Serum Biomarkers in Neuro-Ophthalmology: When to Test

Myasthenia Gravis – LRP4/Agrin

Myasthenia gravis causes fatigable weakness due to an autoimmune disruption of neuromuscular synaptic transmission. Approximately 40% of patients diagnosed with myasthenia gravis present with ocular symptoms including diplopia and ophthalmic signs including fatigable ptosis, Cogan’s lid twitch, and pupil-sparing ophthalmoplegia.⁷ Of these, approximately 38% of cases remain ocular. The mechanism of antibodies targeting the muscle-specific nicotinic acetylcholine receptor (the post-synaptic receptor required for synaptic transmission at the neuromuscular junction) was uncovered by Patrick and Lindstrom in 1973.⁸ Despite a clear pathogenic link, acetylcholine receptor (AChR) antibodies are only detectable in about 50% of patients with ocular myasthenia gravis. Muscle-specific kinase (MuSK), a tyrosine kinase required for formation and maintenance of the neuromuscular junction, was identified as another marker of MG, and according to a retrospective study of 82 patients seropositive for MuSK, only 3 (3.7%) of the patients presented with isolated ocular myasthenia gravis.⁹ Among patients with generalized myasthenia gravis, AChR antibodies are positive in about 80% of patients, and MuSK antibodies are positive in 10%.^9–12

To address the need for additional biomarkers, antibodies targeting proteins that associate with the AChR and its downstream targets have recently been investigated in patients negative for both AChR and MuSK antibodies, termed double-seronegative myasthenia gravis (DNMG). Agrin is a proteoglycan that activates MuSK and aggregates AChRs at the neuromuscular junction.^13–15 0–50% of DNMG patients test positive for agrin antibodies.^16–18 Notably, mutations in the AGRN gene, which encodes agrin, have been implicated in congenital myasthenic syndrome.^19,20 Agrin also binds low-density lipoprotein receptor–related protein 4 (LRP4), which serves to activate MuSK and facilitates clustering and stabilization of the AChR at the neuromuscular junction.²¹ The reported prevalence for LRP4 antibodies among DNMG patients also ranges widely between 0.14% and 50%.^16,17 Rivner et al. tested 181 DNMG patients for LRP4 and agrin antibodies and found 154 (85.1%) were negative for both, 27 (14.9%) were positive for at least one, and 23 (12.7%) patients were positive for both LRP4 and agrin antibodies.¹⁶ LRP4 antibody positivity has a female predominance of 59% and approximately 20% maintain isolated ocular manifestations years after initial presentation.^7,16 LRP4-IgG is less commonly associated with thymus abnormalities (1 of 15 patients who underwent chest imaging in the Rivner et al. study had thymic hyperplasia and 5 underwent thymectomy prior to study entry) and typically portends a good response to pyridostigmine and immunosuppressant therapy.¹⁶ The clinical implications of these markers are still being elucidated, and ongoing work in this area will reveal the diagnostic and management importance of identifying these markers.

When should LRP4 and agrin antibody tests be sent? The LRP4 antibody test should be sent out for patients presenting with isolated ocular myasthenia gravis for whom the AChR antibodies (binding, blocking, and modulating) have already returned negative. That being said, single fiber EMG is the most sensitive test for isolated ocular myasthenia gravis.^22–24 Agrin antibodies are not yet commercially available, and their clinical utility may be limited in DNMG patients.

MOG-IgG

Myelin oligodendrocyte glycoprotein (MOG)-IgG-associated disorder is characterized by optic neuritis (bilateral in 38% of seropositive patients), often in association with optic disc edema, myelitis, and acute disseminated encephalomyelitis (ADEM).⁴⁰ There is a slight female predominance (57%) and wide age range of 2–79 years. Younger adults are more likely to present with unilateral and recurrent disease and older adults (>45 years) with bilateral optic neuritis.⁴⁰ Children can have recurrent optic neuritis and ADEM; in the Chen et al. study, 12/16 MOG-IgG-positive patients with ADEM were under 18 years old.^40,41 MRI often shows enhancement of the intraorbital optic nerve and nerve sheath and is more likely to exhibit extensive, longitudinal (>50% nerve length) retrobulbar nerve lesions than MS or even NMOSD (Figure 1).^30,31 Enhancement of the perineural orbital tissue (“perineuritis”) is common. Lesions of the deep gray nuclei may also be present.^30,31 Compared to NMOSD, spinal cord imaging is more likely to reveal thoracolumbar and conus lesions than cervical lesions. In a study by Sato et al., all six MOG seropositive patients with spinal cord lesions involved the thoracic and/or lumbar region, and only one had a cervical lesion.⁴² A systematic review performed by Vosoughi et al. on 62 patients with MOG-associated disease reported 29 patients with afferent visual manifestations beyond optic neuritis that included uveitis, peripheral ulcerative keratitis, acute macular neuroretinopathy, neuroretinitis, venous stasis retinopathy, large pre-retinal macular hemorrhage, orbital inflammatory syndrome, and orbital apex syndrome.⁴³

