Abstract
Introduction
Multiple sclerosis (MS) is a chronic neuroimmunological condition associated with acute relapses. Differentiating between acute symptom development related to autoimmune inflammatory demyelination versus pseudoexacerbation remains challenging. We aimed to evaluate the utility of a commercially available multi-analyte protein assay in (1) assessing acute patient-reported symptoms suggestive of clinical exacerbations by comparing the acquired proteomic results to the corresponding MRI findings, and (2) examining the role in the surveillance of disease in individuals exposed and non-exposed to FDA-approved disease-modifying therapies.
Methods
A retrospective observational study was conducted within a single academic neuroimmunology clinic among individuals with an existing diagnosis of MS who had paired Octave® Multiple Sclerosis Disease Activity (MSDA) Test and MRI results prior to glucocorticosteroid exposure. Participants were evaluated in two cohorts: (1) individuals with acute clinical symptoms concerning for an exacerbation and (2) individuals seen for routine disease surveillance. The timing of the MSDA Test and MRI studies were quantified along with the association between the MSDA composite score and MRI-confirmed relapses.
Results
Sixty-six individuals with relapsing–remitting MS were included, with 33 being evaluated for acute neurological symptoms and 33 for routine disease surveillance. Among symptomatic individuals, MSDA testing preceded MRI in 79% of cases. Across both cohorts, a Disease Activity Score (DAS) ≥ 6.0–10.0 was highly sensitive and specific for predicting gadolinium-enhancing lesions. Elevated concentrations of myelin oligodendrocyte glycoprotein (p = 0.013) and neurofilament light (NfL) (p < 0.0001), and reduced B-cell activating factor levels (p < 0.0001), were observed in participants with gadolinium enhancement. The DAS predicted enhancement more accurately than NfL alone (area under the curve = 98.06% vs. 82.75%; p = 0.0061).
Conclusion
The use of blood-based biomarkers may enable more timely and precise assessment of acute disease activity, improving clinical decision-making. Additionally, the MSDA Test demonstrated greater accuracy than NfL for identifying MRI-confirmed relapses.
Keywords: Clinical relapse, Multiple sclerosis, Multi-analyte protein assay, Neurofilament, Pseudoexacerbation
Key Summary Points
| Why carry out this study? |
| Multiple sclerosis (MS) is a chronic demyelinating disease where distinguishing true inflammatory relapses from pseudoexacerbations remains difficult. |
| The study compared proteomic results acquired from a commercial blood-based multi-analyte protein assay to MRI findings in individuals with and without clinical symptoms suggestive of an acute relapse. |
| We hypothesized that the proteomic results would discriminate MRI-confirmed relapses better than neurofilament light (NfL) alone. |
| What was learned from the study? |
| The multi-analyte blood-based assay identified significant proteins associated with gadolinium enhancement on MRI, including elevated levels of myelin oligodendrocyte glycoprotein and NfL, and decreased B-cell activating factor. |
| Multi-protein blood-based measures were associated with MRI evidence of acute disease activity and may provide a biologically grounded and temporally relevant measure of acute inflammatory demyelination related to MS. |
Introduction
Multiple sclerosis (MS) represents a spectrum of demyelinating presentations [1] with varying degrees of inflammatory activity and symptom expression, affecting nearly 1 million people in the U.S. [2]. Use of U.S. Food and Drug Administration (FDA) approved disease-modifying therapy (DMT), within differing therapeutic classes [3–11], has been shown to reduce clinical exacerbations, MRI activity associated with autoimmune inflammatory demyelination, and neurological disability. Distinguishing the report of acute symptoms attributable to new MS disease activity within the brain and/or spinal cord [12] from those resulting from a “pseudoexacerbation” or symptoms resulting from recent illness, elevated ambient temperatures, activity, or other external influences remains an important unmet need.
