Current Biomarker Strategies in Autoimmune Neuromuscular Diseases

Current Biomarker Strategies in Autoimmune Neuromuscular Diseases: History

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Subjects: Neurosciences

Contributor:

Menekse Oeztuerk

Antonia Henes

Christina B. Schroeter

Christopher Nelke

Paula Quint

Lukas Theissen

Sven G. Meuth

Tobias Ruck

Inflammatory neuromuscular disorders encompass a diverse group of immune-mediated diseases with varying clinical manifestations and treatment responses. The identification of specific biomarkers has the potential to provide valuable insights into disease pathogenesis, aid in accurate diagnosis, predict disease course, and monitor treatment efficacy.

CIDP
biomarkers
GBS
myasthenia gravis
neuromuscular diseases
IIM
inflammation
myositis

1. Introduction: The Need for Biomarkers in Inflammatory Neuromuscular Disorders

Inflammatory neuromuscular disorders are a heterogeneous group of immune-mediated diseases with diverse underlying pathomechanisms. Epidemiology, clinical manifestations, treatment strategies, and responses vary across the spectrum of disease. Common to all is a potential severe burden of disease with conceivable long-lasting disability. Designated criteria for categorisation of affected individuals into corresponding subgroups are well-established [1]. Over the last few years, our pathophysiological understanding of autoimmune inflammatory neuromuscular disorders has steadily improved. However, essential pathogenic processes remain to be studied. In this regard, the recognition of specific biomarkers could confer additional insights while informing treatment decisions. Biomarkers are characteristic features of biological processes and are detectable and quantifiable in body fluids and tissues [2]. As valuable indicators, they serve, inter alia, diagnostic, prognostic, and therapeutic purposes in diseases.

Considering the rarity and diversity of clinical manifestation of neuromuscular disorders (NMDs), the identification of specific biomarkers for each of them is essential, particularly regarding disease course prediction and improvement of daily clinical practice. In recent years, a considerable development on this matter has emerged. However, there is still a lack of objective biomarkers suitable in NMDs.

2. On the Concept of Biomarkers

The appliance of biomarkers has become increasingly relevant over the last decade. As useful tools, they serve various aspects in disease management. Biomarkers are indicators of both physiological mechanisms and pathogenic processes or responses to various interventions and treatment regimens in general [2,3]. Particularly in diagnostic, prognostic, and predictive aspects, biomarkers can contribute as helpful tools. The detection of the disease of interest is achieved by diagnostic biomarkers. The presence or alteration of a predictive biomarker forecasts probabilities of incidents following the exposure to an intervention or environmental factor [2]. Prognostic biomarkers aid in the estimation of clinical course and severity in the observed condition. Correspondingly, monitoring biomarkers can be employed in longitudinal disease assessment, detecting the status of a condition or measuring treatment effects. Detection of biomarkers may offer insights into causative pathomechanisms. Hence, biomarkers are crucial to the development of treatment strategies including targeted therapies, assisting healthcare for affected individuals and the population.

3. Biomarkers in GBS and CIDP

3.1. Current Biomarkers in GBS and CIDP

Few recognized biomarkers of GBS and CIDP are presently integrated in diagnostics and monitoring of disease courses and treatment responses. An overview of relevant biomarkers in use is given in Table 1.

Table 1. Current biomarkers in autoimmune neuromuscular diseases.

