Sjogren’s Syndrome: An Unmet Therapeutic Need Robert I. Fox, M.D., Ph.D. Scripps Memorial Hospital Scripps/XiMED Medical Center La Jolla, California

All slides are available on my website robertfoxmd.com 2

A new Approach to Therapy is Necessary in Sjogren’s Syndrome – Why? At present, we have unhappy patients and frustrated physicians. The symptoms of ocular/oral pain and fatigue rarely correlate with either our objective exam or our acute phase reactants. After years of failure of trials of biologics, it is time for a new approach.

Goals-1 What are current issues in diagnosis? 1.Is it SS or is it SLE? 1.Do we use traditional criteria (American- European) or the new SICCA criteria? 1.How does SS overlap with the newly described IgG4-related disease?

Specific Goals-2 How can we convey information about treatment of dry eyes/dry mouth to our patients in the limited time allowed in a patient revisit? This is a practical limitation that is very important aspect of patient care. Will deal with this problem in afternoon session.

Goals-3 Update on Pathogenesis including Genetics and Cytokines 1.Acquired immune system: HLA-DR and autoantibody 2.Innate immune system: type 1 IFN signature and IFN-g/IL-17 But we are still missing how the immune process influences the midbrain, cortical and hypothalamic axis aspects of SS.

Goals-4 1.Why have trials at systemic therapies (i.e., rituximab, belimumab) and other biologic agents failed? They have not improved fatigue! 1.New targets must include the immune- neural targets, including the influence of cytokines on micro-glial cells and their transmitters.

Background-1 SS has “benign” and “systemic” manifestations. FDA licensing will require improvement in both manifestations.

Background-2 Benign manifestations include: Dry and painful eyes Dry and painful mouth Myalgias, fatigue Impaired cognition (executive function)— trying to distinguish “fibromyalgia” from “depression”

Patients would: Equate SS with impact similar to moderate angina. Trade 2 years of “life expectancy” to not have SS symptoms.

SS Related Health Care Costs- 1: Dry or painful eyes are now most common cause of visits to Ophthalmologists in Japan. Lost productivity (over $160 billion/year just for dry eyes (especially in computer users where decreased blink rate is 90%.

Direct healthcare costs in Great Britain (NHS) are second only to RA, and exceed SLE. RA £2693 (not including TNFs) pSS £2188 (not including OTC cost of artificial tears or dental costs) Age Matched NHS Controls £849 SS-Related Health Care Costs-2:

SS-Related Health Care Costs-3: Despite these costs of health care, patient and physician dissatisfaction with clinical outcomes is higher in pSS than in SLE or RA.

Classification-1 Is Sjogren’s just SLE with 4/5 Criteria? Different antibody profile (anti- SSA/B) are not criteria for SLE; SS is more organ specific – (salivary/lacrimal gland) and more lymphoproliferative.

Classification-2 Why is Sjogren’s not just SLE with 4/5 Criteria? 1.Interstitial pneumonitis (not pleurisy), interstitial nephritis (not glomerulonephritis) 2.Higher frequency of lymphoma 3.Genome Screens support this with Homing receptors found in SS but not SLE (CXCR5)

Classification-3 What is the relationship of IgG4-related Disease and SS? Mikulicz described salivary gland swelling and lymphoid infiltrate. Initial reports by Mikulicz were TBC or lymphoma. Today, pathology of Mikulicz Disease is most frequently part of IgG4-related disease. Female-Male incidence (9:1 in SS)

Mikulicz (part of IgG4 RD) For many years, Mikulicz was considered the characteristic biopsy of SS. Now, we recognize that they are different processes. Confusion still exists in salivary gland and in extraglandular tissues (i.e., AIP, interstitial pneumonitis).

IgG4-Related Disease Different pattern on biopsy from SS. IgG4 plasma cells plus “storiform” fibrosis. Think of IgG4 RD as a plasma cell dyscrasia where B-cells release particular cytokines. The overlap of cytokines with SS may be confusing.

History and Classification-3 Old vs. New SS Criteria 1984: San Diego Classification (Fox) to include objective criteria for clinical trials 1998: American/European Consensus Criteria (Vitali et al.) included ocular and oral symptoms (validated in Europe) and lab studies (minor salivary gland biopsy or SS-A)

Most important “child” of AECC was the ESSDAI (European Sjogren’s Syndrome Activity Index) which includes 16 domains of activity score including glandular and extraglandular activity The ESSDAI ranges from Practically the range is 1-41 A clinically meaningful change is 3.5 units Constitutional Skin Lung, Renal Neurologic

ESSDAI- European SS Activity Index Weighted domains to give a total score— the Sjogren’s equivalent to ACR-50 for RA. The validated ESSDAI activity score has been the accepted outcome measure of FDA clinical trials.