Beyond the primary goal of preventing neurological disability related to MS, clinicians aim to reduce the development of acute exacerbations to prevent cumulative inflammatory events, decrease the risk of incomplete recovery from a relapse, and minimize the likelihood of persistent negative symptoms that impair quality of life. Exacerbations may occur as experienced symptoms, new MRI activity (asymptomatic or symptomatic) within the brain and/or spinal cord, or both. As new asymptomatic inflammatory changes on MRI outnumber the report of clinical symptoms [13], the application in the surveillance of disease has been essential in guiding both acute and long-term treatment decisions. However, the financial costs associated with testing along with scheduling access in the context of acute symptoms creates challenges for timely recommendations for care. The management of clinical symptoms is more challenging, at times requiring immediate clinical visits, evaluations in urgent care or an emergency room setting, and laboratory testing involving blood and urine.
The use of neurofilament light chain (NfL), a protein that is released in the setting of axonal damage, is an established biomarker in MS, with elevations detected prior to clinical symptom onset [14] and prior to the development of a gadolinium-enhancing lesions on MRI [15, 16]. Recent literature also suggests an association with disability progression [17, 18]. Intuitively, the inclusion of other proteins would add to this foundation, offering improvements over single protein strategies and serving as a complimentary or potentially independent technology to conventional MRI techniques.
In this study, we evaluated the utility of a commercial blood-based multi-analyte protein assay in (1) assessing acute patient-reported symptoms suggestive of clinical exacerbations through comparison of proteomic results with MRI-confirmed disease activity, and (2) examining the role in the routine surveillance for individuals exposed and non-exposed to an FDA-approved DMT. We hypothesize that distinct circulating proteins are associated with MRI-confirmed relapses. Additionally, we evaluated if a multi-analyte approach was superior in predicting gadolinium enhancement when compared to a single protein measure.
Methods
Research Participants
Following implementation of Octave® Multiple Sclerosis Disease Activity (MSDA) testing in April 2025, people with MS seen in the Multiple Sclerosis and Neuroimmunology Clinic at The University of Texas Southwestern Medical Center between April 2025 and February 2026 were assessed for eligibility. Inclusion criteria were comprised of (1) males and females ≥ 18 years of age having (2) an established diagnosis of MS fulfilling 2017 McDonald criteria [19] provided by Board Certified Neurologists with sub-specialty training in clinical neuroimmunology, (3) availability of paired MSDA Test results and MRI data acquired in the setting of reported acute neurological symptoms lasting at least 24 h or for disease surveillance, (4) MSDA Test results and MRI data acquired prior to glucocorticosteroid exposure or other acute treatments, and (5) availability of clinical follow-up data extending at least 30 days following para-clinical testing. Individuals with incomplete medical records and/or compromised biomarker samples were excluded. Two groups were created that represented (1) individuals with acute clinical symptoms concerning for an exacerbation related to MS and (2) individuals seen for routine disease surveillance (Fig. 1). Both groups contained the consecutive inclusion of all eligible subjects. The study was performed in accordance with the Strengthening the Reporting of Observational Studies in Epidemiology guidelines.
Fig. 1.
Flowchart demonstrating the inclusion process for those included in the analysis
The study was approved by The University of Texas Southwestern Medical Center Institutional Review Board (STU-072016-012) and has, therefore, been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki and later amendments. A waiver for consent was granted based on the retrospective study design.
Octave® Multiple Sclerosis Disease Activity Test
The MSDA Test is an analytically [20] and clinically validated [21] commercially available blood-based multi-analyte assay composed of 18 unique proteins developed to characterize biological disease activity in MS. Twelve of the 18 proteins are correlated to increased disease activity related to MS and the remaining 6 are inversely correlated (Table 1). The composite Disease Activity Score (DAS) is generated using an algorithm that incorporates individual protein concentrations with corrections for age and sex, when applicable [20]. The DAS ranges from 1.0 to 10.0 in 0.5-increments and is categorized as low (1.0–4.0), moderate (4.5–7.0), or high (7.5–10.0) disease activity.
Table 1.
