Prevalence and adverse consequences of delayed diagnosis and misdiagnosis in thrombotic antiphospholipid syndrome. An observational cohort study and a review of the literature

Obiectives This study aims to prospectively evaluate the frequency and adverse consequences of diagnostic delay and misdiagnosis in a cohort of patients with thrombotic antiphospholipid syndrome (TAPS). In addition, a systematic review of the literature concerning the diagnostic delay and misdiagnosis of TAPS was carried out. Methods Patient enrollment occurred between 1999 and 2022. The study group was formed by TAPS patients whose diagnosis was delayed and those who were misdiagnosed. The control group was made up of patients who were timely and correctly diagnosed with TAPS. Results The literature review showed 42 misdiagnosed patients, 27 of them were in one retrospective cohort study and 15 in 13 case reports. One hundred sixty-one out of 189 patients (85.2%) received a timely, correct diagnosis of TAPS; 28 (14.8%) did not. The number of patients with diagnostic issues was significantly higher for the first period (1999–2010), and the number of patients with a correct diagnosis was significantly higher for the second one (2011–2022). When the clinical and laboratory characteristics of the patients with delayed diagnosis were compared with those with misdiagnosis, there was a significantly higher number of severe adverse consequences characterized by permanent disability or death in the latter group. The two most common types of misdiagnoses were systemic lupus erythematosus (6 cases, 46.1%) and cardiovascular diseases (4 cases, 30.8%). Conclusions The study demonstrates that although knowledge about TAPS has improved over time, diagnostic delays and errors remains to be addressed as they are strongly associated to adverse consequences. Key Points •Although knowledge of thrombotic antiphospholipid syndrome has improved over time, it is still limited. •Diagnostic delay and misdiagnosis are still an important issue that remains to be addressed as they are strongly associated to adverse consequences. •The three more frequent misdiagnoses are multiple sclerosis, systemic lupus erythematosus and cardiovascular diseases.


Introduction
Antiphospholipid syndrome (APS) is a hypercoagulable disorder characterized by the persistent presence in the blood of antiphospholipid antibodies (aPL) which include lupus anticoagulant (LAC) positivity and/or medium-high levels of anticardiolipin (aCL) and/or anti-beta2-glycoprotein I (anti-β2GPI) antibodies in patients with venous, arterial or micro vessels thrombosis and/or pregnancy morbidity [1].Deep veins of the lower extremities are the most frequent sites of venous thrombosis, while the brain is the site most involved in arterial thrombosis [2].However, atypical sites such as the abdominal aorta can also be involved [3].In addition, APS patients are at risk for small vessel thrombosis which can be localized or spread to multiple organs [2].APS can exist either alone or in association with other disorders, most frequently with systemic lupus erythematosus (SLE).While several proposals for the classification criteria of APS were periodically evaluated, a consensus statement was finally formulated at an international workshop held in Sapporo, Japan, in 1998 [4].The statement defined APS as a condition that must meet at least one clinical criterion (thrombosis or pregnancy morbidity) and a laboratory one (the persistent presence of aCL antibodies and/or LAC).In conjunction with persistent anti-β2GPI antibody positivity to other laboratory tests, those two criteria were confirmed at the consensus conference held in Sydney, Australia, in 2006 [1].For the time being, the diagnostic criteria for APS have not been defined, although the classification criteria outlined here are for the most part also used for diagnostic purposes.
APS is generally considered to fall within the group of rare diseases, being affecting ≤ 50 persons per 100,000 population [5,6].Recent studies have highlighted that most clinicians have only a limited knowledge about rare diseases [7,8].Given its rarity, patients suffering from thrombotic APS (TAPS) pose a great diagnostic challenge and are at high risk of receiving a delayed or incorrect diagnosis [9], a circumstance that can directly impact their chance of recovery and survival as they are exposed to the risk of thrombotic events with life-altering consequences during that interlude.Furthermore, a possible consequence of the delay in diagnosis can be the increase over time of the organ damage which can be determined by the damage index for thrombotic APS (DIAPS) [10,11].It is also important to remember that some neuropsychological or cardiac disorders, SLE or microangiopathies have clinical and/or laboratory features that overlap with those of TAPS, causing even more risk of diagnostic delay and misdiagnosis.Finally, the treatment of overlapping diseases, which is generally quite different from the antithrombotic therapy prescribed to TAPS patients, could worsen the original health problem or even cause severe harm [12,13].
The current study presents a literature review as well as an evaluation of the frequency of diagnostic delay and misdiagnosis occurring in a cohort of TAPS patients followed up prospectively.The adverse consequences linked to delays and errors are also examined.

