Seropositive test, When to treat? Prof. Khaled Al-Jarallah Consultant Internist, Rheumatologist FRCPC, FACR, FACP, FRCP
Disclosures I have no conflict of interest to declare
References Clinical Reasoning Series , Rahul patwari, 2017. • Medow MA, Lucey CR.BMJ Evidence-Based Medicine. A • qualitative approach to Bayes theorem. 2011;16:163-167. Schur Peter, Laboratory testing for Diagnosis, Management • of Patients with Rheumatic Disease. The Rheumatologist, Dec 1,2014.
Referral Scenarios • 48-year-old female referred from polyclinic with 2 weeks history of generalized body ache & +RF 1:80 R/O Rheumatoid arthritis? • Asymptomatic 25-year-old female, referred from polyclinic after employment medical check-up with lab tests showing ANA 1: 640 • R/O SLE ?
Lab test ❑ For informing us of an emerging disease ❑ Diagnose a specific disease ❑ Predict prognosis ❑ Biomarker of disease ❑ Monitor of disease activity
Why was the test requested? !!
Good referral ❑ 34-year-old female ❑ 12 week history of fatigue, joints pain&swelling in both hands .
Sensitivity, Specificity Autoantibody positivity alone does not make a diagnosis Similarly, antibody negativity does not exclude a diagnosis ❑ If highly sensitive, diagnosis can be excluded in case of negativity ❑ If test is highly specific, diagnosis confirmed in case of positivity
Pre and Post test probability Birtane M. et al 2017 The likelihood nomogram used in SLE with an antinuclear antibody test
Categorical probabilities A qualitative approach to Bayes theorem Categorical probability Numerical probability Very unlikely Less likely than 10% Unlikely Between 10% and 33% Uncertain Between 34% and 66% Likely Between 67% and 90% Very likely More likely than 90% Catherine R Lucey, Medow MA, Lucey CR.BMJ Evidence-Based Medicine 2011;16:163-167
Clinical reasoning approach • Construct patient script (data gathering) • Construct disease script (disease population studies) • Match= treat • Mismatch= rule-out • Uncertain= test
Clinical reasoning approach Clinical Reasoning , Rahul patwari, 2017
Catherine R Lucey, Medow MA, Lucey CR.BMJ Evidence-Based Medicine 2011;16:163-167
Clinical Reasoning , Rahul patwari, 2017
Clinical Reasoning , Rahul patwari, 2017
Clinical Reasoning , Rahul patwari, 2017
Lupus in Arab world - disease script Adwan M, Arch Rheumatol 2018;33(4);455-463
Lupus in Arab world – disease script Percentage Ribonucleoprotein Smith ANA ds DNA SSA /Ro Lupus Anti SSB/La RF aCL total Autoantibody Frequency of autoantibodies in Arab patients with systemic lupus erythematosus Adwan M, Arch Rheumatol 2018;33(4);455-463
Percentage Frequency of systemic lupus erythematosus manifestations in Arab world. Arthralgia/arthritis Lupus in Arab world – disease script Anemia Fatigue Malar rash Renal Hemolytic anemia Photosensitivity Fever Alopecia Manifestation Leucopenia Serositis Oral ulcers Neuropsychiatric Thrombocytopenia Adwan M, Arch Rheumatol 2018;33(4);455-463 Lymphadenopathy Pulmonary Cardiac APS Discoid rash Raynaud’s phenomenon Vasculitis Thrombosis Myositis
Traditional approach to lab study ❑ Why was the test requested? Diagnostic ❑ What was the lab test method? Methodology ❑ What was the lab results? Error in interpretation
Critical interpretation of serology test ❑ Sensitivity (proportion of patients with the target disorder who have positive test) ❑ Specificity (proportion of patients who are free of the target disorder who have negative or normal test) ❑ Positive and negative predictive value (based on pretest probability) ❑ Look for high Sensitive & high Specific diagnostic test!