Proteins included in the Octave® Multiple Sclerosis Disease Activity Test and relationship to disease activity
| Proteins correlated with disease activity | Proteins inversely correlated with disease activity |
|---|---|
| Cluster of differentiation 6 (CD6) | Amyloid beta precursor like protein 1 (APLP1) |
| Contactin 2 (CNTN2) | B-cell Activating Factor (TNFSF13B) |
| C-X-C motif chemokine ligand 13 (CXCL13) | CUB domain-containing protein 1 (CDCP1) |
| Glial fibrillary acidic protein (GFAP) | Interleukin 12 B (IL-12B) |
| Leucine-rich repeat transmembrane (FLRT2) | Osteoprotegerin (OPG) |
| MIP 3-Alpha (CCL20) | TRAIL-R1 (TNFRSF10A) |
| Monokine induced by gamma interferon (CXCL9) | |
| Myelin oligodendrocyte glycoprotein (MOG) | |
| Neurofilament light (NfL) | |
| Osteopontin (OPN) | |
| Protogenin (PRTG) | |
| Serpin family a member 9 (SERPINA9) |
Acute Multiple Sclerosis Exacerbation Versus Pseudoexacerbation Definition
An acute MS relapse was defined by the presence of MRI-confirmed gadolinium-enhancing lesions in the brain and/or spinal cord, based on formal interpretations by board-certified neuroradiologists blinded to MSDA results, in the presence or absence of acute symptoms suggestive of autoimmune inflammatory demyelination. Episodes in which individuals reported acute neurological symptoms but lacked MRI confirmation of disease activity were classified as pseudoexacerbations. For all individuals with reported symptoms, standard medical evaluations were performed to assess for acute infection, fever, recent illness, and psychosocial stressors.
Statistical Analysis
All analyses were performed in R (version 4.5.3), and all figures were generated using the ggplot2 package.
The time between the MSDA Test and MRI was calculated as the number of days between the two assessments, with positive values indicating that the MSDA Test preceded the MRI and negative values indicating that the MRI preceded the MSDA Test. To determine if there were differences in the time between MSDA Test and MRI between those with acute symptoms versus those undergoing testing for disease surveillance, a Mann–Whitney U test was performed.
To test for differences in MSDA between those with gadolinium enhancement and those without gadolinium enhancement, a Mann–Whitney U test was performed. A receiver operating curve was constructed to estimate the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of various DAS cut-offs to classify the presence/absence of gadolinium enhancement, and 95% Clopper–Pearson confidence intervals were computed. Lastly, the accuracy of the DAS and 18 proteins of interest to predict gadolinium enhancement was estimated, in the form of the area under the curve (AUC) and the corresponding 95% confidence interval (CI). Interest was then in determining if the AUCs of the DAS and proteins of interest were different than that of NfL. To test this, differences in the AUCs were computed and corresponding p values were estimated using Delong’s approach. A positive difference in AUCs indicated that a given protein had a greater AUC than NfL and improved predictive accuracy. The age- and sex-adjusted values, when applicable, for all 18 proteins were used to evaluate the ability of each protein to predict the presence or absence of gadolinium enhancement.
Significance was defined as a p value < 0.05 and no adjustments for multiple testing were performed.
Results
A total of 66 individuals with relapsing–remitting MS served as the study cohort. The majority of included subjects were non-Hispanic (61/66, 92.4%), white (55/66, 83.3%), and female (53/66, 80.3%). Table 2 provides a summary of the baseline and clinical information by group.
Table 2.
