Background

Connective Tissue Diseases
Systemic Lupus Erythematosus

SLE is an autoimmune CTD. It is one of several types of lupus, the others being cutaneous and drug-induced lupus. About 90% of lupus patients are women between the ages of 15 and 44 years. SLE causes inflammation and can affect any part of the body, most commonly the skin, heart, joints, lungs, blood vessels, liver, kidneys, and nervous system. Although generally not fatal, SLE can increase mortality, most commonly from cardiovascular disease due to accelerated atherosclerosis. SLE can also lead to kidney failure, which may reduce survival. The survival rate in the U.S. is approximately 95% at 5 years and 78% at 20 years.^1, The morbidity associated with SLE is substantial. Symptoms such as joint and muscle pain can impact the quality of life and functional status. SLE also increases patients' risk of infection, cancer, avascular necrosis (bone death), and pregnancy complications (eg, preeclampsia, preterm birth). The course of the disease is variable, and patients generally experience flares of mild-to-severe illness and remission.

Other Connective Tissue Diseases

Several other CTDs may require a differential diagnosis from SLE (eg, rheumatoid arthritis, thyroid disease, Sjögren syndrome, antiphospholipid syndrome, and polymyositis).

Rheumatoid arthritis is a chronic inflammatory peripheral polyarthritis. Rheumatoid arthritis can lead to deformity through stretching of tendons and ligaments and destruction of joints through erosion of cartilage and bone. Rheumatoid arthritis can also affect the skin, eyes, lungs, heart, and blood vessels.

Graves disease is an autoimmune disorder that leads to overactivity of the thyroid gland. The disease arises from thyroid-stimulating hormone receptor antibodies. It is the most common cause of hyperthyroidism. Blood tests may show raised thyroid-stimulating immunoglobulin antibodies.

Hashimoto disease, also known as chronic lymphocytic thyroiditis, is an autoimmune disorder and is the most common cause of hypothyroidism second to iodine insufficiency. It is characterized by an underactive thyroid gland and gradual thyroid failure. Diagnosis is confirmed with blood tests for thyroid-stimulating hormone (T4) and antithyroid antibodies.

Sjögren syndrome is an autoimmune disorder characterized by dryness of the eyes and mouth due to diminished lacrimal and salivary gland function. Affected individuals may also have symptoms of fatigue, myalgia, and cognitive dysfunction, which may be difficult to distinguish clinically from fibromyalgia or medication side effects. Typical antibodies include antinuclear antibody (ANA), anti-Sjögren-syndrome-related antigen, anti-Sjögren syndrome type B, or rheumatoid factor.

Antiphospholipid syndrome is a systemic autoimmune disorder characterized by venous or arterial thrombosis and/or pregnancy morbidity. Antiphospholipid antibodies are directed against phospholipid-binding proteins.

Polymyositis and dermatomyositis are inflammatory myopathies characterized by muscle weakness and inflammation. Dermatomyositis may also have skin manifestations.

Regulatory Status

Clinical laboratories may develop and validate tests in-house and market them as a laboratory service; laboratory-developed tests must meet the general regulatory standards of the Clinical Laboratory Improvement Amendments. The Avise® tests (Exagen Diagnostics) are available under the auspices of the Clinical Laboratory Improvement Amendments. Laboratories that offer laboratory-developed tests must be licensed by the Clinical Laboratory Improvement Amendments for high-complexity testing. To date, the U.S. Food and Drug Administration has chosen not to require any regulatory review of this test.

Related Policies

• Vectra DA Blood Test for Rheumatoid Arthritis (Policy #107 in the Pathology Section)

The first step in assessing a medical test is to formulate the clinical context and purpose of the test. The test must be technically reliable, clinically valid, and clinically useful for that purpose. Evidence reviews assess the evidence on whether a test is clinically valid and clinically useful. Technical reliability is outside the scope of these reviews, and credible information on technical reliability is available from other sources.

Systemic Lupus Erythematosus And Other Connective Tissue Diseases
Clinical Context and Test Purpose

The purpose of serum biomarker panel testing is to provide a diagnostic option that is an alternative to or an improvement on existing tests, such as established SLE classification systems and individual serum biomarker tests, in patients with signs and/or symptoms of SLE.

The question addressed in this policy is does the use of a serum biomarker panel improve the net health outcome in patients with signs and/or symptoms of SLE or other CTDs?

The following PICO was used to select literature to inform this policy.

Patients

The population of interest is individuals with signs and/or symptoms of SLE. Patients with SLE often present with nonspecific symptoms such as fever, fatigue, joint pain, and rash, which can make the disease difficult to diagnose. In some patients, the diagnosis of SLE can be made with certainty (eg, when there are typical symptoms of rash and joint symptoms, and laboratory testing shows a high-titer abnormal ANA in a pattern specific for SLE). However, in many other patients, the symptom patterns of SLE are less clear, and ANA testing is equivocal; as a result, cascade testing with additional serologic tests may be ordered. In addition, ANA testing alone can result in false-positives due to low specificity.

Interventions

The test being considered is serum biomarker panel testing. Patients with signs and/or symptoms of SLE are actively managed by rheumatologists, cardiologists, pulmonologists, nephrologists, and primary care providers in an outpatient clinical setting.

SLE is an autoimmune CTD that can be difficult to diagnose because patients often present with diverse, nonspecific symptoms that overlap with other CTDs; to further complicate matters, commonly used laboratory tests are not highly accurate. Moreover, similar symptoms may also present themselves in patients with fibromyalgia. Currently, differential diagnosis depends on a combination of clinical signs and symptoms and individual laboratory tests. More accurate laboratory tests for SLE and other CTDs could facilitate the diagnosis of the disease. Recently, laboratory-developed, diagnostic panel tests with proprietary algorithms and/or index scores for the diagnosis of SLE and other autoimmune CTDs have become commercially available.

