Haploinsufficiency of immune checkpoint receptor CTLA4 induces a distinct neuroinflammatory disorder

Abstract

BACKGROUNDCytotoxic T lymphocyte antigen 4 (CTLA4) is essential for immune homeostasis. Genetic mutations causing haploinsufficiency (CTLA4h) lead to a phenotypically heterogenous, immune-mediated disease that can include neuroinflammation. The neurological manifestations of CTLA4h are poorly characterized.METHODSWe performed an observational natural history study of 50 patients with CTLA4h who were followed at the NIH. We analyzed clinical, radiological, immunological, and histopathological data.RESULTSEvidence for neuroinflammation was observed in 32% (n = 16 of 50) of patients in this cohort by magnetic resonance imaging (MRI) and/or by cerebrospinal fluid analysis. Clinical symptoms were commonly absent or mild in severity, with headaches as the leading complaint (n = 13 of 16). The most striking findings were relapsing, large, contrast-enhancing focal lesions in the brain and spinal cord observed on MRI. We detected inflammation in the cerebrospinal fluid and leptomeninges before the parenchyma. Brain biopsies of inflammatory lesions from 10 patients showed perivascular and intraparenchymal mixed cellular infiltrates with little accompanying demyelination or neuronal injury.CONCLUSIONSNeuroinflammation due to CTLA4h is mediated primarily by an infiltrative process with a distinct and striking dissociation between clinical symptoms and radiological findings in the majority of patients.FUNDINGNIAID, NIH, Division of Intramural Research, NINDS, NIH, Division of Intramural Research, and the National Multiple Sclerosis Society-American Brain Foundation.TRIAL REGISTRATIONClinicalTrials.gov NCT00001355.

Keywords: Autoimmune diseases; Genetic diseases; Inflammation; Neuroimaging; Neuroscience.

Conflict of interest statement

Conflict of interest: The authors have declared that no conflict of interest exists.

Figures

Figure 1. Flowchart summarizing participant categorization in this study.

The diagram shows the inclusion and categorization of patients included for study in this analysis. Probands with systemic disease include individuals with CTLA4 mutations and any immune-mediated disease.

Figure 2. Representative CTLA4h MRI findings in patients with parenchymal inflammatory lesions.

Patient numbers given here correspond to the data in Table 2. (A and B) Patient 2 showed worsening migrainous headaches with papilledema. T2-FLAIR image (A); T1-weighted contrast-enhanced image (B). (C and D) Patient 7 showed a focal motor seizure with secondary generalization and trace left triceps weakness. T2-FLAIR image (C); T1-weighted contrast-enhanced image (D). (E and F) Patient 5 had new headaches and a normal neurological examination. T2-FLAIR image (E); T1-weighted contrast-enhanced image (F). (G and H) Patient 2 had a return of migrainous headaches following cessation of chronic immune-suppressive medication and a normal examination. Images in G are T2-weighted spinal cord images, and images in H are T1-weighted contrast-enhanced spinal cord images. (I–K) Serial MRI scans of patient 4. Time from scan in I to scan in J: 1 month; time from scan in J to scan in K: 1 year. (L) Change in lesion volume from maximum volume (MRI 1) to last available MRI (MRI 2).

Figure 3. CTLA4h neuroinflammation can relapse at high frequency.

Patient numbers given here correspond to the data in Table 2. Serial MRI scans were compared for new and contrast-enhancing lesions. Black circles indicate MRI scans with new T2-FLAIR or contrast-enhancing lesions; white circles indicates MRI scans without new T2-FLAIR or contrast-enhancing lesions; and orange circles indicate MRI scans without a new lesion but with persistent contrast enhancement.

Figure 4. CTLA4h-associated neuroinflammation often starts in the leptomeninges.

LME on MRI can precede parenchymal lesion formation. Patient numbers given here correspond to the data in Table 2. (A–D) Patient 9 developed headaches and a complex partial seizure attributed to a mesial temporal lobe lesion (not shown). MRI scans at clinical onset (A and B) and 11 days later (C and D) show a fast temporal evolution of LME (yellow chevron) to a parenchymal lesion (blue chevron). T2-FLAIR images (A and C); T1-weighted images after contrast (B and D). (E–L) Patient 13 was clinically asymptomatic and had extensive LME (yellow chevrons) on sequential MRIs, each done approximately 1 month apart. T2-FLAIR images (E, G, I, and K); T2-FLAIR images after contrast (F, H, J, and L).

Figure 5. CSF is inflamed in CTLA4h.

Flow cytometric data comparing patients with CTLA4h with active neuroinflammatory lesions on MRI (black), a prior history of neuroinflammatory lesions (green), and no history of inflammatory lesions (orange) on MRI with a cohort of healthy controls (NDs, white circles). All graphs depict values from blood (left half of each panel) and CSF (right half). (A) Total frequency of CD4+ and CD8+ T cells within total lymphocyte counts and the ratio of CD4+ to CD8+ T cells. (B) Percentage of memory T cell subtypes (CD45RA–CD27+) in CD4+ T cells and CD8+ T cells, and memory Tfh cells (CD45RA–CXCR5+) in CD4+ T cells (right). (C) Frequency of total B cells and memory B cell subtypes, including unswitched memory (IgD+CD27+) and switched memory (IgD–CD27+) B cells. For all flow cytometric data, the Mann-Whitney U test was performed for comparisons between ND and CTLA4 cohorts, within each compartment (i.e., peripheral blood and CSF). Significance was set at P < 0.05. Each flow cytometric experiment for lymphocyte subsets from peripheral blood that was paired with a CSF sample obtained at the same time represents a single experiment, given the nature of the specimen (CSF obtained by LP) and the limited amount of specimen that could be obtained, as well as the sensitivity for the timing of the procedure according to the clinical status and the treatment course of the patient.

Figure 6. Brain biopsy shows inflammation in CTLA4h brain lesions.

(A–D) Right frontal deep white matter lesion showing cellular infiltrate on H&E staining, with infiltration of CD20+ B cells (C) and CD3+ T cells (D). Original magnification, ×100 (A) and ×400 (B–D). (E–J) Left parietal leukocortical white matter lesion showing cellular infiltrate on H&E staining, with evidence of CD138+ plasma cells (H) and both IgG (I) and IgM (J) production. Original magnification, ×100 (E), ×200 (F and H–J), and ×400 (G). (K–M) Right frontal white matter lesion and (N–P) right occipital white matter lesion showing cellular infiltrate on H&E staining (K and N), with variable degrees of astrocytosis (L and O) and preserved neuropil (M and P). Original magnification, ×200.