Baseline demographic and clinical data from all study participants
| Report of acute clinical symptoms | Disease surveillance | |
|---|---|---|
| Number of subjects (n) | 33 | 33 |
| Median (range) age (years) | 49.3 (31.7–73.0) | 47.1 (19.3–78.3) |
| Females, n (%) | 26 (78.8) | 27 (81.8) |
| Males, n (%) | 7 (21.2) | 6 (18.2) |
| Ethnicity, n (%) | ||
| Hispanic | 4 (12.5) | 1 (2.9) |
| Non-Hispanic | 29 (87.5) | 32 (97.1) |
| Race, n (%) | ||
| White | 30 (90.9) | 25 (75.8) |
| African American | 0 (0.0) | 4 (12.1) |
| Asian | 2 (6.1) | 2 (6.1) |
| Asian Indian | 1 (3.0) | 2 (6.1) |
| Median (range) disease durationa (years) | 6.4 (0.11–31.1) | 10.1 (0.05–36.2) |
| Number exposed to disease modifying therapy (%) | 12 (36.4) | 12 (36.4) |
| Median (range) treatment duration (years) | 1.7 (0.26–7.6) | 0.9 (0.14–4.9) |
| Disease modifying therapy types (n, %) | ||
| Cladribine | 0 (0.0) | 2 (16.7) |
| Diroximel fumarate | 1 (8.3) | 0 (0.0) |
| Generic dimethyl fumarate | 1 (8.3) | 1 (8.3) |
| Generic fingolimod | 0 (0.0) | 1 (8.3) |
| Generic teriflunomide | 0 (0.0) | 1 (8.3) |
| Glatiramer acetate | 0 (0.0) | 1 (8.3) |
| Natalizumab | 2 (16.7) | 0 (0.0) |
| Ocrelizumab | 3 (25.0) | 3 (25.0) |
| Ofatumumab | 1 (8.3) | 2 (16.7) |
| Ozanimod | 2 (16.7) | 1 (8.3) |
| Ublituximab-xiiy | 2 (16.7) | 0 (0.0) |
aSince diagnosis
Of the 33 individuals with reported acute clinical symptoms possibly associated with new inflammatory CNS demyelinating events, a range of reported clinical difficulties were described by participants (Table 3). A total of 15 symptom categories were identified with participants reporting a total of 50 symptoms, while 13 individuals described having more than one symptom. The most common symptoms were unilateral sensory (10/50, 20.0%), followed by gait abnormalities (6/50, 12.0%) and cognitive difficulty (5/50, 10.0%) complaints. Table 3 provides additional details involving symptom categories and frequency of events. A total of 64 region-specific MRI studies were performed—28 brain, 24 cervical spine, and 12 thoracic spine—with 25 of 33 subjects having MRI studies of the brain and at least one region within the spinal cord. For individuals evaluated for acute neurological symptoms, the MSDA Test was completed prior to the MRI in 79% (26/33) of individuals with DAS ranging from 6.0 to 9.5.
Table 3.
Symptom categories and frequency of events
| Symptoms | Frequency (n, %) |
|---|---|
| Unilateral sensory | 10 (20.0) |
| Gait abnormalities | 6 (12.0) |
| Cognitive difficulties | 5 (10.0) |
| Bilateral sensory | 4 (8.0) |
| Unilateral motor | 4 (8.0) |
| Bilateral motor | 4 (8.0) |
| Fatigue | 3 (6.0) |
| Unilateral sensorimotor | 3 (6.0) |
| Coordination difficulties | 2 (4.0) |
| Impaired balance | 2 (4.0) |
| Increased neuropathic pain | 2 (4.0) |
| Truncal spasticity | 2 (4.0) |
| Bilateral sensorimotor | 1 (2.0) |
| Dizziness | 1 (2.0) |
| Visual disturbance | 1 (2.0) |
In addition, 33 individuals without acute clinical symptoms underwent both MRI and MSDA testing as part of routine disease surveillance. In this group, a total of 31 brain, 19 cervical spine, and 18 thoracic spine MRI studies were performed. Asymptomatic gadolinium-enhancing lesions were observed in six individuals. In the routine clinical surveillance of disease, the MSDA Test was typically performed closer to or after the MRI [median time: – 4 days (range: – 19, 24; interquartile range (IQR): – 10, – 1)] in contrast to those with reported acute symptoms where the median time for testing preceded the MRI by nearly a week [median time: 6 days (range: – 33, 94; IQR: 0.0, 21.5)] (p = 0.0002).
For individuals being evaluated for acute disease activity, gadolinium-enhancing lesions were observed in 10 individuals. Seven of ten individuals reported complete resolution of acute symptoms following acute glucocorticosteroid treatment. Figure 2 illustrates the significant difference in the DAS between those with and without gadolinium enhancement (p < 0.0001), and Table 4 summarizes the performance of various MSDA Test DAS thresholds in detecting gadolinium enhancement in the setting of acute symptoms or routine disease surveillance.
Fig. 2.
Box plots demonstrating significant differences in the Disease Activity Score between all individuals with and without gadolinium enhancement
Table 4.
