Sundararajan Jayaraman1*
Arathi Jayaraman2- 1Department of Surgery, University of Illinois College of Medicine, Peoria, IL, United States
- 2Xavier University School of Medicine, Oranjestad, Aruba
Multiple sclerosis is a progressive demyelinating central nervous system disorder with unknown etiology. The condition has heterogeneous presentations, including relapsing-remitting multiple sclerosis and secondary and primary progressive multiple sclerosis. The genetic and epigenetic mechanisms underlying these various forms of multiple sclerosis remain elusive. Many disease-modifying therapies approved for multiple sclerosis are broad-spectrum immunomodulatory drugs that reduce relapses but do not halt the disease progression or neuroaxonal damage. Some are also associated with many severe side effects, including fatalities. Improvements in disease-modifying treatments especially for primary progressive multiple sclerosis remain an unmet need. Several experimental animal models are available to decipher the mechanisms involved in multiple sclerosis. These models help us decipher the advantages and limitations of novel disease-modifying therapies for multiple sclerosis.
Introduction
Clinical manifestations of multiple sclerosis
More than 2.8 million people live with multiple sclerosis (MS) worldwide, and the prevalence has been increasing (1). The mean age of diagnosis of MS is 32 years, with twice the number of female patients compared with male patients afflicted with this disease. However, the basis of sexual dimorphism in MS manifestation remains elusive, as in other autoimmune diseases. MS is a prototypical organ-specific autoimmune disease of the central nervous system (CNS), affecting the brain and spinal cord (2–4). Most (85%) patients with MS manifest relapsing-remitting MS (RRMS), characterized by alternate periods of relapses and remissions for decades after an initial episode of neurological dysfunction, clinically isolated syndrome. Relapses accompany CNS inflammation and demyelination detectable as white matter lesions by magnetic resonance imaging. Accumulating disabilities during relapses in most (80%) patients with MS leads to secondary progressive MS (SPMS), characterized by decreased brain volume and increased axonal loss without associated inflammatory lesions. A minor fraction (10%) of patients with MS continue to decline progressively from the beginning of diagnosis without relapses. Variations of MS include progressive-relapsing and pediatric disease and severe Marburg variant. The hallmark of MS is sharply demarcated demyelinating plaque with axons relatively preserved, whereas in neuromyelitis optica (MNO), both axons and myelin are involved, resulting in necrotic cavitation. Severe involvement of optic nerves and the spinal cord is a characteristic of the opticospinal MS (OSMS) subtype, which is more prevalent in African Americans (5, 6). Compared with Whites, African Americans had an older age at onset, experienced greater disability, progressed faster, had increased risk for SPMS, experienced transverse myelitis more often, and were likely to have motor symptoms and the OSMS subtype. The classic multifocal MS is rare in Japanese, who manifest OSMS with features similar to those of the relapsing form of NMO in Western populations, and was proposed to be the same as the NMO disorder, rather than a form of MS (7). However, in Brazilian patients, OSMS is recognized as a milder MS phenotype distinct from NMO (8). While antibody-dependent aquaporin four loss occurred in some patients with NMO, antibody-independent astrocytopathy was found in several demyelinating conditions, including Baló's disease, NMO, and MS (9). In addition to these complexities, MS is also rare among Samis, Turkmen, Uzbeks, Kazakhs, Kyrgyzis, native Siberians, North and South Amerindians, Chinese, Japanese, African blacks, and New Zealand Maoris, in contrast to a high propensity of Sardinians, Parsis, and Palestinians to develop MS (10). The different susceptibilities of distinct racial and ethnic groups are essential determinants of the uneven geographic distribution of MS.
The clinical manifestations of MS include temporary vision loss, sensory and motor problems, fatigue, impaired bowel and sexual functions, cognitive deficits, and paralysis (2–4). Distinct forms of MS appear to correlate with the spatiotemporal dissemination of lesional sites within the CNS (2–4, 11). The hallmarks of MS pathology include the breakdown of the blood–brain barrier, accumulation of immune cell infiltrates, oligodendrocyte loss, demyelination, astrogliosis, axonal degeneration, and disruption of neuronal signaling (Figure 1). Substantial T-cell infiltration occurs in patients with acute and relapsing disease but is spared during later stages of MS, despite an unabated neuronal disability. Intrinsic neuronal deficits such as those associated with Alzheimer's disease are thought to play a role, especially during the advanced stage of MS (11).
Figure 1. Events involved in induction and amelioration of EAE.
Genetics of MS
Although the etiology of MS remains elusive, genes within the human leukocyte antigenic (HLA) loci, such as HLA-A*02:01, HLA-DRB1*15:01, HLA-DRB5, HLA-C, and TNF, have been firmly associated with MS susceptibility (12). In African Americans, classic/multifocal MS is associated with DRB1*15 alleles, whereas OSMS is not (5). Not only the DRB*1501 allele but also the extended DRB1*1501-DQB1*0602 haplotype is commonly found in northern Europeans with MS (5). The HLA-DPB1*0501 haplotype is not uniquely associated with the OSMS subtype, which is relatively more common in Japan (13). Interleukin-2 (IL-2) and its receptor IL-2R play a crucial role in MS and are also crucial for T-cell tolerance (14). In addition, the soluble form of the IL-2R (sIL-2R) plays a role in MS. IL-7 and IL-7Rα form a non-redundant ligand–receptor system and plays a critical role in T-cell activation. Peripheral blood mononuclear cells of patients with MS display deletion of exon 4 of the IL-7 transcript and splice variants lacking exons 5, 6, and 7 (15). A closer analysis of the impact of these genetic variations is necessary for a better understanding of MS pathogenesis.
The pivotal role of T helper cells in MS
Cerebrospinal fluid (CSF)-infiltrating CD4+ T cells of patients with MS proliferated and secreted interferon- γ (IFN- γ), a characteristic of the Th1 subset, but not IL-17 when challenged with the myelin oligodendrocyte glycoprotein 35–55 (MOG35 − 55) peptide in vitro (16), a proposed candidate CNS determinant in MS (17). However, others reported the abundance of IL-17-expressing Th17 cells in the peripheral blood, CSF, and brain lesions of patients with MS, which increased during relapses (18). Increasing evidence also indicates a role of central memory Th17.1 (Th1/17) cells, which share the hallmarks of Th1 and Th17 cells, respectively, in IFN- γ and IL-17 production, in MS (18). In addition to Th17 cells, follicular helper T cells that promote the germinal center formation, B-cell differentiation, and antibody production are also implicated in several autoimmune diseases, including MS (19). The intrathecal inflammatory environment in patients with RRMS promotes the recruitment of peripheral follicular helper T cells to the CNS without increasing their ability to migrate (20). Since the follicular helper T cells failed to transfer demyelinating disease in mice (21), it is unlikely that they have pathological consequences in patients with MS. The role of follicular T helper cells in MS remains to be proven. Although MHC class I-restricted CD8+ cells were found in the brain lesions of patients with MS, they were also present in patients with infections and other brain diseases, providing inconclusive evidence for their involvement in MS (22).
Epstein–Barr virus (EBV) infection and MS
Infection with EBV is associated with monoclonal or oligoclonal B-cell expansion in many autoimmune diseases, including Hashimoto's thyroiditis, Grave's disease, Sjögren's syndrome, rheumatoid arthritis, systemic lupus erythematosus syndrome, and MS (23). Whereas infectious mononucleosis increases the risk of MS, the vast majority (90–95%) of the world population infected with EBV at some point in life does not develop MS (24). Although elevated EBV nuclear antigen 1 IgG titers are associated with gadolinium-enhancing brain lesions, the lack of correlation between acute viral reactivation in the peripheral blood and MS lesions suggests a limited role for EBV infection in driving the disease activity (25). Despite the increased level of EBV viral load in patients with RRMS compared with controls, there was no statistically significant difference in EBV and human herpes virus-6 (HHV-6) copy numbers between the patients and controls (26). In addition, the frequency of NK and CD8+ T cells increased during relapse, which was not associated with EBV and HHV6 plasma viral loads. Although EBV infection has been hypothesized to contribute to MS development in the context of other predisposing conditions, such as the HLA genotype, vitamin D deficiency, smoking, and altered T-cell responses (23), evidence for this hypothesis remains to be garnered.
