Abstract
Natalizumab is an effective monoclonal antibody therapy for the treatment of relapsing- remitting multiple sclerosis (RRMS) and interferes with immune cell migration into the central nervous system by blocking the α4 subunit of very-late activation antigen-4 (VLA-4). Although well tolerated and very effective, some patients still suffer from relapses in spite of natalizumab therapy or from unwanted side effects like progressive multifocal leukoencephalopathy (PML). In search of a routine-qualified biomarker on the effectiveness of natalizumab therapy we applied flow cytometry and analyzed natalizumab binding to α4 and α4 integrin surface levels on T-cells, B-cells, natural killer (NK) cells, and NKT cells from 26 RRMS patients under up to 72 weeks of therapy. Four-weekly infusions of natalizumab resulted in a significant and sustained increase of lymphocyte-bound natalizumab (p<0.001) which was paralleled by a significant decrease in detectability of the α4 integrin subunit on all lymphocyte subsets (p<0.001). We observed pronounced natalizumab accumulations on T and B cells at single measurements in all patients who reported clinical disease activity (n = 4). The natalizumab binding capacity of in vitro saturated lymphocytes collected during therapy was strongly diminished compared to treatment-naive cells indicating a therapy-induced reduction of α4. Summing up, this pilot study shows that flow cytometry is a useful method to monitor natalizumab binding to lymphocytes from RRMS patients under therapy. Investigating natalizumab binding provides an opportunity to evaluate the molecular level of effectiveness of natalizumab therapy in individual patients. In combination with natalizumab saturation experiments, it possibly even provides a means of studying the feasability of patient-tailored infusion intervals. A routine-qualified biomarker on the basis of individual natalizumab saturation on lymphocyte subsets might be an effective tool to improve treatment safety.
Highlights
Recruitment of activated immune cells across the blood-brain barrier (BBB) into the central nervous system (CNS) is considered essential for the initiation of inflammatory brain lesions in multiple sclerosis (MS) [1,2]
After 12 weeks, we observed a significant increase in mean anti-natalizumab rfi levels (p,0.001) on all lymphocyte subsets compared to background levels obtained at baseline
We found that additional in vitro treatment with natalizumab only caused a minor increase of anti-natalizumab rfi levels compared to therapy-derived in vivo natalizumab levels which was 1.360.2 fold in case of CD3+ T cells, and 1.260.1 fold for CD19+ B cells, natural killer (NK) cells, and natural killer T cells (NKT) cells
Summary
Recruitment of activated immune cells across the blood-brain barrier (BBB) into the central nervous system (CNS) is considered essential for the initiation of inflammatory brain lesions in multiple sclerosis (MS) [1,2]. In contrast to other IgG subclasses, IgG4-antibodies are mere blocking antibodies with minor affinity to immune cell Fc receptors, and they do not bind complement. They are neither involved in antibodydependent cell-mediated nor in complement-dependent cellular cytotoxicity [6]. Natalizumab blocks immune cell extravasation into the CNS by selectively binding to the a4 subunit of VLA-4 [7] It is the first monoclonal antibody therapy approved for treatment of MS and was shown to impressively reduce relapse frequency and disease progression in patients with relapsingremitting MS (RRMS) [8,9]. The overall dimension of the pharmacological activity of natalizumab is unsolved and clinical effectiveness is counteracted by the risk to develop progressive multifocal leukoencephalopathy (PML)
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