At least 1 multibiomarker test to aid diagnosis of SLE and other CTDs is commercially available. This panel, Avise CTD (Exagen Diagnostics), contains 22 different tests. It combines 2 smaller panels, a 10-marker panel that includes common SLE tests, as well as cell-bound complement activation products (known as Avise Lupus) and a 12-marker panel that focuses on CTDs other than SLE (known as Avise CTD). Avise CTD includes nuclear antigen antibodies markers to help distinguish CTD, a rheumatoid arthritis panel to rule-in or rule-out rheumatoid arthritis, an antiphospholipid syndrome panel to assess risk for thrombosis and cardiovascular events, and a thyroid panel to help rule-in or rule-out Graves disease and Hashimoto's disease. Specific biomarkers in the panel are listed in Table 1.

Table 1. Avise Systemic Lupus Erythematosus Tests

Systemic Lupus Erythematosus Tests

10-marker Avise Lupus test

Auto-antibodies: ANA, anti-dsDNA, antimutated citrullinated vimentin, C4d erythrocyte-bound complement fragment, C4d lymphocyte-bound complement, anti-Sm, Jo-1, Sci-70, CENP, SS-B/La

Avise CTD test

Avise Lupus test plus the following:

Auto-antibodies: U1RNP, RNP70, SS-A/Ro

Rheumatoid arthritis auto-antibodies: rheumatoid factor IgM, rheumatoid factor IgA, anti-cyclic citrullinated peptide IgG

Anti-phospholipid syndrome auto-antibodies: cardiolipin IgM, cardiolipin IgG, β2-glycoprotein 1 IgG, β2-glycoprotein 1 IgM

Thyroid auto-antibodies: thyroglobulin IgG, thyroid, thyroid peroxidase

ANA: antinuclear antibody; anti-dsDNA: antibodies to double-stranded DNA; anti-Sm: antibodies to Smith nuclear antigen; CTD: connective tissue disease; Ig: immunoglobulin.

The Avise CTD test assesses all 22 markers. Avise CTD uses a 3 step process.^2, The 10-marker panel is done in 2 tiers, and the add-on 12-marker panel is done in a third step to further assist with the differential diagnosis of CTD. In addition, ANA testing is done by enzyme-linked immunosorbent assay and by indirect immunofluorescence. The 2-tiered testing approach to the 10-marker panel is described next.

Tier 1: Tests for anti-Sm, EC4d, BC4d, and anti-ds DNA. If any tests are positive, the result is considered suggestive of SLE and no further testing is done. Cutoffs for positivity are greater than 10 U/mL for anti-Sm, greater than 75 U/mL for EC4d, greater than 200 U/mL for BC4d, and greater than 301 U/mL for anti-dsDNA. Positive findings for anti-dsDNA are confirmed with a Crithidia luciliae assay.

Tier 2: If the tier 1 tests are negative, an index score is created, consisting of results of tests for ANA, EC4d and BC4d, antimutated citrullinated vimentin, anti-Jo-1, anti-Sci-70, anti-CENP, and anti-Ss-B/La. In other words, there are 6 additional markers and the ratio of EC4d to BC4d, both of which were measured in tier 1.

The index score (tier 2), calculated using a proprietary algorithm, rates how suggestive test results are of SLE. Although there is information on cutoffs used to indicate positivity for individual markers, information is not available on how precisely the index score is calculated. The score can range from -5 (highly nonsuggestive of SLE) to 5 (highly suggestive of SLE), and a score of -0.1 to 0.1 is considered indeterminate.

Exagen also offers the Avise Lupus Prognostic test, a 10-marker panel that can be ordered with the Avise Lupus and Avise CTD panels. The prognostic test focuses on patients' risk of lupus nephritis, neuropsychiatric SLE, thrombosis, and cardiovascular events. The test includes anti-C1q, anti-ribosomal P, anti-phosphatidylserine/prothrombin immunoglobulin (Ig) M and IgG, anti-cardiolipin IgM, IgG, and IgA and anti-β2-glycoprotein 1 IgM, IgG, and IgA. Four of the 10 markers are included in both panel tests.

Comparators

Comparators of interest include established SLE classification systems (eg, American College of Rheumatology [ACR], Systemic Lupus International Collaborating Clinics [SLICC]) and clinical diagnosis based on clinical and laboratory findings, such as individual serum biomarker tests, with exclusion of alternative diagnoses. Comparators are actively managed by rheumatologists, cardiologists, pulmonologists, nephrologists, and primary care providers in an outpatient clinical setting.

The diagnosis of SLE has been based on a combination of clinical symptoms and laboratory results. Previously, the ACR published an updated 1982 criteria for classifying SLE. In 1997 the ACR updated 1982 criteria for the classification of SLE.^3,4, In 2019, new classification criteria endorsed by the European League Against Rheumatism (EULAR) and the ACR were developed and validated.^5, The 2019 EULAR/ACR classification criteria requires a positive ANA as an entry criterion. For those with a positive ANA, additive criteria are assessed in 7 clinical and 3 immunological domains. Weighted criteria (ranging from 2 to 10 points) are evaluated within each domain, with the only the highest weighted criterion in a specific domain counting towards the total score. The weighted feature allows for criteria that are more tightly associated with SLE to contribute more heavily to the overall score. A classification of SLE requires a total score of ≥10 points.

The EULAR/ACR classification criteria are as follows:

• Entry criterion: ANA at a titer of ≥1:80 on HEp-2 cells or an equivalent positive test
• If entry criterion is present, apply additive criteria (weight):
  • Constitutional: fever (2)
  • Hematologic: leukopenia (2), thrombocytopenia (4), autoimmune hemolysis (4)
  • Neuropsychiatric: delirium (2), psychosis (3), seizure (5)
  • Mucocutaneous: non-scarring alopecia (2), oral ulcers (2), subacute cutaneous or discoid lupus (4), acute cutaneous lupus (6)
  • Serosal: pleural or pericardial effusion (5), acute pericarditis (6)
  • Musculoskeletal: joint involvement (6)
  • Renal: proteinuria >0.5 g/24 h (4), renal biopsy Class II or V lupus nephritis (8), renal biopsy Class III or IV lupus nephritis (10)
  • Antiphospholipid antibodies: anti-cardiolipin antibodies or anti-β2GP1 antibodies or lupus anticoagulant (2)
  • Complement proteins: low C3 or low C4 (3), low C3 and low C4 (4)
  • SLE-specific antibodies: antibodies to double stranded DNA (anti-dsDNA) or antibodies to Smith antigen (anti-sm) (6)