Disease-modifying therapies for MS
Several disease-modifying therapies (DMTs) were approved for MS treatment by the Food and Drug Administration (FDA) [reviewed in (27, 28)]. These include self-injectables such as the anti-virals IFN-β-1a and b, first-line treatment, and peginterferon beta-1a provided moderate protection against RRMS (29, 30). However, the production of antibodies against IFN-β and the lack of the effect on Th17 cells, considered encephalitogenic, (31) remained a major concern. Glatiramer acetate designed based on four amino acids from myelin basic protein (MBP) was designed to induce clinical disease in animals but was well tolerated with low/moderate efficacy on RRMS (32). Several orally administered drugs, including teriflunomide, provided moderate effects on RRMS (33, 34). Dimethyl fumarate and diroximel fumarate (35, 36), and fingolimod/FTY720 (37), the first approved oral drug for MS, had moderate beneficial effects on RRMS but with several side effects, including progressive multifocal leukoencephalopathy (PML). Modulators of sphingosine-1-phosphate receptor 1 (S1PR1) and S1PR5, and siponimod decreased oligodendrocyte and axonal loss (38). Ozanimod and SIPR1 and 5 agonist reduced plasma neurofilament light-chain concentrations (39), and the selective S1PR1 modulator ponesimod (40) and cladribine, a deoxyadenosine analog (41), provided moderate benefits to patients with MS. Notably, many intravenous infusion strategies were implemented for MS treatment. Mitoxantrone, a general immunosuppressant, was the first-line treatment with high efficacy for MS (42). The first humanized monoclonal antibody (mAb) used for MS treatment, natalizumab (43), is directed against anti-α4β1-integrins and blocks the entry of immune cells into the CNS. Although it is highly effective, it causes PML in John Cunningham virus-seropositive patients. The first humanized mAb, anti-CD52 antibody (campath-1/alemtuzumab), originally used for treating graft vs. host disease proved to be highly efficacious for MS treatment but associated with significant side effects (44, 45). Several B-cell-depleting anti-CD20 mAbs, such as rituximab, ocrelizumab, ofatumumab, and ublituximab, were highly efficacious for MS treatment but with PML occurrence in some cases (46–50). Most of these drugs were designed to regulate adaptive immune cells prominent during the early, but not the late, stage of MS (3, 27, 28). Some of these therapies reduce relapses but do not prevent the progression of MS and the accumulation of disabilities. The first-line treatments for MS, such as glatiramer acetate (32), dimethyl fumarate (35), and natalizumab (anti-IFN-β-1b) (43), affect T cells variously. Whereas IFN-β-1a/b reduced relapses without affecting Th17 cells (31), glatiramer acetate (32) and dimethyl fumarate suppress Th1 while upregulating Th17 cells (27). Alemtuzumab decreases central memory T cells (27). Fingolimod targets the SIPR and blocks T-cell transmigration into the CNS. This treatment results in cardiac complications, varicella–zoster, and herpes simplex virus reactivation, and exacerbation of MS (27, 28, 37). Natalizumab, a humanized monoclonal antibody, selectively targets the α4 subunit of the cell adhesion molecule, very late antigen 4, and prevents leukocyte adhesion and diapedesis at the blood–brain barrier, leading to PML in John Cunningham-virus seropositive patients (2, 3, 27, 28, 43). Systemic administration of anti-CD20 monoclonal B-cell-depleting antibody rituximab in patients with PPMS reduced gadolinium-enhancing lesions and relapses for 48 weeks (46). However, long-term therapy with ocrelizumab, a humanized depleting anti-CD20 mAb, provided modest protection against PPMS (47). Earlier and continuous treatment of patients with PPMS with ocrelizumab over 6.5 years provided sustained benefits on measures of disease progression (48). Since CNS B cells residing in meningeal ectopic lymphoid follicles are associated with subpial inflammation in patients with SPMS, inadequate penetration of the anti-CD20 antibody across the blood–brain barrier into the CNS could explain the lack of protection observed in some studies. Rituximab administered intrathecally also failed to provide clinical benefits in the phase 1b clinical trial on progressive MS (49). Other B-cell-depleting antibodies including ofatumumab (50) and ublituximab, a novel glycoengineered anti-CD20 mAb (51) that was administered SC unlike other mAbs, induced modest protection against MS.
Since 2018, several second-generation molecules with reduced gastrointestinal side effects have been approved for the treatment of MS by the FDA (52). Diroximel fumarate, the second-generation version of dimethyl fumarate, is lymphopenic and modifies monocytes. Oral formulations of S1PR modulators such as siponimod, ozanimod, and ponesimod target S1PR1 and SIPR5 have potentially better safety profiles. Ofatumumab, an anti-CD20 antibody administered subcutaneously, and glycoengineered anti-CD20 antibody, ublituximab, and oral compounds such as teriflunomide and cladribine were also approved for MS treatment (52).
Several other DMTs outnumbering those approved for MS treatment failed to meet the primary study endpoint and progress to a subsequent clinical trial because of commercial decisions. These include antibodies against the IL-12/23 p40 subunit (53), anti-CD25 (54, 55), CTLA-4-Ig (56), and anti-IL-17A (57). The mAbs targeting different subsets of B cells, tabalumab inhibited B-cell activation factor (BAFF), and atacicept induced depletion of mature B cells and suppressed antibody formation (58). However, they failed to deplete memory B cells and inhibit relapsing MS. Moreover, GNbAC1, a humanized mAb directed against an endogenous retroviral protein (59), and raltegravir (Isentress), the HIV integrase strand inhibitor (60), did not have an impact on MS disease activity. Interestingly, natalizumab failed to demonstrate a significant protective effect in patients with SPMS (61, 62). In addition, the anti-CD20 antibody, rituximab, shown to have superior protection in RRMS, has been abandoned due to the expiry of the patent (61, 62).
In addition to these non-specific drug therapies, several attempts were made to induce antigen-specific tolerance in encephalitogenic T cells, which would ensure stable and adequate protection against autoimmune diseases without off-target effects [reviewed in Refs. (63, 64)]. These include the administration of synthetic peptides corresponding to the T-cell epitopes mapped within myelin components such as MBP, MOG, proteolipid proteins (PLP), and altered ligand peptides. Moreover, T-cell receptor (TCR) vaccination constituting attenuated autologous antigen-specific T cells and autologous peripheral blood mononuclear cells chemically coupled with myelin peptides were also undertaken. None of these maneuvers induced T-cell tolerance as assessed by the ability of peripheral blood T cells to proliferate and produce IFN-γ in response to a challenge with the corresponding immunizing peptide in vitro. Significantly, they also did not improve the clinical outcome in patients with MS. Thus, effective methods of inducing antigen-specific tolerance in encephalitogenic T cells without causing adverse reactions remain an unmet need.
Experimental models of MS
Myelin antigen-induced experimental autoimmune encephalomyelitis
The MS-like disease, experimental autoimmune encephalomyelitis (EAE), has been successfully induced in monkeys, guinea pigs, rats, and mice, following immunization with the whole-brain and spinal cord extracts and peptides derived from myelin proteins, such as MBP, PLP, and MOG [reviewed in Ref. (65)]. The mouse is a popular choice for studying MS variations primarily due to the availability of genetically defined inbred strains and transgenic and gene knockout mice. SJL/J mice immunized with the PLP139 − 151 peptide or peptides derived from MBP exhibited relapsing-remitting EAE (RR-EAE) (66), and this model would allow the development of novel DMTs for RRMS. Immunization with rat MOG induced classic EAE in congenic C3H.SW (H-2b) mice, while causing atypical EAE characterized by ataxia, proprioception defects, and axial rotary clinical presentation in C3HeB/Fej (H-2k) mice (66, 67). Atypical EAE was also induced in IFN-γ knockout mice on the BALB/c background immunized with MBP-derived peptides (68). In one study, granulocytes were implicated in atypical EAE (66), while others found the participation of granulocytes in both classic EAE and atypical EAE (68). The brain seems primarily involved in atypical EAE, while the spinal cord is considered the primary target of classic EAE and RR-EAE (66, 68). Since the brain is primarily involved in MS (2–4), atypical EAE models may provide valuable tools for further understanding the mechanisms of brain lesions and their prevention.
MOG is a member of the immunoglobulin superfamily expressed exclusively in the CNS myelin. The MOG35 − 55 region proved to be an immunodominant epitope eliciting T- and B-cell responses and EAE in most strains of mice (65, 69–80). MOG35 − 55 was identified as an autoantigen in patients with MS (17). Immunization of C57BL/6 (H-2b) mice with the MOG35 − 55 peptide elicited EAE (78–80). However, MOG35 − 55 peptide immunization induced a robust and long-lasting progressive EAE (PEAE) in non-obese diabetic (NOD) (H-2g7) mice (70–77). Interestingly, pronounced remissions were observed in some (70–72), but not in other, studies (73–77), indicating variations in PEAE. Genetic drift and gene deletions could be attributed to the inconsistency in remissions in NOD mice bred in different geographical locations—Oceania, Europe, and the United States. The detection of T cells recognizing MOG35 − 55 peptide in patients with MS (17) provided an impetus to explore EAE specifically induced by this peptide autoantigen, although other myelin peptide antigens also elicited EAE in multiple strains of mice (65, 69) (Table 1). Moreover, NOD mice develop several autoimmune diseases, including type 1 diabetes (81) and other endocrine gland-related autoimmune conditions, such as thyroiditis, sialitis, and Sjögren's syndrome (82–84). Thus, NOD mice offer a unique opportunity to study the mechanisms of self-reactive T-cell-mediated neurodegeneration in an autoimmune environment. Significantly, PEAE induced in NOD mice lasts throughout the life of the mice with increasing disabilities (70–77), unlike the non-autoimmune-prone C57BL/6 mice (Table 1) (78–80). Biozzi ABH mice also develop PEAE when immunized with the whole spinal cord homogenate (85). Immunization of Lewis rats with gpMBP68 − 84 (86) and dark Agouti rats with MOG1 − 125 also induced classic EAE (87). Thus, EAE is a well-studied model system of MS and is amenable to investigating the efficacy of novel treatment options.
Table 1. Regulation of EAE by HDAC inhibitors.