Performance characteristics and 95% Clopper–Pearson confidence intervals (CI) of varying Octave® Disease Activity Score thresholds for predicting gadolinium-enhancing lesions
| Octave multiple sclerosis disease activity test score | Sensitivity (95% CI) |
Specificity (95% CI) |
Positive predictive value (95% CI) |
Negative predictive value (95% CI) |
Accuracy (95% CI) |
|---|---|---|---|---|---|
| > 6.0–10.0 |
100.00% (79.41%, 100.00%) |
94.00% (83.45%, 98.75%) |
84.21% (60.42%, 96.62%) |
100.00% (92.45%, 100.00%) |
95.45% (87.29%, 99.05%) |
| > 6.5–10.0 |
87.50% (61.65%, 98.45%) |
96.00% (86.29%, 99.51%) |
87.50% (61.65%, 98.45%) |
96.00% (86.29%, 99.51%) |
93.94% (85.20%, 98.32%) |
| > 7.0–10.0 |
75.00% (47.62%, 92.73%) |
96.00% (86.29%, 99.51%) |
85.71% (57.19%, 98.22%) |
92.31% (81.46%, 97.86%) |
90.91% (81.26%, 96.59%) |
| > 7.5–10.0 |
56.25% (29.88%, 80.25%) |
98.00% (89.35%, 99.95%) |
90.00% (55.50%, 99.75%) |
87.50% (75.93%, 94.82%) |
87.88% (77.51%, 94.62%) |
| > 8.0–10.0 |
43.75% (19.75%, 70.12%) |
98.00% (89.35%, 99.95%) |
87.50% (47.35%, 99.68%) |
84.48% (72.58%, 92.65%) |
84.85% (73.90%, 92.49%) |
| > 8.5–10.0 |
43.75% (19.75%, 70.12%) |
100.00% (92.89%, 100.00%) |
100.00% (59.04%, 100.00%) |
84.75% (73.01%, 92.78%) |
86.36% (75.69%, 93.57%) |
For individuals with gadolinium enhancement when compared to those without, significant elevations in proteins correlated with acute disease activity were observed for myelin oligodendrocyte glycoprotein (MOG) (p = 0.013) and NfL (p < 0.0001). An equivocally significant decrease for contactin 2 (CNTN2) (p = 0.048) was also observed. Additionally, a reduction in concentration for a protein inversely correlated with acute disease activity, B-cell activating factor (TNFSF13B) (p < 0.0001) was identified (Fig. 3).
Fig. 3.
Box plots demonstrating the concentrations of 18 individual proteins of interest in all individuals with and without gadolinium enhancement. BAFF = B-cell activating factor
The DAS was superior in predicting gadolinium enhancement (AUC = 98.06%, 95% CI = 95.33%, 100%), as compared to NfL alone (AUC = 82.75%, 95% CI = 71.42%, 94.08%), with a statistically significant difference of 15.3% (p = 0.0061). Although TNFSF13B had an AUC comparable to that of NfL (AUC = 84.50%, 95% CI = 72.70%, 96.30%), statistical significance was not achieved. Table 5 provides the performance data for the DAS and all 18 proteins.
Table 5.