The SLICC, an international research group, developed revised criteria for diagnosing SLE in 2012.^7, These criteria include more laboratory tests than the 1997 ACR criteria, including elements of the complement system. Patients are classified as having SLE if they satisfy 4 or more of the 18 criteria below, including at least 1 clinical criterion and 1 immunologic criterion, or they have biopsy-confirmed nephritis compatible with SLE and with ANA or anti-dsDNA antibodies. In a sample of 690 patients, the SLICC criteria had a sensitivity of 97% and a specificity of 84% for diagnosing SLE, whereas the ACR criteria applied to the same sample had a sensitivity of 83% and a specificity of 96%. It is not clear how well-accepted the SLICC recommendations are in the practice setting. Table 2 outlines the SLICC criteria.

Table 2. Clinical and Immunologic Criteria

Clinical Criteria

Acute cutaneous lupus (including but not limited to lupus malar rash)

Chronic cutaneous lupus (including but not limited to discoid rash)

Oral ulcers

Nonscarring alopecia in the absence of other causes

Synovitis involving ≥2 joints, characterized by swelling or effusion or and ≥30 min of morning stiffness

Serositis

Renal: excessive protein in the urine or cellular casts in the urine

Neurologic disorder: seizures, psychosis, mononeuritis complex, or peripheral, or cranial neuropathy

Seizures

Hemolytic anemia

Leukopenia or lymphopenia

Thrombocytopenia

Immunologic Criteria

Antinuclear antibody above laboratory reference range

Antibodies to double-stranded DNA above laboratory reference range

Antibodies to Smith nuclear antigen

Antiphospholipid antibody

Low complement (low C3, low C4, or low CH150)

Direct Coombs tests in the absence of hemolytic anemia

To date, the most common laboratory tests performed in the diagnosis of SLE are serum ANA, and, if positive, tests for anti-dsDNA and anti-Sm. ANA tests are highly sensitive (ie, with a high negative predictive value) but have low specificity and relatively low positive predictive value, particularly when the ANA is positive at a low level. Specificity of testing can be increased by testing for specific antibodies against individual nuclear antigens (extractable nuclear antigens) to examine the "pattern" of ANA positivity. These include antigens against single- and dsDNA, histones, Sm, Ro, La, and RNP antibodies. The presence of anti-dsDNA or anti-Sm is highly specific for SLE because few patients without SLE test positive; however, neither test has high sensitivity.^8, The presence of other antibody patterns may indicate the likelihood of other diagnoses. For example, the presence of Ro and La antibodies suggests Sjögren syndrome, while the presence of antihistone antibodies suggests drug-induced lupus.

Outcomes

General outcomes of interest are test accuracy, symptoms, and quality of life.

Table 3. Outcomes of Interest for Individuals With Signs and/or Symptoms of SLE

Outcomes	Details
Test accuracy	Sensitivity and specificity in detecting biomarkers for SLE [FU for several years to assess accuracy of diagnosis]
Symptoms	Malar rash, discoid rash, photosensitivity, mouth or nose ulcers, arthritis (nonerosive), among others [≥ 2 weeks]
Quality of life	Relief of symptoms [≥ 3 years] Reduction in joint and organ damage [≥ 3 years]

FU: follow-up; SLE: systemic lupus erythematosus.

More specifically, outcomes of interest for SLE include a disease activity indices, organ damage, reduction in flares, and reduction in concomitant corticosteroids.^9, Patient reported outcomes are also encouraged, particularly ones that measure fatigue as most experts agree that it is one of the most important symptoms of SLE. However, the U.S. Food and Drug Administration (FDA) has not identified an existing instrument optimal for measuring fatigue in patients with SLE. Both fatigue and pain are the most consequential and frequent symptoms in SLE and these contribute significantly to physical functioning, sleep, and the ability to complete daily tasks, among other quality of life measures.^10, Validated instruments for measuring quality of life in SLE are mainly used in clinical trials. SLE-specific measures include the Lupus-quality-of-life and SLE-specific quality-of-life (SLEQOL) instruments; additionally general quality of life measures are also used to measure health-related quality of life (eg, Short Form 36 (SF-36). Recommended health outcome measures for disease activity and organ damage per FDA guidance is summarized in Table 4.^9,11,

Table 4. Health Outcome Measures Relevant to SLE.

Outcome	Measure (Units)	Assessment	Description	Clinical Interpretation (if available)
Disease activity index
BILAG 200412,	Disease activity is scored from A to E	Disease activity within last month	Ordinal scale index that assesses 9 individual organ systems. Disease activity is scored and converted into 5 levels from A to E. Grade A is very active disease requiring anticoagulation therapy, while Grade E is no current or previous disease activity.	Major clinical response as defined by the FDA as BILAG C scores or better at 6 months with no new BILAG A or B scores with maintenance of response between 6 to 12 months.
SLEDAI-2K13,	Scale from 0 to 105	Disease activity within last 10 days	A 24-item assessment of 16 clinical symptoms and 8 laboratory results that covers 9 organ systems. Items are weighted giving individual item scores ranging from 1 to 8. Categories of activity range from inactive (score of 0) to very active (score > 12).	A score of 6 is considered clinically important and effects the decision to treat.
SLAM-R14,	Scale from 0 to 81	Disease activity within last month	Evaluates 9 organ systems plus 7 laboratory features. Each organ item is scored 0 to 3 points. Laboratory categories can score a maximum of 21 points. Higher scores indicate higher disease activity.	A score of 7 is considered clinically important and affects the decision to treat.
ECLAM15,	Scale from 0 to 17.5	Disease activity within last month	A 33-item assessment that is organized into 12 categories, including 10 organ symptoms plus ESR and complement levels. Individual item scores range from 0.5 to 2. Higher scores indicate higher disease activity.	-
Organ damage assessment
SLICC/ACR damage index 16,	Scale from 0 to 46	Disease damage present for ≥6 months or after irreversible event	Captures items of permanent change after a diagnosis of SLE that covers specific manifestations in 12 organ systems. The 41-item assessment scores the presence of organ damage from 1 to 3 points. Higher scores indicate higher damage.	Organ damage is considered if the score is ≥1. Cumulative damage is a poor prognostic sign and a predictor of mortality.

