A. Bertolotto

Persistent neutralizing antibodies abolish the interferon β bioavailability in MS patients

Background: MxA is an antiviral protein exclusively induced by type I interferons (IFN) and some viruses, and MxA gene expression is one of the most appropriate markers for measuring the biologic activity of exogenous IFNβ. Methods: A new quantitative-competitive PCR method was used to quantify MxA mRNA in peripheral blood mononuclear cells of 99 treatment-naïve and 92 IFNβ-treated patients with MS (22 Avonex, 17 Betaferon, and 53 Rebif-22). Every 3 months, IFNβ-induced neutralizing antibodies (NAb) were evaluated in sera using a cytopathic effect assay. Three categories of patients were identified: NAb negative (NAb−), persistent NAb positive (NAb+, ≥2 consecutive positive samples), and isolated NAb+ (one positive sample). Results: Treatment-naïve patients expressed detectable MxA mRNA levels (mean = 36 ± 32 fg MxA/pg glyceraldehyde-3-phosphate dehydrogenase (GAPDH); range 1 to 160) and an upper normal threshold was established (mean + 3 SD = 132 fg MxA/pg GAPDH). IFNβ-treated patients exhibited more than 11-fold higher levels (mean = 412 ± 282 fg MxA/pg GAPDH; range 16 to 1,172). However, 17 patients did not exhibit an increase in MxA mRNA level; 15 of these 17 patients showed a concurrent Nab+ titer. Moreover, 13 were persistent NAb+. Isolated NAb+ patients did not show a decrease in bioavailability of IFNβ (n = 9; mean = 567 ± 366 fg MxA/pg GAPDH; range 83 to 1,120). In NAb− patients, bioavailability was comparable among the three different IFNβ preparations 12 hours after injection. Conclusion: During IFNβ therapy, the presence of NAb reduced or abolished bioavailability in a relevant percentage of patients. These data could be important for the early detection of patients with MS who are not responsive to IFNβ therapy.

Neutralizing antibodies reduce the efficacy of βIFN during treatment of multiple sclerosis

Objective: To analyze the impact of neutralizing antibodies (NAbs) on the clinical efficacy of IFNβ. Methods: This was an open-label study involving 78 patients with multiple sclerosis treated with Betaferon 8 million IU (MIU) subcutaneously (SC) every other day (n = 20), Rebif 22 μg SC 3 times weekly (n = 25), or Avonex 30 μg IM once weekly (n = 33). Every 3 months, blood samples were collected and sera were analyzed for NAbs using an antiviral cytopathic effect assay. Patients were categorized according to their NAb status: NAb negative (NAb−); isolated NAb positive (NAb+), patients with ≥1 positive sample (titer ≥ 20); and persistent NAb+, patients with ≥2 consecutive positive samples (titer ≥ 20). Patients who were NAb− and persistent NAb+ were compared for relapse rate, time between first and second relapse, percentage of relapse-free patients, and percentage of patients who had a sustained progression of ≥1 point on the Expanded Disability Status Scale (EDSS). Results: The incidence of persistent NAb+ patients was 35% for Betaferon, 20% for Rebif, and 3% for Avonex. During IFNβ treatment, both NAb+ and NAb− patients showed a reduction in relapse rate; this reduction (25%) was not significant in NAb+ patients but was significant (67%; p < 0.0001) in NAb− patients. In addition, the mean relapse rate was higher (p = 0.039), mean time between first and second relapse was shorter (13 vs 21 months; p = 0.0064), and there was a trend suggesting that NAbs affected the probability of remaining relapse free (p = 0.08). A higher percentage of NAb+ patients versus NAb− patients had worsening of EDSS scores during follow-up (p = 0.013). Conclusion: Persistent NAbs significantly reduce the clinical efficacy of IFNβ.

Predictive markers for response to interferon therapy in patients with multiple sclerosis.

Background: Prolonged therapy with interferon β (IFNβ) often leads to the development of anti-IFNβ binding antibodies (BAbs). A subset of the BAbs is of a neutralizing nature (neutralizing antibodies, NAbs) and is associated with reduced clinical efficacy of therapy. Myxovirus-resistance-protein A (MxA) has proven to be a reliable biomarker of IFNβ bioactivity. We analyzed the prognostic value of MxA mRNA, NAbs, and BAbs on the risk of having a new relapse in IFNβ-treated patients. Methods: A 3-year study was conducted in 137 IFNβ-treated patients. Blood samples for BAbs, NAbs, and MxA mRNA measurements were taken after 12 ± 3 months of therapy. Analysis of relapse-free survival (RFS) was performed for all measures by using known thresholds, generating “positive” and “negative” groups. Also, time between sampling and following relapse and risk of new relapses were calculated. Results: The MxA-negative group showed poorer RFS rates than the MxA-positive group [p < 0.0001, hazard ratio (HR) = 2.87]. Likewise, the NAb-positive group showed poorer RFS rates than the NAb-negative group (p =0.0013; HR = 2.49). On the contrary, BAb measurement did not show a clear clinical significance. Conclusions: Findings indicate that measurements of both myxovirus-resistance-protein A (MxA) and neutralizing antibodies (NAbs) predict the risk of new relapses; however, the slightly stronger prognostic significance of MxA mRNA and the easier method for it measurement make MxA mRNA the preferred biomarker for monitoring interferon β (IFNβ)-treated patients. This information can be used to better tailor treatment to the individual patient with MS. GLOSSARY: BAb = binding antibody; CPE = cytopathic effect; EDSS = Expanded Disability Status Scale; GAPDH = glyceraldehyde phosphate dehydrogenase; HR = hazard ratio; IFN = interferon; MxA = myxovirus-resistance-protein A; NAb = neutralizing antibody; RFS = relapse-free survival; ROC = receiver operating characteristic; TRU = 10-fold reduction units.

