Identifying functional defects in patients with immune dysregulation due to LRBA and CTLA-4 mutations.
Heterozygous CTLA-4 deficiency has been reported as a monogenic cause of common variable immune deficiency with features of immune dysregulation. Direct mutation in CTLA-4 leads to defective regulatory T-cell (Treg) function associated with impaired ability to control levels of the CTLA-4 ligands, CD80 and CD86. However, additional mutations affecting the CTLA-4 pathway, such as those recently reported for LRBA, indirectly affect CTLA-4 expression, resulting in clinically similar disorders. Robust phenotyping approaches sensitive to defects in the CTLA-4 pathway are therefore required to inform understanding of such immune dysregulation syndromes. Here, we describe assays capable of distinguishing a variety of defects in the CTLA-4 pathway. Assessing total CTLA-4 expression levels was found to be optimal when restricting analysis to the CD45RA-Foxp3+fraction. CTLA-4 induction following stimulation, and the use of lysosomal-blocking compounds, distinguished CTLA-4 from LRBA mutations. Short-term T-cell stimulation improved the capacity for discriminating the Foxp3+Treg compartment, clearly revealing Treg expansions in these disorders. Finally, we developed a functionally orientated assay to measure ligand uptake by CTLA-4, which is sensitive to ligand-binding or -trafficking mutations, that would otherwise be difficult to detect and that is appropriate for testing novel mutations in CTLA-4 pathway genes. These approaches are likely to be of value in interpreting the functional significance of mutations in the CTLA-4 pathway identified by gene-sequencing approaches.
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Tie Zheng
TZ
Hou
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Nisha
N
Verma
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Jennifer
J
Wanders
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Alan
A
Kennedy
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Blagoje
B
Soskic
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Daniel
D
Janman
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Neil
N
Halliday
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Behzad
B
Rowshanravan
Immunology Department, Great Ormond Street Hospital for Children National Health Service (NHS) Foundation Trust, London, United Kingdom.
Austen
A
Worth
Immunology Department, Great Ormond Street Hospital for Children National Health Service (NHS) Foundation Trust, London, United Kingdom.
Waseem
W
Qasim
Papworth and Addenbrooke's Hospital NHS Foundation Trusts, Cambridge, United Kingdom.
Helen
H
Baxendale
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Hans
H
Stauss
Clinical Immunology Department, Royal Free Hospital, London, United Kingdom.
Suranjith
S
Seneviratne
Sección de Infectología e Inmunopatología, Unidad de Pediatría, Hospital Virgen del Rocío/Instituto de Biomedicina de Sevilla (IBiS), Sevilla, Spain.
Olaf
O
Neth
Sección de Infectología e Inmunopatología, Unidad de Pediatría, Hospital Virgen del Rocío/Instituto de Biomedicina de Sevilla (IBiS), Sevilla, Spain.
Peter
P
Olbrich
Primary Immunodeficiency Group, Institute of Cellular Medicine, Medical School, Newcastle University, Newcastle upon Tyne, United Kingdom; and.
Sophie
S
Hambleton
Department of Immunology, University of Manchester, Royal Manchester Children's Hospital, Manchester, United Kingdom.
Peter D
PD
Arkwright
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Siobhan O
SO
Burns
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
Lucy S K
LS
Walker
Institute of Immunity and Transplantation, Division of Infection & Immunity, School of Life and Medical Sciences, University College London, Royal Free Hospital, London, United Kingdom.
David M
0000-0001-6506-3115
DM
Sansom
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics*; D060908:CTLA-4 Antigen / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D017074:Common Variable Immunodeficiency / Q000235:genetics; D051858:Forkhead Transcription Factors / Q000032:analysis; D006801:Humans; D055633:Immune System Phenomena / Q000235:genetics; D008024:Ligands; D009154:Mutation; D050378:T-Lymphocytes, Regulatory / Q000276:immunology / Q000473:pathology
D048868:Adaptor Proteins, Signal Transducing; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D008024:Ligands; C428703:LRBA protein, human