Cinzia Cordioli

IFNβ bioavailability in multiple sclerosis patients: MxA versus antibody-detecting assays ☆

Anti-IFNbeta antibodies are related to IFNbeta bioavailability loss in multiple sclerosis. We investigated the reliability of radioimmunoprecipitation and cytopathic assays in detecting binding (BAbs) and neutralizing (NAbs) antibodies and the correlation of these antibodies to MxA mRNA production. Eleven percent of IFNbeta-treated patients showed a lack of MxA induction, with an inverse correlation between MxA mRNA and the presence of BAbs and NAbs. Some patients had contemporary MxA induction in the presence of high NAb titres, thus calling into question the reliability of cytopathic assay. Since anti-IFNbeta antibodies well correlated with MxA induction loss, MxA assay is an appropriate test to determine IFNbeta bioavailability.

Renewal of the T-cell compartment in multiple sclerosis patients treated with glatiramer acetate

The immunomodulating activity of glatiramer acetate on T-cells of multiple sclerosis patients has only been partially clarified. The objective of this work was to investigate whether glatiramer acetate modifies thymic release of newly produced T-cells and the peripheral composition of the T-cell repertoire. T-cell receptor excision circles, thymic naive (CD4+CD45RA+CCR7 +CD31+) T helper cells, and central (CD4+CD45RA -CCR7+) and effector (CD4+CD45RA-CCR7 -) memory T-cells were evaluated in 89 untreated patients, 84 patients treated for at least 1 year, and 31 patients beginning treatment at the time of inclusion in the study and then followed-up for 12 months; controls were 81 healthy donors. The T-cell repertoire was analysed in selected samples. The percentage of thymicnaive T helper cells was diminished in untreated patients, but rose to control values in treated subjects; a decrease in central memory T-cells was also observed in treated patients. Follow-up patients could be divided into two subgroups, one showing unmodified thymicnaive T helper cells and T-cell diversity, the other in which the increased release of new T-cells was accompanied by modifications of the T-cell repertoire. Glatiramer acetate modifies the peripheral T-cell pool by activating a thymopoietic pathway of T-cell release that leads to a different setting of T-cell diversity and, likely, to a dilution of autoreactive T-cells.

Modulation of IFNAR1 mRNA expression in multiple sclerosis patients.

Interferon-beta receptor (IFNAR) is composed of 2 subunits, IFNAR1 and IFNAR2, the latter of which is expressed as functional (IFNAR2.2), non-functional (IFNAR2.1) and soluble (IFNAR2.3) isoform. Real-Time PCR analysis of mRNA for all IFNAR components in multiple sclerosis patients naïve for therapy and undergoing long-term treatment with interferon-beta shows that IFNAR1 mRNA level is lower than in healthy controls. If long-term treated patients are divided according to the production of mRNA for Myxovirus protein-A, a marker of interferon-beta bioactivity, IFNAR1 mRNA reaches the values observed in controls only in Myxovirus protein-A-induced patients. Since chronic cell stimulation by interferon-beta induces IFNAR protein down-regulation, we suggest that the increase of IFNAR1 mRNA might serve as a mechanism for counterbalancing the loss of protein receptor, enhancing, at least in this sub-group of patients, cell responsiveness to interferon-beta.

MxA mRNA Quantification and Disability Progression in Interferon Beta-Treated Multiple Sclerosis Patients

Even though anti-interferon beta (IFNβ) antibodies are the main determinants of IFNβ bioactivity loss and Myxovirus-resistance protein A (MxA) is the most established marker of IFNβ biological activity in IFNβ-treated multiple sclerosis patients, their usefulness in the routine clinical practice is still debated. Therefore, 118 multiple sclerosis patients naïve for treatment were enrolled for a 3-year longitudinal observational study mimicking the conditions of a real-world setting. In order to evaluate the kinetics of bioactivity loss in blood samples obtained every 6 months after therapy initiation, MxA and interferon receptor isoform/subunit mRNA were quantified by real-time PCR, anti-IFNβ binding antibodies were detected by radioimmunoprecipitation, and neutralizing antibodies by cytopathic effect inhibition assay. Clinical measures of disease activity and disability progression were also obtained at all time points. We found that, at the individual-patient level, the response to IFNβ therapy was extremely heterogeneous, including patients with stable or transitory, early or late loss of IFNβ bioactivity, and patients with samples lacking MxA mRNA induction in spite of absence of antibodies. No interferon receptor isoform alterations that could explain these findings were found. At the group level, none of these biological features correlated with the measures of clinical disease activity or progression. However, when MxA mRNA was evaluated not at the single time point as a dichotomic marker (induced vs. non-induced), but as the mean of its values measured over the 6-to-24 month period, the increasing average MxA predicted a decreasing risk of short-term disability progression, independently from the presence of relapses. Therefore, a more bioactive treatment, even if unable to suppress relapses, reduces their severity by an amount that is proportional to MxA levels. Together with its feasibility in the routine laboratory setting, these data warrant the quantification of MxA mRNA as a primary tool for a routine monitoring of IFNβ therapy.

