Pharmaceutical Market Europe • January 2022 • 26-27
AUTOIMMUNE AND CHRONIC INFLAMMATORY DISEASES
Advances in DHODH inhibition: better activity, better selectivity, better safety
Dihydroorotate dehydrogenase selectively targets cells with high metabolic activity in multiple autoimmune and chronic inflammatory diseases
By Hella Kohlhof and Evelyn Peelen
Dihydroorotate dehydrogenase (DHODH) inhibitors are currently in development for different indications including cancer, autoimmune diseases and even COVID-19. How can one target be important for so many different diseases and on the other hand harbour this natural selectivity?
DHODH was discovered in 1953 and its relevance in autoimmune diseases was first described many years later. The only DHODH inhibitors approved to date are leflunomide (Arava) for the treatment of rheumatoid arthritis and psoriatic arthritis in 1998 and teriflunomide (Aubagio), its active metabolite, for the treatment of multiple sclerosis (MS) in 2012. Given the validated mechanism of inhibiting DHODH, several companies followed this track, providing DHODH inhibitors with different chemical structures, aiming for better activity, better selectivity and a better safety profile.
Currently, only one new DHODH inhibitor (vidofludimus calcium) is in clinical development for the treatment of autoimmune diseases such as ulcerative colitis and multiple sclerosis (Table 1 – see page 28). All other DHODH inhibitors in development, including Brequinar, farudostat, AG-636, JNJ-7485665, orludostat, are in development to target cancer – mainly leukaemia and lymphoma – or are being tested in smaller COVID-19 trials, including emvodostat, Brequinar or RP7214 (Table 1 – see page 28). One exception is PP-001 which is in development for non-infectious uveitis and keratoconjunctivitis (Table 1 – see page 28).
Along with the ongoing clinical trials, DHODH is expected to be a valuable target in other autoimmune/inflammatory diseases. Preclinical work indicates that DHODH inhibition might be beneficial in the treatment of diseases such as diabetes, acute allograft rejection, graft versus host disease, Guillain-Barré syndrome (GBS), myasthenia gravis and systemic lupus erythematosus. In addition to autoimmune diseases and cancer, DHODH inhibitors seem to have a broad-spectrum antiviral activity. Plasmodial and fungal DHODH is also used as a drug target for treating malaria or fungal infection.
‘DHODH inhibitors are currently in development for different indications including cancer, autoimmune diseases and even COVID-19, and
are also expected to be a valuable target in other autoimmune/ inflammatory diseases’
DHODH inhibition: only targeting cells with a high metabolic active status
The mechanism of inhibiting human DHODH is based on the high metabolic active status of the cell, which is induced upon a ‘disease trigger’.
These cells have a high nucleotide demand for proliferation, migration, production of pro-inflammatory cytokines or the viral replication and formation of virus particles. Here, DHODH plays a crucial role as it is the rate-limiting enzyme in the de novo pyrimidine synthesis pathway, which offers a cell the possibility of quickly reacting to an increased need for nucleotides, within minutes or hours.
For a long time, it was thought that the only outcome of DHODH inhibition would be the depletion of nucleotides for DNA replication and, therefore, inhibition of proliferation of cancer or immune cells. Nowadays, this picture is outdated. Further investigation revealed that inhibition of DHODH has a much bigger impact on mRNA production than on DNA production, as metabolically active cells produce high amounts of mRNA in order to produce functional signalling molecules, receptors or enzymes. For this mRNA production, up to 100-fold higher amounts of nucleotides are needed, compared to just doubling the DNA in case of a proliferating cell. Of course, these activated cells need to proliferate as well. For example, an effector immune cell producing high amounts of cytokines only impacts the immune response if a full armada of effector cells is active. Therefore, proliferation inhibition of immune cells is often used as a read-out in cellular test systems to describe the activity of a DHODH inhibitor, even though this is more a marker and not the real reason for the anti-inflammatory or other outcome.