Other demyelinating disease models
Infection of mice with the neurotropic picornavirus Theiler's murine encephalomyelitis virus (TMEV) induces a disease similar to PPMS involving the brain, brainstem, and spinal cord (88). The TMEV infects macrophage/microglia, oligodendrocytes, and astrocytes during the chronic phase. Axonal damage in MS and EAE occurs secondary to inflammatory demyelination (outside-in model) (89). By contrast, TMEV infection induces demyelinating lesions that develop from the axon to the myelin (inside-out model) (90). Although TMEV infection cannot occur naturally in rodents or humans (91), it is a valuable model for studying the efficacy of drugs to prevent axonal degeneration independent of immune mechanisms. Feeding of C57BL/6 mice with the copper-chelating agent cuprizone induced demyelination, oligodendrocyte death, and profound activation of astrocytes and microglia (91). Removing cuprizone from the diet led to the regeneration of oligodendrocytes from the pool of oligodendrocyte progenitors and the formation of myelin sheaths, indicating the reversible nature of the disease. Interestingly, lysolecithin injection produced focal areas of demyelination in SJL/J mice, rats, and rabbits due to direct toxic effects on myelin sheath without affecting other cells and axons (91). These models help study the process of de- and remyelination independent of the involvement of immune mechanisms.
EAE models for investigation of MS therapeutics
EAE models have traditionally been used to benchmark the efficacy of various disease-modifying therapies. However, several inconsistencies between mice and humans concerning the outcome of these attempts have been intensely debated (91–93). A few established MS therapies, including glatiramer acetate (copolymer 1), mitoxantrone, and natalizumab, were tested in animal models, which turned out to be potent non-specific suppressors and unsuitable for all patients with MS (27). Some DMTs were investigated in EAE models retrospectively after disappointing outcomes in human trials (91, 92). The failures of translational therapies for MS treatment could be due to differences in genetics, the extent of blood–brain barrier disruption, and individual variability in the responsiveness of patients to treatment. Emphasis has also been placed on discovering reliable biomarkers of MS and improving the design of CNS drug delivery (93). Most of the multifocal symptoms of classic MS have not been reproduced in rodent models. This limitation should be kept in mind when discussing the lack of efficacy of the DMTs for MS treatment since this disease is highly heterogenous and sometimes manifest with other comorbidities.
Epigenetic approaches to control EAE
In EAE, adaptive immune T and B cells, the innate immune granulocytes, and the CNS-resident cells such as microglia, astrocytes, and oligodendrocytes collectively contribute to neurodegeneration. Gene expression is a highly regulated process, and aberrant expression of mRNA encoding cytokines and chemokines contributes to pathological manifestations. Although the genome-wide association studies have implicated genes encoding human leukocyte antigens in MS pathogenesis (94), environmental factors such as Epstein–Barr virus infection, smoking, and vitamin D deficiency may influence gene expression via epigenetic mechanisms (95). Epigenetics is the heritable changes in gene expression without altering the DNA sequence, which can provide a mechanism by which external factors, including drugs, produce various phenotypic variations with identical genotypes (96). Discordance in the rate of MS among monozygotic twins suggests that susceptible genes alone are not enough to manifest the neuronal disease, implying the participation of epigenetic mechanisms in disease manifestation (97). DNA methylation (98) and microRNAs (99) have been proposed to play a role in MS. However, direct evidence supporting the contention that modulation of these epigenetic mechanisms can result in neuroprotection is lacking.
Histone acetylation is the most well-characterized posttranslational mechanism of histone modifications, facilitating an open chromatin configuration and gene transcription (96) (Figure 1). The balance between acetylation by histone acetyltransferases and their regulation by histone deacetylases (HDACs) dictates the outcome of transcription of many protein-coding genes (96) and, interestingly, a non-coding microRNA (100). Trichostatin A (TSA), a hydroxamate member, was initially developed for cancer treatment (101) and is the most potent broad-spectrum HDAC inhibitor (102). TSA inhibits the transcription of class I, IIa, IIb, and IV HDACs (76). When C57BL/6 mice were immunized with MOG35 − 55 and treated with large doses of TSA s.c throughout the investigation, a modest reduction in the EAE score was noted (78) (Table 1). Similarly, daily oral administration of vorinostat, another hydroxamate that inhibits class I and IIa HDACs (102) throughout the period of investigation, also reduced the acute EAE in C57BL/6 mice (79). Interestingly, the class I HDAC inhibitor and the antiepileptic drug valproic acid when administered prophylactically or therapeutically reduced acute EAE in C57BL/6 mice (80). Notably, s.c administration of a lower dose of TSA prophylactically up to 45 days on alternate days provided irreversible and prolonged protection against PEAE in NOD mice (74). Consistent with these encouraging results of HDAC inhibitors to treat neurodegenerative diseases in mice, oral treatment of Lewis rats (86) or i.p administration of DA rats (87) with valproic acid reduced EAE induced by immunization with gpMBP68 − 84 and MOG1 − 125 peptides, respectively. In contrast to the success of reducing the clinical scores by TSA and valproic acid in C57BL/6 and NOD mice, oral administration of another hydroxamate panobinostat, givinostat, a pan-lysine inhibitor, or entinostat therapeutically from day 20 onward failed to afford protection against PEAE (77). These data indicate that not all HDAC inhibitors can serve as potent DMTs for ongoing neurodegeneration.
Neuroprotection provided by TSA, vorinostat (SAHA), and valproic acid corroborated with reduced CNS inflammation and demyelination in mice (74, 75, 79, 80). Significantly, inhibition of axonal degeneration during PEAE was also prominently mediated by TSA (74). Reduced T-cell proliferation and suppression of Th17 cells were noted in HDAC inhibitor-treated rodents (74, 79, 86, 87). Neuroprotection was also accompanied by decreased CD4+CD44+ cells, a characteristic of activated/memory cells (103), and reduced ability of T cells to produce IFN-γ, IL-17A, and GM-CSF in response to a challenge with MOG35 − 55 in vitro (74). Histone hyperacetylation rendered T cells unresponsive to the MOG35 − 55 antigen challenge while retaining their ability to respond to polyclonal stimulation (74), akin to anergy (104). By contrast, daily oral administration of HDAC inhibitors such as panobinostat, givinostat, and entinostat from the start of clinical signs (day 20) failed to protect NOD mice from PEAE or fatality, despite reduced T-cell proliferation in vitro and diminished transcription of Tbet and Rorγt (77). However, the antiepileptic drug valproic acid (54) and the anti-cancer drug, TSA (74), administered therapeutically (after the disease onset, Table 1) provided robust neuroprotection and thus may be useful in a clinical setting.
Regulation of the innate immune system in EAE by HDAC inhibitors
In MS, innate immune cells, such as infiltrating macrophages and dendritic cells, and CNS-resident microglia, have been implicated in the reactivation of T cells during the effector phase of neurodegeneration (2, 3). In NOD mice, PEAE development was associated with the expansion of mature (MHC class II+) CD11b+Ly-6G+ neutrophils and, to a lesser extent (MHC class II+) CD11b+Ly-6C+ mature monocytes in the peripheral lymphoid compartment before the onset of the peak clinical disease (75). Participation of neutrophils in monophasic EAE of C57BL/6 mice was indicated by increased neutrophils in the bone marrow, blood, and spleen during the early phase of the disease (105). Studies suggested a role for neutrophils in MS during the initial formation of lesions in the brain, but not during the advanced stages of the disease, probably owing to the short-lived nature of neutrophils (106). Treatment with TSA concurrently afforded neuroprotection and diminished the frequency of neutrophils in secondary lymphoid organs and their influx into the spinal cord (75), indicating a role for these cells in the PEAE model (Figure 1). Thus, in addition to myelin-specific T-cell tolerance induction, selective regulation of the innate immune system appears to be an integral part of the regulation of neurodegeneration by the HDAC inhibitor TSA.
Implications of HDAC inhibitor-induced regulation of EAE to MS treatment
Impact of immune regulation
Immune responses elicited by immunization with the whole spinal cord homogenate or various peptides derived from the CNS-associated MBP, PLP, and MOG have been extensively studied in mice and rats that develop monophasic EAE, PEAE, and atypical EAE (65–80, 86, 87). Various methodologies such as ELISA, Western blot, flow cytometry, and quantitative reverse transcriptase-mediated polymerase chain reaction (RTq-PCR) have provided significant insights into the underlying immune mechanisms of EAE. However, consensus on whether any given immune mediator can serve as a biomarker indicating the stage and severity of the chronic disease remains enigmatic. Most studies focused on immune mediators typically at the peak of the clinical disease after in vitro activation with T-cell ligands. A systematic and comprehensive analysis of basal levels of 41 genes frequently implicated in neurodegeneration and their regulation by TSA treatment was assessed using RTq-PCR in the CNS and secondary lymphoid organs longitudinally during the prolonged course of PEAE (27 weeks) without overt activation in vitro (76). These studies indicated that immunization of NOD mice with MOG35 − 55 increased the expression levels of mRNAs encoding IL-4 and IL-17A in the CNS during the chronic phase, days 21–54. The reduction in the level of IL-17A gene in TSA-treated mice is consistent with the proposed role of IL-17A in EAE (107). Prolonged expression of Nos2 in the CNS (76) is in line with the association of iNOS-positive macrophages, astrocytes, and granulocytes in demyelinating pathology (108). Increased numbers of neutrophils in the spleen and spinal cord and their downregulation by the histone modifier treatment support this contention (75).
On the other hand, in the peripheral lymphoid tissues, genes encoding the heterodimeric chains of IL-27, IL-27p28, and IL-27EBi3, implicated in EAE (109), were overexpressed in PEAE mice, which were reduced by TSA treatment. Augmentation of the transcriptional repressors by histone acetylation could indirectly cause a reduction in gene expression. Notably, in vitro activation of peripheral lymphoid cells from TSA-treated mice exhibited compromised expression of both intracellular and secreted IL-17A and IFN-γ (74). Interestingly, TSA treatment reduced the infiltration of Th1 and Th17 cells from the periphery into the spinal cord (74) (Figure 1). This is similar to the suppressive effect of valproic acid on the influx of T cells into the spinal cord of EAE Lewis rats (86). These data demonstrate that the infiltration of T lymphocytes into the CNS is crucial for neurodegeneration, and their retardation by HDAC inhibitors facilitates neuroprotection.