ACR: American College of Rheumatology; BILAG: British Isles Lupus Assessment Group; ECLAM: European Consensus Lupus Activity Measure; ESR: erythrocyte sedimentation rate; FDA: U.S. Food and Drug Administration; SLAM-R: Systemic Lupus Erythematosus Activity Measure revised; SELENA: Safety of Estrogen in Lupus Erythematosus National Assessment Trial; SLE: systemic lupus erythematosus; SLEDAI-2K: Systemic Lupus Erythematosus Disease Activity Index 2000; SLICC: Systemic Lupus Erythematosus International Collaborating Clinics.

Lastly, a quicker diagnosis of SLE could allow the initiation of treatments for SLE sooner. Treatments for SLE can ameliorate symptoms, reduce disease activity, and slow progression of organ damage; however, there is no cure. Muscle and joint pain, fatigue, and rashes are generally treated initially with nonsteroidal anti-inflammatory drugs. Antimaltimes drugs such as hydroxychloroquine can relieve some symptoms of SLE including fatigue, rashes, and joint pain. Patients with more severe symptoms (eg, heart, lung, or kidney involvement) can be treated with corticosteroids or immune suppressants. There are also biologic treatments (eg, rituximab) approved by the U.S. Food and Drug Administration for the treatment of rheumatoid arthritis and are being evaluated for SLE.

Study Selection Criteria

Below are selection criteria for studies to assess whether a test is clinically valid.
• The study population represents the population of interest. Eligibility and selection are described.
• The test is compared with a credible reference standard.
• If the test is intended to replace or be an adjunct to an existing test; it should also be compared with that test.
• Studies should report sensitivity, specificity, and predictive values. Studies that completely report true- and false-positive results are ideal. Studies reporting other measures (eg, receiver operating characteristic [ROC], area under receiver operating characteristic [AUROC], c-statistic, likelihood ratios) may be included but are less informative.
• Studies should also report reclassification of diagnostic or risk category.

Technically Reliable

Assessment of technical reliability focuses on specific tests and operators and requires a review of unpublished and often proprietary information. Review of specific tests, operators, and unpublished data are outside the scope of this policy and alternative sources exist. This policy focuses on the clinical validity and clinical utility.

Clinically Valid

A test must detect the presence or absence of a condition, the risk of developing a condition in the future, or treatment response (beneficial or adverse).

Review of Evidence
Serum Biomarker Panel Tests

Putterman et al (2014) published data from a large cross-sectional, industry-sponsored study evaluating serum biomarkers for the diagnosis of SLE.^20, They analyzed the 10 markers in the Avise Lupus (plus ANA) using a 2-tier testing logic similar to that employed in the commercially available panel (see the Background section). The study evaluated 2 cohorts (N =794 patients); 593 participants were enrolled between April and August 2010, and 201 participants enrolled between June 2011 and September 2013. Together, the 2 cohorts consisted of 304 patients who met ACR classification criteria for SLE, 161 patients diagnosed with other rheumatic diseases and 205 healthy volunteers. Results of serum testing were available for 764 (96%) of 794 participants.

The diagnostic accuracy of the CB-CAP EC4d and BC4d were compared with reduced complement (C3, C4) and anti-dsDNA. The AUROC curve was significantly higher for EC4d (0.82) and BC4d (0.84) than for C3 (0.73) and C4 (0.72) (p<0.001). The AUROC curve was significantly higher for BC4d than for anti-dsDNA (0.79; p=0.009) but the difference was not statistically significant between EC4d and anti-dsDNA.

A total of 140 (46%) patients with SLE, 9 (3%) patients with other diseases, and 1 healthy volunteer tested positive for at least 1 of the 4 tier 1 markers. Patients testing negative for tier 1 tests underwent tier 2 testing and an index score was calculated. A total of 102 (62%) of 164 patients with SLE analyzed in tier 2 had an index score greater than 0 (ie, suggestive of SLE). Moreover, 245 of 276 patients with other rheumatic diseases had an index score of less than 0 (ie, not suggestive of SLE). When the results of tier 1 and 2 testings were combined, the overall sensitivity for SLE was 80% (242/304) and the overall specificity for distinguishing SLE from other diseases was 86% (245/285). The specificity for distinguishing between SLE and healthy volunteers was 98% (201/205).

As shown in Table 5, the specificity and area under the curve were higher for models including CB-CAPs than in those without these markers; sensitivity was slightly lower.

Table 5. Diagnostic Accuracy of Various Combinations of Markers

Measures	dsDNA, Sm, and ANA	dsDNA, Sm, ANA, Plus Antibody Specificity Components But Not CB-CAPs	Two-Tiered Testing Using All Markers, Including CB-CAPs EC4d and BC4d
Sensitivity, %	89	83	80
Specificity, %	53	76	86
Area under the curve	0.78	0.80	0.91

ANA: antinuclear antibodies; CB-CAP: cell-bound complement activation product; dsDNA: double-stranded DNA; Sm: Smith nuclear antigen.

A limitation of Putterman et al (2014) is that the study sample population included patients with SLE who met ACR classification criteria, but not patients with symptoms suggestive of SLE who failed to meet ACR criteria. It is not known how the diagnostic accuracy of the panel test compares with the ACR classification criteria or with concurrent clinician diagnosis (the mean time since SLE diagnosis was 11 years).

A subsequent industry-sponsored study by Wallace et al (2016) analyzed serum biomarkers as well as an algorithm for diagnosing SLE.^21, This study analyzed markers in the Avise Lupus (plus ANA) test using a 2-tier testing logic to evaluate SLE patients who met ACR criteria (n=75) and patients with primary fibromyalgia (n=75). High expression of CB-CAP EC4d or BC4d had 43% sensitivity and 96% specificity for the diagnosis of SLE. Use of a multianalyte assay with the algorithm, including CB-CAP levels, generated indeterminate results in 12 of the 150 subjects enrolled. For the remainder of patients, use of the algorithm to diagnosis SLE was 60% sensitive and 100% specific. Study limitations included a selection of patients with well-established diagnosis and long duration of disease.