Literature review
In accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) checklist protocol [14], a systematic review of full text manuscripts in the English language was carried out.Studies focusing on the diagnostic delay and misdiagnosis of TAPS with or without pregnancy morbidity published between July 1999 (the time that the Sapporo updated classification criteria of TAPS were published) and November 2022 were included in the search.The diagnostic delay due to delayed diagnosis or misdiagnosis was defined as the time between the onset of clinical manifestations suggestive of TAPS and the formulation of a correct diagnosis.The papers exclusively regarding purely obstetric APS were excluded from the study.Two authors (AR and MT) reviewed the literature and made the final decision independently and blindly.The records were retrieved by searching Medline via Pubmed, Scopus and Web of Science Databases.The references of relevant articles were also hand-searched to identify other potentially relevant studies.The online search was limited to observational studies (cohort, case-control and case series studies), but given the rarity of TAPS, the search strategy also included case reports.Positioned in different combinations in order to improve the sensitivity of the search strategy, the keywords entered into the search engine were: thrombotic antiphospholipid syndrome, antiphospholipid syndrome, primary antiphospholipid syndrome, secondary antiphospholipid syndrome, antiphospholipid syndrome related to other diseases, diagnostic delay, misdiagnosis and adverse consequences.The titles and abstracts of the articles originally identified were screened and those needing further examination were pinpointed.Once that phase was completed, all of the full-text articles identified were evaluated and the studies eligible for inclusion were determined.

Study population
Study group: was formed by the patients with onset of clinical manifestations suggestive for TAPS between July 1999 and November 2022.The inclusion criteria were the following: detection in outpatient clinic of the Rheumatology Unit of the Padua University Hospital of diagnostic issues such as the delay of a TAPS diagnosis or a misdiagnosis in patients with the clinical and laboratory classification criteria for TAPS as established by Sapporo or Sydney Consensus Conferences [1,4].The adverse consequences of the diagnostic delay or of the misdiagnosis were identified and registered.Both in patients with diagnostic delay and in those with misdiagnosis the time between the first manifestation and the correct TAPS diagnosis always was greater than six months.
Control group: included patients in whom the onset of clinical manifestations of TAPS occurred between July 1999 and November 2022, who received timely a correct diagnosis of TAPS according to the clinical and laboratory criteria formulated during the Sapporo or Sydney consensus conferences [1,4].In these patients the interval between the first manifestation and TAPS correct diagnosis varied between 3 and 6 months, the time necessary to have the confirmation of aPL positivity and to perform adequate blood tests and instrumental examinations.

Autoantibody detection
ACL and anti-β2GPI antibodies were determined by ELISA assays using a home-made method described elsewhere [15].ACL antibody values were expressed in IgG phospholipid (GPL) and IgM phospholipid (MPL) units, respectively.The results of anti-β2GPI antibodies were expressed in arbitrary units.The cut-off values for the medium-high levels of aCL and anti-β2GPI antibodies were calculated as > the 99 th percentile.LAC was assessed using a three-step procedure carried out utilizing platelet-poor plasma samples following updated guidelines and utilizing diluted Russell Viper Venom and diluted Activated Partial Thromboplastin Times as screening tests [16].

Statistical analysis
The categorical variables were expressed as frequencies and percentages; the continuous variables were expressed as mean and standard deviation.Univariate analysis was performed to evaluate the association between the categorical variables using Fischer's exact test, and between the continuous variables using a nonparametric Mann-Whitney U test.A < 0.05 p value was considered significant.All statistical analyses were performed using GraphPad Prism statistical software (San Diego, CA, USA).
No observational studies concerning a diagnostic delay were identified.However, there was an Italian Regional Rare Disease Registry referring to 740 patients with a definite diagnosis of APS characterized by thrombosis and/or pregnancy morbidity, registered between 1983 and 2015, where a mean diagnostic delay of 4.7 years ± 8.3 SD was reported; the figure resulted significantly reduced over time when data were stratified by time period [9].