Disease	Type of Biomarker	Biomarker	Detection Method	Correlation	Occurence	References	Limitations/Comment
GBS/CIDP	Diagnostic	Lipooligosaccharides (LOS) with A, B, C, E, F and H loci; Serotype and sequence type of Campylobacter jejuni	PCR screening, genes from published LOS loci and sequencing Serum and CSF	Identification of C. jejuni-associated GBS	GBS and Miller-Fisher syndrome (MFS)	[4,5]
	Diagnostic	Antibodies against Campylobacter jejuni DNA-binding protein (C-Dps)	ELISA, Western Blot Serum	Detection of anti-C-Dps-IgG indicates C. jejuni related GBS	Campylobacter jejuni-related GBS	[6]	Also detected in patients with C. jejuni enteritis (rarely)
	Diagnostic	Metallo-proteinases (MMPS) (MMP-9, TIMP-1)	ELISA Serum	No correlation	Described in CIDP	[7]	Not disease-specific
	Diagnostic	Antibodies to peptides from myelin proteins P0, P2_14–25, PMP22 and connexin 32	Antigen-specific proliferation assay, Immunoprecipitation, Western Blot Serum	No correlation	Described in GBS and CIDP	[8,9,10,11,12,13,14]
	Diagnostic Monitoring	Sphingomyelin (SM)	Fluorescence-based assay CSF	Correlation with disease activity, elevated in active CIDP.	Increased in GBS and CIDP	[15]
	Diagnostic	Cystatin C (cysteine protease inhibitor)	ELISA CSF	Decrease may be linked to higher cathepsin B activity (cathepsin B levels increased in CSF)	Significant decrease of cystatin c levels in GBS and CIDP patients	[16,17,18,19]	Decrease also observed in MS patients
	Diagnostic	Protein 14-3-3	Immunoblot assay CSF	Early detection (12 to 48 h after disease onset) in GBS	Elevated in CSF of GBS and CIDP patients	[20,21]	Not disease-specific
	Diagnostic	IL-8	Multiplex bead immunoassays CSF	Aid in differentiation between CIDP and GBS, including acute-onset CIDP. CSF IL-8 in GBS > CIDP Optimal IL-8 cutoff → 70 pg/mL	Should be measured initially during diagnostical process High specificity and positive predictive value	[22]	Not disease-specific
	Predictive Prognostic	Autoantibodies to gangliosides (GM1, GA1, GD1a, GD1b, GalNAc-GD1a, 9-O-Acetyl GD1b, GD3, GM1, GT1a, GT1b, GT3, GQ1b, 0-Acetyl GT3, LM-1, GD1a/GD1b, GM1/GalNac-GD1a, GM1/PA, GM1/GD1a, GM1/GT1b, LM1/GA1) IgG and IgM	ELISA Serum	Correlation with clinical phenotypes and specific symptoms of GBS e.g., ophthalmoplegia and Anti-GQ1b IgG Anti-GM1 linked to Campylobacter jejuni, titers correlate with clinical recovery and therapy response GM1/GalNac-GD1a linked to respiratory infection	High prevalence in GBS of Anti-GM1 and Anti-GT1a	[23,24,25,26,27,28,29]
	Predictive Prognostic	Antibodies against nodal and paranodal proteins: Neurofascin (Nfasc155 and Nfasc140/186) Contactin-1 (CNTN1) Contactin- associated protein-1 (Caspr1) Caspr1/CNTN1 complex Gliomedin	ELISA, Immunoprecipitation, cell-based Assays Serum	Associated with specific clinical manifestation e.g., ataxia and tremor Poorer response to IVIg Anti-CNTN1 seem to benefit from corticosteroids	Nfasc155 in 4–18% of CIDP cases	[30,31,32,33,34,35,36,37,38]
	Predictive Prognostic	Neurofilaments Phosphorylated neurofilament heavy protein (pNFH) Neurofilament light chain (Nfl)	ELISA, Electrochemiluminescence (ECL) based immunoassay Serum and CSF	Indicator for neurodegeneration Positive correlation with clinical and electrophysiological presentation in GBS Association with disease progression and therapy outcome in CIDP	Elevated in both serum and CSF of GBS and CIDP patients	[25,39,40,41,42,43]	General indicators of axonal damage, also detectable in other patient groups with evidence of structural CNS damage
	Predictive Prognostic	Tau-proteins	ELISA CSF	Correlation with clinical manifestation and poorer clinical outcome	Elevated in CSF of GBS and CIDP patients	[25,42,44,45]	Also detectable in other patient groups with evidence of structural CNS damage e.g., Alzheimer’s Disease
	Prognostic	Autoantibodies against galactocerebroside (Gal-C)	ELISA Serum	Association with sensory deficits and autonomic disruption in GBS Association with Mycoplasma pneumoniae infection	GBS	[46,47]
	Prognostic	Neuron-specific enolase (NSE)	Enzyme immunoassay methods CSF	Higher levels correlate with a longer duration of disease	Elevated in CSF of GBS and CIDP patients	[48,49,50]	CSF-NSE is not GBS or CIDP specific; Elevation is also observed other conditions e.g., Creutzfeldt-Jakob disease [51]