Example of Part of the ESSDAI Scoring

Although the Consensus AECC was used for over over 20 years in over 1300 publications and 50 clinical trials approved by FDA and EMA … the AECC were never “formally” submitted to ACR for US validation and formal ACR acceptance since everyone seemed comfortable using these criteria. Further, formal ACR validation would require an additional study in a US cohort.

History and Classification-4 SICCA Preliminary Criteria (Shiboski, 2012) Requires 2 out of 3 criteria: 1. Positive Anti-SS A/B or ANA >320; 2. Ocular Staining Score >3; 3. Positive labial gland biopsy: focus score >1.

There is about an 82% overlap between the SICCA criteria and the AECC criteria

Does it matter? Our outcome measures ESSDAI may not be true in new SICCA system, so there is no way to judge study outcomes. Literature search and prognosis are all based on old AECC criteria. The 5 published studies comparing both systems indicate IT DOES make a difference.

Problems with New SICCA Classification-1 ACR tentatively accepting these, pending “validation” on independent cohort. Independent validation has not been performed and the grant denied. At least 5 published studies have shown important differences in the patients identified by AECC and SICCA. Although 82% similar, the discordant group greatly affect the outcome in clinical trials.

Other Problems with new SICCA Classification Few Rheumatologists do SG biopsies. Few Ophthalmologists perform the ocular exam as specified by SICCA (because it is time-consuming and much harder than a simple Lissamine green). Committees in process of a new “combined” criteria.

Pathogenesis-1 Concordance of SS among identical twins only about 20% Thus, genetic sequence is not enough and over 80% is epigenetic— environmental factor or gene regulation. Distinct histone acetylation pattern upstream of key genes.

Pathogenesis-2 Large sequences of untranslated mRNA. Novel miRNA, some with sequence similar to EBV fragments. Genetics in GWAS recently published and only SS (not SLE) has homing receptor (CXCR5) as a strong “hit.”

To briefly summarize PATHOGENESIS … Acquired Immune System-- HLA DR and Associated T-cell directed B- cell antibodies; IFN-g and IL-17 pathways Innate immune system— Type I IFN signature NK like cells link acquired and innate

T ime course of autoimmune response* 1. Genetic factors predispose to Sjogren’s. 2. Environmental factors such as a viral infection may lead to formation of autoantibodies. 3. Antibodies precede disease (however, presence of antibody does not necessarily mean disease). Genetic Factors (including sex) (HLA-DR) Genetic Factors (including sex) (HLA-DR) Genetic Factors (including sex) (HLA-DR) Genetic Factors (including sex) (HLA-DR) Auto- antibodies Acquired Immune system (HLA-DR) T/B-cells Disease Manifestations * Time period of years Innate Immune system (Toll receptor) Genetic Factors (including sex) (HLA-DR) Environmental Factor (virus-such as EBV ) (apoptotic fragment) Type I IFN Immune complex

Gene expression profiling of minor salivary glands clearly distinguishes primary Sjögren's syndrome patients from healthy control subjects The gene signature is type I IFN Arthritis & Rheumatism Volume 52, Issue 5, pages , 5 MAY 2005 DOI: /art Volume 52, Issue 5,

The main cytokine targets match those identified in genome wide screens* HLA-DR (T-cell), CTLA and IFN-  NF-K /IkB Homing receptor (CXCR5) Type I IFN –IRF5, STAT4, TLR3/7/9 and pkR (cytoplasmic sensor) B-cell activation –BLK, BAFF, IL12, and A20 (TNFAIP3) * Most of these targets do not map to the encoded protein but to upstream sites of RNA transcription that are not translated (presumed epigenetic sites such as methylation)

Pathogenesis-3 Genome-Wide Association Parallels-- our studies on cytokine profiles Strongest is HLA-DR and acquired immune system leading to T-cell/B-cell production of autoantibody. Next strongest are Innate Immune markers associated with Type 1 IFN production. As noted above, also find a homing receptor (CXCR5) which goes with the tissue-specific homing receptors. In SS patients with lymphoma, find A20 (member of TNF superfamily) that suppresses NFK-b and B-cell proliferation.

The main cytokine targets match those identified in genome wide screens* HLA-DR (T-cell), CTLA and IFN-  NF-K /IkB Homing receptor (CXCR5) Type I IFN –IRF5, STAT4, TLR3/7/9 and pkR (cytoplasmic sensor) B-cell activation –BLK, BAFF, IL12, and A20 (TNFAIP3) * Most of these targets do not map to the encoded protein but to upstream sites of RNA transcription that are not translated (presumed epigenetic sites such as methylation)

Distinct SS Gland and Blood Methylation Maps

If we know the genes and the cytokines, why have we made so little progress with biologics in Sjogren’s Syndrome? a) Although we can improve the extraglandular manifestations (rash, cytopenias, pneumonitis), none have fulfilled the requirements where patients actually feel better. b) The most frequently used biologic in SS remains rituximab—even though it failed its randomized trials.