Our cohort's data
Between July 1999 and November 2022, 189 patients attending the rheumatology outpatient clinic were ultimately diagnosed with TAPS.One hundred and thirtyeight were women (73.0%) and 51 (27.0%) men; the mean age at the time they were diagnosed was 46.0 years ± 13.9 SD.A timely and correct diagnosis of TAPS, in accordance with the Sapporo or Sidney classification criteria [1,4], was formulated for 161 (85.2%) of them.A diagnostic issue was registered for 28 (14.8%) of the patients; 15 of them (7.9%) had a diagnostic delay and 13 (6.9%) a misdiagnosis.The clinical and laboratory characteristics of the patients who received a correct diagnosis and those who received a tardy diagnosis or were misdiagnosed are outlined in Table 2. Data analysis showed that there was a significantly higher percentage of females in the correctly diagnosed TAPS group and a higher percentage of males in the group with diagnostic issues.It also showed that there was a significantly higher number of patients with diagnostic issues in the first period, and a significantly higher number of patients with correct TAPS diagnosis in the second one.Finally, it was found that in a significantly high number of patients with diagnostic issues, the determination of aPL at the onset of the clinical manifestations was missing.The correctly diagnosed patients and those with diagnostic issues did not show any significant differences in the type of vascular involvement and aPL antibody profile at onset or in the clinical form of TAPS.
The clinical and laboratory characteristics of patients with delayed diagnosis or misdiagnosis are outlined in Table 3 and 4, respectively.The adverse consequences caused by the diagnostic issues and the outcome after the correct diagnosis of TAPS are also outlined.In both groups there was a higher frequency of diagnostic errors between the 1999-2010 period with respect to the 2011-2022 one, a high number/percentage of patients who were not tested for aPL antibodies, and a long delay during which the patients in both groups developed adverse consequences before the correct diagnosis was made.In patients with diagnostic issues the most frequent first clinical manifestation that led to a diagnostic delay or misdiagnosis was deep vein thrombosis (DVT) of lower or upper limbs, present in 40% and 23.1% of the cases, respectively; while, the first manifestations leading to the correct TAPS diagnosis was an adverse consequence of diagnostic delay.In particular, in both patients with diagnostic delay and in those with misdiagnosis the most frequent manifestation leading to correct diagnosis was arterial thrombosis found in 40% and 61.5% of cases, respectively and characterized mainly by ischemic stroke.These manifestations were reported in detail in Table 3 and 4  the most frequent first manifestation of TAPS which led to the diagnosis was arterial thrombosis present in 37.3% of cases (Table 2).In patients with timely and correct TAPS diagnosis aPL were tested at the diagnosis time (Table 2).While in both groups with delayed diagnosis and misdiagnosis aPL were determined at the first clinical manifestations and re-tested after the occurrence of adverse consequences at the time of correct TAPS diagnosis (Tables 3 and 4).The medical consultations of patients with timely and correct diagnosis were on average three, a number certainly much lower than that of patients with diagnostic delay or misdiagnosis.Unfortunately, it was not possible to make a statistical comparison because most of these latter patients came to our tertiary center after a large, but unspecified number of medical consultations made by doctors of other centers.Importantly, there were no thrombotic events after the correct diagnosis of TAPS was made and the appropriate antithrombotic treatment started.The main misdiagnoses formulated were: SLE in 6 cases (46.1%) and cardiovascular diseases in 4 (30.8%).The characteristics of the two groups are compared in Table 5: the TAPS patients who were misdiagnosed were significantly younger with respect to the patients with a delayed diagnosis.Furthermore, there was a significantly higher percentage of severe adverse consequences characterized by permanent disability or death in the patients who were misdiagnosed.There were two deaths: a patient diagnosed with SLE in whom the diagnosis of secondary TAPS was formulated 8 years late (Table 4, case 7) who died at the age of 47 of sepsis due to severe ischemic lesions in the inferior and superior limbs and a patient who was diagnosed with a mitral valve infectious endocarditis who died at the age 52 of catastrophic APS (Table 4, case 10).As far as the clinical forms of TAPS were concerned, there was a significantly higher percentage of primary TAPS in the patients with a delayed diagnosis, and there was a higher percentage of secondary TAPS in the misdiagnosed patients (as SLE in 75% of the cases).There were no significant differences between the patients with delayed diagnosis and those with misdiagnosis as far as the mean age at the time of onset of manifestations suggestive of TAPS, the type of vascular involvement, the aPL antibody profile, the failure to order aPL antibody testing and the mean delay in the TAPS correct diagnosis were concerned.

