As more treatment options emerge that have a significant impact on the peripheral immune system, the evaluation of lymphocyte count, and that of specific lymphocyte subsets, become more important in the treatment selection and management of patients with multiple sclerosis (MS).
1. Introduction
As more treatment options emerge that have a significant impact on the peripheral immune system, the evaluation of lymphocyte count, and that of specific lymphocyte subsets, become more important in the treatment selection and management of patients with multiple sclerosis (MS)
[1,2][1][2]. A greater understanding of the underlying pathophysiological mechanisms of MS has led to the development of therapeutics that address the cell count, migration, or functional state of lymphocytes. Though helpful in combatting the disease, changes in lymphocyte physiology can also be treatment-limiting. In addition, the measurement of peripheral lymphocyte counts appears to be important for treatment sequencing and planning of wash-out periods
[3]. Pharmacological effects on lymphocytes in the peripheral blood can serve as markers of patient compliance and can also assist in understanding the mechanism of action of MS therapies
[4,5][4][5].
Peripheral blood lymphocytes are frequently monitored in clinical practice as blood is easily accessible
[6]: lymphocytes continuously enter and exit the lymphoid and non-lymphoid organs via the blood
[7]. The assessment of lymphocyte subsets in the blood may provide useful information on immune system status
[8]. The measurement of physiological parameters of lymphocyte subsets has been used for some time to assist the selection of treatment regimens in specific diseases, e.g., human immunodeficiency virus (HIV) infection
[9]. However, blood lymphocytes can also be influenced by many conditions other than a disease or its treatment, including stress, smoking, sports, and aging
[8]. The extent of variation caused by these different factors can easily obscure alterations that have diagnostic value in pathogenic conditions.
2. Disease-Modifying Drugs and Their Effects on Lymphocyte Count
2.1. Mechanism of Action: Immunomodulation
2.1.1. Glatiramer Acetate
General Facts and Clinical Trial Data
Glatiramer acetate (GA) was the first DMT for MS successfully evaluated in humans (1977) and was approved by the US Food and Drug Administration in December 1996 and by the European Medicines Agency in 2001 for daily (20 mg/day) or triweekly (40 mg) subcutaneous application in patients with MS. Initially developed as a chemical and immunological analog of the major myelin antigen (myelin basic protein, MBP) to induce experimental autoimmune encephalopathy (EAE), GA did not work as intended. Instead of promoting encephalitic changes, GA was revealed as an efficient suppressor of encephalitic modulation. This effect and could even prevent EAE, which should normally be induced by myelin antigens such as GA
[34][10]. Across five randomized controlled clinical trials, GA 20 mg has consistently demonstrated efficacy in reducing the annualized relapse rate (ARR 29%) and MRI disease activity (33% reduction in the total number of enhancing lesions) and slowing of disability progression in patients with relapsing-remitting MS (RRMS)
[35,36][11][12]. Due to its favorable and well-characterized safety profile, GA is still often prescribed in patients with mild or moderate forms of MS.
Mechanism of Action and Impact on Lymphocyte Count
GA cross-reacts with MBP in a humoral and cellular respect and serves as an altered peptide ligand that promotes regulatory T cells instead of stimulating autoimmune T cell reactivity
[34,37][10][13]. The immunological effect underlies a strong and effective binding of MHCII molecules on antigen-presenting cells (APC). They compete with MBP and other myelin proteins for binding sites
[38,39][14][15]. This binding effectively replaces MBP, proteolipid protein (PLP), and MOG-derived peptides on their MHCII binding sites. This results in an altered T cell response, leading to suppression of myelin reactive T cells
[39,40][15][16] and the emergence of regulatory Th2 cells, which are able to recognize GA as well as MBP to cross the blood-brain barrier and secrete anti-inflammatory cytokines
[41,42][17][18]. These GA-specific Th2 cells additionally secrete high amounts of brain-derived neurotrophic factor (BDNF), which promotes neuroprotective effects
[43][19]. Furthermore, GA functionally inactivates T cells by antagonism on the T cell receptor and can induce regulatory CD4+, CD25+ cells by activating the regulatory pathway protein FOXP3 (
Figure 31B)
[44][20].
Figure 1. Association of proposed mechanism of action (MoA) of disease-modifying therapies (DMTs) and effects on lymphocytes. Categorized: oral therapies (A), injectables (B), and infusion therapies (C). CNS, central nervous system; COX-1, Cyclooxygenase-1; GSH, DHODH, dihydroorotate dehydrogenase; Glutathione; HCA2, hydroxy-carboxylic acid receptor 2, HIF-1α, hypoxia-inducible factor -1α; IL, interleukin; JAK/STAT, Janus kinases/signal transducer and activator of transcription proteins; nrf2, nuclear factor erythroid-derived 2-like 2; TNF-α, tumor necrosis factor-α; PGE2, prostaglandin E2; S1P, sphingosine-1-phosphate; Th1/2/17, T helper 1/2/17 cells; VLA, very late antigen.
While the total number of T cells in the blood compartment remains stable, studies have shown that GA treatment is associated with a reduction of B cells, plasma blasts, memory B cells, and a shift from pro- to anti-inflammatory B cell phenotypes
[45][21]. This may be driven by the cross-reactivity of B cell receptors for GA with antigens that are expressed in MS lesions
[45][21]. In contrast with interferon beta, GA is only associated with leukopenia or leukocytosis in exceptional cases
[46][22].