Comparative receiver operator curve analysis and corresponding 95% confidence intervals (CI) of the Octave® MSDA Test Disease Activity Score and 18 proteins for predicting gadolinium enhancement, with area under the curve differences relative to neurofilament light chain
| Measure | Area under the curve (95% CI) | Difference in area under the curve (%) (relative to neurofilament light chain) | |
|---|---|---|---|
| Estimate (95% CI) | p value | ||
| Disease activity score | 98.06% (95.33%, 100.00%) | 15.31% (4.37%, 26.25%) | 0.0061 |
| B-cell activating factor (TNFSF13B) | 84.50% (72.70%, 96.30%) | 1.75% (–17.14%, 20.64%) | 0.8522 |
| Neurofilament light (NfL) | 82.75% (71.42%, 94.08%) | 0.00% (0.00%, 0.00%) | – |
| Myelin oligodendrocyte glycoprotein (MOG) | 70.75% (56.41%, 85.09%) | –12.00% (–27.88%, 3.88%) | 0.1385 |
| Contactin 2 (CNTN2) | 66.56% (52.08%, 81.04%) | –16.19% (–33.25%, 0.87%) | 0.0585 |
| CUB domain-containing protein 1 (CDCP1) | 66.00% (49.49%, 82.51%) | –16.75% (–40.92%, 7.42%) | 0.1774 |
| Interleukin 12 B (IL-12B) | 65.37% (48.55%, 82.20%) | –17.38% (–41.78%, 7.03%) | 0.1519 |
| Amyloid beta precursor like protein 1 (APLP1) | 64.25% (47.42%, 81.08%) | –18.50% (–38.16%, 1.16%) | 0.0651 |
| C-X-C motif chemokine ligand 13 (CXCL13) | 63.62% (49.64%, 77.61%) | –19.13% (–36.40%, –1.85%) | 0.0300 |
| Monokine induced by gamma interferon (CXCL9) | 62.50% (45.29%, 79.71%) | –20.25% (–45.39%, 4.89%) | 0.1065 |
| Protogenin (PRTG) | 61.87% (46.68%, 77.07%) | –20.88% (–39.39%, –2.36%) | 0.0256 |
| TRAIL-R1 (TNFRSF10A) | 59.12% (43.87%, 74.38%) | –23.63% (–38.20%, –9.05%) | 0.0015 |
| Osteoprotegerin (OPG) | 58.31% (43.70%, 72.92%) | –24.44% (–37.87%, –11.00%) | 0.0004 |
| MIP 3-Alpha (CCL20) | 57.62% (41.97%, 73.28%) | –25.13% (–48.48%, –1.77%) | 0.0316 |
| Glial fibrillary acidic protein (GFAP) | 56.87% (41.86%, 71.89%) | –25.88% (–42.08%, –9.67%) | 0.0017 |
| Serpin family a member 9 (SERPINA9) | 55.12% (38.87%, 71.38%) | –27.62% (–48.34%, –6.91%) | 0.0077 |
| Leucine-rich repeat transmembrane (FLRT2) | 52.88% (34.93%, 70.82%) | –29.87% (–49.83%, –9.92%) | 0.0034 |
| Cluster of differentiation 6 (CD6) | 49.25% (32.48%, 66.02%) | –33.50% (–51.11%, –15.89%) | 0.0002 |
| Osteopontin (OPN) | 45.62% (29.03%, 62.22%) | –37.13% (–59.38%, –14.87%) | 0.0010 |
Discussion
We have demonstrated that a commercially available, blood-based, multi-analyte proteomic assay provided patient specific information associated with an MRI-confirmed acute relapse. We have further characterized the real-world timing of testing relative to MRI, demonstrating that, in the setting of acute neurological symptoms suggestive of an MS relapse, the MSDA Test was frequently obtained prior to imaging. In the setting of acute neurological symptoms and in the routine clinical surveillance of individuals, a DAS ranging between ≥ 6.0 and 10.0 was identified to be highly sensitive and specific in the prediction of a gadolinium-enhancing lesion within the CNS. Additionally, significant elevations in MOG and NfL were observed along with a reduction in concentration for TNFSF13B in association with MRI-confirmed clinical relapses. We also identified that the multi-analyte DAS was superior to NfL alone in the prediction of gadolinium enhancement in all studied subjects.
The utility of serum biomarkers has been studied in traumatic brain injury [22], Parkinson’s Disease [23], and Alzheimer’s Disease [24]. The identification of disease-specific proteins represents a remarkable advancement towards understanding early disease biology. Such learnings may translate into an earlier diagnosis, provide prognostic information, or serve as a guide to therapy initiation or optimizing regimens depending on the current management approach. The MSDA Test is a commercially available laboratory test that has been both analytically [20] and clinically [21] validated, demonstrating the value of the 18-protein multi-analyte panel in association with objective acute MRI disease activity. A higher DAS ranging from 4.5 to 10.0 increases the odds of having ≥ 1 gadolinium-enhancing lesion on MRI by 4.49 compared to scores ranging from 1.0 to 4.0 [21]. Additionally, scores ranging from 7.5 to 10.0 increase the odds for ≥ 2 gadolinium-enhancing lesions on MRI by 20.99, relative to scores ranging from 1.0 to 7.0 [21].
