- Molecular MoA: o Bioactivation: Fingolimod requires bioactivation for activity. Phosphorylation of a

hydroxyl group establishes Fingolimod-Phosphate as a Sphingosine-1-Phosphate mimic. It therefore can be recognized by the sphingosine phosphate (SP) receptor.

§ SP Receptors are 7-transmembrane GPCR with an extra N-terminal loop. Sphingolamide-Phosphate, the natural endogenous agonist, causes functional antagonism through negative feedback. Receptor is internalized, slowing/inhibiting Leukocyte migration.

o Receptor Agonism: Fingolimod-Phosphate is an agonist at S1P1/3/4/5 but not S1P2. § S1P1: Fingolimod-Phosphate agonism of this receptor, similar to Shingolamide-Phosphate,

causes functional antagonism and internalization of the receptor. This inhibits leukocyte egress and protects the CNS from accumulation and further attack. Also minor cytoskeleton disruption.

• These receptors are located in the lymph nodes. § S1P3: Agonism at the Type 3 Receptor disrupts the cytoskeleton, inducing

cardiac side effects like Bradycardia. This is regarded as toxicity § S1P5: Agonism at the Type 5 Receptor elicits neuroprotective properties

- Fingolimod-mods: The next generation of optimized natural products. Cardiotoxicity in the form of prolonged QT, is suspected to be due in part by Fingolimod action at S1P3 receptors. Recent molecular adaptations have been made to alter affinity for these off-target sites at the 1,2 diol.

o Ozanimod (Celgene): S1PR1 > S1PR3 x10,000 – Showing improved cardiac safety o Ponesimod (Actelion): S1PR1 > S1PR3 x650 o Siponimod (trials): S1PR1 > S1PR3 x10,000

Teriflunomide: A oral disease modifying agents (DMARD) - MoA: Teriflunomide is essentially 3 molecules in 1, existing as tautomers through stereoisomeric relationships. It

inhibits dihydroorotate dehydrogenase, thus preventing de novo pyrimidine synthesis in T and B cells. As a result, Teriflunomide is antiproliferative and produces an anti-inflammatory effect.

o Teriflunomide is more effective against activated lymphocytes - Fun Fact: Teriflunomide is the active metabolite of leflunomide

(11/28) Federle Lecture: Medchem and Pharmacology of DMARDs in Rheumatology Perpetuation of an Autoimmune Disease: Rheumatic diseases like Rheumatic arthritis are often trapped in a damaging loop of autoimmune reactions. Initiated by an environmental component, such as injury or infection, stimulation of the immune system may propel an individual into this vicious cycle. There may be genetic factors at play, imparting a greater tendency leaning towards the pro-inflammatory side of things. Tissue damage will re-stimulate T cells, developing auto-aggressive T cells, and more cytokines

- The goal of treatment is to break this cycle. Targets of the Cycle: Mast cells release inflammatory mediators, in the form of:

- Membraned-Derived Lipid Mediators: Lipid metabolism, over the course of minutes, will release Prostaglandins, such as Leukotrienes, Platelet Activation Factor

o à Intervention: Cox Inhibitors - Cytokine Production: Over the course of hours, new mRNA and protein synthesis of: IL-1,3,4,5,6,8, TNF

o à Intervention: DMARDS, Synthetic (MTX, LFN, SSZ, HCQ) and Biological (mAb) - Re-tipping the Balance: Cytokines play a major role in RA, and adjusting pro/anti-inflammatory balance is the

basis for biological therapeutics o Methods:

§ (1) Neutralize Cytokines § (2) Receptor Blockade § (3) Induction of Anti-Inflammatory pathways

o Pro-Inflammatory Cytokines: Primarily TNF and IL-1b § Want to INHBIIT these

o Auto-Inflammatory Cytokines: IL10 and TGFb § Want to PROMOTE these

Combinatorial therapy is the best way to treat RA, and the base of this therapy is MTX.