Mossell et al (2016) reported on an industry-sponsored retrospective case-control study of 23 patients who had a positive Avise Lupus test result and 23 patients who had a negative result.^22, All patients were ANA-positive but negative for auto-antibodies specific for SLE, representing cases difficult to diagnose. Each positive Avise test case was matched to a control (negative test) from the same clinic with the same ANA level. A chart review was performed by a nonblinded rheumatologist approximately 1 year after the test results were available. Of the cases with a positive Avise Lupus test, 20 (87%) were diagnosed with SLE during follow-up. This compared with 4 (17%) individuals who had a negative result on the Avise Lupus test, resulting in a sensitivity of 83.3% and specificity of 86.4%. Interpretation of this study is limited due to its retrospective design, relatively short follow-up to monitor the progression of the disease, and the lack of an independent reference standard, because the diagnosis was based in part on the results of that test. The authors noted that prospective studies would be performed.

Liang et al (2020) conducted a retrospective single-center study of 117 patients in a rheumatology clinic without a confirmed SLE diagnosis who had received an Avise CTD test as part of their clinical care between April 2014 and November 2016.^23, The study aimed to determine whether the Avise test would aid in assessing the risk of developing SLE in patients who had undifferentiated findings presenting in a real-world setting. At the clinic, patients who had inflammatory arthritis, undifferentiated CTD, or other diagnoses or features suggestive of SLE received Avise testing. In this cohort of patients without a diagnosis of SLE at baseline, the diagnosis at 2 years from baseline changed in 80% (16/20) of patients who had a positive test as opposed to only 28.9% (28/97) who had a non-positive test. Of the 20 patients who had a positive test, 13 (65%) had their diagnosis changed to SLE at 2 years. The Avise test was associated with a specificity of 93%, with a sensitivity of 57%, positive predictive value of 65% and negative predictive value of 90%. The study also observed that patients with a positive Avise test had a significant accrual of clinical features, as defined by SLICC and ACR criteria, as well as organ damage, as defined by the SLICC Damage Index, compared to those without a positive test over the 2 year period. Additionally, there were no significant differences in medication regimens received by positive versus non-positive patients at baseline or at 2 years, except for more frequent use of mycophenolate mofetil in positive patients at year 2. Limitations of the study include its retrospective design and the potential for confirmation bias as treating physicians were aware of the Avise results and were potentially less likely to diagnose SLE in a patient with a negative Avise test. The authors concluded that the Avise CTD may be useful in predicting the development of SLE.

Ramsey-Goldman et al (2020) evaluated the usefulness of CB-CAPs and a multianalyte assay in patients with suspected SLE to predict progression to SLE as classified by ACR criteria in an industry-sponsored prospective observational study at 7 academic institutions.^24, Patients with probable SLE as suspected by lupus experts who also met 3 ACR criteria (n=92) were enrolled along with patients with established SLE based on ACR and SLICC criteria (n=53). A control group of patients with primary Sjogren's syndrome and other rheumatic diseases (n=101) were also included. The multianalyte panel with algorithm evaluated was the Avise Lupus test. The sensitivity of CB-CAPs and MAP at enrollment was higher compared to anti-dsDNA levels or low complement levels. The MAP was more sensitive and specific than CB-CAPs in patients with probable SLE (40% vs 28% and 96% vs 86%, respectively). The ability of positive CB-CAPs and MAPs to predict fulfillment of the ACR criteria at 9 to 18 months after enrollment was also analyzed. In the subgroup of 20 patients with probable SLE who fulfilled ACR criteria within 18 months, 8 (40%) had a MAP score >0.8 at enrollment. Kaplan-Meier estimates found that a MAP score >0.8 was predictive of progression to classifiable SLE (hazard ratio 3.11, 95% confidence interval 1.26 to 7.69). A limitation of the study was the relatively small population of patients with probable SLE.

Clinically Useful

A test is clinically useful if the use of the results informs management decisions that improve the net health outcome of care. The net health outcome can be improved if patients receive correct therapy, or more effective therapy, or avoid unnecessary therapy, or avoid unnecessary testing.

Review of Evidence
Direct Evidence

Direct evidence of clinical utility is provided by studies that have compared health outcomes for patients managed with and without the test. Because these are intervention studies, the preferred evidence would be from randomized controlled trials.

Chain of Evidence

Indirect evidence on clinical utility rests on clinical validity. If the evidence is insufficient to demonstrate test performance, no inferences can be made about clinical utility.

A more accurate and timelier diagnosis of SLE (ie, before multiorgan system involvement) and other CTDs could lead to better patient management (eg, more appropriate medical treatment). This, in turn, could improve health outcomes (eg, less joint or organ damage, improved survival).

Randomized Controlled Trials

Serum biomarker panel tests should be compared with usual clinical diagnosis assessments. Clinical diagnosis for SLE is not standardized, but generally consists of assessments of individual biomarkers in patients with signs and symptoms suspicious of SLE. One randomized controlled trial (RCT) is available directly comparing serum biomarker panel tests to standard diagnosis laboratory testing.^25, Characteristics of the trial are shown in Table 6.

Table 6. Summary of RCT Characteristics

Study; Trial	Countries	Sites	Dates	Participants	Interventions
					Active	Comparator
Wallace et al (2019); CARE for Lupus trial25,	United States	32	July 2017 to December 2018	145 patients who were referred to a rheumatologist with a clinical suspicion for SLE, including a history of ANA positivity	Avise Lupus test (n=72)	Standard diagnosis laboratory testing (n=73)

ANA: antinuclear antibody; CARE: Clinical Laboratory Assessment and Recommendations for Lupus; RCT: randomized controlled trial; SLE: systemic lupus erythematosus; US: United States.