Recurrent attacks of optic neuritis occur in 80% of MOG-IgG-positive patients within 3 years of their first attack and are frequently associated with severe vision loss (often to the count-fingers level). Optical coherence tomography may demonstrate peripapillary retinal nerve fiber layer thinning and ganglion cell layer/complex thinning that worsens with each recurrence.⁴⁰ Significant CSF pleocytosis (>100 cells/mm³) and the absence of oligoclonal bands can be suggestive of MOG-IgG-associated disease. Patients are both corticosteroid-responsive and frequently suffer recurrent flares with attempted tapering. Corticosteroid-sparing agents including azathioprine, rituximab, and mycophenolate mofetil are recommended.^41,44

When should MOG antibody testing be sent? MOG antibody testing should be sent in patients with atypical optic neuritis who have optic disc edema, corticosteroid-responsive vision loss, with CSF pleocytosis, and the aforementioned neuroimaging findings. Given its more recent discovery, the utility of routine testing for MOG-IgG in all patients with clinical optic neuritis is unclear.

Paraneoplastic Autoimmune Syndromes

Clinical manifestations of paraneoplastic syndromes result from autoimmune attack on normal tissue that is triggered by ectopic protein expression by an underlying neoplasm. Neuro-ophthalmic manifestations of paraneoplastic conditions often precede a cancer diagnosis, highlighting the importance of early recognition and timely work-up.

Opsoclonus–Myoclonus Syndrome

Opsoclonus (high-frequency, conjugate, and chaotic multidirectional saccadic movements) and myoclonus (involuntary sudden jerking movements of the limbs) are hallmark features of opsoclonus–myoclonus syndrome (OMS), which is strongly associated with malignancy.^45–48 In the pediatric population, OMS manifests in the context of neuroblastoma in 50% of cases (with the remainder believed to be post-infectious).^45,49,50 In adults, OMS is most commonly associated with: small cell lung cancer, ovarian cancer, and breast cancer.^51,52 Opsoclonus is suspected to result from antibody-mediated dysfunction of omnipause neurons that gate saccadic eye movement).^51,53

Paraneoplastic antibodies associated with OMS were discovered in 1985 and 1988, respectively: antineuronal nuclear antibody type 1 (ANNA-1) (seen with SCLC and neuroblastoma) and antineuronal nuclear antibody type 2 (ANNA-2) (seen with small cell lung cancer and breast carcinoma).^51,54 The pathologic mechanism by which ANNA-1 and 2 mediate their effects remains unclear, but postmortem studies have uncovered an associated loss of cerebellar Purkinje cells and pontine neurons in the parapontine reticular formation.⁵⁵ ANNA-1-positive OMS also manifests with limbic encephalitis, encephalomyelitis, polyneuropathy and sensory neuronopathies, and dysautonomia. In the Lucchinetti et al. study, 2/162 (1.25%) ANNA-1-positive patients had OMS. ANNA-2-positive OMS is associated with jaw dystonia, larygospasm, and brainstem encephalitis. In the work-up of OMS, urine catecholamine levels should be sent for homovanillic and vanillylmandelic acids that may suggest neuroblastoma. Even if these antibodies are negative, a comprehensive evaluation for cancer should be performed, including imaging of the chest, abdomen and pelvis, mammography, and (if unrevealing) whole-body fludeoxyglucose positron emission tomography (FDG-PET).

Treatment is targeted to the underlying malignancy, in addition to IV corticosteroids, intravenous immune globulin, or plasma exchange for the acute manifestations, with subsequent corticosteroid-sparing immunotherapy long term.⁵⁵

When should ANNA-1 and ANNA-2 be sent? When patients present with clinical manifestations that are suspected to represent OMS. Negative results do not exclude the possibility of malignancy.

Paraneoplastic Optic Neuropathy

Paraneoplastic optic neuropathies are typically seen in middle-aged and older patients presenting with painless, subacute visual loss that can be bilaterally simultaneous or rapidly sequential. The characteristic features vary according to the underlying autoantibody, many of which are actively being studied.

