Cancer Immunology, Immunotherapy

, Volume 66, Issue 3, pp 319–332 | Cite as

Targeting of the WT191–138 fragment to human dendritic cells improves leukemia-specific T-cell responses providing an alternative approach to WT1-based vaccination

  • Nergui Dagvadorj
  • Anne Deuretzbacher
  • Daniela Weisenberger
  • Elke Baumeister
  • Johannes Trebing
  • Isabell Lang
  • Carolin Köchel
  • Markus Kapp
  • Kerstin Kapp
  • Andreas Beilhack
  • Thomas Hünig
  • Hermann Einsele
  • Harald Wajant
  • Götz Ulrich Grigoleit
Original Article
First Online:
Received:
Accepted:

Abstract

Due to its immunogenicity and overexpression concomitant with leukemia progression, Wilms tumor protein 1 (WT1) is of particular interest for immunotherapy of AML relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT). So far, WT1-specific T-cell responses have mainly been induced by vaccination with peptides presented by certain HLA alleles. However, this approach is still not widely applicable in clinical practice due to common limitations of HLA restriction. Dendritic cell (DC) vaccines electroporated with mRNA encoding full-length protein have also been tested for generating WT1-derived peptides for presentation to T-cells. Alternatively, an efficient and broad WT1 peptide presentation could be elicited by triggering receptor-mediated protein endocytosis of DCs. Therefore, we developed antibody fusion proteins consisting of an antibody specific for the DEC205 endocytic receptor on human DCs and various fragments of WT1 as DC-targeting recombinant WT1 vaccines (anti-hDEC205-WT1). Of all anti-hDEC205-WT1 fusion proteins designed for overcoming insufficient expression, anti-hDEC205-WT110–35, anti-hDEC205-WT191–138, anti-hDEC205-WT1223–273, and anti-hDEC205-WT1324–371 were identified in good yields. The anti-hDEC205-WT191–138 was capable of directly inducing ex vivo T-cell responses by co-incubation of the fusion protein-loaded monocyte-derived mature DCs and autologous T-cells of either healthy or HSCT individuals. Furthermore, the DC-targeted WT191–138-induced specific T-cells showed a strong cytotoxic activity by lysing WT1-overexpressing THP-1 leukemia cells in vitro while sparing WT1-negative hematopoietic cells. In conclusion, our approach identifies four WT1 peptide-antibody fusion proteins with sufficient production and introduces an alternative vaccine that could be easily translated into clinical practice to improve WT1-directed antileukemia immune responses after allo-HSCT.

Keywords

Wilms tumor protein 1 Tumor-associated antigen Immunotherapy against high-risk AML Anti-hDEC205-WT1 antibody fusion protein Cytotoxic T-cells Tumor vaccine 

Abbreviations

aa

Amino acid

AML

Acute myeloid leukemia

CAR

Chimeric antigen receptor

CHO

Chinese hamster ovary

CHO-hDEC205

hDEC205-stably expressing CHO

CMV

Cytomegalovirus

CTL

Cytotoxic T lymphocyte

ET

Effector to target ratio

ELISPOT

Enzyme-linked immuno spot

FCS

Fetal calf serum

GM-CSF

Granulocyte–macrophage colony stimulating factor

GpL

Gaussia princeps luciferase

GvL

Graft-versus-leukemia

hDEC205

Human DEC205 endocytic receptor

HEK293

Human embryonic kidney

HLA

Human leukocyte antigen

HSCT

Hematopoietic stem cell transplantation

IC50

50% inhibitory concentration

ICS

Intracellular cytokine staining

IFN

Interferon

IL

Interleukin

Kd

Dissociation constant

Ki

Dissociation constant of inhibitor

LAL

Limulus amebocyte lysate

MDS

Myelodysplastic syndrome

moDCs

Monocyte-derived dendritic cells

NBT/BCIP

Nitro blue tetrazolium/5-Bromo-4-chloro-3-indolyl phosphate

PBMCs

Peripheral blood mononuclear cells

PHA-L

Phytohemagglutinin-L

pM

Picomolar

PN

Patient number

RT-PCR

Reverse transcriptase polymerase chain reaction

scFv

Single chain variable fragment

SDS-PAGE

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

TAA

Tumor-associated antigen

TCR

T-cell receptor

TNF

Tumor necrosis factor

WT1

Wilms tumor protein

WT1_D

Isoform D of WT1 protein

WT1full

WT1 in full-length

WT1major and WT1small

Major and small fragments of WT1 protein

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Notes

Acknowledgements

We thank our colleagues in the Division of Molecular Internal Medicine for their assistance and Professor Matthias Eyrich for his critical discussion. We also thank Dr. Torsten Steinbrunn for proofreading of the manuscript. Nergui Dagvadorj was supported by scholarships from the Government of Mongolia, the Fritz Thyssen Foundation of Germany, the Interdisciplinary Center for Clinical Research (IZKF) of the University of Würzburg, Germany and the German Academic Exchange Service (DAAD), Germany.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

262_2016_1938_MOESM1_ESM.pdf (623 kb)
Supplementary material 1 (PDF 623 kb)

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Nergui Dagvadorj
    • 1
    • 5
  • Anne Deuretzbacher
    • 1
  • Daniela Weisenberger
    • 2
  • Elke Baumeister
    • 1
  • Johannes Trebing
    • 2
  • Isabell Lang
    • 2
  • Carolin Köchel
    • 1
  • Markus Kapp
    • 1
  • Kerstin Kapp
    • 1
  • Andreas Beilhack
    • 3
  • Thomas Hünig
    • 4
  • Hermann Einsele
    • 1
  • Harald Wajant
    • 2
  • Götz Ulrich Grigoleit
    • 1
  1. 1.Laboratory for Immunotherapy, Department of Internal Medicine IIUniversity Hospital of WürzburgWürzburgGermany
  2. 2.Division of Molecular Internal Medicine, Department of Internal Medicine IIUniversity Hospital of WürzburgWürzburgGermany
  3. 3.Division of Experimental Stem Cell Transplantation, Interdisciplinary Center for Clinical ResearchUniversity of WürzburgWürzburgGermany
  4. 4.Institute for Virology and ImmunobiologyUniversity of WürzburgWürzburgGermany
  5. 5.Mongolian National University of Medical SciencesUlaanbaatarMongolia

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