Although migration inhibitory factor (MIF) has been proposed to be crucial for EAE (110), surprisingly, it was not transcriptionally upregulated in the CNS and lymphoid tissues of NOD mice manifesting PEAE (76). Yet, TSA treatment repressed the constitutive expression of Mif in protected mice. Surprisingly, several other genes implicated in EAE, including GM-CSF (111), prominent chemokine CCL2 (112), transcription factors T-bet (113), and RORγt (114), were neither overexpressed in the PEAE mice nor downregulated by TSA treatment (76). However, in EAE rats, valproic acid treatment suppressed the mRNA levels of IFN-γ, TNF-α, IL-1β, MMP9, iNos, and Tbet and increased IL-4 in the spinal cord (86). The transcription factor FoxP3 mRNA was neither upregulated in the PEAE model nor modulated by chromatin modifier treatment (76), similar to the lack of suppression of FoxP3 transcription in another study (77). TSA treatment also did not alter the numbers of FoxP3+ T regulatory cells in NOD mice (74, 76). Although the transcription factor FoxP3 is essential for the generation of T regulatory cells (115), it is contentious whether these cells are involved in the regulation of EAE (116, 117). Studies in mice indicated the upregulation of genes encoding anti-oxidants, neuroprotection, and neuronal differentiation by TSA treatment (78), while the expression of Sox8 and Mog was upregulated in valproic acid-treated rat brains (87). Valproic acid administration also reduced the genes crucial for apoptosis, and caspase-3,−8, and−9 in T cells (78). Collectively, these data indicate that the HDAC inhibitors modulate the transcription of several genes crucially involved in neurodegeneration.
The role of histone deacetylases in EAE and their modulation by TSA
Surprisingly, immunization of NOD mice with MOG35 − 55 upregulated the transcription of Hdac11 in the CNS, but none of the 11 Hdacs in the peripheral lymphoid cells (76). The wide-spectrum HDAC inhibitor, TSA, did not diminish the Hdac11 enzymatic activity in vitro (118), indicating the lack of correlation between Hdac expression and Hdac activity. Nevertheless, the data demonstrating the selective upregulation of Hdac11 in the spinal cord of PEAE mice and its downregulation by TSA treatment have implications to the control of MS by histone modifiers. The use of high-resolution in situ hybridization and imaging revealed abundant expression of Hdac11 in the hippocampus and Purkinje cells of rat brains, suggesting a role in locomotor activity and ataxic syndromes, respectively (119). However, it is unclear whether in PEAE mice, Hdac11 expression is localized to these cells and downregulated by TSA treatment. Knockout of Hdac11 reduced the infiltration of monocytes and myeloid DC into the CNS, expression of CCL2, clinical severity, and demyelination (120). Although both TSA treatment and Hdac11 gene knockout resulted in amelioration of EAE, the protective effect of Hdac11 deletion observed may be secondary to the absence of Hdac11 in the CNS and unrelated to the impact on monocytes and CCL2 expression (120). Nevertheless, by extrapolation, repression of Hdac11 could be beneficial in treating patients with MS with broad-spectrum HDAC inhibitors, such as TSA. Although Hdacs other than Hdac11 was not regulated by the histone modifier either in the peripheral lymphoid tissues or in the CNS (74), HDAC3 mRNA was reportedly increased in the peripheral blood mononuclear cells of patients with RRMS (121). However, another study failed to validate this observation (122), indicating uncertainty of the role of HDAC3 in MS. Interestingly, TSA treatment prevented the manifestation of type 1 diabetes in NOD mice associated with the transcriptional repression of Hdac4, Hdac8, and Hdac9, but not Hdac11, in the spleen (123). However, TSA administration did not influence the transcription of Hdac genes expressed in the target organ pancreas. These data suggest that the overexpression of specific Hdac is tissue- and disease-specific, which could be utilized to manipulate hard-to-treat diseases, including MS.
Implications of HDAC inhibition to MS treatment
Targeting multiple HDAC isoforms might be necessary for specific indications and proof-of-concept studies. The involvement of specific HDACs crucial for various forms of MS has not yet been delineated. Studying the expression level of different HDAC genes in particular cell types in the secondary lymphoid organs and the CNS is essential for designing selective HDAC inhibitors for MS treatment. Based on the data obtained, it is possible to create more selective compounds that could prove safer by reducing off-target effects. In addition to the downregulation of many genes, the expression of the transcription factor Dec1 (Bhlhe40) was upregulated in the CNS of TSA-treated mice (76). Thus, HDAC inhibitors such as TSA with broad specificity might provide benefits against complex neurodegenerative diseases by concurrently repressing and increasing the transcription of multiple genes. The wide range of the action of the broad-spectrum HDAC inhibitor is likely to provide protection against complex neurodegenerative diseases like MS. Consistently, therapeutic intervention with HDAC inhibitors has been proposed to enhance synaptic plasticity, learning, and memory in Alzheimer's disease, Huntington's disease, and Parkinson's disease (124). Lysine acetylation of non-histones constitutes a significant portion of the acetylome in mammalian cells and is involved in several cellular functions, including gene transcription (125). However, it is unclear whether HDAC inhibitors can also acetylate non-histones and alter gene transcription in conjunction with gene regulation mediated by acetylation of histone tails. Nevertheless, changes in gene expression due to inhibition of HDACs by small-molecule inhibitors could have substantial impact on regulating disease pathogenesis.
Recent work has unraveled the inheritance of non-DNA sequence-based epigenetic information, epimutations, across several generations in yeast to humans (126). The signals that underpin these epimutations, including DNA methylation, histone modification, and non-coding RNAs, and the underlying mechanisms are beginning to be understood (127). Treatment of the nematode Auanema freiburgensis with class I HDAC inhibitors butyrate and valproic acid, and the broad-spectrum HDAC inhibitor TSA increased the acetylation of histones 3 and 4 (128). Notably, they also exerted transgenerational effects on the offspring by producing increased numbers of hermaphrodites, suggesting that histone acetylation represents the histone code. The HDAC inhibitors have successfully ameliorated several diseases, including type 1 diabetes (123, 129–132), EAE (74–76), asthma (133), lupus (134, 135), and colitis (136), in animal models, indicating their usefulness to treat a variety of diseases. Accumulating data indicate that histone modifier-mediated hyperacetylation in lymphoid cells and the target tissues is associated with the amelioration of type 1 diabetes (129) and PEAE (74), and selective regulation of genes. It remains to be seen whether the changes in gene expression observed following treatment with HDAC inhibitors have transgenerational consequences.
Conclusion
This review discusses the effects of HDAC inhibitors on EAE regulation (Table 1) and, by extrapolation, their utility in treating MS. Neuroprotection in mice was accompanied by the repression of mostly non-overlapping sets of genes induced by immunization with myelin antigens and a few constitutively expressed genes in the peripheral lymphoid system and the CNS. Notably, TSA administration contrived the expansion of granulocytes and induced T-cell tolerance in the periphery while reducing the influx of immune cells into the CNS (Figure 1). Lessons learned from the EAE models require validation, which may provide impetus to investigate the efficacy of histone modifiers for treating MS variants efficiently. Since HDAC inhibitors such as valproic acid and hydroxamates are currently used in patients for ailments unrelated to MS and are well tolerated, these small-molecule inhibitors may be used for treating MS.
Author contributions
SJ conceived and executed the project and wrote the first draft of the manuscript. AJ conducted most of the experiments reported in this article and edited the manuscript. All authors contributed to the article and approved the submitted version.
Funding
The work described in this article was supported by the University of Illinois at Chicago.
Acknowledgments
We thank all the co-investigators who participated in the study discussed in this article for their contribution.