Health outcome results for RCTs are summarized in Table 7. Wallace et al (2019) reported quality of life measures with the 5-level EuroQOL-5 Dimension index, however, outcomes were not reported by treatment group.

Table 7. Summary of RCT results

	Disease activity	Initiation of SLE-specific treatment	Quality of life
Wallace (2019)25,	Change in PGA from baseline to week 12	Initiation of hydroxychloroquine	Change from baseline to week 12 for EQ5D-5L
N	145	145	145
Avise Lupus test	-0.39 ±0.08	25%	Not reported by treatment group
Standard diagnosis laboratory testing	-0.29 ±0.06	14%
Difference (95% CI)	Not reported (p=0.39)	Not reported (p=0.14)

CI: confidence interval; EQ5D-5L: 5-level EuroQOL-5 Dimension; PGA: physician global assessment; RCT: randomized controlled trial; SLE: systemic lupus erythematosus

Wallace et al (2019) evaluated the clinical utility of the Avise Lupus test for the diagnosis of lupus as compared to standard diagnosis laboratory testing.^25, The primary endpoint of the trial was the change in the physicians' estimate of likelihood of SLE before and after testing (12 weeks after enrollment). Physicians estimated the likelihood on a 5-point Likert scale ranging from 0 (very low) to 4 (very high). At baseline, pretest likelihood was similar between the standard diagnosis laboratory testing group and the Avise Lupus test group and the likelihood of SLE decreased in both groups after testing, but the magnitude of the decrease was greater in the Avise Lupus test group. The change in likelihood of SLE from randomization to post-test was -0.44 ± 0.10 in the Avise Lupus test group versus -0.19 ± 0.07 in the standard diagnosis laboratory testing group (p=0.027). The corresponding changes from baseline to end of study at week 12 was -0.31 ± 0.10 versus -0.61 ± 0.10 (p=0.025), for each group respectively.

Table 8. Study Relevance Limitations

Study	Populationa	Interventionb	Comparatorc	Outcomesd	Follow-Upe
Wallace (2019)25,			2. In the standard diagnosis laboratory group, physicians were not directed to order any specific laboratory test.	1. Formal diagnosis, or fulfillment of classification of SLE not included	1. Short follow-up did not allow for confirmation of SLE diagnosis or impact on longer term health outcomes

The evidence limitations stated in this table are those notable in the current review; this is not a comprehensive gaps assessment.

a Population key: 1. Intended use population unclear; 2. Clinical context is unclear; 3. Study population is unclear; 4. Study population not representative of intended use.
b Intervention key: 1. Not clearly defined; 2. Version used unclear; 3. Delivery not similar intensity as comparator; 4.Not the intervention of interest.
c Comparator key: 1. Not clearly defined; 2. Not standard or optimal; 3. Delivery not similar intensity as intervention; 4. Not delivered effectively.
d Outcomes key: 1. Key health outcomes not addressed; 2. Physiologic measures, not validated surrogates; 3. No CONSORT reporting of harms; 4. Not establish and validated measurements; 5. Clinical significant difference not prespecified; 6. Clinical significant difference not supported.
e Follow-Up key: 1. Not sufficient duration for benefit; 2. Not sufficient duration for harms.

Study	Allocationa	Blindingb	Selective reportingc	Data completenessd	Powere	Statisticalf
Wallace (2019)25,		1. No blinding was used in the study 3. Post-test likelihood of SLE assessed by the treating physician	2. Between group differences in quality of life measures were not reported		1. Power calculations were not performed	4. Median differences and 95% confidence intervals between treatment groups for outcomes was not reported

a Allocation key: 1. Participants not randomly allocated; 2. Allocation not concealed; 3. Allocation concealment unclear; 4. Inadequate control for selection bias.
b Blinding key: 1. Not blinded to treatment assignment; 2. Not blinded outcome assessment; 3. Outcome assessed by treating physician.
c Selective Reporting key: 1. Not registered; 2. Evidence of selective reporting; 3. Evidence of selective publication.
d Data Completeness key: 1. High loss to follow-up or missing data; 2. Inadequate handling of missing data; 3. High number of crossovers; 4. Inadequate handling of crossovers; 5. Inappropriate exclusions; 6. Not intent to treat analysis (per protocol for noninferiority trials).
e Power key: 1. Power calculations not reported; 2. Power not calculated for primary outcome; 3. Power not based on clinically important difference.
f Statistical key: 1. Analysis is not appropriate for outcome type: (a) continuous; (b) binary; (c) time to event; 2. Analysis is not appropriate for multiple observations per patient; 3. Confidence intervals and/or p values not reported; 4. Comparative treatment effects not calculated.

The diagnostic accuracy of the serum biomarker panel test was initially established in observational studies that evaluated the sensitivity and specificity of the test in patients with established SLE. However, the more relevant question of whether the Avise test can aid in the diagnosis or exclusion of SLE in a population with suspected SLE or undifferentiated findings in a real-world setting was examined in more recent studies. One retrospective evaluation found that in patients with undifferentiated CTD positive Avise test results increased the likelihood of a SLE diagnosis within 2 years. Another prospective observational study reported similar results in that patients with probable SLE were more likely to fulfill ACR criteria for SLE within 9 to 18 months after a positive Avise test result. These studies are limited by a lack of a comparator. One randomized controlled trial evaluated the influence of test results from Avise and standard diagnosis laboratory testing on rheumatologists’ likelihood of diagnosing SLE, which found that physicians were less likely to diagnosis SLE in a patient with a negative Avise test. The short follow-up period of the study limits an assessment on how this information would impact health outcomes. Additionally, the comparator arm in the trial, which was not standardized, may not be reflective of current practice where classification criteria are used widely.

Connective Tissue Diseases Other Than Systemic Lupus Erythematosus
Clinical Context and Test Purpose

The purpose of serum biomarker panel testing is to provide a diagnostic option that is an alternative to or an improvement on existing tests, such as clinical diagnosis and individual serum biomarker tests, in patients with signs and/or symptoms of CTD (besides SLE).

The question addressed in this policy is does the use of a serum biomarker panel improve the net health outcome in patients with signs and/or symptoms of SLE or other CTDs?

