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Adherence to thrombophilia testing guidelines and its influence on anticoagulation therapy: A single-center cross-sectional study

Open AccessPublished:December 14, 2022DOI:https://doi.org/10.1016/j.thromres.2022.12.001

      Highlights

      • The therapeutic impact of recommended thrombophilia testing strategies is uncertain.
      • We conducted a cross-sectional study of 3686 patients during a 10-year period.
      • Thrombophilia work-up was adherent to guidelines only in 33 % tested patients.
      • Adherence was not associated with influence on anticoagulation treatment.
      • Testing in weakly provoked VTE was associated with prolonged anticoagulation.

      Abstract

      Introduction

      The collected evidence on thrombophilia guidelines is scarce and data about their impact on clinical decisions are unknown. We aimed to investigate the adherence to thrombophilia testing guidelines, its therapeutic impact in patients with guideline-adherent and non-adherent testing and identify the patients' clinical characteristics mostly associated with treatment decisions.

      Materials and methods

      We conducted a single-center cross-sectional study of patients referred for thrombophilia testing at the outpatient clinic of a tertiary hospital between 01/2010–10/2020. We systematically evaluated the adherence of thrombophilia testing to internal guidelines and the influence of test results on anticoagulation therapy. Using multivariable logistic regression, we evaluated the association between clinical characteristics and influence of thrombophilia tests on anticoagulation therapy in the entire cohort and by indication for referral.

      Results

      Of 3686 included patients, mostly referred for venous thromboembolism (2407, 65 %) or arterial thrombosis (591, 16 %), 3550 patients (96 %) underwent thrombophilia testing. Indication for testing was according to guidelines in 1208 patients (33 %). Test results influenced treatment decisions in 56 of 1102 work-ups (5.1 %) that were adherent to guidelines, and in 237 of 2448 (9.7 %) non-adherent work-ups (absolute difference, 4.3 %; 95 % confidence interval, 2.9–6.3 %). Age < 50 years, female sex, absence of risk factors and co-morbidities, weakly provoked venous thromboembolism and referral indication other than venous thromboembolism were associated with influence on anticoagulation therapy.

      Conclusions

      Adherence to guidelines for thrombophilia testing was poor and did not have an impact on treatment decisions. Refinement of selection criteria is needed to increase the therapeutic impact of thrombophilia testing.