Conflict of interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
Publisher's note
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References
1. Walton C, King R, Rechtman L, Kaye W, Leray E, Marrie RA, et al. Rising prevalence of multiple sclerosis worldwide: Insights from the Atlas of MS, third edition. Mult Scler. (2020) 26:1816–21. doi: 10.1177/1352458520970841
2. Weissert R. The immune pathogenesis of multiple sclerosis. J Neuroimmune Pharmacol. (2013) 8:857–66. doi: 10.1007/s11481-013-9467-3
3. Dendrou CA, Fugger L, Friese MA. Immunopathology of multiple sclerosis. Nat Rev Immunol. (2015) 15:545–58. doi: 10.1038/nri3871
4. Kearney H, Miller DH, Ciccarelli O. Spinal cord MRI in multiple sclerosis–diagnostic, prognostic and clinical value. Nat Rev Neurol. (2015) 11:327–38. doi: 10.1038/nrneurol.2015.80
5. Cree BA, Reich DE, Khan O, De Jager PL, Nakashima I, Takahashi T, et al. Modification of multiple sclerosis phenotypes by African ancestry at HLA. Arch Neurol. (2009) 66:226–33. doi: 10.1001/archneurol.2008.541
6. Kister I, Bacon T, Cutter GR. How multiple sclerosis symptoms vary by age, sex, and race/ethnicity. Neurol Clin Pract. (2021) 11:335–41. doi: 10.1212/CPJ.0000000000001105
7. Kira J. Neuromyelitis optica and Asian phenotype of multiple sclerosis. Ann N Y Acad Sci. (2008) 1142:58–71. doi: 10.1196/annals.1444.002
8. Papais Alvarenga RM, Araújo ACRAE, Nascimento ACB, Araujo NEC, Meneguette NS, Neri VC, et al. Is Asian type MS an MS phenotype, an NMO spectrum disorder, or a MOG-IgG related disease? Mult Scler Relat Disord. (2020) 42:102082. doi: 10.1016/j.msard.2020.102082
9. Kira J. Autoimmunity in neuromyelitis optica and opticospinal multiple sclerosis: astrocytopathy as a common denominator in demyelinating disorders. J Neurol Sci. (2011) 311:69–77. doi: 10.1016/j.jns.2011.08.043
10. Rosati G. The prevalence of multiple sclerosis in the world: an update. Neurol Sci. (2001) 22:117–39. doi: 10.1007/s100720170011
11. Frischer JM, Bramow S, Dal-Bianco A, Lucchinetti CF, Rauschka H, Schmidbauer M, et al. The relation between inflammation and neurodegeneration in multiple sclerosis brains. Brain. (2009) 132:1175–89. doi: 10.1093/brain/awp070
12. Hollenbach JA, Oksenberg JR. The immunogenetics of multiple sclerosis: a comprehensive review. J Autoimmun. (2015) 64:13–25. doi: 10.1016/j.jaut.2015.06.010
13. Fukazawa T, Kikuchi S, Miyagishi R, Miyazaki Y, Yabe I, Hamada T, et al. HLA-dPB1*0501 is not uniquely associated with opticospinal multiple sclerosis in Japanese patients. Important role of DPB1*0301. Mult Scler. (2006) 12:19–23. doi: 10.1191/135248506ms1252oa
14. Peerlings D, Mimpen M, Damoiseaux J. The IL-2–IL-2 receptor pathway: Key to understanding multiple sclerosis. J Transl Autoimmun. (2021) 4:100123. doi: 10.1016/j.jtauto.2021.100123
15. Rane L, Vudattu N, Bourcier K, Graniar E, Hillert J, Seyfert V, et al. Alternative splicing of interleukin-7 (IL-7) and interleukin-7 receptor alpha (IL-7Ralpha) in peripheral blood from patients with multiple sclerosis (MS). J Neuroimmunol. (2010) 222:82–6. doi: 10.1016/j.jneuroim.2010.02.014
16. Cruciani C, Puthenparampil M, Tomas-Ojer P, Jelcic I, Docampo MJ, Planas R, et al. T-cell specificity influences disease heterogeneity in multiple sclerosis. Neurol Neuroimmunol Neuroinflamm. (2021) 8:e1075. doi: 10.1212/NXI.0000000000001075
17. Bernard CC, Johns TG, Slavin A, Ichikawa M, Ewing C, Liu J, et al. Myelin oligodendrocyte glycoprotein: a novel candidate autoantigen in multiple sclerosis. J Mol Med (Berl). (1997) 75:77–88. doi: 10.1007/s001090050092
18. Moser T, Akgün K, Proschmann U, Sellner J, Ziemssen T. The role of TH17 cells in multiple sclerosis: therapeutic implications. Autoimmun Rev. (2020) 9:102647. doi: 10.1016/j.autrev.2020.102647
19. Ueno H, Banchereau J, Vinuesa CG. Pathophysiology of T follicular helper cells in humans and mice. Nat Immunol. (2015) 16:142–52. doi: 10.1038/ni.3054
20. Holm Hansen R, Talbot J, Højsgaard Chow H, Bredahl Hansen M, Buhelt S, Herich S, et al. Increased intrathecal activity of follicular helper T cells in patients with relapsing-remitting multiple sclerosis. Neurol Neuroimmunol Neuroinflamm. (2022) 9:e200009. doi: 10.1212/NXI.0000000000200009
21. Quinn JL, Kumar G, Agasing A, Ko RM, Axtell RC. Role of TFH cells in promoting T helper 17-induced neuroinflammation. Front Immunol. (2018) 9:382. doi: 10.3389/fimmu.2018.00382
22. Machado-Santos J, Saji E, Tröscher AR, Paunovic M, Liblau R, Gabriely G, et al. The compartmentalized inflammatory response in the multiple sclerosis brain is composed of tissue-resident CD8+ T lymphocytes and B cells (2018). Brain. (2018) 141:2066–82. doi: 10.1093/brain/awy151
23. Pender MP. The essential role of Epstein-Barr virus in the pathogenesis of multiple sclerosis. Neuroscientist. (2011) 17:351–67. doi: 10.1177/1073858410381531
24. Thacker EL, Mirzaei F, Ascherio A. Infectious mononucleosis and risk for multiple sclerosis: a meta-analysis. Ann Neurol. (2006) 59:499–503. doi: 10.1002/ana.20820
25. Farrell RA, Antony D, Wall GR, Clark DA, Fisniku L, Swanton J, et al. Humoral immune response to EBV in multiple sclerosis is associated with disease activity on MRI. Neurology. (2009) 73:32–8. doi: 10.1212/WNL.0b013e3181aa29fe
26. Salehi Z, Beheshti M, Nomanpour B, Khosravani P, Naseri M, Sahraian MA, et al. The association of EBV and HHV-6 viral load with different NK and CD8+ T cell subsets in the acute phase of relapsing-remitting multiple sclerosis. Cell J. (2021) 23:626–32.
27. Wingerchuk DM, Carter JL. Multiple sclerosis: current and emerging disease-modifying therapies and treatment strategies. Mayo Clin Proc. (2014) 89:225–40. doi: 10.1016/j.mayocp.2013.11.002
28. Hauser S. L., Cree B. A. C. (2020). Treatment of multiple sclerosis: a review. Am J Med. 133:1380–90.e2. doi: 10.1016/j.amjmed.2020.05.049
29. The IFNB Multiple Sclerosis Study Group and The University of British Columbia MS/MRI Analysis Group. Interferon beta-1b in the treatment of multiple sclerosis: final outcome of the randomized controlled trial. Neurology. (1995) 45:1277–85. doi: 10.1212/WNL.45.7.1277
30. Calabresi P, Kieseier B, Arnold D, Balcer L, Boyko A, Pelletier J, et al. Clinical efficacy and safety of peginterferon beta-1a in relapsing multiple sclerosis:data from the pivotal phase 3 ADVANCE Study. Neurology. (2013) 80 (Meeting Abstracts 1):S31–006.
31. Axtell RC, Raman C, Steinman L. Type I interferons: beneficial in Th1 and detrimental in Th17 autoimmunity. Clin Rev Allergy Immunol. (2013) 44:114–20. doi: 10.1007/s12016-011-8296-5
32. Dhib-Jalbut S. Mechanisms of action of interferons and glatiramer acetate in multiple sclerosis. Neurology. (2002) 58:S3–9. doi: 10.1212/WNL.58.8_suppl_4.S3
33. Miller A, Kappos L, Comi G, Confavreux C, Freedman M, Olsson T, et al. Teriflunomide efficacy and safety in patients with relapsing multiple sclerosis: results from TOWER, a second, pivotal, phase 3 placebo-controlled study. Neurology. (2013) 80 (Meeting Abstracts 1): S01.004.