The following PICO was used to select literature to inform this policy.

Patients

The population of interest is individuals with signs and/or symptoms of CTD (other than SLE). Presenting clinical features of CTD are highly variable and can be non-specific, which contribute to the difficulty in diagnosis.

Interventions

The test being considered is serum biomarker panel testing.

Comparators

Comparators of interest include clinical diagnosis and individual serum biomarker tests.

Comparators are actively managed by rheumatologists, cardiologists, pulmonologists, nephrologists, and primary care providers in an outpatient clinical setting.

Outcomes

General outcomes of interest are test accuracy, symptoms, and quality of life.

Table 6. Outcomes of Interest for Individuals With Signs and/or Symptoms of CTD (Besides SLE)

Outcomes	Details
Test accuracy	Sensitivity and specificity in detecting biomarkers for CTDs other than SLE [FU for several years to assess accuracy of diagnosis]
Symptoms	Dry eyes and mouth, fatigue, cognitive dysfunction, muscle weakness and inflammation [≥ 2 weeks]
Quality of life	Symptom relief [≥ 3 years] Reduction in joint and organ damage [≥ 3 years]

CTD: connective tissue disease; FU: follow-up; SLE: systemic lupus erythematosus.
Study Selection Criteria

Below are selection criteria for studies to assess whether a test is clinically valid.
• The study population represents the population of interest. Eligibility and selection are described.
• The test is compared with a credible reference standard.
• If the test is intended to replace or be an adjunct to an existing test; it should also be compared with that test.
• Studies should report sensitivity, specificity, and predictive values. Studies that completely report true- and false-positive results are ideal. Studies reporting other measures (eg, receiver operating characteristic [ROC], area under receiver operating characteristic [AUROC], c-statistic, likelihood ratios) may be included but are less informative.
• Studies should also report reclassification of diagnostic or risk category.

A study by Liu et al (2009) evaluated lymphocyte-bound CAPS (LB-CAPS).^18, Of the 517 participants in this cross-sectional study, 179 patients had autoimmune or inflammatory diseases other than SLE. Not all of these diseases were CTDs but several CTDs were included in the study. Levels of LB-CAPs, T-cell bound C4d (TC4d) and C3d (TC3d), and B-cell bound C4d (BC4d), and C3d (BC3d) were measured in all participants. The diagnostic accuracy of these markers was assessed using ROC analysis. The area under the curve was 0.727 for TC4d and 0.770 for BC4d. TC4d was estimated to be 56% sensitive and 80% specific for differentiating SLE from other diseases. BC4d had 56% sensitivity and 80% specificity.

A cross-sectional study of platelet C4d by Navratil et al (2006) included 420 total participants, 115 of whom had rheumatic inflammatory/autoimmune or hematologic diseases other than SLE, several of which were CTDs.¹⁰ The authors reported that the marker was 98% specific for a diagnosis of SLE compared to the patients with other diseases.

Serum Biomarker Panel Tests for Connective Tissue Diseases Other Than Systemic Lupus Erythematosus

As previously discussed, Putterman et al (2014) published data from a large cross-sectional, industry-sponsored study evaluating serum biomarkers for the diagnosis of SLE.^20, They analyzed the 10 markers in the Avise Lupus (plus ANA) using a 2-tier testing logic similar to that employed in the commercially available panel (see the Background section). Of the 794 patients in the study, 161 were diagnosed with rheumatic diseases other than SLE.

A total of 140 (46%) patients with SLE, 9 (3%) patients with other diseases, and 1 healthy volunteer tested positive for at least 1 of the 4 tier 1 markers. Patients testing negative for tier 1 tests underwent tier 2 testing and an index score was calculated. A total of 245 of 276 patients with other rheumatic diseases had an index score of less than 0 (ie, not suggestive of SLE). When the results of tier 1 and tier 2 testings were combined, the overall specificity for distinguishing SLE from other diseases was 86% (245/285).

An earlier study by Kalunian et al (2012) reported on the first cohort of 593 individuals included in the Putterman et al (2014) analysis.^17, Out of 593 participants, 178 patients had rheumatic diseases, 210 had SLE, and 205 were healthy volunteers. Authors evaluated the performance of a 7-marker biomarker panel for the diagnosis of SLE; some markers are included in a commercially available panel test. The biomarkers included ANA, anti-dsDNA, antimutated citrullinated vimentin, and the CB-CAPs (EC4d, PC4d, BC4d). In relation to SLE, the combination of anti-dsDNA and the multivariate logistic regression analysis index score yielded 87% specificity against other rheumatic diseases.

Summary of Evidence for Diagnosing Connective Tissue Diseases Other Than Systemic Lupus Erythematosus

All studies found centered around diagnosing SLE with other CTDs as comparators and did not assess the sensitivity of the biomarker tests to detect CTDs other than SLE. For individuals with signs and/or symptoms of CTD (besides SLE) who receive serum biomarker panel testing, more studies are needed. Relevant outcomes are test accuracy, symptoms, and quality of life. The evidence is insufficient to determine the effects of the technology on health outcomes.

Summary of Evidence

For individuals with signs and/or symptoms of SLE who receive serum biomarker panel testing, the evidence includes several diagnostic accuracy studies and 1 prospective evaluation of clinical utility that compared the impact of the test results on physicians' evaluation of patients with a clinical suspicion for SLE. Relevant outcomes are test accuracy, symptoms, and quality of life. One case-control study found high sensitivity and specificity for a commercially available test for diagnosing SLE. More recent evaluations have tested how a panel test can aid in the diagnosis or exclusion of SLE in a population with suspected SLE or undifferentiated findings. Two observational studies found that patients with a positive Avise test were more likely to have classifiable SLE after 9 months to 2 years of follow-up. Additionally, a randomized controlled trial evaluated the influence of test results from Avise and standard diagnosis laboratory testing on rheumatologists’ likelihood of diagnosing SLE, which found that physicians were less likely to diagnosis SLE in a patient with a negative Avise test. The evidence is insufficient to determine the effects of the technology on health outcomes.