	Prognostic Predictive Monitoring	Cytokines Interferon gamma (IFN γ), Tumor necrosis factor α (TNF α), Transforming growth factor β1 (TGF β1), IL-1β, IL-4, IL-6, IL-10, IL-12, IL-16, IL-17, IL-18, IL-22, IL-23, IL-37	ELISA based assays (multiplexed fluorescent bead-based immunoassay Serum and CSF	TNF α and IFN γ are elevated in GBS and correlate with clinical severity TGF β1 levels are decreased in the early course of GBS, downregulation correlates with clinical disability Serum levels are positively correlated with GBS disease severity and decreased after IVIg treatment (IL-17A, IL-37) CSF IL-17A levels were positively correlated with clinical manifestation of GBS Upregulation of IL-4 and IL-10 are linked to recovery phase in GBS CSF and serum levels of interleukins declined after IVIg treatment	Cytokine elevation is described in CIDP and GBS	[52,53,54,55,56,57,58,59,60,61,62,63,64,65]	Not disease-specific.
	Prognostic Monitoring	Serum complement proteins C3, C3a, C5a, C5b-9	Nephelometry Serum	Upregulation predicts poor prognosis High C3 correlates with complement activation with high C3a und C5a Correlation with disease activity	Increased in GBS and CIDP	[66,67,68]	Not disease-specific
	Prognostic Monitoring	S100B protein (calcium-binding astroglial protein)	ELISA Serum and CSF	Elevated in CSF and serum Association with clinical severity and poor prognosis Decrease in stable disease course	Elevated in GBS and CIDP	[42,49,50]	Expression of S100B is not restricted to neural tissue; Serum levels can be increased after e.g., bone fractures or hepatic injury [69,70]
	Prognostic Diagnostic	Stem cell factor (SCF) Hepatocyte growth factor (HGF)	Multiplex bead-based ELISA CSF	Increased	Elevated levels in CSF CIDP > GBS Correlation with chronicity	[71]	Value of this examination is still uncertain
	Monitoring	Chemokines CCR2, CCL7,CCL3, CCL27, CCR1, CCR5, CXCL10, CXCR3, CXCL9, CXCL12, monocyte chemoattractant protein 1 (MCP-1)	Multiplex bead-based ELISA Serum and CSF	CCR2 and MCP-1 decreased in the recovery stage	Increased in GBS and CIDP	[25,72,73,74]	Not disease-specific
	Monitoring	Intercellular adhesion molecule 1 (ICAM-1) Vascular cell adhesion molecule 1 (VCAM-1) Vascular Endothelial Growth Factor (VEGF)	Multiplex bead-based ELISA CSF	Decrease after therapy in studies Correlation with repair processes is currently being studied	Increased in GBS and CIDP	[71,75,76]	Not disease-specific
	Monitoring	MicroRNAs has-miR4717-5p (GBS) has-miR-642b-5p (GBS) miR-31-5p (CIDP)	Microarray, droplet digital PCR Serum	High levels of miR-31-5p correlate with longer disease duration Potential in improving personalized patient care	Detectable in GBS and CIDP	[77,78,79,80]
MG	Diagnostic Monitoring	Anti-AChRs (muscle nicotinic acetylcholine receptors) IgG subtype 1 and 3	Radioimmunprecipitation assay (RIPA) with high specificity and sensitivity, fixed cell-based assays Serum	Monitoring in patients with immunosuppressive treatment Higher levels in ocular MG are associated with conversion to generalized MG Higher levels in late-onset MG	85% in generalized MG, highly specific for MG	[81,82,83,84,85,86,87,88,89,90,91,92,93,94,95,96,97,98,99]	Lower titers in ocular MG Inconsistent studies regarding correlation with disease severity and treatment response
	Diagnostic Prognostic Predictive	Anti-MuSK (Muscle-specific kinase) IgG subtype 4	Radioimmunoprecipitation assay (RIPA), ELISA, cell-based assay (CBA) Serum	Correlation with disease severity Affection of facial-bulbar muscles Early crises and challenging treatment Worse outcome Association with early onset MG and better response to rituximab	5–8% of MG 30–50% of AChR-negative MG	[100,101,102,103,104,105,106,107,108,109,110]	More often in female patients Highest sensitivity in detection via CBA
	Diagnostic	Anti-LRP4 (low-density lipoprotein receptor-related protein 4) IgG subtype 1 and 2	Cell-based assay (CBA) Serum	Stronger clinical manifestation than in seronegative MG	2% of MG; higher in non-AChR and non-MuSK cases	[111,112,113,114,115,116,117]	Higher prevalence in female patients Not specific for MG (e.g., found in ALS as well)
	Diagnostic Prognostic	Anti-Titin	ELISA, Cell-based assay (CBA) Serum	Thymoma-associated MG More frequent hospitalization	20–40% in Anti-AChR-positive MG	[95,118,119,120]	Screening for thymoma presence should follow positive testing