      Keywords

      1. Introduction

      The clinical utility of testing for hereditary and acquired thrombophilia and the best selection criteria for testing remain uncertain. Because confirmation of high-risk thrombophilia influences treatment decisions to a bigger extent than confirmation of low-risk thrombophilia, identification of these patients is essential to obtain a high therapeutic yield of thrombophilia testing [
      • Moran J.
      • Bauer K.A.
      Managing thromboembolic risk in patients with hereditary and acquired thrombophilias.
      ]. Due to changing practice over time and inconsistent recommendations in current guidelines [
      • Stevens S.M.
      Role of thrombophilia testing: con.
      ,
      • Vrotniakaite-Bajerciene K.
      • Tritschler T.
      • Jalowiec K.A.
      • Broughton H.
      • Brodard J.
      • Porret N.A.
      • Haynes A.
      • Rovo A.
      • Kremer Hovinga J.A.
      • Aujesky D.
      • Angelillo-Scherrer A.
      Thrombophilia impact on treatment decisions, subsequent venous or arterial thrombosis and pregnancy-related morbidity: a retrospective single-center cohort study.
      ], the adherence to guidelines in clinical practice is variable [
      • Somma J.
      • Sussman I.I.
      • Rand J.H.
      An evaluation of thrombophilia screening in an urban tertiary care medical center: a "real world" experience.
      ,
      • Meyer M.R.
      • Witt D.M.
      • Delate T.
      • Johnson S.G.
      • Fang M.
      • Go A.
      • Clark N.P.
      Thrombophilia testing patterns amongst patients with acute venous thromboembolism.
      ].
      Baglin et al. reported the first clinical guidelines to test for thrombophilia, suggesting no role of unselected thrombophilia testing and discussing the role of testing in patients with a strong positive family history for venous thromboembolism (VTE) or recurrent VTE [
      • Baglin T.
      • Gray E.
      • Greaves M.
      • Hunt B.J.
      • Keeling D.
      • Machin S.
      • Mackie I.
      • Makris M.
      • Nokes T.
      • Perry D.
      • Tait R.C.
      • Walker I.
      • Watson H.
      Clinical guidelines for testing for heritable thrombophilia.
      ]. Subsequent guidelines proposed even more restrictive selection criteria suggesting a very limited role of testing in persons without prior VTE and in patients with pregnancy-related morbidity [
      • Chong L.-Y.
      • Fenu E.
      • Stansby G.
      • Hodgkinson S.
      Management of venous thromboembolic diseases and the role of thrombophilia testing: summary of NICE guidance.
      ,
      • Kearon C.
      • Akl E.A.
      • Comerota A.J.
      • Prandoni P.
      • Bounameaux H.
      • Goldhaber S.Z.
      • Nelson M.E.
      • Wells P.S.
      • Gould M.K.
      • Dentali F.
      • Crowther M.
      • Kahn S.R.
      Antithrombotic therapy for VTE disease: antithrombotic therapy and prevention of thrombosis, 9th ed: American College of Chest Physicians Evidence-Based Clinical Practice Guidelines.
      ,
      • Hicks L.K.
      • Bering H.
      • Carson K.R.
      • Kleinerman J.
      • Kukreti V.
      • Ma A.
      • Mueller B.U.
      • O'Brien S.H.
      • Pasquini M.
      • Sarode R.
      • Solberg Jr., L.
      • Haynes A.E.
      • Crowther M.A.
      The ASH choosing Wisely® campaign: five hematologic tests and treatments to question.
      ,
      • Hillis C.M.
      • Schimmer A.D.
      • Couban S.
      • Crowther M.A.
      The Canadian choosing wisely campaign: the Canadian Hematology Society's top five tests and treatments.
      ,
      • Stevens S.M.
      • Woller S.C.
      • Bauer K.A.
      • Kasthuri R.
      • Cushman M.
      • Streiff M.
      • Lim W.
      • Douketis J.D.
      Guidance for the evaluation and treatment of hereditary and acquired thrombophilia.
      ,
      • Carroll B.J.
      • Piazza G.
      Hypercoagulable states in arterial and venous thrombosis: when, how, and who to test?.
      ,
      • Ashraf N.
      • Visweshwar N.
      • Jaglal M.
      • Sokol L.
      • Laber D.
      Evolving paradigm in thrombophilia screening.
      ,
      • Konstantinides S.V.
      • Meyer G.
      • Becattini C.
      • Bueno H.