Assessing association of comorbidities with treatment choice and persistence in MS: A real-life multicenter study

Objective: To assess whether the presence of concomitant diseases at multiple sclerosis (MS) diagnosis is associated with the choice and the treatment persistence in an Italian MS cohort. Methods: We included newly diagnosed patients (2010–2016) followed in 20 MS centers and collected demographic and clinical data. We evaluated baseline factors related to the presence of comorbidities and the association between comorbidities and the clinical course of MS and the time to the first treatment switch. Results: The study cohort included 2,076 patients. Data on comorbidities were available for 1,877/2,076 patients (90.4%). A total of 449/1,877 (23.9%) patients had at least 1 comorbidity at MS diagnosis. Age at diagnosis (odds ratio 1.05, 95% confidence interval [CI] 1.04–1.06; p < 0.001) was the only baseline factor independently related to the presence of comorbidities. Comorbidities were not significantly associated with the choice of the first disease-modifying treatment, but were significantly associated with higher risk to switch from the first treatment due to intolerance (hazard ratio 1.42, CI 1.07–1.87; p = 0.014). Association of comorbidities with risk of switching for intolerance was significantly heterogeneous among treatments (interferon β, glatiramer acetate, natalizumab, or fingolimod; interaction test, p = 0.04). Conclusions: Comorbidities at diagnosis should be taken into account at the first treatment choice because they are associated with lower persistence on treatment.

Early detection and favourable outcome of natalizumab-related progressive multifocal leukoencephalopathy (PML) in two multiple sclerosis patients.

A 42-year-old woman with RRMS and EDSS level of 7, previously treated with interferon (IFN)-b-1a and mitoxantrone, was started with natalizumab. JCV antibodies status test was not available at that time. After 31 infusions a routine MRI showed right parieto-occipital T2WI-hyperintense lesion with subcortical gadolinium (Gd) enhancement (Fig. 1), thus natalizumab treatment was immediately stopped. Right neglect, apraxia and alexia were present and EDSS increased to 8.5. CSF JCV-PCR performed at National Institute of Health Laboratory of Molecular Medicine and Neuroscience-Maryland, USA, (NINDS) was positive (24 DNA copies). She received only two of the seven scheduled PLEX cycles because of thrombosis on the site of venous catheterization. Oral prednisone (32.5 mg/day) was started and continued for the next 18 months. Cognitive functions gradually improved during 23-month follow-up as well as neurological examination (EDSS decreased to 7.5). Serial brain MRI scans showed stabilization of lesion size with progressive gadolinium enhancement disappearance (Fig. 2). Case 2

Assessing long-term prognosis improvement as a consequence of treatment pattern changes in MS

Objective: To assess whether the age at which multiple sclerosis (MS) patients reach Expanded Disability Status Scale (EDSS) milestones changed as long as new drugs for the treatment of MS became available. Methods: We evaluated the long-term impact of therapies on disability progression assessing whether there is a detectable delay in the age at which patients reached EDSS milestones in more recent years. We used data collected over more than 30 years in the Center of Brescia, Italy. We compared the age at EDSS = 6 among patients diagnosed with relapsing-remitting MS in different time periods, adjusting for age at diagnosis and median interval among EDSS visits, by a multivariate Cox model. Results: A total of 1324 MS patients were included. Patients diagnosed in more recent periods reached EDSS = 6 at an older age: the rate at which patients reached EDSS = 6 in those diagnosed in 1991–1995 was similar to those diagnosed in 1980–1990 (hazard ratio (HR) = 1.09, p = 0.68) and to those diagnosed in 1996–2000 (HR = 0.85, p = 0.44), it was reduced by 37% in patients diagnosed in 2001–2005 (HR = 0.63, p = 0.05), by 46% in patients diagnosed in 2006–2010 (HR = 0.54, p < 0.02). Conclusion: A clear modification of MS course is observed after 2000; among other causes, this can be associated to the changes in the treatment patterns experienced in those years.