Methotrexate (MTX): Folic acid analog. Most important anti-rheumatic drug, the cornerstone of combinatorial therapy

- MoA: MTX enters cell via the Reduced Folate Carrier (RFC) and is poly-glutamated by Folylpolyglutamate Hydrolase (FPGH) to MTX-PG. Polyglutamation stabilizes MTX allowing it to stay within the cell. MTX then exhibits its action by inhibiting 3 separate enzymes:

o ATIC: Increases free adenosine (Adenosine component considered to be the major source of efficacy) § Adenosine is a potent anti-inflammatory molecule, therefore, MTX inhibits inflammation and

induces vasodilation, promoting the transition from neutrophil-mediated inflammation to a more efficient and highly specific dendritic cell-mediated response.

§ à Suppresses pro-inflammatory mediators TNF, IFG, IL2,6,8, NO. § à Enhances production of anti-inflammatory mediators, IL10, IL4, IL1

o TYMS: Decreases pyrimidine synthesis § Downregulation of Thymidylate Synthetase (TS) disrupts the production of pyrimidine

nucleotides, thereby preventing proliferation of anti-inflammatory cells. (Because the fastest replicating cells are those of the immune system)

o DHFR: Inhibits transmethylation reaction § Dihydrofolate Reductase (DHFR) is responsible for methylating RNA and DNA, thus, its

inhibition leads to further disruption of DNA synthesis and proliferation - Folate Supplementation: Folate Deficiency is a risk factor of MTX toxicity, must be added to therapy to help

reduce severity of side effects. Dose on a day not taking MTX. - Pharmacokinetics: Must monitor kidney function to determine if it can be used.

o Half-Life: MTX = 6 hours but MTX-PG = 60 hours o Elimination: 50-100% excreted in the urine, except for MTX-PG

- AE: N/V/D, Alopecia Women, Pregnancy? à BAD. Men, Fertility? à BAD. - Efficacy: MTX alone rarely induces remissions of RA – though it is the cornerstone of Combination DMARDs

o No trial has ever suggested that any other synthetic DMARD is superior to MTX Hydroxychloroquine (Plaquenil, HCQ) - An antimalarial. +Chloroquine (CQ)

- MoA: Slightly basic molecule that raises the pH in phagosomes, decreasing the efficiency of antigen loading on MHC. Less antigen presentation = Less inflammation

o Photoprotective effect: Additionally, it blocks the arachidonic acid pathway by inhibiting phospholipase A2/C reducing [prostaglandins] and lipid peroxidation

- Place in Therapy: Due to their limited ability to prevent joint damage on their own, reserved for only very mild or non-erosive RA disease. Often combined with MTX for additive benefits for S/Sx as part of triple therapy (Sulfasalazine/MTX/HCQ)

- AE: Relatively safe and well-tolerated. The half-life is 40-50 days, and most is excreted unchanged, however, it may distribute, accumulate, and crystallize in the Skin and Retina (melanin-containing cells)

o Retinal Toxicity: Have eyes examined at least once a year, even more frequently if elderly. Sulfasalazine (Azulfidine, SSZ) Prodrug, metabolized by intestinal microbial to Mesalamine + Sulfapyridine

MoA: Its mechanism has not been fully elucidated, but relates to its metabolites - Anti-Inflammatory: Exhibited by Mesalamine (5-ASA analog)

§ Inhibits pro-inflammatory effects of Arachidonic Acid cascade § Inhibits ATIC and DHFR, increasing Adenosine (like MTX!) § Downregulates Neutrophil activation, chemotaxis and migration

- Immunomodulatory: Represses cell division of T, B, and NK cells § Decreasing Ig and Autoantibody (RF) production

PK: The antibiotic component, Sulfapyridine, is extensively metabolized in the Liver & excreted via Urine as glucuronide conjugates. (Consider Liver and Renal function) Toxicity: GI discomfort and D early in use. Category B/C, safe in preggers.

Leflunomide (Arava, LFN) Prodrug, bioactivated to its active metabolite A77-1726 - MoA: Active metabolite (A77-1726) inhibits Dihydroorotate dehydrogenase

(DHODH), which reduces production of Orotate used for nucleotide synthesis. This decreases pyrimidine biosynthesis, thus blocking mitogen-stimulated activation of T cells.

- Toxicity: GI (N/D, Dyspepsia), HTÝ, CholesterolÝ, Hepatotoxic, Pregnancy Category X

Glucocorticoids (GC): GC are among the most effective anti-inflammatory and immunosuppressive therapeutic options, and have thus become a cornerstone of therapy for many rheumatic disorders – but check place-in-therapy.

