Contributions
Abstract: P652
Type: Poster
Abstract Category: Therapy - disease modifying - Immunomodulation/Immunosuppression
Background: The mechanisms underlying both the therapeutic effect of dimethyl fumarate (DMF), as well as its ability to differentially decrease counts of particular lymphocyte subsets in treated patients, are not fully elucidated. Here, using a combination of in vitro and in vivo approaches, we considered whether these DMF effects are mediated by selective apoptosis of distinct T cell subsets.
Methods: Total lymphocyte counts (TLC) as well as flow cytometry analysis of effector and regulatory
T cell subsets within peripheral blood mononuclear cells (PBMC), were carried out on samples obtained from MS patients prior to and at 3-month intervals following DMF treatment initiation. In vitro, PBMC were cultured with addition of DMF, monomethyl fumarate (MMF) or appropriate vehicle controls, subsequently stained for markers of T cell subsets and apoptosis (Annexin V/propidium iodide), then analyzed by flow cytometry.
Results: As previously reported DMF therapy reduced TLC, particularly over the first 6 months (mean decrease 0.79 x 109 cells/L; n=13; p=0.0009), with a preferential reduction in CD8+ T cells resulting in increased CD4:CD8 ratios (mean fold increase 1.4; p=0.002). Greater decreases were noted for memory versus naïve T cell subsets, and for conventional versus regulatory T cell subsets. These differential effects resulted in increased ratios of regulatory to effector T cells, and of anti- to pro-inflammatory cytokine-expressing T cells. In vitro, DMF induced dose-dependent apoptosis of CD3+ T cells (increase in late apoptotic cells vs. vehicle: 11% for 10µM DMF, p=0.0181; 51% for 50µM DMF, p=0.0001). Greater DMF-induced apoptosis was seen for CD8+ versus CD4+ T cells (79% vs. 39%, p=0.0001); for memory versus naïve CD4+ and CD8+ T cell subsets (CD4+: 73% vs. 18%, p=0.0004; CD8+: 92% vs. 8%, p=0.0064), and for conventional versus regulatory T cell subsets
(42% vs. 6%, p=0.0004).
Conclusions: In vitro, DMF induces T cell apoptosis that disproportionately affects CD8+ T cells, memory T cells, and non-regulatory T cells. This differential in vitro susceptibility of distinct T cell subsets to DMF-induced apoptosis mirrors the observed differential in vivo effects of DMF on the same T cells subsets and may thus provide insights both into the therapeutic mode of action of DMF in MS, as well as the mechanism underlying DMF-induced lymphopenia.
Disclosure:
Dr Mahtab Ghadiri is a recipient of the BMRI/McGill University Multiple Sclerosis scholarship, funded by Novartis.
Ayman Rezk: nothing to disclose.
Dr Rui Li reports: nothing to disclose.
Dr Felix Luessi has received travel grants from Teva Pharma and Merck Serono.
Dr Frauke Zipp has received research grants from Teva, Merck Serono, Novartis and Bayer as well as consultation funds from Teva, Merck Serono, Novartis, Bayer Healthcare, Biogen Idec Germany, ONO, Genzyme, Sanofi-Aventis and Octapharma. Her travel compensation has been provided for by the aforementioned companies.
Dr Jack Antel serves on advisory/safety monitoring boards for Novartis, Sanofi-Genzyme, Biogen Idec, EMD Serono and Medday Pharmaceuticals, and as editor of the Americas, Multiple Sclerosis Journal.
Dr Paul Giacomini has received personal compensation for speaking, consulting, and advisory board participation from Allergan, Bayer HealthCare, Biogen Idec, EMD Serono, Genzyme, Novartis, Roche and Teva Neuroscience, has received research support from Biogen Idec and Teva Neuroscience, has been a consultant for NeuroRx Research, an imaging Contract Research Organization, and has acted as a principal investigator or sub-investigator for clinical trials for Alexion, Bayer HealthCare, Biogen Idec, Elan, EMD Serono, GlaxoSmithKline, Novartis, Ono, Roche-Genentech, Sanofi-Aventis and Teva Neuroscience.
Dr Amit Bar-Or has participated as a speaker at meetings sponsored by, received consulting fees and/or received grant support from: Amplimmune, Bayhill Therapeutics, Berlex/Bayer, Biogen Idec, Diogenix, Eli-Lilly, Genentech, GlaxoSmithKline, Guthy-Jackson/GGF, Merck/EMD Serono, Medimmune, Mitsubishi Pharma, Novartis, Ono Pharma, Receptos, Roche, Sanofi-Genzyme, Teva Neuroscience, Wyeth.