Previously Studied in SS Anti-CD20 –glandular, extraglandular and fatigue BAFF (Blys)-ACR 2012 abstracts* Abatacept (CD40 L)-ACR 2012 Allogeneic mesenchymal cells- ACR 2012 abstracts and article in Blood

Other Inhibitors of IFN a. Initial trials of anti-type 1 IFN had infusion reactions and only modest efficacy. b. Medi 546 (type 1 IFN-R antagonists) now in phase 1 (scleroderma) and juvenile SLE phase 2 trial.

But we are still missing key targets in the pathogenesis of fatigue and the adrenal-hypothalmic axis. In both SS and SLE, we can lower the cytokine with biologics, but the patient still feels little improvement. This will be the focus of future direction for therapy.

The functional Circuit To understand “benign symptoms” and develop better therapies, we must review the concept of the functional circuit in SS. This “circuit” starts with unmyelinated nerves at the mucosal surface. These nerves travel to the “midbrain” (the salvitory and lacrimatory nuclei of cranial nerve V) Have reflections to the cortical surface that sense pain or dryness.

Background-3 The Functional Circuit in SS 1. Mucosal Surface (inflammatory cytokines and metalloproteinase) 1. Mucosal Surface (inflammatory cytokines and metalloproteinase) 2. Midbrain Vth Nucleus (lymphocytes and glial cells) 2. Midbrain Vth Nucleus (lymphocytes and glial cells) 4. Gland (lymphs, cytokines, metalloproteinase) 4. Gland (lymphs, cytokines, metalloproteinase) 3. Vascular (iNOS, CAMs, Chemokines) 3. Vascular (iNOS, CAMs, Chemokines) Brain Cortex Nociception (pain) glial cells and corticcal neurons Brain Cortex Nociception (pain) glial cells and corticcal neurons These sites and their cytokines correlate with systemic manifestations We must understand these sites to treat “benign” symptoms.

Recent studies in multiple sclerosis and animal models of “flu” Have led to at least 5 approved therapies; Have highlighted the importance of cytokines on microglial cells; Have identified new targets including mTor, AKT, IKDelta, and Sphingosine (the fingolimod receptor).

Does this apply to Sjogren’s syndrome? Patients with early SS had corneal pain that decreased completely with topical anesthesia*. Patients with chronic SS showed only a partial (30% decrease) in eye pain after topical anesthetic* Functional MRI (fMRI) showed nocioceptive pattern— called phantom pain amplification *Rosenthal et al

Neuro-plasticity is a new concept for most rheumatologists A stimulus that is not painful (innocuous) in a normal model may be painful in its “autoimmune” congenic. Events in the CNS are not detected in the peripheral blood. Models such as MS model or murine flu.

Thrombospondin (-/-) Mouse at 24 wks. Where a trivial stimuli Causes pain response Wild type A pain stimuli that is innocuous in Wild Type does cause nociceptive pain in tsp (-/-) mouse model. The Pain Threshold is Lowered in the Tsp (-/-) mouse.

To study the mechanism of neurogenic or nociceptive pain we must use animal model-1 The thrombospondin (-/-) mouse (TSP null) or the TGF-  receptor mutation both develop SS like disease. The mouse develops both oral and ocular lesions. The mouse develops ANA and SS-A antibodies.

To study the mechanism of neurogenic or nociceptive pa2n we must use animal model-2 Thrombospondin is a matrix protein that plays a role in activation of latent TGF-  Activated TGF-  promotes Treg and inhibits Th-17 (IFN-  Thus, TSP (null) has high levels of Th-17, IL- 17 and IFN- 

Thrombospondin (-/-) mouse model of SS 4 wks. Lacrimal gland biopsies The mouse has ANA+, SS-A+ TSP null can not activate TGF-  In absence TGF-  continuous Th-17 TGF-  and cytokine activation stimulates mTor/AKT WT Tsp-/- 24 wks

At the level of the Vth nerve (Tsp -/- mouse) Microglial cells translate inflammatory signals that go to nociceptive cortex WT TSP (-/-) mTor and AKT activated in response to “lower stimuli” in the tsp (-/-) mouse

The tsp-null mouse allows us to look at the interaction of peripheral inflammation and microglial cells-1 Activation of microglial cells through mTor/AKT. In absence of thrombospondin, constitutive activation of Th17 and IFN-  activates microglial cells.

Take home lesson for Rheumatologists: There is more to pain in SS than peripheral inflammation. Nociceptive (pain) pathway occurs through smad3 and non-smad pathways that involve mTor/AKT pathways in cranial nerve V.