- Place in Therapy: Used as adjunctive therapy in combination with other DMARDs. RA is the only disease in which GC therapy is often started and maintained at a low dose as additional therapy. Highly effective,<10mg/day

- Joint-Sparing Effect: Theorized to be based on the inhibition of pro-inflammatory cytokines IL-1 and TNF TNFa: Major player in pro-inflammatory response, induces more cytokine production, expression of adhesion molecules

- Forms: The two forms of TNFa are related by TACE, the TNFa Converting Enzyme o Membrane-Bound: TNFa (26kDa) o Soluble TNFa (17kDa)

- Receptors: Though similar, the two TNFa receptors differ in their intracellular domain and signaling pathway o TNFR-1: Constitutively expressed in all tissues, capable of inducing apoptosis via Death Domain.

§ Internalized TNF recruits TRADD, leading to DNA degradation and cell death. o TNFR-2: Only expressed on Immune cells, capable of inducing NFkB signaling pathways

§ Activation recruits TRAF2, activating NFkBà Production of cytokines, inhibition of apoptosis § Produces: pro-inflammatory TNFa and IL-1b, Upregulation of Adhesion molecules (ICAMs)

- Targets for Inhibition of TNFa: TNFa molecule, TNFR, TACE, Signaling pathway, NFkB Infliximab (Remicade): Biological DMARD - Chimeric (25% murine, 75% human) mAb

- MoA: “Neutralize Cytokines”. Bind and neutralize both Soluble and Membrane-bound TNFa - AE: Infusion Rxn (itch, flush, N), HA, Abdominal pain.

o Increased risk of infection due to immunosuppression. (URTI – Tuberculosis) o Increased risk of non-Hodgkins lymphoma o Immunogenicity: Patient develops HAMA (human anti-nouse Ab) towards Infliximab. Thus, we efficacy

has a limited timeline, which can be extended by concurrent immunosuppressive therapy. Tocilizumab (Actemra): Biological DMARD – Humanized mAb

- First anti-IL6R mAb approved for arthritis, used for moderate to severe RA for those who have failed an anti-TNF. IL-6 is a chemical messenger involved in destructive immune responses, found overproduced in the joints

- MoA: “Receptor Blockade”, targets the IL-6 receptor Abatacept (Orencia): Biological DMARD – Fusion Protein Checkpoint Inhibitor

- MoA: Modified IgG domain recognizes CD80/86, binds and prevents CD28 interaction, producing negative feedback to inhibit T cell activation and inhibiting inflammatory cytokine production

Janus Kinase Inhibitors: The first oral DMARD (2012) – newest drugs to combat RA - Drug: Tofacitinib (Xeljanz) - MoA: Jakinhibs interrupt signaling downstream of a multiplicity of cytokines by inhibiting cytokine receptors,

rather than blocking one cytokine at a time. AE: Increased serious infections (Infection-risk) (12/1) Chevalier Lecture: Rheumatoid Arthritis Epidemiology: Affecting 1-2% of people world-wide, this disease affects 3x as many women as men and is one of the most common causes of disability. It has been associated with decreased life expectancy

- Risk Factors: Smoking (x3, and worse prognosis!), Genetic Susceptibility (mom/dad? +6%), Gender specific factors (hormones?), Occupational exposure, infectious triggers, presence of autoantibodies (RF, anti-CCP)

Pathophysiology: Autoimmune disease characterized by an imbalance of pro-/anti-inflammatory cytokines. Patients present with chronic inflammation and proliferation of synovial tissue (Synovitis), which can invade the cartilage and bone producing erosions ~ breakdown of bone in the joint space. Once damaged, it is irreversible Clinical Presentation: RA is usually a disease of the small joints

- Symmetrical Joint Swelling (Synovitis) - Morning Stiffness lasting ³ 1 hour - Low-grade fever, weight loss, fatigue, weakness, loss of

appetite - Joint pain, tenderness, and muscle aches - In the hands, the Proximal interphalangeal and

Metacarpophalangeal joints are most commonly affected - Late Complications: Ulnar Deviation, Boutonniere

deformities, and Muscular atrophy. Bony erosions. Significant difficulty performing Activities of Daily Living (ADLs)