      • Geersing G.J.
      • Harjola V.P.
      • Huisman M.V.
      • Humbert M.
      • Jennings C.S.
      • Jiménez D.
      • Kucher N.
      • Lang I.M.
      • Lankeit M.
      • Lorusso R.
      • Mazzolai L.
      • Meneveau N.
      • Áinle F.N.
      • Prandoni P.
      • Pruszczyk P.
      • Righini M.
      • Torbicki A.
      • Van Belle E.
      • Zamorano J.L.
      2019 ESC guidelines for the diagnosis and management of acute pulmonary embolism developed in collaboration with the European Respiratory Society (ERS): the task force for the diagnosis and management of acute pulmonary embolism of the European Society of Cardiology (ESC).
      ,
      • Nicolaides A.N.
      • Fareed J.
      • Kakkar A.K.
      • Comerota A.J.
      • Goldhaber S.Z.
      • Hull R.
      • Myers K.
      • Samama M.
      • Fletcher J.
      • Kalodiki E.
      • Bergqvist D.
      • Bonnar J.
      • Caprini J.A.
      • Carter C.
      • Conard J.
      • Eklof B.
      • Elalamy I.
      • Gerotziafas G.
      • Geroulakos G.
      • Giannoukas A.
      • Greer I.
      • Griffin M.
      • Kakkos S.
      • Lassen M.R.
      • Lowe G.D.
      • Markel A.
      • Prandoni P.
      • Raskob G.
      • Spyropoulos A.C.
      • Turpie A.G.
      • Walenga J.M.
      • Warwick D.
      Prevention and treatment of venous thromboembolism–international consensus statement.
      ]. Guidance for thrombophilia in patients with arterial thrombosis is even more sparse [
      • Carroll B.J.
      • Piazza G.
      Hypercoagulable states in arterial and venous thrombosis: when, how, and who to test?.
      ,
      • May J.E.
      • Moll S.
      How I treat unexplained arterial thrombosis.
      ].
      The adherence to thrombophilia testing guidelines is low in acute hospital care and disciplines other than hematology [
      • Somma J.
      • Sussman I.I.
      • Rand J.H.
      An evaluation of thrombophilia screening in an urban tertiary care medical center: a "real world" experience.
      ,
      • Alidoost M.
      • Conte G.A.
      • Gupta V.
      • Patel S.
      • Patel I.
      • Shariff M.
      • Gor S.
      • Levitt M.J.
      • Asif A.
      • Hossain M.A.
      Trends of ordering hypercoagulability work-up at an academic medical center.
      ,
      • Cox N.
      • Johnson S.A.
      • Vazquez S.
      • Fleming R.P.
      • Rondina M.T.
      • Kaplan D.
      • Chauv S.
      • Fontaine G.V.
      • Stevens S.M.
      • Woller S.
      • Witt D.M.
      Patterns and appropriateness of thrombophilia testing in an academic medical center.
      ,
      • Virparia R.
      • Brunetti L.
      • Vigdor S.
      • Adams C.D.
      Appropriateness of thrombophilia testing in patients in the acute care setting and an evaluation of the associated costs.
      ]. Better adherence was shown for hematologists in a small retrospective study [
      • Kwon A.J.
      • Roshal M.
      • DeSancho M.T.
      Clinical adherence to thrombophilia screening guidelines at a major tertiary care hospital.
      ]. Previous studies mainly focus on adherence for thrombophilia testing in patients with VTE [
      • Somma J.
      • Sussman I.I.
      • Rand J.H.
      An evaluation of thrombophilia screening in an urban tertiary care medical center: a "real world" experience.
      ,
      • Alidoost M.
      • Conte G.A.
      • Gupta V.
      • Patel S.
      • Patel I.
      • Shariff M.
      • Gor S.
      • Levitt M.J.
      • Asif A.
      • Hossain M.A.
      Trends of ordering hypercoagulability work-up at an academic medical center.
      ], whereas studies of patients with arterial thrombosis or pregnancy-related morbidity, and asymptomatic persons with a family history of VTE are lacking.
      Therefore, we aimed to evaluate the adherence to thrombophilia testing guidelines for any type of indications, the influence of test results on subsequent guideline-recommended treatment decisions according to guideline adherence and investigate the patients' characteristics mostly associated with further anticoagulation treatment after the thrombophilia work-up in a cross-sectional study of patients referred to a tertiary thrombophilia center.