Emotional stressors potential the role of cytokines in pain pathways Emotional Physiological Similar pattern of Fos-ir in cortical neurons in response to distinct stressors

Summary-1 1.Functional circuit needs to be considered when assessing “benign” symptoms of corneal or oral pain. 2.Symptoms of oral/ocular pain do not correlate with markers of systemic inflammation (ESR/CRP) because the events are contained within the brainstem and cortex.

Summary-2 3. Afferents go to midbrain regions of Cranial Vth. 4.Microglial cells are site of cytokine/neurokine interaction 5. Receptors and neurokines from microglial cells are therapeutic targets.

Summary-3 6. Novel targets include mTor and AKT pathways 7. These mTor/AKT pathways also implicated in chronic pain and depression—so we must collaborate with these neurochemists.

Summary-4 8. Cortical “memory” of nociceptive pain is well described in neurologic literature. 9. fMRI indicates that nociceptive pain is the cause of benign symptoms in SS that do not correlate with acute phase reactants

Moulton et*. Al used fMRI in SS patients with chronic ocular pain using fMRI of nociceptive pain have been studied Cortical regions that activate with ocular pain signal at “benign stimuli levels” occur only in chronic SS patients with severe pain *Moulton EA, Becerra L, Rosenthal P, Borsook D. An Approach to Localizing Corneal Pain Representation in Human Primary Somatosensory Cortex. PloS one 2012;7:e44643.

Summary We have made advances in “systemic inflammation” and these are encouraging. 11. For “drug licensing” we will also need to improve the patients “quality of life” symptoms of dryness, pain and fatigue 12. We need for “autoimmune” divisions to work with “neuro-chemistry” research divisions.

Thank you for inviting me to participate in this wonderful meeting. We will discuss treatment of dryness of eyes and mouth during afternoon session.

Clinical Key Points : Dry Mouth

Dry and Painful Mouth-1 If you thought that Dentists did not care about SS, then wait until you see their Dental Care Plans -- The answer to all problems is a $25,000 tooth implant.

Dry and Painful Mouth-2 Must treat underlying oral candida (which is erythematous spots on roof of mouth) before anything will work. Candida often lurks under dentures– Patients would rather run naked through clinic than remove a denture.

Dry and Painful Mouth-3 Angular cheilitis the most obvious hint. Treatment of oral candida is a slow process involving multiple steps. Use website for education.

High Risk of Lymphoma

EYE DRYNESS results in the clinical appearance of keratoconjunctivitis sicca (KCS) characteristic of Sjogren’s Syndrome The upper lid literally sticks to the Epithelial surface and pulls surface mucin layers off. The Rose Bengal dye retention test is like “rain water pooling in a street pothole” This test can be done at bedside and allows “triage” and rapid referral of patients to Ophthalmology

Rash distinct from SLE (erythema annulare)

Arthritis distinct from RA

We are also looking at Additional Targets of Interests Chemokines and their receptors (CCR) on vascular cells and lymphocytes TLR receptors: SLAC-15 that links Toll receptor and type 1 IFN Methylation modulators and siRNA Neural mediator circuits: Receptors on cornea--substance P (TRPV1), VIP and CGRP pain receptors TRPM8, TRPA1, and CGRP in trigeminal ganglion neurons Trigeminal ganglion neurons- MCP-1, MIP-2, CCR and CCL at the blood brain barrier

CCR and Blood Brain Barrier

The tsp-null mouse allows us to look at the interaction of peripheral inflammation and microglial cells Activation of microglial cells through mTor/AKT In absence of thrombospondin, constitutive activation of Th17 and IFN-  activates microglial cells Nociceptive (pain) pathway occurs through smad3 and non-smad pathways that involve mTor/AKT pathways in cranial nerve V

The main cytokine targets match those identified in genome wide screens* HLA-DR (T-cell), CTLA and IFN-  NF-K /IkB Homing receptor (CXCR5) Type I IFN –IRF5, STAT4, TLR3/7/9 and pkR (cytoplasmic sensor) B-cell activation –BLK, BAFF, IL12, and A20 (TNFAIP3) * Most of these targets do not map to the encoded protein but to upstream sites of RNA transcription that are not translated (presumed epigenetic sites such as methylation)

Previously Studied in SS Anti-CD20 –glandular and extraglandular BAFF (Blys)-ACR 2012 abstracts* Abatacept (CD40 L)-ACR 2012 Allogeneic mesenchymal cells-ACR 2012 abstracts and article in Blood &st=nocache&actn=search&dt=12/24/2012%202:29:59%20P

Other Inhibitors of IFN a. Initial trials of anti-type 1 IFN had infusion reactions and only modest efficacy b. Medi 546 (type 1 IFN-R antagonists) now in phase 1 (scleroderma) and juvenile SLE phase 2 trial

Dry and Painful Mouth-4 Different toothpastes and mouth rinses (again, use websites for education). Avoid mouth breathing by keeping sinuses open. Use cool water humidifiers, air purifiers at bedside at night.