Autoantibody profiles in patients with immune checkpoint inhibitor-induced neurological immune related adverse events

Abstract

Background: Neurological immune-related adverse events (irAE-n) are severe and potentially fatal toxicities of immune checkpoint inhibitors (ICI). To date, the clinical significance of neuronal autoantibodies in irAE-n is poorly understood. Here, we characterize neuronal autoantibody profiles in patients with irAE-n and compare these with ICI-treated cancer patients without irAE-n.

Methods: In this cohort study (DRKS00012668), we consecutively collected clinical data and serum samples of 29 cancer patients with irAE-n (n = 2 pre-ICI, n = 29 post-ICI) and 44 cancer control patients without irAE-n (n = 44 pre- and post-ICI). Using indirect immunofluorescence and immunoblot assays, serum samples were tested for a large panel of neuromuscular and brain-reactive autoantibodies.

Results: IrAE-n patients and controls received ICI treatment targeting programmed death protein (PD-)1 (61% and 62%), programmed death ligand (PD-L)1 (18% and 33%) or PD-1 and cytotoxic T-lymphocyte-associated protein (CTLA-)4 (21% and 5%). Most common malignancies were melanoma (both 55%) and lung cancer (11% and 14%). IrAE-n affected the peripheral nervous system (59%), the central nervous system (21%), or both (21%). Prevalence of neuromuscular autoantibodies was 63% in irAE-n patients, which was higher compared to ICI-treated cancer patients without irAE-n (7%, p <.0001). Brain-reactive autoantibodies targeting surface (anti-GABA_BR, -NMDAR, -myelin), intracellular (anti-GFAP, -Zic4, -septin complex), or unknown antigens were detected in 13 irAE-n patients (45%). In contrast, only 9 of 44 controls (20%) presented brain-reactive autoantibodies before ICI administration. However, seven controls developed de novo brain-reactive autoantibodies after ICI initiation, therefore, prevalence of brain-reactive autoantibodies was comparable between ICI-treated patients with and without irAE-n (p = .36). While there was no clear association between specific brain-reactive autoantibodies and clinical presentation, presence of at least one of six selected neuromuscular autoantibodies (anti-titin, anti-skeletal muscle, anti-heart muscle, anti-LRP4, anti-RyR, anti-AchR) had a sensitivity of 80% (95% CI 0.52-0.96) and a specificity of 88% (95% CI 0.76-0.95) for the diagnosis of myositis, myocarditis, or myasthenia gravis.

Conclusion: Neuromuscular autoantibodies may serve as a feasible marker to diagnose and potentially predict life-threatening ICI-induced neuromuscular disease. However, brain-reactive autoantibodies are common in both ICI-treated patients with and without irAE-n, hence, their pathogenic significance remains unclear.

Keywords: autoimmunity; cancer immunotherapy; immune checkpoint inhibitors; immune related adverse events; myositis; neuronal autoantibodies; neurotoxicity; paraneoplastic syndromes.

Figure 1

CONSORT diagram. Between September 2017 and January 2022, a total of 86 cancer patients were enrolled for the study. After application of inclusion and exclusion criteria, 73 patients were included. As two patients that were originally allocated to the control cohort developed irAE-n, a total of 29 irAE-n patients and 44 cancer control patients were analyzed. CSF, cerebrospinal fluid; ICI, immune checkpoint inhibitor; irAEs, immune related adverse events; irAE-n, neurological immune related adverse event.

Figure 2

Serum autoantibody profiles in cancer patients with irAE-n and controls. (A) Frequency and specification of neuromuscular autoantibodies of 41 cancer patients before (upper left) and six weeks after (middle left) ICI treatment initiation who did not develop high-grade (= CTCAE ≥ 3) irAEs and 24 ICI-treated cancer patients with irAE-n (lower left). (B) Frequency and specification of brain-reactive autoantibodies of 44 cancer patients before (upper right) and six weeks after (middle right) ICI treatment initiation who did not develop high-grade (CTCAE ≥ 3) irAEs and 29 ICI-treated cancer patients with irAE-n (lower right). Ab, autoantibody; AchR, acetylcholine receptor; CASPR2, contactin-associated protein-like 2; GABA_BR, gamma-aminobutyric-acid B receptor; GFAP, glial fibrillary acidic protein; Homer-3, Homer protein homolog 3; ICI, immune checkpoint inhibitor; irAEs, immune related adverse events; irAE-n, neurological immune related adverse event; ITPR1, inositol 1,4,5-trisphosphate receptor 1; LRP4, low-density lipoprotein receptor-related protein 4; MuSK, muscle-specific tyrosine kinase; NMDAR, N-methyl-D-aspartate receptor; P/Q VGCC, P/Q-type voltage gated calcium channel; RyR, ryanodine receptor; Zic4, zinc finger 4.

Figure 3

Histopathological and clinical findings in patients with ICI-induced myositis. (A-F) Hematoxylin-eosin (HE) and immunohistochemical staining in a female patient with fatal ICI-induced myositis and myocarditis and positive anti-titin, anti-heart muscle and anti-skeletal muscle autoantibodies. Frozen sections of HE-stained (A) cardiac muscle (D) and skeletal muscle showing necrotic myofibers and lymphocyte infiltration (original magnification x 100). CD8 (large panel, original magnification x 200) and major histocompatibility complex (MHC) class I (inset, original magnification x 100) staining revealing cytotoxic T cell invasion and sarcolemmal overexpression of MHC class I by (B) cardiac and (E) skeletal muscle fibers. CD68 (large panel, original magnification x 200) and MHC class II (inset, original magnification x 100) staining showing macrophage invasion and sarcolemmal as well as sarcoplasmic overexpression of MHC class II by (C) cardiac and (F) skeletal muscle fibers. (G) Comparison of peripheral blood creatine kinase (CK) levels in patients with ICI-induced myositis shows a trend towards higher CK levels in patients with multiple neuromuscular autoantibodies compared to patients without neuromuscular autoantibodies. (H) Sagittal and (I) axial contrast-enhanced magnetic resonance imaging of the lumbar spine with T1-weighted turbo spin echo sequence showing paravertebral contrast enhancement (arrows) as a sign of muscle edema in a patient with ICI-induced myositis and myasthenia. Ab, autoantibody; ICI, immune checkpoint inhibitor; U/l, units per liter.

Figure 4

Brain-directed autoantibodies in ICI-treated cancer patients with irAE-n. (A, B) Cell-based assays of fixed recombinant HEK293-cells (original magnification x 200) expressing single neuronal antigens showing anti-Zic4 autoantibodies (1:100 dilution; A) and anti-GABA_BR autoantibodies (1:10, B) in a patient with small cell lung cancer and Guillain-Barré syndrome (GBS) after pembrolizumab treatment. However, anti-ganglioside autoantibodies, which are associated with GBS, were tested negative during clinical routine. (C) Tissue-based assay using indirect immunofluorescence screening on monkey cerebellum sections (original magnification x 200) showing anti-GFAP autoantibodies (1:100 dilution) in a melanoma patient with encephalitis after treatment with ipilimumab and nivolumab. GABA_BR, gamma-aminobutyric-acid B receptor; GFAP, glial fibrillary acidic protein; ICI, immune checkpoint inhibitor; irAE-n, neurological immune related adverse event; Zic4, zinc finger 4.