34. Coyle PK, Khatri B, Edwards KR, Meca-Lallana JE, Cavalier S, Rufi P, et al. Teriflunomide real-world evidence: global differences in the phase 4 Teri-PRO study. Mult Scler Relat Disord. (2019) 31:157–64. doi: 10.1016/j.msard.2019.03.022
35. van Oosten BW, Killestein J, Barkhof F, Polman CH, Wattjes MP. PML in a patient treated with dimethyl fumarate from a compounding pharmacy [published correction appears in N Engl J Med 368:1950]. N Engl J Med. (2013) (2013) 368:1658–9. doi: 10.1056/NEJMc1215357
36. Wray S, Then Bergh F, Wundes A, Arnold DL, Drulovic J, Jasinska E, et al. Efficacy and safety outcomes with diroximel fumarate after switching from prior therapies or continuing on DRF: results from the phase 3 EVOLVE-MS-1 Study. Adv Ther. (2022) 39:1810–31. doi: 10.1007/s12325-022-02068-7
37. Montalban X, Comi G, O'Connor P, Gold S, de Vera A, Eckert B, et al. Oral fingolimod (FTY720) in relapsing multiple sclerosis: impact on health-related quality of life in a phase II study. Mult Scler. (2011) 17:1341–50. doi: 10.1177/1352458511411061
38. Kappos L, Li DK, Stüve O, Hartung HP, Freedman MS, Hemmer B, et al. Safety and efficacy of siponimod (BAF312) in patients with relapsing-remitting multiple sclerosis: dose-blinded, randomized extension of the Phase 2 BOLD Study. JAMA Neurol. (2016) 73:1089–98. doi: 10.1001/jamaneurol.2016.1451
39. Harris S, Comi G, Cree BA, Arnold DL, Steinman L, Sheffield JK, et al. Plasma neurofilament light chain concentrations as a biomarker of clinical and radiologic outcomes in relapsing multiple sclerosis: post hoc analysis of phase 3 ozanimod trials. Eur J Neurol. (2021) 28:3722–30. doi: 10.1111/ene.15009
40. Kappos L, Fox RJ, Burcklen M, Freedman MS, Havrdová EK, Hennessy B, et al. Ponesimod compared with teriflunomide in patients with relapsing multiple sclerosis in the active-comparator phase 3 OPTIMUM Study: a randomized clinical trial. JAMA Neurol. (2021) 78:558–67. doi: 10.1001/jamaneurol.2021.0405
41. Giovannoni G, Soelberg Sorensen P, Cook S, Rammohan K, Rieckmann P, Comi G, et al. Safety and efficacy of cladribine tablets in patients with relapsing-remitting multiple sclerosis: results from the randomized extension trial of the CLARITY study. Mult Scler. (2018) 24:1594–604. doi: 10.1177/1352458517727603
42. Foo EC, Russell M, Lily O, Ford HL. Mitoxantrone in relapsing-remitting and rapidly progressive multiple sclerosis: ten-year clinical outcomes post-treatment with mitoxantrone. Mult Scler Relat Disord. (2020) 44:102330. doi: 10.1016/j.msard.2020.102330
43. Perumal J, Balabanov R, Su R, Chang R, Balcer L, Galetta S, et al. Natalizumab in early relapsing-remitting multiple sclerosis: a 4-year, open-label study. Adv Ther. (2021) 38:3724–42. doi: 10.1007/s12325-021-01722-w
44. Cohen JA, Coles AJ, Arnold DL, Confavreux C, Fox EJ, Hartung HP, et al. Alemtuzumab versus interferon beta 1a as first-line treatment for patients with relapsing-remitting multiple sclerosis: a randomised controlled phase 3 trial. Lancet. (2012) 380:1819–28. doi: 10.1016/S0140-6736(12)61769-3
45. Baker D, Ali L, Saxena G, Pryce G, Jones M, Schmierer K, et al. The irony of humanization: alemtuzumab, the first, but one of the most immunogenic, humanized monoclonal antibodies. Front Immunol. (2020) 11:124. doi: 10.3389/fimmu.2020.00124
46. Hauser SL, Waubant E, Arnold DL, Vollmer T, Antel J, Fox RJ, et al. B-cell depletion with rituximab in relapsing-remitting multiple sclerosis. N Engl J Med. (2008) 358:676–88. doi: 10.1056/NEJMoa0706383
47. Montalban X, Hauser SL, Kappos L, Arnold DL, Bar-Or A, Comi G, et al. Ocrelizumab versus placebo in primary progressive multiple sclerosis. N Engl J Med. (2017) 376e:209–20. doi: 10.1056/NEJMoa1606468
48. Wolinsky JS, Arnold DL, Brochet B, Hartung HP, Montalban X, Naismith RT, et al. Long-term follow-up from the ORATORIO trial of ocrelizumab for primary progressive multiple sclerosis: a post-hoc analysis from the ongoing open-label extension of the randomised, placebo-controlled, phase 3 trial. Lancet Neurol. (2020) 19:998–1009. doi: 10.1016/S1474-4422(20)30342-2
49. Bergman J, Burman J, Gilthorpe JD, Zetterberg H, Jiltsova E, Bergenheim T, et al. Intrathecal treatment trial of rituximab in progressive MS: an open-label phase 1b study. Neurology. (2018) 91:e1893–901. doi: 10.1212/WNL.0000000000006500
50. Gärtner J, Hauser SL, Bar-Or A, Montalban X, Cohen JA, Cross AH, et al. Efficacy and safety of ofatumumab in recently diagnosed, treatment-naive patients with multiple sclerosis: results from ASCLEPIOS I and II. Mult Scler. (2022) 28:1562–75. doi: 10.1177/13524585221078825
51. Fox E, Lovett-Racke AE, Gormley M, Liu Y, Petracca M, Cocozza S, et al. A phase 2 multicenter study of ublituximab, a novel glycoengineered anti-CD20 monoclonal antibody, in patients with relapsing forms of multiple sclerosis. Mult Scler. (2021) 27:420–9. doi: 10.1177/1352458520918375
52. Faissner S, Gold R. Efficacy and safety of multiple sclerosis drugs approved since 2018 and future developments. CNS Drugs. (2022) 36:803–17. doi: 10.1007/s40263-022-00939-9
53. Segal BM, Constantinescu CS, Raychaudhuri A, Kim L, Fidelus-Gort R, Kasper LH, et al. Repeated subcutaneous injections of IL12/23 p40 neutralising antibody, ustekinumab, in patients with relapsing-remitting multiple sclerosis: a phase II, double-blind, placebo-controlled, randomised, dose-ranging study. Lancet Neurol. (2008) 7:796–804. doi: 10.1016/S1474-4422(08)70173-X
54. Gold R, Giovannoni G, Selmaj K, Havrdova E, Montalban X, Radue EW, et al. Daclizumab high-yield process in relapsing-remitting multiple sclerosis (SELECT): a randomised, double-blind, placebo-controlled trial. Lancet. (2013) 381:2167–75. doi: 10.1016/S0140-6736(12)62190-4
55. Kappos L, Cohan S, Arnold DL, Robinson RR, Holman J, Fam S, et al. Safety and efficacy of daclizumab beta in patients with relapsing multiple sclerosis in a 5-year open-label study (EXTEND): final results following early termination. Ther Adv Neurol Disord. (2021) 14:1756286420987941. doi: 10.1177/1756286420987941
56. Viglietta V, Bourcier K, Buckle GJ, Healy B, Weiner HL, Hafler DA, et al. CTLA4Ig treatment in patients with multiple sclerosis: an open-label, phase 1 clinical trial. Neurology. (2008) 71:917–24. doi: 10.1212/01.wnl.0000325915.00112.61
57. Havrdová E, Belova A, Goloborodko A, Tisserant A, Wright A, Wallstroem E, et al. Activity of secukinumab, an anti-IL-17A antibody, on brain lesions in RRMS: results from a randomized, proof-of-concept study. J Neurol. (2016) 263:1287–95. doi: 10.1007/s00415-016-8128-x
58. Baker D, Pryce G, James LK, Schmierer K, Giovannoni G. Failed B cell survival factor trials support the importance of memory B cells in multiple sclerosis. Eur J Neurol. (2020) 27:221–8. doi: 10.1111/ene.14105
59. Curtin F, Porchet H, Glanzman R, Schneble HM, Vidal V, Audoli-Inthavong ML, et al. A placebo randomized controlled study to test the efficacy and safety of GNbAC1, a monoclonal antibody for the treatment of multiple sclerosis - Rationale and design. Mult Scler Relat Disord. (2016) 9:95–100. doi: 10.1016/j.msard.2016.07.002
60. Gold J, Marta M, Meier UC, Christensen T, Miller D, Altmann D, et al. A phase II baseline versus treatment study to determine the efficacy of raltegravir (Isentress) in preventing progression of relapsing remitting multiple sclerosis as determined by gadolinium-enhanced MRI: the INSPIRE study. Mult Scler Relat Disord. (2018) 24:123–8. doi: 10.1016/j.msard.2018.06.002
61. Rolfes L, Pawlitzki M, Pfeuffer S, Huntemann N, Wiendl H, Ruck T, et al. Failed, interrupted, or inconclusive trials on immunomodulatory treatment strategies in multiple sclerosis: update 2015-2020. BioDrugs. (2020) 34:587–610. doi: 10.1007/s40259-020-00435-w
62. Krämer J, Wiendl H. What have failed, interrupted, and withdrawn antibody therapies in multiple sclerosis taught us? Neurotherapeutics. (2022) 19:785–807. doi: 10.1007/s13311-022-01246-3
63. Lutterotti A, Martin R. Antigen-specific tolerization approaches in multiple sclerosis. Expert Opin Investig Drugs. (2014) 23:9–20. doi: 10.1517/13543784.2014.844788
64. Steinman L. The re-emergence of antigen-specific tolerance as a potential therapy for MS. Mult Scler. (2015) 21:1223–38. doi: 10.1177/1352458515581441