Biological responsiveness to first injections of interferon-beta in patients with multiple sclerosis

This study is the first to evaluate biological response to first injections of interferon-beta (IFNbeta) in patients with multiple sclerosis. MxA mRNA was measured in 96 patients receiving IFNbeta-1a (Avonex, n=32), IFNbeta-1b (Betaferon, n=19), IFNbeta-1a (Rebif) 22 microg (n=30), or IFNbeta-1a 44 microg (n=15). Patients were analysed before, 3 and 24 h after the first injection, and 12 h after the second administration. Results showed that up-regulation was evident within 3 h of IFNbeta injection, peaked 12 h after injection, and progressively declined 24 h after administration. The cumulative responses were similar after a single administration, regardless of product/dose. Moreover, data indicate that the abolition of the biological activity detected during IFNbeta therapy is due to underlying phenomena (e.g., neutralizing antibodies), because all patients were constitutively responders to IFNbeta at treatment initiation.

Variable responses to rituximab treatment in neuromyelitis optica (Devic's disease).

We have described two cases of Devic’s disease patients treated with rituximab with different outcomes. The results indicate that there may be early unresponsiveness in very aggressive cases. Well designed clinical trials are needed to assess treatment effects in such a rare disease.SommarioAbbiamo descritto due casi di Malattia di Devic trattati con Rituximab con differente risposta terapeutica. I risultati indicano la possibilità di una non risposta precoce nei casi molto aggressivi. Sono necessari trial clinici ben strutturati per la valutazione dell’efficacia dei trattamenti in questa rara malattia.

Expression and regulation of IFNα/β receptor in IFNβ-treated patients with multiple sclerosis

Background: The cytokine interferon beta (IFNβ) is successfully used in the treatment of multiple sclerosis (MS), although there is a high degree of variability in the response. A common mechanism involved in the modulation of responsiveness to cytokines is represented by regulation of their receptor expression through autocrine ligand–mediated loops. The present study is aimed at investigating the regulation of IFNα/β receptor (IFNAR) during IFNβ therapy in patients with MS and at correlating it with the biologic responsiveness to the cytokine. Methods: Quantitative PCR measurements of IFNAR-1 and the three IFNAR-2 isoforms were performed in 141 patients after short-term and long-term treatment. Patients were also regularly screened for anti-IFNβ neutralizing antibodies (NAbs). IFN-inducible myxovirus resistance protein A messenger RNA was used as an indicator of bioactivity. Results: Pretreatment levels of IFNAR-2 in patients were lower overall than in controls (p = 0.038), and high levels correlated with greater bioactivity. Upon prolonged treatment, NAb-negative patients displayed a state of decreased transmembrane IFNAR-2 expression (p ≤ 0.025), whereas levels of soluble IFNAR-2 were slightly increased (p < 0.0001). The presence of NAbs reversed these effects (p ≤ 0.0056). In NAb-positive patients, pretreatment expression levels of both transmembrane IFNAR-2 isoforms were significantly lower than in NAb-negative patients (p ≤ 0.0089). Conclusions: Findings show that interferon-α/β receptor (IFNAR)-2 isoforms are important regulators of the responsiveness to endogenous and systemically administered interferon beta (IFNβ). They show a dual action, agonistic and antagonistic, that influences both the magnitude and the nature of the biologic response to IFNβ. Levels of IFNAR-2 are regulated with the aim of keeping the body in a state of equilibrium, even when nonphysiologic stimuli are present.

Expression and regulation of IFNalpha/beta receptor in IFNbeta-treated patients with multiple sclerosis.