Abstract: P652
Type: Poster
Abstract Category: Therapy - disease modifying - Immunomodulation/Immunosuppression
Background: The mechanisms underlying both the therapeutic effect of dimethyl fumarate (DMF), as well as its ability to differentially decrease counts of particular lymphocyte subsets in treated patients, are not fully elucidated. Here, using a combination of in vitro and in vivo approaches, we considered whether these DMF effects are mediated by selective apoptosis of distinct T cell subsets.
Methods: Total lymphocyte counts (TLC) as well as flow cytometry analysis of effector and regulatory
T cell subsets within peripheral blood mononuclear cells (PBMC), were carried out on samples obtained from MS patients prior to and at 3-month intervals following DMF treatment initiation. In vitro, PBMC were cultured with addition of DMF, monomethyl fumarate (MMF) or appropriate vehicle controls, subsequently stained for markers of T cell subsets and apoptosis (Annexin V/propidium iodide), then analyzed by flow cytometry.
Results: As previously reported DMF therapy reduced TLC, particularly over the first 6 months (mean decrease 0.79 x 109 cells/L; n=13; p=0.0009), with a preferential reduction in CD8+ T cells resulting in increased CD4:CD8 ratios (mean fold increase 1.4; p=0.002). Greater decreases were noted for memory versus naïve T cell subsets, and for conventional versus regulatory T cell subsets. These differential effects resulted in increased ratios of regulatory to effector T cells, and of anti- to pro-inflammatory cytokine-expressing T cells. In vitro, DMF induced dose-dependent apoptosis of CD3+ T cells (increase in late apoptotic cells vs. vehicle: 11% for 10µM DMF, p=0.0181; 51% for 50µM DMF, p=0.0001). Greater DMF-induced apoptosis was seen for CD8+ versus CD4+ T cells (79% vs. 39%, p=0.0001); for memory versus naïve CD4+ and CD8+ T cell subsets (CD4+: 73% vs. 18%, p=0.0004; CD8+: 92% vs. 8%, p=0.0064), and for conventional versus regulatory T cell subsets
(42% vs. 6%, p=0.0004).
Conclusions: In vitro, DMF induces T cell apoptosis that disproportionately affects CD8+ T cells, memory T cells, and non-regulatory T cells. This differential in vitro susceptibility of distinct T cell subsets to DMF-induced apoptosis mirrors the observed differential in vivo effects of DMF on the same T cells subsets and may thus provide insights both into the therapeutic mode of action of DMF in MS, as well as the mechanism underlying DMF-induced lymphopenia.
Disclosure:
Dr Mahtab Ghadiri is a recipient of the BMRI/McGill University Multiple Sclerosis scholarship, funded by Novartis.
Ayman Rezk: nothing to disclose.
Dr Rui Li reports: nothing to disclose.
Dr Felix Luessi has received travel grants from Teva Pharma and Merck Serono.
Dr Frauke Zipp has received research grants from Teva, Merck Serono, Novartis and Bayer as well as consultation funds from Teva, Merck Serono, Novartis, Bayer Healthcare, Biogen Idec Germany, ONO, Genzyme, Sanofi-Aventis and Octapharma. Her travel compensation has been provided for by the aforementioned companies.
Dr Jack Antel serves on advisory/safety monitoring boards for Novartis, Sanofi-Genzyme, Biogen Idec, EMD Serono and Medday Pharmaceuticals, and as editor of the Americas, Multiple Sclerosis Journal.
Dr Paul Giacomini has received personal compensation for speaking, consulting, and advisory board participation from Allergan, Bayer HealthCare, Biogen Idec, EMD Serono, Genzyme, Novartis, Roche and Teva Neuroscience, has received research support from Biogen Idec and Teva Neuroscience, has been a consultant for NeuroRx Research, an imaging Contract Research Organization, and has acted as a principal investigator or sub-investigator for clinical trials for Alexion, Bayer HealthCare, Biogen Idec, Elan, EMD Serono, GlaxoSmithKline, Novartis, Ono, Roche-Genentech, Sanofi-Aventis and Teva Neuroscience.
Dr Amit Bar-Or has participated as a speaker at meetings sponsored by, received consulting fees and/or received grant support from: Amplimmune, Bayhill Therapeutics, Berlex/Bayer, Biogen Idec, Diogenix, Eli-Lilly, Genentech, GlaxoSmithKline, Guthy-Jackson/GGF, Merck/EMD Serono, Medimmune, Mitsubishi Pharma, Novartis, Ono Pharma, Receptos, Roche, Sanofi-Genzyme, Teva Neuroscience, Wyeth.