For individuals with signs and/or symptoms of CTD (besides SLE) who receive serum biomarker panel testing, more studies are needed. Relevant outcomes are test accuracy, symptoms, and quality of life. The evidence is insufficient to determine the effects of the technology on health outcomes.

SUPPLEMENTAL INFORMATION
Practice Guidelines and Position Statements

No guidelines or statements were identified.

U.S. Preventive Services Task Force Recommendations

Not applicable.

Ongoing and Unpublished Clinical Trials

A search of ClinicalTrials.gov in May 2020 did not identify any ongoing or unpublished trials that would likely influence this policy.]
________________________________________________________________________________________

Horizon BCBSNJ Medical Policy Development Process:

This Horizon BCBSNJ Medical Policy (the “Medical Policy”) has been developed by Horizon BCBSNJ’s Medical Policy Committee (the “Committee”) consistent with generally accepted standards of medical practice, and reflects Horizon BCBSNJ’s view of the subject health care services, supplies or procedures, and in what circumstances they are deemed to be medically necessary or experimental/ investigational in nature. This Medical Policy also considers whether and to what degree the subject health care services, supplies or procedures are clinically appropriate, in terms of type, frequency, extent, site and duration and if they are considered effective for the illnesses, injuries or diseases discussed. Where relevant, this Medical Policy considers whether the subject health care services, supplies or procedures are being requested primarily for the convenience of the covered person or the health care provider. It may also consider whether the services, supplies or procedures are more costly than an alternative service or sequence of services, supplies or procedures that are at least as likely to produce equivalent therapeutic or diagnostic results as to the diagnosis or treatment of the relevant illness, injury or disease. In reaching its conclusion regarding what it considers to be the generally accepted standards of medical practice, the Committee reviews and considers the following: all credible scientific evidence published in peer-reviewed medical literature generally recognized by the relevant medical community, physician and health care provider specialty society recommendations, the views of physicians and health care providers practicing in relevant clinical areas (including, but not limited to, the prevailing opinion within the appropriate specialty) and any other relevant factor as determined by applicable State and Federal laws and regulations.

___________________________________________________________________________________________________________________________

Index:
Serum Biomarker Panel Testing for Systemic Lupus Erythematosus and Other Connective Tissue Diseases
Serum Biomarker Panel Testing for Systemic Lupus Erythematosus
Systemic Lupus Erythematosus, Serum Biomarker Panel Testing
SLE Serum Biomarker Panel Testing
Avise SLE 2.0 Test
Avise SLE + Connective Tissue 2.0 Test
Avise SLE Prognostic Test

References:
1. Kasitanon N, Magder LS, Petri M. Predictors of survival in systemic lupus erythematosus. Medicine (Baltimore). May 2006;85(3):147-156. PMID 16721257

2. American College of Rheumatology (ACR). 1997 Update of the 1982 American College of Rheumatology Revised Criteria for Classification of Systemic Lupus Erythematosus. n.d.; https://www.rheumatology.org/Portals/0/Files/1997%20Update%20of%201982%20Revised.pdf. Accessed June 1, 2018.

3. Hochberg MC. Updating the American College of Rheumatology revised criteria for the classification of systemic lupus erythematosus. Arthritis Rheum. Sep 1997;40(9):1725. PMID 9324032

4. Guidelines for referral and management of systemic lupus erythematosus in adults. American College of Rheumatology Ad Hoc Committee on Systemic Lupus Erythematosus Guidelines. Arthritis Rheum. Sep 1999;42(9):1785-1796. PMID 10513791

5. Gill JM, Quisel AM, Rocca PV, et al. Diagnosis of systemic lupus erythematosus. Am Fam Physician. Dec 1 2003;68(11):2179-2186. PMID 14677663

6. Petri M, Orbai AM, Alarcon GS, et al. Derivation and validation of the Systemic Lupus International Collaborating Clinics classification criteria for systemic lupus erythematosus. Arthritis Rheum. Aug 2012;64(8):2677-2686. PMID 22553077

7. Suresh E. Systemic lupus erythematosus: diagnosis for the non-specialist. Br J Hosp Med (Lond). Oct 2007;68(10):538-541. PMID 17974296

8. J C-V, Chitkara P, Christianakis S, et al. Finding the best approach to autoimmune connective tissue disease diagnosis (Paid supplement supported by Exagen Diagnostics). Rheumatology News. 2014; August:1-8.

9. Liu CC, Kao AH, Hawkins DM, et al. Lymphocyte-bound complement activation products as biomarkers for diagnosis of systemic lupus erythematosus. Clin Transl Sci. Aug 2009;2(4):300-308. PMID 20161444

10. Navratil JS, Manzi S, Kao AH, et al. Platelet C4d is highly specific for systemic lupus erythematosus. Arthritis Rheum. Feb 2006;54(2):670-674. PMID 16447243

11. Putterman C, Furie R, Ramsey-Goldman R, et al. Cell-bound complement activation products in systemic lupus erythematosus: comparison with anti-double-stranded DNA and standard complement measurements. Lupus Sci Med. Nov 2014;1(1):e000056. PMID 25396070

12. Kalunian KC, Chatham WW, Massarotti EM, et al. Measurement of cell-bound complement activation products enhances diagnostic performance in systemic lupus erythematosus. Arthritis Rheum. Dec 2012;64(12):4040-4047. PMID 22932861

13. Wallace DJ, Silverman SL, Conklin J, et al. Systemic lupus erythematosus and primary fibromyalgia can be distinguished by testing for cell-bound complement activation products. Lupus Sci Med. 2016;3(1):e000127. PMID 26870391

14. Mossell J, Goldman JA, Barken D, et al. The Avise Lupus Test and cell-bound complement activation products aid the diagnosis of systemic lupus erythematosus. Open Rheumatol J. Oct 2016;10:71-80. PMID 27867431

Codes:
(The list of codes is not intended to be all-inclusive and is included below for informational purposes only. Inclusion or exclusion of a procedure, diagnosis, drug or device code(s) does not constitute or imply authorization, certification, approval, offer of coverage or guarantee of payment.)

CPT*

81599
84999
0039U
0062U