The results of this observational study confirm the efficacy of NTZ treatment in reducing disease activity and disability progression in RRMS patients already demonstrated in NTZ pivotal trials 
. In addition they show that a short-term NTZ treatment may significantly improve cognitive performances and fatigue in these patients. A decrease of the proportion of cognitively impaired MS patients and an improvement of sustained attention, information processing speed and visuo-spatial memory, as measured by the SDMT, PASAT 2 and SPART, were evident since the first year of NTZ treatment. Most of these beneficial effects on cognition were still present after two years of treatment. These findings are consistent with previously published Italian 
and Swedish 
observational studies suggesting a beneficial effect of NTZ on cognitive performances. In the Italian study, even if a smaller number of patients (n
39 at 1 year; n
11 at 2 years) than in the current study was evaluated and the cognitive impairment was assessed by a neuropsychological battery different from the BRB we used, an improvement in tests which explore information processing speed, executive function and memory tests were also found 
. Data from the Swedish post-marketing surveillance program of MS patients treated with NTZ for up to 2 years, clearly demonstrate an improvement in SDMT 
It is noteworthy that in the current study we used, also, a measure of the global change of cognitive performances that is the CII. We found a significant improvement in the mean CII value after 1 year NTZ treatment and this value was furtherly improved in the second year of treatment. The CII 
may be considered a more suitable tool to evaluate changes in cognitive performances than considering the individual tests, because CII is independent by the number of cognitive tests failed at the BRB and the ST (i.e. if an individual patient was impaired in six tests at baseline and failed four tests at follow-up, that patient would be still considered cognitively impaired, despite the fact that his or her performances were objectively improved), therefore its use strengthens the results of this study. Moreover in this study we used alternate versions of cognitive tests, and adequate time intervals between them to reduce the major methodological issue when neuropsychological tests are repeated over time that is the practice effect 
The significant improvement of fatigue measures after 1 and 2 years of NTZ treatment that we have found, is also in line with a previous German study 
that showed a decrease of fatigue in RRMS treated with NTZ for 6 months.
The effect of potential clinical and demographic confounders which could affect the changes of neuropsychological performances and fatigue scores at the end of the 1st year of treatment was ruled out by a logistic regression analysis that demonstrated that both cognitive impairment and fatigue were independent from other clinical variables, in this cohort. The only covariate with a positive predictive effect on the neuropsychological performances was the educational level, as measured by the time spent in formal school education, in accordance with recent reports 
showing a significant beneficial effect of the so-called brain reserve on cognition and on the brain atrophy progression in MS.
The degree and the time-trend of the improvement induced by NTZ on cognitive functions and fatigue in the current and previous studies 
seem to suggest that these two phenomena could be, at least in mild disabled patients, more related to inflammation than to neurodegeneration mechanisms. A growing evidence seems to support this view. In the experimental autoimmune encephalomyelitis (EAE), the rodent model of MS, it was recently demonstrated that the activation of microglia and the inflammatory cytokines released from infiltrating lymphocytes, especially TNF-α, are able to alter synaptic transmission 
. This induced synaptopathy is related to cognitive dysfunction in this experimental model of MS 
. The significant reduction of cerebrospinal fluid (CSF) and plasma levels of pro-inflammatory cytokines and of CSF levels of the light-chain neurofilament, a marker of axonal loss, during the NTZ treatment 
may be responsible of the cognitive dysfunction improvement induced by NTZ. Moreover 1 year NTZ treatment has been demonstrated to prevent the accumulation of active cortical lesions in RRMS, an MRI measure that is correlated with cognitive decline 
We recognize that the main limit of this observational study is the lack of a concurrent control group. However it must be considered that many barriers may prevent the execution of a controlled observational study aimed to evaluate the effects of a second line drug (i.e. NTZ) on a population of MS patients with high disease activity. The inclusion of a placebo group, that remains untreated for up to 2 years, is not feasible and ethical. Moreover a comparison between patients treated with a second line treatment (NTZ) and patients treated with a first line treatment (IFN β or GA) is also difficult to carry out, because the unbalanced baseline measures of disease activity (i.e. ARR, EDSS, MRI) between the two groups (indication bias) may influence the measured association between exposure and outcome. Within-subject methods, including self-controlled case-series method 
as we used in this study, in which outcomes are compared between periods before and after treatment exposure within the same individuals, are less susceptible to confounding by indication and may eliminate between-person confounding 
Nevertheless an analysis is ongoing aimed to identify a suitable active drug control group among the DMD treated MS patients collected, in the same period of time, in the MS databases (Imed-web) in Bari and Florence Centers. We are using a propensity score matching technique 
, which is the most common method currently used to reduce bias in treatment comparisons in observational studies 
to select patients whose demographics and disease characteristics overlap with those of our cohort of NTZ-treated patients, and that were evaluated for cognitive impairment at the same time-point of the NTZ-treated cohort. A preliminary analysis [data not shown] on a sample of 20 RRMS patients (mean ± SD: age: 35.12±10.99; disease duration: 10.32±9.92; mean EDSS 3.5±1.14; mean ARR in the year before the treatment initiation: 1.95±0.85) treated with different formulations of DMDs was performed. The results showed that DMD treatment reduced the baseline mean CII values over a two year follow-up (mean [SD] Baseline CII: 14.61 [6.62]; mean [SD] 1 Year CII: 13.06 [6.03]; mean [SD] 2 Years CII: 12.78 [6.06] in these sample of patients, but this effect did not reach a statistical significance, Friedmann two-way test: p
0.241). However the lack of statistical significance could be due to the small sample size of this DMD treated group. So far, few studies reported some beneficial effects of IFN beta treatment on cognitive functions in RRMS patients 
, whereas GA treatment failed to demonstrate any significant effect on cognition 
, but only some significant effects on fatigue measures 
Although no direct comparison could be made, our findings allow to speculate that the effects of NTZ on both cognition and fatigue measures might be greater than first line DMDs.
A longer follow-up in a larger population, including a propensity score matched control group, is needed to confirm whether the early beneficial effects of NTZ in improving cognitive performances and fatigue, we found in this study, hold-up in the long-term.