	Diagnostic Prognostic	Anti-Kv1.4 (voltage gated potassium channel)	Cell-based assay (CBA), Radioimmunoprecipitation assay (RIPA) Serum	Association with myasthenic crises and thymoma Association with bulbar manifestation, myocarditis and QT-Time prolongation (Japanese population)	11–18% (Japanese MG population)	[95,121,122]	More frequent in female patients Expensive detection
	Diagnostic	Anti-Rapsyn	ELISA Serum	No correlations	15% of MG	[95,123]	Not MG-specific
	Diagnostic Prognostic	Anti-Cortactin	Western Blot, ELISA Serum	Mild symptoms	10–25% of MG	[95,124,125,126]	Not MG specific
	Diagnostic Prognostic Predictive	Anti-Agrin IgG subtype 1 and 3	ELISA; CBA; serum	Correlation with limited therapeutic response and mild to severe clinical manifestation	2–5% of MG, mainly seropositive MG	[95,127,128]	More frequent in male patients
	Diagnostic Prognostic Predictive	Micro-RNAs miR-150-5p miR-21-5p miR-30e-5p let-7 miRNA family	Microarray, droplet digital PCR Serum	Correlation with treatment response High miR-30e-5p levels are associated with risk of generalization in ocular MG	Studied in AchR and MuSK-positive MG	[129]
IIM	Diagnostic Predictive Prognostic	Anti-Mi-2	ELISA, Immunoblot Serum	Classical DM, associated with beneficial prognosis, mild myositis, lower risk of ILD, better treatment response especially to rituximab Correlation with disease activity Association with HLA-DR7	MSA 2–45% prevalence in DM	[130,131,132,133,134,135]	Positive sera may also be found by ELISA in PM patients [136] Prevalence can only be estimated (varying among different countries)
	Diagnostic Predictive Prognostic	Anti-ARS (aminoacyl-tRNA synthetases) Jo-1, PL-7, PL-12, EJ, OJ, KS, ZO, YRS	RNA-Immunoprecipitation, ELISA, Line Blots, Serum	Association with ASyS Higher mortality and interstitial lung disease (ILD) incidence in non-Anti-Jo-1-ARS (+) Higher treatment dosage required	MSA Anti-Jo-1 15–30% in DM/PM Others < 5%	[134,137,138,139,140,141]	Low prevalence of non-Anti-Jo-1-ARS RNA-Immunoprecipitation not widely available Rates of false-positive cases higher in Line Blots [140]
	Diagnostic Predictive Prognostic	Anti-NXP2 (anti-nuclear matrix protein 2)	Immunoprecipitation, Western Blot Serum	Association with calcinosis and severe myositis, cancer development Correlation with disease activity	MSA Adult and juvenile DM 1–5%	[134,142,143,144,145]	Immunoassays have been released and are currently discussed
	Diagnostic Predictive Prognostic Monitoring	Anti-MDA-5 (Melanin differentiation-associated protein-5)/CADM140	Immunoprecipitation, Western Blot, ELISA Serum	Associated with clinically amyotrophic DM (CADM), ILD, poor prognosis, sever skin manifestation Titer levels linked to disease severity and outcome	MSA 15–20% in IIM, mainly CADM	[133,134,146,147]	Higher prevalence in Asia More frequent in women
	Diagnostic Prognostic	Anti-TIF1γ/α (transcription factor 1 γ/α)	ELISA, Immunoprecipitation Serum	Malignancy-associated DM	MSA 10–15%, higher prevalence in cancer-associated DM, rare in PM	[134,148,149,150,151,152]	Cancer association is applied to adults [151]
	Diagnostic Predictive Prognostic	Anti-SAE (small ubiquitin-like modifier activating enzyme)	Immunoprecipitation, Indirect Immunofluorescence test Serum	Cancer association Serum levels correlate with disease activity	MSA 1–5% in DM	[153,154,155]
	Diagnostic Prognostic	Anti-SRP (Anti-signal recognition particle)	RNA Immunoprecipitation, ELISA Serum	Associated with a rapidly progressive disease course with severe weakness Cancer-associated SRP-IMNM	MSA 20–25% in IMNM	[156,157,158]	More frequent in women Primarily in adults
	Diagnostic Prognostic	Anti-HMGCR (3-hydroxy-3-methylglutaryl-coenzyme A reductase)	Immunoprecipitation, ELISA Serum	Significant association with serum creatine kinase Higher serum muscle enzymes than in other IIM Correlation with disease activity Cancer association	MSA 6–12% in IIM	[158,159,160,161,162,163,164,165]	Cave: Statin therapy!
	Predictive Prognostic	Serum soluble CD163	ELISA Serum	Biomarker for macrophage activation Correlation with disease severity Association with Anti-MDA5 (+) cases	PM/DM	[166,167,168]	Not IIM-specific; supporting
	Diagnostic	Anti-cN1A (cytosolic 5′-nucleotidase 1A)	Addressable laser bead immunoassay (ALBIA), ELISA Serum	No correlation with disease severity	Around 50% in IBM	[169,170]	Moderate sensitivity, high specificity in ALBIA [171] Not IBM-specific, found also in known autoimmune diseases e.g., SLE [172]
	Diagnostic	Micro-RNAs miR-96-5p	RTqPCR Serum	No correlation described	Upregulation in PM, DM and Anti-Jo1 positive cases	[173]

