG2A is increased. The degranulation of NK cells towards melanoma cells, which express both VLA-4 and VCAM-1, was not affected when NTZ was added to the co-culture or when both NK cells and melanoma cells were each pre-exposed to NTZ for over 12h. In contrast, degranulation was significantly inhibited after 48h of pre-incubation indicating that NTZ can influence NK-cell degranulation towards melanoma cells only after a prolonged exposure. Using a migration chamber assay, we observed that the migration of NK cells towards melanoma cells was dependent upon the concentration of melanoma cells in the lower chamber, and that it was significantly reduced in presence of NTZ. Our results show that upon exposure to NTZ both cytolytic activity and migration toward melanoma cells were affected, suggesting that binding of NTZ to NK cells affects pathways involved in these NK-cell functions. We analyzed the expression of CD49d on NK cells from MS patients treated with NTZ and observed that it decreases with time of treatment. These data suggest that blockade of VLA-4 on NK-cell surface alters some key functions involved in the immune surveillance toward melanoma by NK cells and may provide a mechanistic explanation for the reported occurrence of melanoma in MS patients treated with NTZ.

Original languageEnglish
Pages (from-to)109-115
Number of pages7
JournalImmunology Letters
Volume181
DOIs
Publication statusPublished - Jan 2017

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Integrin alpha4beta1
Natural Killer Cells
Melanoma
In Vitro Techniques
Vascular Cell Adhesion Molecule-1
Natalizumab
Cell Degranulation
Relapsing-Remitting Multiple Sclerosis

Keywords

  • Adult
  • Cell Degranulation
  • Cell Line, Tumor
  • Cell Movement
  • Cross-Sectional Studies
  • Cytotoxicity, Immunologic
  • Female
  • Gene Expression
  • Humans
  • Immunologic Surveillance
  • Integrin alpha4beta1
  • Killer Cells, Natural
  • Lymphocyte Activation
  • Male
  • Melanoma
  • Middle Aged
  • Natalizumab
  • Receptors, Natural Killer Cell
  • Young Adult
  • Journal Article

Cite this

Gandoglia, I., Ivaldi, F., Carrega, P., Armentani, E., Ferlazzo, G., Mancardi, G., ... Laroni, A. (2017). In vitro VLA-4 blockade results in an impaired NK cell-mediated immune surveillance against melanoma. Immunology Letters, 181, 109-115. https://doi.org/10.1016/j.imlet.2016.11.015

In vitro VLA-4 blockade results in an impaired NK cell-mediated immune surveillance against melanoma. / Gandoglia, Ilaria; Ivaldi, Federico; Carrega, Paolo; Armentani, Eric; Ferlazzo, Guido; Mancardi, Gianluigi; Kerlero de Rosbo, Nicole; Uccelli, Antonio; Laroni, Alice.

In: Immunology Letters, Vol. 181, 01.2017, p. 109-115.

Research output: Contribution to journalArticle

Gandoglia, I, Ivaldi, F, Carrega, P, Armentani, E, Ferlazzo, G, Mancardi, G, Kerlero de Rosbo, N, Uccelli, A & Laroni, A 2017, 'In vitro VLA-4 blockade results in an impaired NK cell-mediated immune surveillance against melanoma', Immunology Letters, vol. 181, pp. 109-115. https://doi.org/10.1016/j.imlet.2016.11.015
Gandoglia, Ilaria ; Ivaldi, Federico ; Carrega, Paolo ; Armentani, Eric ; Ferlazzo, Guido ; Mancardi, Gianluigi ; Kerlero de Rosbo, Nicole ; Uccelli, Antonio ; Laroni, Alice. / In vitro VLA-4 blockade results in an impaired NK cell-mediated immune surveillance against melanoma. In: Immunology Letters. 2017 ; Vol. 181. pp. 109-115.
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AU - Gandoglia, Ilaria