Discussion
This is the first observational cohort study to evaluate the frequency and adverse consequences of delayed diagnosis and misdiagnosis in TAPS patients.According to data from the literature cases [17][18][19][20][21][22][23][24][25][26][27][28][29][30] and the Italian Regional Rare Disease Registry [9], the number of patients with diagnostic issues was significantly higher over the first part of the study period (1999-2010), and the number of patients with a correct diagnosis was significantly higher in the second one (2011-2022).The increasing availability of medical information and the easier access to diagnostic tests over time could have apparently contributed to preparing clinicians to address the challenge of recognizing TAPS patients.Despite the lower frequency of TAPS in the males, there was a significantly higher percentage of men in the group of patients with diagnostic issues with respect to those who were correctly diagnosed.The result could have some explanations: as autoimmune diseases affect mainly females, the differential diagnosis with autoimmune conditions is less obvious when dealing with male patients; on the other hand, cardiovascular diseases are more common among males, thus physicians could underestimate other possible differential diagnoses, including TAPS.As there were no significant differences in the age, type of vascular involvement, aPL antibody profile, and the clinical form of TAPS between the correctly diagnosed patients and those with diagnostic issues, presumably the delays or misdiagnoses were linked to an inadequate preparation of clinicians as far as TAPS is concerned.The hypothesis is consistent with the finding of a significant high number of patients with diagnostic issues who were not tested for aPL antibodies at the onset of clinical manifestations (Table 2).It is also important to note that in clinical practice the manifestations suggestive of TAPS including venous, arterial or micro vessel thrombosis can also be indicative of other more frequently observed disorders such as cardiovascular diseases, hypertension, diabetes, SLE or other systemic autoimmune diseases, therefore it could be difficult for several doctors attributing them to a rare and little known disease such as TAPS and activating the diagnostic workup also including the determination of the aPL antibodies.
In this experience the clinical manifestation that most frequently led to the diagnostic issues was DVT both in patients with diagnostic delay and in those with misdiagnosis.DVT is a disease that can be due to several disorders such as neoplasms, haematological diseases or discoagulopathies, which are commonly found in the general population and which could mislead the doctor from the diagnosis of a rare disease such as TAPS.Indeed, in 66.7% of these patients with DVT aPL determination was not performed.Instead, the most frequent adverse consequence that led both patients with diagnostic delay and those with misdiagnosis to the correct diagnosis was arterial thrombosis and mainly ischemic stroke, a severe disease that requires an in-depth diagnostic procedure that also includes aPL testing.An analysis of the study's data uncovered a long time lag between the onset of clinical manifestations and the correct diagnosis of TAPS in both the patients with delayed diagnosis and in those who were misdiagnosed (Table 5); this could explain the adverse consequences found in all the patients with diagnostic issues.Just as in those cases reported in the literature (Table 1), our TAPS patients who were misdiagnosed with SLE (Table 4, cases 1, 2, 5, 7) or with cardiovascular diseases (Table 4, cases 6, 9, 10, 11) suffered from more severe adverse consequences associated to permanent disability or death.Notably the secondary form of TAPS was found only in the misdiagnosed patients; in 75% of this group of patients TAPS was associated with SLE, which presumably delayed diagnosis and treatment.
In accordance with the cases reported in the literature, the diagnoses frequently formulated prior to a TAPS diagnosis were SLE [18,19,25] and cardiovascular diseases [21,22,24,[28][29][30], which were recorded overall in 76.9% of our patients.Although originally described in connection to SLE, APS was recognized as a primary disease in the late 1980s [31].Since APS and SLE share several clinical and immunological features such as hematological, cardiac, renal and neurological manifestations as well as aPL antibodies, it can be quite challenging to distinguish between the two disorders.In a recent study [32] the Systemic Lupus International Collaborating Clinics' (SLICC) classification system for SLE [33] was assessed in a cohort of 100 patients with primary APS.The study found that 28% of the patients could have been mistakenly classified as SLE.Although the