Introduction
Multiple sclerosis (MS) is a
chronic inflammatory demyelinating disease of the central nervous system
(CNS)
that
afflicts approximately 2.5 million people worldwide(Baecher-Allan,
Kaskow & Weiner, 2018; Goverman, 2009). MS lesions can emerge in the
whole CNS and are most easily observed in the white matter as focal
regions of inflammation and demyelination(Reich, Lucchinetti &
Calabresi, 2018). The well-defined cause of MS still remains elusive
now, although genetic susceptibility and environmental factors have been
associated with the MS occurrence(Ascherio, 2013; Dendrou, Fugger &
Friese, 2015a). MS is the leading cause of disability in young adults,
resulting in serious socioeconomic consequences for the individual
patient and for society (approximately $10 billion in United
States)(Adelman, Rane & Villa, 2013; Holland, Schneider, Rapp & Kalb,
2011; Jennum, Wanscher, Frederiksen & Kjellberg, 2012). Therefore,
development of innovative therapeutic is an urgent need to improve the
clinical benefit and life quality for MS patients.
MS is widely considered to be an inflammatory T-cell-mediated autoimmune
disease of CNS(Ridderstad Wollberg et al., 2014). In clinical MS and EAE
model, immune system T leukocytes transmigrate from the bloodstream into
the CNS to assault the myelin sheath surrounding nerve fibers(Zozulya &
Wiendl, 2008). Autoreactive T cells indiscriminately recognize CNS
antigens play a critical pathogenic role in the induction of CNS
inflammation and demyelination, and thereby in the progression of
neurological function deficit and physical disability(T Hart &
Massacesi, 2009).
In
clinical, increased population of activated T cells against the myelin
were observed in MS patients compared to healthy human. Autoreactive
CD4+ T cells were considered as the key driver events
in EAE models, indicating important immunopathologic features of
MS(Dendrou, Fugger & Friese, 2015a). Currently, increasing animal
experiment results and clinical trials data of MS demonstrate that
CD8+ T-cells may also contribute to inflammation
sustaining and tissue destruction in MS(Friese & Fugger, 2005; Lassmann
& Bradl, 2017). Collectively, T lymphocytes have emerged as important
players in the pathogenic mechanism of CNS autoimmune inflammation,
representing a promising therapeutic target of MS.
Human Dihydroorotate dehydrogenase
(DHODH) is a rate-limiting enzyme in the de novo pyrimidine
biosynthesis pathway. DHODH binds to flavin mononucleotide (FMN)
cofactor in conjunction with ubiquinone to catalyze the oxidation of
dihydroorotate to orotate(Löffler, Jöckel & Schuster, 1997;
Munier-Lehmann, Vidalain, Tangy & Janin, 2013; Sykes et al., 2016).
Human DHODH is located in the intracellular mitochondrial inner membrane
and
thereby
can be categorized into class 2 DHODH family. As an essential protein
that catalyzes the conversion of DHO to orotate, DHODH is a critical
player in the pyrimidine de novo biosynthesis of DNA and RNA. In
high proliferative cells, such as activated T lymphocytes, increasedde novo pyrimidine biosynthesis can confer their superior growth
capacity(Quemeneur, Gerland, Flacher, Ffrench, Revillard & Genestier,
2003). Blockade of de novo pyrimidine biosynthesis by
pharmacological targeting of DHODH has been considered as a promising
therapeutic strategy for autoimmune diseases through inhibition of T
lymphocytes activation. Teriflunomide (A77 1726) have been approved for
the treatment of MS through blocking pyrimidine de novobiosynthesis at the level of DHODH(Dimitrova, Skapenko, Herrmann,
Schleyerbach, Kalden & Schulze-Koops, 2002; Ruckemann et al., 1998).
While, its observed hepatotoxicity in clinical and potential
teratogenicity in preclinical model limit its benefit.
In the present study, we
identified piperine, a main bioactive constituent
of
traditional medicine black pepper, as a natural inhibitor of human DHODH
using a panel of biochemical and biophysical methods, including
enzymatic assay, isothermal titration calorimetry and X-ray crystal
diffraction. We further demonstrated that DHODH inhibition by piperine
ameliorated
MOG-induced
EAE mouse model accompanied with reduced inflammation as well as
lessened myelin and BBB destruction. Taken together, our data revealed
that piperine targets T cells in EAE by inhibition of DHODH and provided
a potential treatment strategy for MS patients.