Inhibition of DHODH has even greater capabilities than just direct depletion of nucleotides. Its inhibition impacts the production of phospholipids, the glycosylation of certain proteins, by O-linked N-acetylglucosaminylation (O-GlcNAc), oxidative phosphorylation and glycolysis, as well as mRNA translation. Interestingly, the inhibition of the latter leads via nucleotide stress to the upregulation of Stress Protein 1 (SP1) and subsequently induction of hexamethylene-bis-acetamide (HMBA)-inducible protein 1 (HEXIM1). HEXIM1 binding to the transcription complex, containing the positive transcription elongation factor b (P-TEFb), finally leads to sequestration of the transcriptional machinery and stopping of the transcriptional elongation of certain pro-tumorigenic genes, while stabilising others, for example, anti-tumoral genes. These findings demonstrate an important role for DHODH and HEXIM1 in coupling nucleotide metabolism with transcriptional regulation (Figure 1).
Figure 1. Targeting DHODH leads to metabolic stress in metabolically active cells
Broad applicability of DHODH inhibition with multiple sclerosis of special interest
Although DHODH inhibition appears to be multipotent, multiple sclerosis (MS) seems to be one of the main autoimmune diseases of interest. Successful MS trials (phase 2 and 3) showed a reduction in Expanded Disability Status Scale (EDSS) progression and serum neurofilament light chain levels (a biomarker for axonal damage) as well as slowing of brain atrophy. This indicates, in addition to the well-described anti-inflammatory properties of DHODH inhibitors on peripheral immune cells, an additional neuroprotective effect.
This appears to be supported by preclinical studies showing that DHODH inhibition also targets central nervous system-resident cells, preventing neurodegeneration. Effects have been observed for DHODH inhibition on inflamed rat primary microglia or co-cultures of HIV-vector infected monocytes with microglia. It has been observed that DHODH inhibition of microglia:
- Reduces proliferation
- Increases IL-10 expression
- Reduces reactivation of T cells
- Decreases pro-inflammatory chemokine expression.
In addition to these in vitro models, there are also in vivo models, showing reduced microglia inflammation or a reduction in number of microglia. However, in these studies it is difficult to elucidate a possible direct effect, since inflammation, infiltration of peripheral immune cells and pro-inflammatory microglia responses cannot be uncoupled. Furthermore, DHODH presence and high enzymatic activity has been shown in rat motor neurons.
DHODH inhibition in peripheral nerves has been shown to restore mitochondrial dynamics upon oxidative stress, which reduces axonal damage. Oxidative stress has been implicated in many neurodegenerative diseases such as cerebral ischaemia, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis (ALS) and MS. The effect of DHODH inhibition on brain resident cells hints at an additional application for DHODH inhibition for neuroprotection in neurodegenerative diseases.
‘Inhibition of DHODH has a much bigger impact on mRNA production than on DNA production, as metabolically active cells produce high amounts of mRNA in order to produce functional signalling molecules, receptors or enzymes’
Another positive impact of DHODH inhibitors for the treatment of autoimmune diseases is the described antiviral effect. Literature implicates a role of viral infections in pathogeneses or disease progression of autoimmune diseases. For MS especially, Epstein-Barr virus (EBV) infection or reactivation has been implicated in disease pathogenesis and disease progression, respectively. Preclinical and clinical data shows that DHODH inhibitors prevent the reactivation of EBV.
Another rare but detrimental virus for MS patients and patients with other autoimmune diseases under immunosuppressive treatment is the John Cunningham virus (JCV). Reactivation of JCV has the potential to induce progressive multifocal leukoencephalopathy (PML), which can be fatal. In vitro assays indicted that DHODH inhibition can prevent this reactivation and subsequently the risk for PML as well.
The de novo pyrimidine pathway and, therefore, the rate-limiting enzyme, DHODH, is only important in cells that have a high demand for nucleotides and other macromolecules such as phospholipids and glycosylated proteins. Since this demand is mainly associated with a disease trigger, such as malignant transformation in cancer, highly active immune cells in autoimmune diseases or virus replication during virus infections, targeting DHODH may be a successful strategy to treat a variety of diseases. As the level of nucleotide demand is positively associated with the trigger strength, natural selectivity and, therefore, good safety is a given for highly specific DHODH inhibitors. For the first time, advances in DHODH inhibition enable small molecule oral medications with few, if any, side effects for treating autoimmune and chronic inflammatory diseases.
Hella Kohlhof is Chief Scientific Officer Immunic Therapeutics
Evelyn Peelen is Senior Manager Translational Pharmacology at Immunic Therapeutics