In the current study, we conducted a quantitative in-depth proteome and deglycoproteome analysis of cerebrospinal fluid (CSF) from relapsing-remitting multiple sclerosis (RRMS) and neurological controls using mass spectrometry and pathway analysis. More than 2000 proteins and 1700 deglycopeptides were quantified, with 484 proteins and 180 deglycopeptides significantly changed between pools of RRMS and pools of controls. Approximately 300 of the significantly changed proteins were assigned to various biological processes including inflammation, extracellular matrix organization, cell adhesion, immune response, and neuron development. Ninety-six significantly changed deglycopeptides mapped to proteins that were not found changed in the global protein study. In addition, four mapped to the proteins oligo-myelin glycoprotein and noelin, which were found oppositely changed in the global study. Both are ligands to the nogo receptor, and the glycosylation of these proteins appears to be affected by RRMS. Our study gives the most extensive overview of the RRMS affected processes observed from the CSF proteome to date, and the list of differential proteins will have great value for selection of biomarker candidates for further verification.

Immune checkpoint inhibitors have shown efficacy against metastatic triple-negative breast cancer (mTNBC) but only for PD-L1positive disease. The randomized, placebo-controlled ALICE trial (NCT03164993, 24 May 2017) evaluated the addition of atezolizumab (anti-PD-L1) to immune-stimulating chemotherapy in mTNBC. Patients received pegylated liposomal doxorubicin (PLD) and low-dose cyclophosphamide in combination with atezolizumab (atezo-chemo; n = 40) or placebo (placebo-chemo; n = 28). Primary endpoints were descriptive assessment of progression-free survival in the per-protocol population (>3 atezolizumab and >2 PLD doses; n = 59) and safety in the full analysis set (FAS; all patients starting therapy; n = 68). Adverse events leading to drug discontinuation occurred in 18% of patients in the atezo-chemo arm (7/40) and in 7% of patients in the placebo-chemo arm (2/28). Improvement in progression-free survival was indicated in the atezo-chemo arm in the per-protocol population (median 4.3 months versus 3.5 months; hazard ratio (HR) = 0.57; 95% confidence interval (CI) 0.33–0.99; log-rank P = 0.047) and in the FAS (HR = 0.56; 95% CI 0.33–0.95; P = 0.033). A numerical advantage was observed for both the PD-L1positive (n = 27; HR = 0.65; 95% CI 0.27–1.54) and PD-L1negative subgroups (n = 31; HR = 0.57, 95% CI 0.27–1.21). The progression-free proportion after 15 months was 14.7% (5/34; 95% CI 6.4–30.1%) in the atezo-chemo arm versus 0% in the placebo-chemo arm. The addition of atezolizumab to PLD/cyclophosphamide was tolerable with an indication of clinical benefit, and the findings warrant further investigation of PD1/PD-L1 blockers in combination with immunomodulatory chemotherapy.

The rapidly growing number of biomedical studies supported by mass spectrometry based quantitative proteomics data has made it increasingly difficult to obtain an overview of the current status of the research field. A better way of organizing the biomedical proteomics information from these studies and making it available to the research community is therefore called for.

Multiple sclerosis (MS) is a chronic disease characterized by inflammation, demyelination, and neurodegeneration of the central nervous system (CNS). There is no cure. Current treatments target the autoimmune aspects of MS but do not directly improve CNS remyelination. Pro-remyelinating treatment might optimize the treatment of MS patients. In this project, we aimed to investigate different strategies to improve remyelination and mitigate axonal damage in the cuprizone model, an animal model for de-and remyelination. Our goal was to determine the effect of biologically active vitamin D (calcitriol) on remyelination (Paper I), and axonal damage (Paper II).Moreover, we investigated the effect of the MS-medication fingolimod on remyelination and axonal damage in the cerebellum (Paper III). Finally, we assessed the impact of fingolimod in the cerebrum (Paper IV).C57Bl/6 mice were exposed to the neurotoxicant cuprizone. In the vitamin D experiment, high-dose calcitriol or placebo was given by intraperitoneal injections twice a week. In the fingolimod experiment, fingolimod or placebo was given by oral gavage daily. In both experiments, mice were investigated at several time points during remyelination. Histochemistry and immunohistochemistry were used to investigate remyelination, axonal damage, and loss. We analyzed the brain proteome by proteomic analysis to further determine the CNS effects of fingolimod exposure. Treatment with high-dose calcitriol improved the remyelination process (paper I).Vitamin D given before, but not after cuprizone-induced demyelination prevented acute axonal damage and axonal loss (paper II). Given after cuprizone-induced demyelination, fingolimod did not affect cerebellar remyelination, the number of oligodendrocytes, microglia or astrocyte activation, or acute axonal damage at any time point (paper III). Fingolimod was functionally active during remyelination, resulting in a downregulation of sphingosine-1-phosphate receptor 1 protein levels in the brain. We found, however, no difference in the degree of remyelination, oligodendrocyte numbers, nor the degree of axonal damage or loss in the corpus callosum (paper IV).In the cuprizone model, high-dose calcitriol given during remyelination improved remyelination. However, axonal damage was only prevented if vitamin D was given before demyelination occurred. Fingolimod modulated the sphingosine-1-phosphate receptor 1 levels in the cerebrum but did not increase remyelination, nor protect against axonal injury or loss in the cerebellum or cerebrum when given after cuprizone-induced demyelination.