Interventional Not Pharmacological Study to Investigate the Role of ADA in Multiple Sclerosis PatientsTreated With an Immune-Reconstitution Therapy (2-CdA)

The therapeutic landscape for multiple sclerosis (MS) is rapidly evolving. For the past 25 years, there has been an accelerating inclusion of new immunomodulating drugs. MS immunotherapies may also be classified in a different way, into treatments that are given continuously (chronic treatments) and medications that are applied intermittently (immune reconstitution therapies [IRTs]). The principle behind the latter is depletion of the immune system that allows it to rebuild itself. An IRT by definition is given at short intermittent courses and not continuously. IRT modalities were shown to induce long-term remission of MS that, in some cases, is close to the definition of a "cure".

Most importantly, IRT using these modalities causes substantial changes in the lymphocyte repertoire after the reconstitution phase.

This is particularly true for Cladribine (2-CdA), a high efficacy therapy that selectively depletes peripheral lymphocytes, recently approved by the European Medicine Agencies for RMS patients with high activity of disease.

Lymphocytes have a high intracellular ratio of DCK to 5'-NTs compared with other cell types. Specifically, levels of DCK and the ratio of DCK to 5'-NT are high in T cells (CD4+ and CD8+), B cells, and dendritic cells but are very low in numerous non-hematologic cell types (http://biogps.gnf.org). This makes lymphocytes, mainly memory B cells, particularly sensitive to the accumulation of 2-CdA nucleotides. The intracellular accumulation of 2-CdATP leads to the incorporation of 2-CdATP into cellular DNA, disrupting the double helix structure and leading to a failure of DNA repair and synthesis. The resulting DNA strand breaks alter the cell cycle progression inducing cell death mediated by apoptosis. Although apoptosis is the most prominent mechanism of action of 2-CdA, additional mechanisms cannot be excluded. Many evidences shown that nucleotide signaling governs some of the most essential responses in immunity, ranging from antigen-driven T lymphocyte proliferation to T helper 1 (Th1) and Th2 cell differentiation, from neutrophil and macrophage chemotaxis to intracellular pathogen killing, and from NADPH-oxidase activation to IL-1β maturation and release. In addition, an analysis of lymphocytes subsets collected in the pivotal phase III trial of Cladribine showed that while T cells reduction shows some Cladribine-dose dependency, B cells have a similar reduction with both dosing schedules in the trial. Moreover, the mean number of T and B cells do not differ between patients remaining free from relapses and patients developing relapses, suggesting that efficacy of Cladribine is not strictly dependent on reduction of lymphocyte count. On the other hand, a lower lymphocyte count has been correlated with a higher risk of developing an Herpes Zoster infection, with all the events restricted and dermatomal, and most of them graded as mild or moderate. Thus, severe lymphopenia is also an important identified risk for safety. In phase III clinical trials (RCTs), 20-25% of the subjects treated with 3.5mg/kg of oral 2-CdA developed transient Grade 3-4 lymphopenia. The rate of patients who discontinued the treatment for lymphopenia was highest (12.4%) in the group that received the greatest cumulative dose of 2-CdA (CC 8.75 mg/kg) at the end of the extension phase of the CLARITY study (22). This effect has been also correlated in others disease with a reduced level of DCK mRNA and a higher level of 5'NT mRNA in lymphocytes. Though the biological activation of 2-CdA needs DCK function independently of ADA action, it cannot be excluded ADA participation in other aspects of 2-CdA mechanism of action, taking in consideration ADA crucial role in lymphocytes activation and function.

In line with this, it has been recently identified an ADA genetic variant (rs244072, non-risk/risk allele: T/C) linked to brain inflammation and MS disease severity (article in preparation), by screening more than 50 SNPs in MS-related genes of 514 MS patients and looking for association with clinical and biochemical parameters. The most striking result observed was a direct correlation between the presence of at least one risk allele and EDSS score, namely CT/CC patients show higher values of EDSS than TT patients (p=0.016).