CRMP5 – Optic Neuropathy with Retinitis and Vitritis

Cross et al. and Sawyer et al. reported the phenotypic overlap between lung cancer patients with carcinoma-associated retinopathy and CRMP5-IgG-associated optic neuropathy.^56,57 CRMP5 is a neuronal cytoplasmic protein that regulatesneurite outgrowth during development and regeneration. CRMP5-IgG-associated disease presents with retinitis and often bilateral optic neuropathy without optic nerve enhancement on MRI.⁵⁶

Cohen et al. reviewed 29 CRMP5-IgG seropositive patients with neuro-ophthalmic manifestations, the majority of which preceded a cancer diagnosis.⁵⁸ Of the 59% with anterior visual pathway or intraocular findings, initial median visual acuity was 20/50, 82% were diagnosed with an optic neuropathy, and all who underwent fundoscopic examination at onset had optic disc edema, often associated with uveitis, retinitis or both. None of the patients showed retrobulbar optic nerve enhancement on MRI,⁵⁸ consistent with prior studies,^56,59–61 suggesting that CRMP5-IgG does not cause optic neuritis as was initially asserted. Ocular motility dysfunction secondary to central nystagmus was present in 41% of patients.⁵⁸ Visual function improved in 50% of the cohort with immunosuppressive therapy. Associated neurologic manifestations (in order of decreasing frequency) included neuropathic disorders (predominantly peripheral neuropathy and polyradiculoneuropathy), cerebellar ataxia, limbic encephalitis, cognitive decline, autonomic dysfunction, choreoathetosis, myelopathy, and Lambert–Eaton syndrome.

Systemic work-up for malignancy is positive in 62–87% of the patients positive for CRMP5-IgG.^56,58 Small-cell carcinoma is the most common underlying malignancy (83%), though it can also be seen in mixed small-cell lung cancer and non-small-cell lung cancer, breast ductal carcinoma in situ, and thymoma.^56,58

When should CRMP5-IgG testing be sent? Serologic testing for CRMP5-IgG should be sent in patients with optic disc edema without optic nerve enhancement on MRI, particularly in conjunction with retinitis and vitritis.

GFAP – Optic Papillitis

IgG antibodies targeting glial fibrillary acidic protein (GFAP), an astrocytic cytoplasmic intermediate filament protein, have been associated with inflammatory optic disc edema in 40% of patients with corticosteroid-responsive GFAP-IgG-positive meningoencephalitis, an entity originally described by Fang et - al.^62–64 Chen et al. reviewed the ophthalmic findings of 10 patients with GFAP IgG-associated meningoencephalitis with bilateral disc edema.⁶⁴ Among those, three (30%) had mild vitritis and preserved central visual acuity, but two (20%) developed arcuate visual field defects with corresponding ganglion cell layer thinning and clinical optic atrophy upon resolution of the disc edema.⁶⁴ Fluorescein angiography revealed venular leakage in one patient.⁶⁴ Two patients (20%) had mildly elevated opening pressures on lumbar puncture (265 and 298 mm H₂O).

Neurologic manifestations of GFAP-IgG-associated disease include headache, encephalopathy, postural tremor, and cerebellar ataxia.⁶⁴ MRI brain classically shows perivascular radial enhancement extending from the ventricles or cerebellum without associated retrobulbar optic nerve signal change.^62–65 Associated malignancies are reported in 14–38% of patients with GFAP-IgG-associated disease, including teratomas, prostate and GI adenocarcinomas, melanomas, and carcinoid tumors.^63,64,66 IV methylprednisolone should be initiated on presentation followed by high-dose oral corticosteroids with a duration based upon clinical judgment. Though we have no intention of conflating the two paraneoplastic syndromes, there have been case reports of optic neuritis with N-methyl-D-aspartate receptor (NMDAR) antibody-associated encephalitis, which was the first cell-surface antibody-mediated autoimmune encephalitis reported.^67–69 There are also reports of efferent manifestations in NMDAR antibody-mediated encephalitis, including ocular bobbing and OMS, but this antibody is better known for its association with neuropsychiatric symptoms, seizures, and movement disorders.^70,71

When should GFAP-IgG testing be sent? Serum and CSF GFAP-IgG should be sent in patients with bilateral optic papillitis without associated retrobulbar optic nerve enhancement on MRI. Radial perivascular enhancement extending from the ventricles or cerebellum on brain MRI, certainly in the setting of meningoencephalitis, should raise suspicion for GFAP-IgG-related disease.