65. Procaccini C, Rosa D, Pucino V, Formisano V, Matarese LG. Animal models of multiple sclerosis. Eur J Pharmacol. (2015) 759:182–91. doi: 10.1016/j.ejphar.2015.03.042
66. Stromnes IM, Cerretti LM, Liggitt D, Harris RA, Goverman JM. Differential regulation of central nervous system autoimmunity by T(H)1 and T(H)17 cells. Nat Med. (2008) 14:337–42. doi: 10.1038/nm1715
67. Pierson ER, Wagner C, Goverman JM. The contribution of neutrophils to CNS autoimmunity. Clin Immunol. (2018) 189:23–8. doi: 10.1016/j.clim.2016.06.017
68. Abromson-Leeman S, Bronson R, Luo Y, Berman M, Leeman R, Leeman J, et al. T-cell properties determine disease site, clinical presentation, and cellular pathology of experimental autoimmune encephalomyelitis. Am J Pathol. (2004) 165:1519–33. doi: 10.1016/S0002-9440(10)63410-4
69. Shetty A, Gupta SG, Varrin-Doyer M, Weber MS, Prod'Homme T, Molnarfi N, et al. Immunodominant T-cell epitopes of MOG reside in its transmembrane and cytoplasmic domains in EAE. Neurol Neuroimmunol Neuroinflamm. (2014) 1:e22. doi: 10.1212/NXI.0000000000000022
70. Slavin A, Ewing C, Liu J, Ichikawa M, Slavin J, Bernard CC, et al. Induction of a multiple sclerosis-like disease in mice with an immunodominant epitope of myelin oligodendrocyte glycoprotein. Autoimmunity. (1998) 28:109–20. doi: 10.3109/08916939809003872
71. Hidaka Y, Inaba Y, Matsuda K, Itoh M, Kaneyama T, Nakazawa Y, et al. Cytokine production profiles in chronic relapsing-remitting experimental autoimmune encephalomyelitis: IFN-γ and TNF-α are important participants in the first attack but not in the relapse. J Neurol Sci. (2014) 340:117–22. doi: 10.1016/j.jns.2014.02.039
72. Baker D, Nutma E, O'Shea H, Cooke A, Orian JM, Amor S, et al. Autoimmune encephalomyelitis in NOD mice is not initially a progressive multiple sclerosis model. Ann Clin Transl Neurol. (2019) 6:1362–72. doi: 10.1002/acn3.792
73. Basso AS, Frenkel D, Quintana FJ, Costa-Pinto FA, Petrovic-Stojkovic S, Puckett L, et al. Reversal of axonal loss and disability in a mouse model of progressive multiple sclerosis. J Clin Invest. (2008) 118:1532–43. doi: 10.1172/JCI33464
74. Jayaraman A, Soni A, Prabhakar BS, Holterman M, Jayaraman S. The epigenetic drug trichostatin A ameliorates experimental autoimmune encephalomyelitis via T cell tolerance induction and impaired influx of T cells into the spinal cord. Neurobiol Dis. (2017) 108:1–12. doi: 10.1016/j.nbd.2017.07.015
75. Jayaraman A, Sharma M, Prabhakar B, Holterman M, Jayaraman S. Amelioration of progressive autoimmune encephalomyelitis by epigenetic regulation involves selective repression of mature neutrophils during the preclinical phase. Exp Neurol. (2018) 304:14–20. doi: 10.1016/j.expneurol.2018.02.008
76. Jayaraman A, Avgush K, Kulam R, Soni A, Khan A, Kerdjoudj M, et al. Treatment of autoimmune encephalomyelitis with a histone deacetylase inhibitor. Analyzing the role of immune-response genes. Free Neuropathol. (2020) 1:19. doi: 10.17879/freeneuropathology-2020-2819
77. Buonvicino D, Ranieri G, Chiarugi A. Treatment with non-specific HDAC inhibitors administered after disease onset does not delay evolution in a mouse model of progressive multiple sclerosis. Neuroscience. (2021) 465:38–45. doi: 10.1016/j.neuroscience.2021.04.002
78. Camelo S, Iglesias AH, Hwang D, Due B, Ryu H, Smith K, et al. Transcriptional therapy with the histone deacetylase inhibitor trichostatin A ameliorates experimental autoimmune encephalomyelitis. J Neuroimmunol. (2005) 164:10–21. doi: 10.1016/j.jneuroim.2005.02.022
79. Ge Z, Da Y, Xue Z, Zhang K, Zhuang H, Peng M, et al. Vorinostat, a histone deacetylase inhibitor, suppresses dendritic cell function and ameliorates experimental autoimmune encephalomyelitis. Exp Neurol. (2012) 241:56–66. doi: 10.1016/j.expneurol.2012.12.006
80. Lv J, Du C, Wei W, Wu Z, Zhao G, Li Z, et al. The antiepileptic drug valproic acid restores T cell homeostasis and ameliorates pathogenesis of experimental autoimmune encephalomyelitis. J Biol Chem. (2012) 287:28656–65. doi: 10.1074/jbc.M112.356584
81. Makino S, Kunimoto K, Muraoka Y, Mizushima Y, Katagiri K, Tochino Y, et al. Breeding of a non-obese, diabetic strain of mice. Jikken Dobutsu. (1980) 29:1–13. doi: 10.1538/expanim1978.29.1_1
82. Damotte D, Colomb E, Cailleau C, Brousse N, Charreire J, Carnaud C, et al. Analysis of susceptibility of NOD mice to spontaneous and experimentally induced thyroiditis. Eur J Immunol. (1997) 27:2854–62. doi: 10.1002/eji.1830271117
83. Noorchashm H, Noorchashm N, Kern J, Rostami SY, Barker CF, Naji A, et al. B-cells are required for the initiation of insulitis and sialitis in nonobese diabetic mice. Diabetes. (1997) 46:941–6. doi: 10.2337/diab.46.6.941
84. Abughanam G, Maria OM, Tran SD. Studying Sjögren's syndrome in mice: What is the best available model? J Oral Biol Craniofac Res. (2021) 11:245–55. doi: 10.1016/j.jobcr.2020.12.001
85. Pryce G, O'Neill JK, Croxford JL, Amor S, Hankey DJ, East E, et al. Autoimmune tolerance eliminates relapses but fails to halt progression in a model of multiple sclerosis. J Neuroimmunol. (2005) 165:41–52. doi: 10.1016/j.jneuroim.2005.04.009
86. Zhang Z, Zhang ZY, Wu Y, Schluesener HJ. Valproic acid ameliorates inflammation in experimental autoimmune encephalomyelitis rats. Neuroscience. (2012) 221:140–50. doi: 10.1016/j.neuroscience.2012.07.013
87. Castelo-Branco G, Stridh P, Guerreiro-Cacais AO, Adzemovic MZ, Falcão AM, Marta M, et al. Acute treatment with valproic acid and l-thyroxine ameliorates clinical signs of experimental autoimmune encephalomyelitis and prevents brain pathology in DA rats. Neurobiol Dis. (2014) 71:220–33. doi: 10.1016/j.nbd.2014.08.019
88. Lipton HL, Twaddle G, Jelachich ML. The predominant virus antigen burden is present in macrophages in Theiler's murine encephalomyelitis virus induced demyelinating disease. J Virol. (1995) 69:2525–33. doi: 10.1128/jvi.69.4.2525-2533.1995
89. Tsunoda I, Kuang L-Q, Libbey JE, Fujinami RS. Axonal injury heralds virus-induced demyelination. Am J Pathol. (2003) 162:1259–69. doi: 10.1016/S0002-9440(10)63922-3
90. Tsunoda I, Tanaka T, Saijoh Y, Robert SF. Targeting inflammatory demyelinating lesions to sites of Wallerian degeneration. Am J Pathol. (2007) 171:1563–75. doi: 10.2353/ajpath.2007.070147
91. Mix E, Meyer-Rienecker H, Hartung HP, Zettl UK. Animal models of multiple sclerosis—potentials and limitations. Prog Neurobiol. (2010) 92:386–404. doi: 10.1016/j.pneurobio.2010.06.005
92. Pachner AR. The neuroimmunology of multiple sclerosis: fictions and facts. Front Neurol. (2022) 12:796378. doi: 10.3389/fneur.2021.796378
93. Villoslada P, Steinman L. New targets and therapeutics for neuroprotection, remyelination and repair in multiple sclerosis. Expert Opin Investig Drugs. (2020) 29:443–59. doi: 10.1080/13543784.2020.1757647
94. International Multiple Sclerosis Genetics Consortium, Wellcome Trust Case Control Consortium 2. Genetic risk and a primary role for cell-mediated immune mechanisms in multiple sclerosis. Nature. (2011) 476:214–19. doi: 10.1038/nature10251
95. Koch MW, Metz LM, Kovalchuk O. Epigenetic changes in patients with multiple sclerosis. Nat Rev Neurol. (2013) 9:35–43. doi: 10.1038/nrneurol.2012.226
96. Allis CD, Jenuwein T. The molecular hallmarks of epigenetic control. Nat Rev Genet. (2016) 17:487–500. doi: 10.1038/nrg.2016.59
97. Ebers GC, Bulman DE, Sadovnick AD, Paty DW, Warren S, Hader W, et al. A population-based study of multiple sclerosis in twins. N Engl J Med. (1986) 315:1638–42. doi: 10.1056/NEJM198612253152603
98. Iridoy Zulet M, Pulido Fontes L, Ayuso Blanco T, Lacruz Bescos F, Mendioroz Iriarte M. Epigenetic changes in neurology: DNA methylation in multiple sclerosis. Neurologia. (2017) 32:463–8. doi: 10.1016/j.nrleng.2015.03.020
99. Mycko MP, Baranzini SE. microRNA and exosome profiling in multiple sclerosis. Mult Scler. (2020) 26:599–604. doi: 10.1177/1352458519879303
100. Jayaraman A, Zhao T, Jayaraman S. Histone modifier differentially regulates gene expression and unravels survival role of microRNA-494 in leukemic T cell. Microrna. (2021) 10:39–50. doi: 10.2174/2211536610666210412153322
101. Marks PA, Richon VM, Breslow R, Rifkind RA. Histone deacetylase inhibitors as new cancer drugs. Curr Opin Oncol. (2001) 13:477–83. doi: 10.1097/00001622-200111000-00010