AU - Ivaldi, Federico

AU - Carrega, Paolo

AU - Armentani, Eric

AU - Ferlazzo, Guido

AU - Mancardi, Gianluigi

AU - Kerlero de Rosbo, Nicole

AU - Uccelli, Antonio

AU - Laroni, Alice

N1 - Copyright © 2016 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

PY - 2017/1

Y1 - 2017/1

N2 - Natalizumab (NTZ) is a monoclonal antibody targeting the α4β1 integrin (CD49d/CD29), very late antigen-4 (VLA-4), which is approved for treatment of relapsing-remitting multiple sclerosis (RR-MS). A possible association between NTZ treatment and a higher risk of melanoma is under debate. Natural Killer (NK) cells, which express VLA-4, represent an innate barrier limiting spreading of melanoma under steady state conditions. Indeed, because of their expression of activating receptors, they are very efficient in recognizing and killing melanoma cells without the need of a previous priming. For this reason, we aimed at assessing whether NK-cell functions might be impaired by sustained exposure to NTZ. To investigate this possibility we isolated NK cells from healthy donors and tested their cytotoxic and migratory functions against primary melanoma cells derived from subcutaneous and lymph node metastases. Flow cytometry analysis demonstrated expression of CD49d on both freshly isolated NK cells and activated NK cells. Moreover, VLA-4 and its receptor, vascular cell adhesion protein-1 (VCAM-1) were similarly expressed on freshly isolated NK cells. However, upon a short exposure to NTZ, expression of VLA-4 on NK cells decreased. Analysis of NK receptor expression upon exposure of NK cells from three healthy donors to NTZ indicated that DNAM-1 and NKp46 are apparently decreased, while NKG2A is increased. The degranulation of NK cells towards melanoma cells, which express both VLA-4 and VCAM-1, was not affected when NTZ was added to the co-culture or when both NK cells and melanoma cells were each pre-exposed to NTZ for over 12h. In contrast, degranulation was significantly inhibited after 48h of pre-incubation indicating that NTZ can influence NK-cell degranulation towards melanoma cells only after a prolonged exposure. Using a migration chamber assay, we observed that the migration of NK cells towards melanoma cells was dependent upon the concentration of melanoma cells in the lower chamber, and that it was significantly reduced in presence of NTZ. Our results show that upon exposure to NTZ both cytolytic activity and migration toward melanoma cells were affected, suggesting that binding of NTZ to NK cells affects pathways involved in these NK-cell functions. We analyzed the expression of CD49d on NK cells from MS patients treated with NTZ and observed that it decreases with time of treatment. These data suggest that blockade of VLA-4 on NK-cell surface alters some key functions involved in the immune surveillance toward melanoma by NK cells and may provide a mechanistic explanation for the reported occurrence of melanoma in MS patients treated with NTZ.

AB - Natalizumab (NTZ) is a monoclonal antibody targeting the α4β1 integrin (CD49d/CD29), very late antigen-4 (VLA-4), which is approved for treatment of relapsing-remitting multiple sclerosis (RR-MS). A possible association between NTZ treatment and a higher risk of melanoma is under debate. Natural Killer (NK) cells, which express VLA-4, represent an innate barrier limiting spreading of melanoma under steady state conditions. Indeed, because of their expression of activating receptors, they are very efficient in recognizing and killing melanoma cells without the need of a previous priming. For this reason, we aimed at assessing whether NK-cell functions might be impaired by sustained exposure to NTZ. To investigate this possibility we isolated NK cells from healthy donors and tested their cytotoxic and migratory functions against primary melanoma cells derived from subcutaneous and lymph node metastases. Flow cytometry analysis demonstrated expression of CD49d on both freshly isolated NK cells and activated NK cells. Moreover, VLA-4 and its receptor, vascular cell adhesion protein-1 (VCAM-1) were similarly expressed on freshly isolated NK cells. However, upon a short exposure to NTZ, expression of VLA-4 on NK cells decreased. Analysis of NK receptor expression upon exposure of NK cells from three healthy donors to NTZ indicated that DNAM-1 and NKp46 are apparently decreased, while NKG2A is increased. The degranulation of NK cells towards melanoma cells, which express both VLA-4 and VCAM-1, was not affected when NTZ was added to the co-culture or when both NK cells and melanoma cells were each pre-exposed to NTZ for over 12h. In contrast, degranulation was significantly inhibited after 48h of pre-incubation indicating that NTZ can influence NK-cell degranulation towards melanoma cells only after a prolonged exposure. Using a migration chamber assay, we observed that the migration of NK cells towards melanoma cells was dependent upon the concentration of melanoma cells in the lower chamber, and that it was significantly reduced in presence of NTZ. Our results show that upon exposure to NTZ both cytolytic activity and migration toward melanoma cells were affected, suggesting that binding of NTZ to NK cells affects pathways involved in these NK-cell functions. We analyzed the expression of CD49d on NK cells from MS patients treated with NTZ and observed that it decreases with time of treatment. These data suggest that blockade of VLA-4 on NK-cell surface alters some key functions involved in the immune surveillance toward melanoma by NK cells and may provide a mechanistic explanation for the reported occurrence of melanoma in MS patients treated with NTZ.

KW - Adult

KW - Cell Degranulation

KW - Cell Line, Tumor

KW - Cell Movement

KW - Cross-Sectional Studies

KW - Cytotoxicity, Immunologic

KW - Female

KW - Gene Expression

KW - Humans

KW - Immunologic Surveillance

KW - Integrin alpha4beta1

KW - Killer Cells, Natural

KW - Lymphocyte Activation

KW - Male

KW - Melanoma

KW - Middle Aged

KW - Natalizumab

KW - Receptors, Natural Killer Cell

KW - Young Adult

KW - Journal Article

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JO - Immunology Letters

JF - Immunology Letters

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