Less Frequent and Less Severe Flu-Like Syndrome in Interferon Beta-1a Treated Multiple Sclerosis Patients with at Least One Allele Bearing the G>C Polymorphism at Position -174 of the IL-6 Promoter Gene.

One of the most common adverse event of interferon beta (IFNβ) therapy for multiple sclerosis is flu-like syndrome (FLS), which has been reportedly related to increased levels of cytokines such as interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-α). Average cytokine levels can be affected by single nucleotide polymorphism in the gene promoter regions. To investigate whether IL-6 -174 G>C and TNF-α -376 G>A polymorphisms could be correlated to the incidence of FLS, and whether an anti-inflammatory/antipyretic therapy may influence FLS development, a prospective observational study was performed in 190 treatment naïve, multiple sclerosis patients who started IM IFNβ-1a 30mcg once weekly. The identification of IL-6 -174 G>C and TNF-α -376 G>A polymorphisms was achieved by performing an amplification-refractory mutation system. Serum IL-6 levels were measured using enzyme-linked immunosorbent assay in blood samples taken before therapy and then after the first and last IFNβ-1a injection of the follow-up. FLS-related symptoms were recorded by patients once per week during the first 12 weeks of therapy into a self-reported diary. We found that patients carrying at least one copy of the C allele at position -174 in the promoter of IL-6 gene produced lower levels of IL-6 and were less prone to develop FLS, which was also less severe. On the contrary, the polymorphism of TNF-α had no effect on FLS. Patients taking the first dose of anti-inflammatory/antipyretic therapy in the peri-injection period (within 1 hour) experienced a reduced FLS severity. In conclusion, the study of IL-6 -174 G>C polymorphism would allow the identification of patients lacking the C nucleotide on both alleles who are at risk of a more severe FLS, and may be addressed to a timely and stronger anti-inflammatory/antipyretic therapy for a more effective FLS prevention.

Clinical activity after fingolimod cessation: disease reactivation or rebound?

There is debate as to whether the apparent rebound after fingolimod discontinuation is related to the discontinuation itself or whether it is due to the natural course of highly active multiple sclerosis (MS). Our aim was to survey the prevalence of severe reactivation and rebound after discontinuation of fingolimod in a cohort of Italian patients with MS.

Determinants of therapy switch in multiple sclerosis treatment-naïve patients: A real-life study:

Background: With many options now available, first therapy choice is challenging in multiple sclerosis (MS) and depends mainly on neurologist and patient preferences. Objectives: To identify prognostic factors for early switch after first therapy choice. Methods: Newly diagnosed relapsing–remitting MS patients from 24 Italian centers were included. We evaluated the association of baseline demographics, clinical, and magnetic resonance imaging (MRI) data to the switch probability for lack of efficacy or intolerance/safety with a multivariate Cox analysis and estimated switch rates by competing risks models. Results: We enrolled 3025 patients. The overall switch frequency was 48% after 3 years. Switch risk for lack of efficacy was lower with fingolimod (hazard ratio (HR) = 0.50; p = 0.009), natalizumab (HR = 0.13; p < 0.001), dimethyl-fumarate (HR = 0.60; p = 0.037), teriflunomide (HR = 0.21; p = 0.031) as compared to interferons. Younger age (HR = 0.96; p < 0.001), diagnosis delay (HR = 1.23; p = 0.021), higher baseline Expanded Disability Status Scale (HR = 1.17; p = 0.001), and spinal cord lesions (HR = 1.46; p = 0.001) were independently associated with higher inefficacy switch rates. We found lower switch for intolerance/safety with glatiramer acetate (HR = 0.61; p = 0.001), fingolimod (HR = 0.35; p = 0.002), and dimethyl-fumarate (HR = 0.57; p = 0.022) as compared to interferons, while it increased with natalizumab (HR = 1.43; p = 0.022). Comorbidities were associated with intolerance switch (HR = 1.28; p = 0.047). Conclusion: Several factors are associated with higher switch risk in patients starting a first-line therapy and could be integrated in the decision-making process of first treatment choice.