Immune-mediated mechanisms following antecedent infections, commonly with a subset of Campylobacter jejuni strains with ganglioside-mimicking lipooligosaccharides (LOS), result in the typical clinical phenotype of progressive ascending symmetrical paresis of the limbs with hypo- to areflexia in GBS [174]. CIDP is an autoimmune neuropathy affecting peripheral nerves. The common clinical hallmark is the symmetrical weakness of distal and proximal portions of the limbs, whereas pure motor, pure sensory, and focal subtypes are described equally. A diagnostic delay occurs frequently in CIDP [174].

Impairments of the blood-nerve barrier and the blood-cerebrospinal fluid (CSF) barrier as barriers of the PNS are concomitant with the pathophysiology underlying GBS and CIDP. Tissue of peripheral nerves, serum, and CSF compose the predominant origins of biomarkers [25]. Biomarkers can also be linked to immediate damage of the PNS.

4. Biomarkers in MG

4.1. Current Biomarkers in MG

MG is a chronic antibody-mediated autoimmune disease leading to focal or generalized muscle fatigability including respiratory symptoms or dysarthria [196,197]. Exclusive ocular symptoms (ocular MG) are possible and often represent the first clinical manifestation. Disease exacerbations inducing myasthenic crisis and ICU admission are still frequently observed. Causative auto-Abs target different components of the neuromuscular junction (NMJ) and disrupt regular transmission [198]. The prevalence is stated to be around 150–300 per million population [199]. The age of 50 years is used to distinguish between early-onset MG (EOMG) and late-onset MG (LOMG), as two peaks of incidence have been recognized [200,201]. In most cases (85%), auto-Abs against the extracellular domain of muscle nicotinic acetylcholine receptors (AChRs) are detected [84,85,86]. Biomarkers applied in MG are primarily disease-underlying auto-Abs and antigenic structures. An overview is given in Table 1. Specifically in the case of MG, the assignment of biomarkers to the initially introduced subgroups is rather ambiguous, as many functions are simultaneously fulfilled.

5. Biomarkers in IIM

5.1. Current Biomarkers in IIM

Idiopathic inflammatory myopathies (IIM) are a rare heterogenous cluster of autoimmune-mediated diseases affecting mainly skeletal muscles. Alongside typical manifestations with muscle weakness and fatiguing, IIMs are often accompanied by specific organ manifestations, including skin and lungs, among others. IIMs can be subclassified into different groups—dermatomyositis (DM), polymyositis (PM), immune-mediated necrotizing myopathy (IMNM), antisynthetase syndrome (ASyS), inclusion body myositis (IBM), and overlap myositis (OM) [223]. Importantly, clinical presentations, treatment responses, and prognoses differ strongly throughout subgroups [224]. Several supporting biomarkers, almost all of them auto-Abs, have been identified in the past and serve understanding causative mechanisms (Table 1). Nevertheless, IIMs are still deeply underdiagnosed.

Non-specific muscle enzymes including creatine kinase (CK), aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), and aldolase are elevated due to muscle damage and do not necessarily correlate with clinical severity or disease activity [225]. Due to the challenges involved in accurately assigning biomarkers to specific subgroups in IIMs, the researchers have chosen to focus separately on myositis-specific Abs (MSAs) and further biomarkers of interest. The respective classifications can be found in Table 1, delineating the different categories.

This entry is adapted from the peer-reviewed paper 10.3390/cells12202456

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