new American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) 2019 classification criteria [34] have not been tested in a primary APS setting, Signorelli et al. [35] reported that the ACR/EULAR 2019 criteria [34] had a higher accuracy with respect to the SLICC 2012 one [33] in differentiating primary TAPS from SLE in 67 patients (misclassification 6.0% vs 35.8%).Another factor that may contribute in clinical practice to the tendency to mistake TAPS for SLE could be lupus anticoagulant's misleading name that could cause inexperienced physicians to lean towards a diagnosis of SLE, a more frequent and better known disease.As has been reported in the literature [18,19,25,32] and observed in our study, the TAPS patients who were diagnosed with SLE were prescribed inappropriate treatments such as corticosteroids and/or immunosuppressive drugs and not the life-saving anticoagulant and/or antiplatelet treatments they necessitated (Table 4, cases 1, 2, 5, 7, 8, 12).As is well known, the heart is a target organ in TAPS.Although not included in the current classification criteria, heart valve disease is considered one of the most frequent cardiac manifestations in patients with TAPS [13,36].The disorder is easily misdiagnosed as rheumatic valve disease or infectious/culture-negative endocarditis.Crucially, early diagnosis and aggressive anticoagulation treatment are considered imperative to avoid thromboembolism, further valvular destruction and/or myocardial dysfunction in these patients [13,36].
Limitations of the study: our experience was gained exclusively in a rheumatological context, therefore it mainly includes patients with misdiagnosis of TAPS as SLE or cardiovascular diseases.It seems probable that a not well defined percent of patients diagnosed with multiple sclerosis do in fact have APS, a condition with a totally different treatment and prognosis.Indeed, the clinical presentation and lesions evidenced by magnetic resonance imaging may be similar and therefore lead to a misdiagnosis [37].However, a misdiagnosis of multiple sclerosis is manly reported in the literature by immunological and neurological centers [17,23,38].Furthermore, until 2005 the diagnosis of TAPS was made according to the Sapporo classification criteria published in 1999 [4], which required the repeated detection of aCL and/or LAC and not of anti-β2GPI antibodies; the latter was subsequently included in the Sydney criteria published in 2006  [1].Therefore, between 1999 and 2005, TAPS patients who were positive only for anti-β2GPI antibodies were not diagnosed correctly.
Our data demonstrate that although more knowledge about TAPS, a rare, life-threatening disease, has become available, diagnostic delays and misdiagnoses continue to be associated to adverse consequences.What have we learned from this study?The findings suggest that medical schools need to train practitioners and specialized physicians to recognize the symptoms and manifestations of TAPS in order to be able to order the diagnostic tests (LAC, aCL and anti-β2GPI antibodies) to detect the syndrome.Physicians need to pay special attention to distinguishing TAPS from its overlappers, such as multiple sclerosis, SLE and cardiovascular diseases.It would also be important that the ACR/EULAR task forces improve the specificity and sensitivity of the current classification criteria by making them more detailed especially with regard to the type of organ damage that characterizes thrombosis in TAPS, something that would help physicians recognize and diagnose TAPS.They could also include other clinical manifestations now considered non-criteria such as nephropathy and cardiac valvulopathy in the ongoing classification criteria of TAPS and recommend the use of new laboratory tests such as anti-prothrombin-phosphatidylserine antibodies [39,40].A correct, timely diagnosis and appropriate treatment can make an important difference in saving lives and improving patients' prognoses.

Fig. 1
Fig. 1 Flowchart showing the article selection process

Table 1
List of the observational studies published between 1999 and 2022 focusing on the misdiagnosis of thrombotic antiphospholipid syndrome

Table 3
The clinical and laboratory features of patients who received a delayed diagnosis of thrombotic antiphospholipid syndrome between 1999 and 2022

Table 4
The clinical and laboratory features of misdiagnosed thrombotic antiphospholipid syndrome patients between 1999 and 2022

Table 5
The clinical and laboratory characteristics of patients who received a delayed diagnosis of thrombotic antiphospholipid syndrome and those who were misdiagnosed APS antiphospholipid syndrome, LAC lupus anticoagulant, IgG/IgM aCL immunoglobulin G/M anticardiolipin antibodies, IgG/IgM anti-β2GPI immunoglobulin G/M anti-β2Glycoprotein I antibodies * significant value