History- Progress in Rheumatology Serology ❑ 1940s (RF and LE cell perp) ❑ 1950s (ANA and anti- DNA) ❑ 1960s (Sm, RNP, Ro, La) ❑ 1982 (ANCA) ❑ 1990s (Anti-CCP)
Hospitals covered by FOM immunology lab • Mubarak Hospital • Amiri Hospital • Jaber Hospital • Adan Hospital • Ahmadi Hospital • Military Hospital
Current methods used in FOM immunology Lab ❑ CRP : Nephelometry ❑ RF : Nephelometry ❑ Anti CCP 1, 2 : Enzyme-linked immune sorbent assay (ELISA) ❑ ANA : Immunofluorescence (IF) ❑ Anti dsDNA : ELISA ❑ ENA: Immunoblot Assays ❑ ANCA : Immunofluorescence (IF) ; MPO: ELISA, PR3: ELISA ❑ Complements(C3,C4) : Nephelometry
Rheumatoid Factor (RF) ❑ What is it? RA and related diseases causes the production of globulin known as RF which is an autoantibody directed against the Fc portion of IgG, can be IgM , IgA, IgG, IgE , IgD ❑ That antibody binds to normal circulating IgG, forming immune complexes that are deposited in the joints which leads to inflammation of the joints
Rheumatoid Factor (RF) ❑ Positive in ~ 70% RA patients ❑ High RF indicator of worse prognosis. Also show aggressive, erosive joint disease, rheumatoid nodules, extra articular involvement ❑ Positive also in Sjogren’s syndrome, SLE, cryoglobulinemia, interstitial fibrosis, malignancy, various infectious disease ❑ Low titrations seen in 5% healthy population ❑ RF not used for monitoring treatment response and disease
Principle of the RF test ❑ RF anti-antibody can be detected in the laboratory by its ability to bind and form clumps with latex particles or red blood cells (Rose-Waaler test) that contain human Immunoglobulin G (IgG). ❑ If the RF is present in the patient’s blood it attaches to the IgG coating the latex particles causing clumps. ❑ Agglutination is considered a positive reaction that indicates the presence of rheumatoid factor at a detectable level. Shows if test is Positive or negative
Principle of the RF test ❑ Nephelometry technique is used in clinical laboratories for qualitative assessment of RF. It is relatively easily automated. ❑ In nephelometry, levels of several blood plasma proteins is made by measuring the light passed through the sample. IMMage Machine, used in FOM immunology lab for Nephylometry.
Anti-Cyclic citrullinated peptide antibodies (anti-CCP) ❑ Anti-CCP are autoantibodies produced by the immune system that are directed against cyclic citrullinated peptides (CCP).Changes happen in structure of CCP which make them a target for IgG antibodies in RA ❑ Detected using ELISA ❑ Anti CCP has higher specificity than RF ❑ Anti-CCP1(96% specificity, 53% sensitivity for RA); Anti-CCP2(99% specificity, 61.6% sensitivity for RA) ❑ Occurs years before development of clinical symptoms of RA; Associated with aggressive and erosive disease
Anti-CCP • Early assays (ELISA) had sensitivity of 67% and specificity of 95% (compared to 69% and 85% for RF). Later generation assays are even better. • However no test is perfect • False positives seen in active TB, Sjogren’s, SLE, scleroderma, and poly-and dermatomyositis. • In these cases, however, titers are lower than those seen in RA
True Positive or False Positive
RF & Anti-CCP in Rheumatic & Other Diseases Schur P H, The Rheumatologists, 2014
Autoantibodies to nuclear antigens ❑ Antibodies developing against DNA, RNA, histones, centromeres, nucleolus and other nucleoproteins in cell nucleus ❑ High sensitivity, low specificity ❑ High titration does not correlate with disease activity or severity, so not used for monitoring disease activity
Antinuclear Antibodies (ANA)
Sensitivity of ANA in Autoimmune and Non Rheumatic Disease Schur P H, The Rheumatologist,2014
Autoantibodies to nuclear antigens ❑ ANA measured in 2 ways ❑ Generic ANA measurement completed with Immunofluorescence (IF) and Enzyme-linked immune sorbent assay (ELISA) ❑ If ANA positive, specific antibodies detected with automated methods ❑ ANA staining patterns has been recognized to have a low sensitivity and specificity for different autoimmune disorders.
Common immunofluorescence antinuclear antibodies associated with specific diseases
ANA Disease Associations Sensitivity Specificity of Antinuclear antibodies Schur P H, The Rheumatologist, 2014
Anti-dsDNA antibodies • Anti-dsDNA antibodies are used in the evaluating and managing patients with SLE. • Anti-dsDNA antibodies are of primary importance in the pathogenesis and disease activity i.e. lupus nephritis
Anti-dsDNA antibodies Disease associations • Anti-dsDNA antibodies was reported in patients with other disorders, including rheumatoid arthritis, Sjögren's syndrome, scleroderma, overlap connective tissue disease, myositis, uveitis, juvenile arthritis, antiphospholipid syndrome, Grave's disease, autoimmune hepatitis, infections and lymphoma. • Anti-dsDNA antibodies have been reported in patients treated with minocycline, etanercept, and infliximab.
Extractable Nuclear Antigens (ENA) ❑ Over 100 different soluble cytoplasmic and nuclear antigens ❑ Example: Ro, La, Sm, RNP, Scl-70 and Jo1 ❑ Detected by immunoblotting techniques Machine used in clinical lab for Immunoblotting.
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