A subset of individuals had a DAS ranging from ≥ 6.0 to 10.0 with no detectable gadolinium-enhancing lesions, contributing to false positives in our sensitivity and specificity calculations. This observation may be the result of MRI negative relapses influenced by technical factors associated with MRI, regions of the CNS being imaged, and variations in scanning protocols [25]. However, all the MRI studies were performed at 3.0 Tesla in this study. Elevations in the DAS may also reflect early biological disease activity that precedes overt structural changes detectable by MRI. By comparison, a DAS ranging from 1.0 to 5.5 may correspond to a pseudoexacerbation as indicated by our data.
Within both academic and community medical practices, the use of empiric glucocorticosteroids in the management of acute symptoms related to MS is frequently employed despite the lack of objective evidence for a current infection, changes in neurological examination findings, or the presence of gadolinium-enhancing lesions or new and/or newly enlarging T2-weighted hyperintense lesions. At times, acute treatment is prescribed in response to patient insistence, to meet patient expectations, to minimize workflow pressures, or to address the mounting pressure of patient satisfaction scores that may be related to reimbursement of services. However, risks are inherent with exposure. Patient-reported adverse effects of high-dose intravenous methylprednisolone treatment are common with change in taste, facial flushing, GI pain, sleep disturbances, appetite changes, agitation, and behavioral changes being reported [26]. The risk for liver injury [27], steroid-induced avascular necrosis [28], mood disorders [29], along with consistent elevations in blood glucose [30] also exists.
Our findings also indicated that the multi-analyte protein assay allowed for the determination of biological activity predictive of a clinical relapse earlier than data provided via MRI. In addition, the data appeared more efficient compared to the high number of region-specific MRI studies ordered and performed to address acute symptoms reported. The anticipated greater accessibility to people with MS relative to MRI availability, combined with the avoidance of cumulative systemic risks associated with gadolinium [31], offers a more rapid and safer solution for disease surveillance. Collectively, these advantages suggest that multi-analyte protein assays may represent a practical first-line method for assessing acute or impending disease activity.
In MRI-confirmed relapses, increased concentrations of MOG and NfL were observed. As MOG is a structural glycoprotein expressed on the outer surface of CNS myelin sheaths, produced by oligodendrocytes [32], and NfL, a protein marker of neuroaxonal injury, the observed elevations in those with gadolinium-enhancement are consistent with known pathophysiology and disease activity [33, 34]. Along with these findings, we also identified a reduction in concentration of TNFSF13B, a protein inversely correlated with acute disease activity. The observed reduction in TNFSF13B when gadolinium-enhancing lesions are present may reflect local consumption of this protein in the setting of acute inflammation [35]. This is further supported by evidence of increased TNFSF13B levels in anti-CD20-treated individuals where B cells are depleted [36, 37]. These findings highlight the potential of MOG, NfL, and TNFSF13B as blood-based markers for detecting acute MS lesions.
Our data also revealed that the multi-analyte assay approach outperformed NfL alone in the prediction of gadolinium enhancement on MRI. This observation is biologically plausible as overlapping inflammatory and neurodegenerative pathways would be more comprehensively represented using a composite biomarker strategy rather than relying on a single measure. Additionally, individual proteins may be dysregulated across multiple pathological conditions, limiting their specificity [38–41].
The provided results should be evaluated in the context of limitations. The study cohort was acquired from a single academic center and the included subjects may not be generalizable to other groups. The number of individuals studied was also of limited size. However, 24% of studied individuals were observed with gadolinium-enhancing lesions. Social determinants of health from participants were not captured; thus, external factors including socioeconomic and geographic barriers to care may have affected the timing of the provided services for those who reported acute symptoms. Body mass index and measures of renal function were not available and may have influenced NfL concentrations. The study was also retrospective in design. However, a prospective, randomized study attempting to demonstrate how the MSDA Test or similar platforms enhance clinical care that would require clinicians to be blinded to results from a commercially available multi-analyte protein assay may raise ethical concerns. In this context, blinding may withhold information capable of identifying current disease activity as well as elevated risk for disease evolution before becoming apparent both clinically and on MRI.