102. Ruijter D, van Gennip AJ, Caron AH, Kemp HN, van Kuilenburg SAB. Histone deacetylases (HDACs): characterization of the classical HDAC family. Biochem J. (2003) 370:737–49. doi: 10.1042/bj20021321
103. Croft M, Duncan DD, Swain SL. Response of naive antigen-specific CD4+ T cells in vitro: characteristics and antigen-presenting cell requirements. J Exp Med. (1992) 176:1431–7. doi: 10.1084/jem.176.5.1431
104. Schwartz RH, Mueller DL, Jenkins MK, Quill H. T-cell clonal anergy. Cold Spring Harb Symp Quant Biol. (1989) 54:605–10. doi: 10.1101/SQB.1989.054.01.072
105. Rumble JM, Huber AK, Krishnamoorthy G, Srinivasan A, Giles DA, Zhang X, et al. Neutrophil-related factors as biomarkers in EAE and MS. J Exp Med. (2015) 212:23–35. doi: 10.1084/jem.20141015
106. Casserly CS, Nantes JC, Whittaker Hawkins RF, Vallières L. Neutrophil perversion in demyelinating autoimmune diseases: Mechanisms to medicine. Autoimmun Rev. (2017) 16:294–307. doi: 10.1016/j.autrev.2017.01.013
107. Nakahama T, Hanieh H, Nguyen NT, Chinen I, Ripley B, Millrine D, et al. Aryl hydrocarbon receptor-mediated induction of the microRNA-132/212 cluster promotes interleukin-17-producing T-helper cell differentiation. Proc Natl Acad Sci USA. (2013) 110:11964–9. doi: 10.1073/pnas.1311087110
108. Tran EH, Hardin-Pouzet H, Verge G, Owens T. Astrocytes and microglia express inducible nitric oxide synthase in mice with experimental allergic encephalomyelitis. J Neuroimmunol. (1997) 74:121–9. doi: 10.1016/S0165-5728(96)00215-9
109. Goldberg R, Wildbaum G, Zohar Y, Maor G, Karin N. Suppression of ongoing experimental autoimmune encephalomyelitis by neutralizing the function of the p28 subunit of IL-27. J Immunol. (2004) 173:6465–71. doi: 10.4049/jimmunol.173.10.6465
110. Powell ND, Papenfuss TL, McClain MA, Gienapp IE, Shawler TM, Satoskar AR, et al. Cutting edge: macrophage migration inhibitory factor is necessary for progression of experimental autoimmune encephalomyelitis. J Immunol. (2005) 175:5611–4. doi: 10.4049/jimmunol.175.9.5611
111. McQualter JL, Darwiche R, Ewing C, Onuki M, Kay TW, Hamilton JA, et al. Granulocyte macrophage colony-stimulating factor: a new putative therapeutic target in multiple sclerosis. J Exp Med. (2001) 194:873–82. doi: 10.1084/jem.194.7.873
112. Moreno M, Bannerman P, Ma J, Guo F, Miers L, Soulika AM, et al. Conditional ablation of astroglial CCL2 suppresses CNS accumulation of M1 macrophages and preserves axons in mice with MOG peptide EAE. J Neurosci. (2014) 34:8175–85. doi: 10.1523/JNEUROSCI.1137-14.2014
113. O'Connor RA, Cambrook H, Huettner K, Anderton SM. T-bet is essential for Th1-mediated, but not Th17-mediated, CNS autoimmune disease. Eur J Immunol. (2013) 43:2818–23. doi: 10.1002/eji.201343689
114. Martinez NE, Sato F, Omura S, Kawai E, Takahashi S, Yoh K, et al. RORγt, but not T-bet, overexpression exacerbates an autoimmune model for multiple sclerosis. J Neuroimmunol. (2014) 276:142–9. doi: 10.1016/j.jneuroim.2014.09.006
115. Hori S, Nomura T, Sakaguchi S. Control of regulatory T cell development by the transcription factor Foxp3. Science. (2003) 299:1057–61. doi: 10.1126/science.1079490
116. Danikowski KM, Jayaraman S, Prabhakar B. Regulatory T cells in multiple sclerosis and myasthenia gravis. J Neuroinflammation. (2017) 14:117. doi: 10.1186/s12974-017-0892-8
117. Jayaraman S, Prabhakar B. Immune tolerance in autoimmune central nervous system disorders. In: Mitoma H, Manto M, editor. Neuroimmune diseases; from cells to the living brain. Berlin: Springer (2019). p. 143–66. doi: 10.1007/978-3-030-19515-1_5
118. Bradner JE, West N, Grachan ML, Greenberg EF, Haggarty SJ, Warnow T, et al. Chemical phylogenetics of histone deacetylases. Nat Chem Biol. (2010) 6:238–43. doi: 10.1038/nchembio.313
119. Broide RS, Redwine JM, Aftahi N, Young W, Bloom FE, Winrow CJ, et al. Distribution of histone deacetylases 1-11 in the rat brain. J Mol Neurosci. (2007) 31:47–58. doi: 10.1007/BF02686117
120. Sun L, Telles E, Karl M, Cheng F, Luetteke N, Sotomayor EM, et al. Loss of HDAC11 ameliorates clinical symptoms in a multiple sclerosis mouse model. Life Sci Alliance. (2018) 1:e201800039. doi: 10.26508/lsa.201800039
121. Zhang F, Shi Y, Wang L, Sriram S. Role of HDAC3 on p53 expression and apoptosis in T cells of patients with multiple sclerosis. PLoS ONE. (2011) 6:e16795. doi: 10.1371/journal.pone.0016795
122. Tegla CA, Azimzadeh P, Andrian-Albescu M, Martin A, Cudrici CD, Trippe R, et al. SIRT1 is decreased during relapses in patients with multiple sclerosis. Exp Mol Pathol. (2014) 96:139–48. doi: 10.1016/j.yexmp.2013.12.010
123. Jayaraman A, Arianas M, Jayaraman S. Epigenetic modulation of selected immune response genes and altered functions of T lymphocytes and macrophages collectively contribute to autoimmune diabetes protection. BBA Adv. (2021) 1:100031. doi: 10.1016/j.bbadva.2021.100031
124. Konsoula Z, Barile FA. Epigenetic histone acetylation and deacetylation mechanisms in experimental models of neurodegenerative disorders. J Pharmacol Toxicol Methods. (2012) 66:215–20. doi: 10.1016/j.vascn.2012.08.001
125. Narita T, Weinert BT, Choudhary C. Functions and mechanisms of non-histone protein acetylation. Nat Rev Mol Cell Biol. (2019) 20:156–74. doi: 10.1038/s41580-018-0081-3
126. Fitz-James MH, Cavalli G. Molecular mechanisms of transgenerational epigenetic inheritance. Nat Rev Genet. (2022) 23:325–41. doi: 10.1038/s41576-021-00438-5
127. Turner BM. Histone acetylation and an epigenetic code. Bioessays. (2000) 22:836–45. doi: 10.1002/1521-1878(200009)22:9<836::AID-BIES9>3.0.CO;2-X
128. Robles P, Turner A, Zuco G, Adams S, Paganopolou P, Winton M, et al. Parental energy-sensing pathways control intergenerational offspring sex determination in the nematode Auanema freiburgensis. BMC Biol. (2021) 19:102. doi: 10.1186/s12915-021-01032-1
129. Patel T, Patel V, Singh R, Jayaraman S. Chromatin remodeling resets the immune system to protect against autoimmune diabetes in mice. Immunol Cell Biol. (2011) 89:640–9. doi: 10.1038/icb.2010.144
130. Jayaraman S, Patel A, Jayaraman A, Patel V, Holterman M, Prabhakar B, et al. Transcriptome analysis of epigenetically modulated genome indicates signature genes in manifestation of type 1 diabetes and its prevention in NOD mice. PLoS ONE. (2013) 8:e55074. doi: 10.1371/journal.pone.0055074
131. Jayaraman S. Type 1 diabetes and epigenetics. In: Zhang R, editor. The epigenetics of autoimmunity. Translational Epigenetics Series, Volume 5. Amsterdam: Elsevier Inc. (2018), p. 188–205. doi: 10.1016/B978-0-12-809912-4.00009-X
132. Patel V, Jayaraman A, Jayaraman S. Epigenetic reprogramming ameliorates type 1 diabetes by decreasing the generation of Th1 and Th17 subsets and restoring self-tolerance in CD4+ T cells. Int Immunopharmacol. (2022) 103:108490. doi: 10.1016/j.intimp.2021.108490
133. Choi J-H, Oh S-W, Kang M-S, Kwon HJ, Oh G-T, Kim D-Y, et al. Trichostatin A attenuates airway inflammation in mouse asthma model. Clin Exp Allergy. (2005) 35:89–96. doi: 10.1111/j.1365-2222.2004.02006.x
134. Mishra N, Reilly CM, Brown DR, Ruiz P, Gilkeson GS. Histone deacetylase inhibitors modulate renal disease in the MRL-lpr/lpr mouse. J Clin Invest111. (2003) 539–52. doi: 10.1172/JCI16153
135. Reilly CM, Thomas M, Gogal Jr R, Olgun S, Santo A, Sodhi R, et al. The histone deacetylase inhibitor trichostatin A upregulates regulatory T cells and modulates autoimmunity in NZB/W F1 mice. J Autoimmun. (2008) 31:123–30. doi: 10.1016/j.jaut.2008.04.020
Keywords: central nervous system, epigenetics, experimental autoimmune encephalomyelitis, histone deacetylases, multiple sclerosis, myelin oligodendrocyte glycoprotein, neutrophils, T helper cells, tolerance
Citation: Jayaraman S and Jayaraman A (2022) Impact of histone modifier-induced protection against autoimmune encephalomyelitis on multiple sclerosis treatment. Front. Neurol. 13:980758. doi: 10.3389/fneur.2022.980758
Received: 28 June 2022; Accepted: 09 September 2022;
Published: 14 October 2022.
Edited by:
Robert Weissert, University of Regensburg, GermanyReviewed by:
Jinming Han, Capital Medical University, ChinaGamaleldin Harisa, King Saud University, Saudi Arabia
Copyright © 2022 Jayaraman and Jayaraman. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Sundararajan Jayaraman, YW51ZTI0NjgmI3gwMDA0MDt1aWMuZWR1