Recommended Monitoring
Considering these rare cases of lymphopenia/leukopenia but also leukocytosis and thrombocytopenia, a regular check of blood counts should be done at least tri-monthly in the course of the first year of therapy (
Table 21 and
Table 32). Subsequently, laboratory intervals can be increased to once or twice per year in the case of normal blood counts. The risk of severe GA-associated infections is low and not clinically meaningfully increased (1–2%)
[47][23].
Although there are no convincing study results regarding immune responses following vaccinations, GA treatment is not considered to limit immune responses
[1]. Verifying sufficient vaccination response via titer recording should be considered. Patients receiving GA should not be vaccinated with attenuated vaccines.
2.1.2. Interferons
General Facts and Clinical Trial Data
Interferons are a family of cytokines and physiologically function as signaling proteins. Since 1993 (US) and 1995 (EU), respectively, interferon-type beta (IFN-β) has played a role in the disease-modifying treatment of MS. Within the scope of the PRISMS study, subcutaneous (three times a week) application of INF-β-1a showed a risk reduction for relapses of 27% (22 µg, three times a week) and 33% (44 µg, three times a week) in a dose-related manner. Furthermore, it proved an effective treatment for RRMS in terms of defined disability and all MRI outcome measures
[48][24]. Today there are various preparations that differ by mode and frequency of administration. In addition to RRMS, interferon is approved for the treatment of clinically isolated syndrome (CIS) and immunomodulation during pregnancy and breastfeeding
[49,50][25][26].
Mechanism of Action and Impact on Lymphocyte Count
The effects of interferons are complex and, even today, are not completely understood. Activation of the JAK-/STAT-pathway via binding of the IFNAR-2 receptor is an established mechanism of action that leads to the expression of various genes (e.g., MX protein, beta2-microglobulin, 2′/5′-oligoadenylate synthetase, neopterin)
[51][27]. The activation of the signal transduction by INF-β results in an antiviral, immunomodulatory, and antiproliferative effect
[52][28].
With respect to the immunomodulatory impact, the following underlying mechanisms are considered (
Figure 31B):
- (a)
-
IFN-β leads to a reduction of dendritic cells and down-regulates the antigen presentation by APCs in peripheral blood and in the CNS by microglia and monocytes.
-
- (b)
-
The expression of toll-like receptor (TLR) 3, TLR7, and myeloid differentiation primary response 88 (MyD88) on dendritic cells increases, which leads to an altered immune response.
-
- (c)
-
INF-β induces CD4+, CD8+, CD25+, FOXP3+, and FOXA1+ T cells (regulatory T cells). A reduced inflammatory T cell response is observed by inhibiting the stimulation and activation of T cells (e.g., by modulation of co-stimulating molecules on dendritic cells), inhibition of the expression of MHCII molecules, and co-stimulating factors like CD80 and CD28 on APC
[53,54][29][30].
-
- (d)
-
The secretion of cytokines and chemokines is altered during IFN-β treatment (interleukin (IL)-10 and IL-4 increased; IL-2 and TNFα decreased). The differentiation of CD4+ cells shift from Th1 to a Th2 phenotype; thereby, resulting in a less pro-inflammatory but more anti-inflammatory cytokine milieu
[55][31].
-
- (e)
-
The number of Th17 cells also decreases, leading to a reduction of IL-17 release and induction of apoptosis of autoreactive T cells
[56,57][32][33].
-
- (f)
-
The effects on cytokines, chemokines, matrix metalloproteinase, and adhesion molecules (especially very late antigen [VLA]-4 on T cells) result in a reduced leukocyte migration via the blood-brain barrier into the CNS
[53,58,59][29][34][35].
-
IFN-β-1a treatment results in selective, time-dependent effects on many cell populations in peripheral blood
[60][36]. The IFN-β-promotes down-regulation of pro-inflammatory CD4+, CD8+ memory T cells, and memory B cells accompanied by an increase in regulatory T cells
[52,53,58,61][28][29][34][37].
The majority of patients treated with IFN-β exhibit a fall in absolute lymphocyte counts of approximately 20–30% compared to the baseline value. About 15% of interferon-treated patients develop lymphocyte decreases below the lower limit of normal, 3.5% below 0.8 GPt/L, and about 1% of patients below 0.5 GPt/L
[62][38]. The drop in lymphocyte count is often transient and recovers to normal levels within months. During a study evaluating the dynamics of lymphopenia during IFN-β treatment, onset of cytopenia occurred within the first 6 months of therapy in at least two-thirds of patients
[62][38]. The majority of events were mild and generally resolved within 3–4 months while continuing therapy. Dose reductions were uncommon, and only a small proportion of patients (6 of 727; 0.8%) discontinued treatment after approximately 2 years because of hematological abnormalities when receiving the highest dose of INF-β-1a (44 μg three times weekly).
Recommended Monitoring
The rate of severe infections during IFN-β treatment does not seem to be significantly increased
[1]. On the contrary, IFN-β has clear antiviral effects. There are no data available with respect to the duration of lymphocyte recovery in the case of lymphopenia. However, if repopulation has not occurred long after treatment discontinuation, hematological diseases should be excluded.
A regular check of blood counts including, leukocyte and lymphocyte counts, should be done at least tri-monthly in the course of the first year of therapy (
Table 21 and
Table 32). Subsequently, laboratory intervals can be increased to once or twice a year in the case of normal blood count levels.
Table 1. Recommended lymphocyte thresholds for disease-modifying therapies.
|
Drug Name |
Recommendations for Lymphocyte Cut-Off Values |