Conclusion
The utilization of multi-analyte blood-based biomarkers in medicine is expected to increase even further, providing rapid availability, disease-specific results, and anticipated lower cost to traditionally-based diagnostic approaches. Here, we identified the association between DAS ranges and gadolinium-enhancing lesions along with the value of multi-analyte measures over a single protein. Envisioning a future where MRI studies may be supplanted by blood-based multi-analyte protein assays, tailored to the desired management target for acute versus progressive disease, is becoming within reach given the scientific advancements in our understanding of these clinical courses. These data will also allow for more precise clinician–patient education using a biologically-informed model of care.
Acknowledgements
We sincerely thank all participants who contributed to this study.
Author Contributions
All authors meet the International Committee of Medical Journal Editors criteria (ICMJE) for authorship. Darin T. Okuda conceptualized the study and contributed to methodology, validation, and drafting of the manuscript. Tatum M. Moog, Isabella J. Huddleston, Crystal M. Wright, Katy W. Burgess, Jose R. Santoyo, and Peter V. Sguigna curated data, participated in investigation, and contributed to reviewing and editing the manuscript. Morgan C. McCreary curated and formally analyzed the data and contributed to drafting as well as reviewing and editing the manuscript. Olaf Stüve contributed to data curation, methodology, and manuscript review. Diem H. Tran contributed to data curation, investigation, methodology, and manuscript review. All authors read and approved the final manuscript.
Funding
No funding was received for this study or publication of this article. The Rapid Service Fee was funded by the authors.
Data Availability
The datasets generated and analyzed during this study are available from the corresponding author on reasonable request following successful completion of a data sharing agreement with The University of Texas Southwestern Medical Center.
Declarations
Conflict of Interest
Darin T. Okuda received personal compensation for consulting and advisory services from Amgen, Biogen Inc., Cortechs.ai, EMD Serono, Genentech, Genzyme/Sanofi, Immunic Therapeutics, Moderna, Octave Bioscience, Pfizer, and Zenas BioPharma along with research support from Alexion and Novartis, has issued national and international patents along with pending patents related to other developed technologies, and received royalties for intellectual property licensed by The Board of Regents of The University of Texas System. Crystal M. Wright received personal compensation from EMD Serono, Genentech, and TG Therapeutics. Katy W. Burgess received personal compensation from EMD Serono and Sanofi. Peter V. Sguigna received personal compensation for consulting services from Genentech, EMD Serono, Horizon Therapeutics, Amgen, and Bristol Myers Squibb, and has received research support from Clene Nanomedicine, Genentech, Novartis, and Zenas Biopharma. Olaf Stüve serves on the editorial boards of Therapeutic Advances in Neurological Disorders, Expert Review of Clinical Immunology, and he is a section editor for Current Treatment Options in Neurology, has served on data monitoring committees for Genentech-Roche, and Novartis without monetary compensation, has advised Cellarity, Lilly, Octave Bioscience, and Zenas BioPharma, receives grant support from EMD Serono. Tatum M. Moog, Morgan C. McCreary, Isabella J. Huddleston, Jose R. Santoyo, and Diem H. Tran report no disclosures.
Ethical Approval
The study was approved by The University of Texas Southwestern Medical Center Institutional Review Board (STU-072016-012) and has, therefore, been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki and later amendments. A waiver for consent was granted based on the retrospective study design.
Footnotes
The original online version of this article was revised to correct the sentence starting "Figure 2 illustrates the..." and to correct the Tables 4 and 5.
Change history
8/15/2026
The original online version of this article was revised to correct the sentence starting "Figure 2 illustrates the..." and to correct the Tables 4 and 5.
Change history
8/14/2026
A Correction to this paper has been published: https://doi.org/10.1007/s40120-026-01005-y
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The datasets generated and analyzed during this study are available from the corresponding author on reasonable request following successful completion of a data sharing agreement with The University of Texas Southwestern Medical Center.