Background: The cytokine interferon beta (IFNβ) is successfully used in the treatment of multiple sclerosis (MS), although there is a high degree of variability in the response. A common mechanism involved in the modulation of responsiveness to cytokines is represented by regulation of their receptor expression through autocrine ligand–mediated loops. The present study is aimed at investigating the regulation of IFNα/β receptor (IFNAR) during IFNβ therapy in patients with MS and at correlating it with the biologic responsiveness to the cytokine. Methods: Quantitative PCR measurements of IFNAR-1 and the three IFNAR-2 isoforms were performed in 141 patients after short-term and long-term treatment. Patients were also regularly screened for anti-IFNβ neutralizing antibodies (NAbs). IFN-inducible myxovirus resistance protein A messenger RNA was used as an indicator of bioactivity. Results: Pretreatment levels of IFNAR-2 in patients were lower overall than in controls (p = 0.038), and high levels correlated with greater bioactivity. Upon prolonged treatment, NAb-negative patients displayed a state of decreased transmembrane IFNAR-2 expression (p ≤ 0.025), whereas levels of soluble IFNAR-2 were slightly increased (p < 0.0001). The presence of NAbs reversed these effects (p ≤ 0.0056). In NAb-positive patients, pretreatment expression levels of both transmembrane IFNAR-2 isoforms were significantly lower than in NAb-negative patients (p ≤ 0.0089). Conclusions: Findings show that interferon-α/β receptor (IFNAR)-2 isoforms are important regulators of the responsiveness to endogenous and systemically administered interferon beta (IFNβ). They show a dual action, agonistic and antagonistic, that influences both the magnitude and the nature of the biologic response to IFNβ. Levels of IFNAR-2 are regulated with the aim of keeping the body in a state of equilibrium, even when nonphysiologic stimuli are present.

Expression and regulation of IFN / receptor in IFN -treated patients with multiple sclerosis

Background: The cytokine interferon beta (IFNβ) is successfully used in the treatment of multiple sclerosis (MS), although there is a high degree of variability in the response. A common mechanism involved in the modulation of responsiveness to cytokines is represented by regulation of their receptor expression through autocrine ligand–mediated loops. The present study is aimed at investigating the regulation of IFNα/β receptor (IFNAR) during IFNβ therapy in patients with MS and at correlating it with the biologic responsiveness to the cytokine. Methods: Quantitative PCR measurements of IFNAR-1 and the three IFNAR-2 isoforms were performed in 141 patients after short-term and long-term treatment. Patients were also regularly screened for anti-IFNβ neutralizing antibodies (NAbs). IFN-inducible myxovirus resistance protein A messenger RNA was used as an indicator of bioactivity. Results: Pretreatment levels of IFNAR-2 in patients were lower overall than in controls (p = 0.038), and high levels correlated with greater bioactivity. Upon prolonged treatment, NAb-negative patients displayed a state of decreased transmembrane IFNAR-2 expression (p ≤ 0.025), whereas levels of soluble IFNAR-2 were slightly increased (p < 0.0001). The presence of NAbs reversed these effects (p ≤ 0.0056). In NAb-positive patients, pretreatment expression levels of both transmembrane IFNAR-2 isoforms were significantly lower than in NAb-negative patients (p ≤ 0.0089). Conclusions: Findings show that interferon-α/β receptor (IFNAR)-2 isoforms are important regulators of the responsiveness to endogenous and systemically administered interferon beta (IFNβ). They show a dual action, agonistic and antagonistic, that influences both the magnitude and the nature of the biologic response to IFNβ. Levels of IFNAR-2 are regulated with the aim of keeping the body in a state of equilibrium, even when nonphysiologic stimuli are present.

Esercizio fisico e salute psicologica in donne affette da Sclerosi Multipla: l’effetto di moderazione del livello di disabilità

Lo studio intende indagare gli effetti di un programma di attivita motoria sullo stato di salute psicologica dei soggetti con Sclerosi Multipla (SM), analizzando gli effetti diretti dell’esercizio fisico sulla qualita di vita (QdV) e sulla depressione, e gli effetti di moderazione del livello di disabilita nella relazione tra allenamento motorio e variabili psicologiche. Sono state coinvolte 31 donne affette da SM recidivante remittente, con un’eta media di 40 anni (DS = 5; min = 22; max = 50) e un livello di disabilita di 2.0 dell’Expanded Disability Status Scale (EDSS). I partecipanti allo studio sono stati suddivisi, secondo specifiche tecniche di appaiamento, in due gruppi (sperimentale, GS - controllo, GC), omogenei al baseline per le variabili di interesse. Il GS ha partecipato a 12 settimane di training motorio bisettimanale, basato su attivita aerobica ed esercizi di rinforzo muscolare; mentre il GC ha mantenuto inalterate le proprie abitudini. La partecipazione all’intervento motorio ha un’influenza positiva sui sintomi depressivi (p < .001) e sulla QdV (p < .001). Il livello di disabilita e risultato un moderatore della relazione tra partecipazione all’esercizio fisico e le variabili psicologiche.

P141: Motor evoked potentials from multiple recording sites of the lower limbs as a monitoring tool of central motor function

Results: When the interval between 2 consecutive stimuli was 0.5 s, the response to the second sound was about half of the size in comparison to the first one. On the other hand, when the interval was longer than 4 s, the amplitudes of both responses were the same. The responses differ significantly in accordance with the interval of the stimuli. Ketamine suppressed the response to the first auditory stimuli that resulted in the amplitudes of both responses becoming the same even in the smaller interval. Conclusion: We therefore conclude that the auditory gating function in common marmosets is similar to humans. In addition, the deficit induced by ketamine suggests that it could be a model for the study of some psychiatric diseases.