      2. Materials and methods

      2.1 Study design

      This cross-sectional study was conducted at the Department of Hematology of the Bern University Hospital in Switzerland. We included consecutive patients who were referred for testing of hereditary and/or acquired thrombophilia by general practitioners or non-hematologist medical specialists between January 2010 and October 2020. Patients medical records were systematically queried in the hospital database system with the support of the hospital data management service using specific internal codes for thrombophilia work-up. Patients were included if they had provided a general consent and a documented history of objectively confirmed VTE and/or arterial thrombosis in any location, a history of pregnancy-related morbidity or were referred for thrombophilia testing due to a positive family history for VTE or hereditary thrombophilia. The study was approved by the Ethics Commission of the Canton of Bern (ID 2019-02102).
      Standard imaging techniques were applied to diagnose a venous or arterial event [
      • Wilbur J.
      • Shian B.
      Diagnosis of deep venous thrombosis and pulmonary embolism.
      ,
      • Hasan T.F.
      • Rabinstein A.A.
      • Middlebrooks E.H.
      • Haranhalli N.
      • Silliman S.L.
      • Meschia J.F.
      • Tawk R.G.
      Diagnosis and management of acute ischemic stroke.
      ,
      • Anderson J.L.
      • Morrow D.A.
      Acute myocardial infarction.
      ,
      • Firnhaber J.M.
      • Powell C.S.
      Lower extremity peripheral artery disease: diagnosis and treatment.
      ,
      • Rajesh S.
      • Mukund A.
      • Arora A.
      Imaging diagnosis of splanchnic venous thrombosis.
      ,
      • Karaosmanoglu A.D.
      • Uysal A.
      • Akata D.
      • Ozmen M.N.
      • Karcaaltincaba M.
      Role of imaging in visceral vascular emergencies.
      ,
      • Ghoneim A.
      • Straiton J.
      • Pollard C.
      • Macdonald K.
      • Jampana R.
      Imaging of cerebral venous thrombosis.
      ]. Pregnancy-related morbidities were defined as pregnancy loss at all gestational ages, placenta failure, preeclampsia [
      • Davenport W.B.
      • Kutteh W.H.
      Inherited thrombophilias and adverse pregnancy outcomes: a review of screening patterns and recommendations.
      ] and HELLP syndrome (hemolysis, elevated liver enzyme levels, low platelet count) according to diagnostic criteria of obstetricians and gynecologists [
      • Gupta M.
      • Feinberg B.B.
      • Burwick R.M.
      Thrombotic microangiopathies of pregnancy: differential diagnosis.
      ]. Categorization as minor and major provoking risk factors of VTE was based on criteria provided by the International Society on Thrombosis and Haemostasis (ISTH) [
      • Kearon C.
      • Ageno W.
      • Cannegieter S.C.
      • Cosmi B.
      • Geersing G.J.
      • Kyrle P.A.
      Categorization of patients as having provoked or unprovoked venous thromboembolism: guidance from the SSC of ISTH.
      ]. In addition to the ISTH based criteria, the presence of intravenous catheter [
      • Lu H.
      • Yang Q.
      • Yang L.
      • Qu K.
      • Tian B.
      • Xiao Q.
      • Xin X.
      • Lv Y.
      • Zheng X.
      The risk of venous thromboembolism associated with midline catheters compared with peripherally inserted central catheters: a systematic review and meta-analysis.
      ] or May-Thurner syndrome (>70 % iliofemoral compression) [
      • Harbin M.M.
      • Lutsey P.L.
      May-thurner syndrome: history of understanding and need for defining population prevalence.
      ] were categorized as major risk factors, whereas immobilization >4 h [
      • Chandra D.
      • Parisini E.
      • Mozaffarian D.
      Meta-analysis: travel and risk for venous thromboembolism.
      ] and heavy smoking (>20 pack years) [
      • Enga K.F.
      • Braekkan S.K.
      • Hansen-Krone I.J.
      • le Cessie S.
      • Rosendaal F.R.
      • Hansen J.B.
      Cigarette smoking and the risk of venous thromboembolism: the Tromsø Study.
      ] as minor risk factors. VTE in the presence of merely an environmental risk factor (male sex and older age) was categorized as unprovoked thromboembolism.
      Objective clinical data were retrospectively collected from structured electronic forms using a standardized case report form and entered in a computerized database (REDCap software) by two persons. In case of a disagreement, a third person was included to reconcile. Data comprised demographic characteristics of patients (age, sex) and their family history for VTE in first- and second-degree relative, details of all previous thrombotic events or pregnancy-related morbidity (date and location), risk factors of most recent VTE and arterial thrombosis (heavy smoking [>20 pack years], immobilization >4 h, infections requiring bedrest >3 days, estrogen-based medications, pregnancy and peripartum period, intravenous catheters, active cancer, obesity [body mass index (BMI) > 30 kg m−2], trauma, surgery, cancer medication, presence of extensive varicose veins, patent foramen ovale or other septal defect) and co-morbidities (diabetes mellitus, arterial hypertension, liver cirrhosis, kidney failure, rheumatic disease, depression, chronic inflammatory disease, dyslipidemia, cardiovascular diseases, pulmonary diseases, neurological diseases).

      2.2 Thrombophilia testing

      Thrombophilia testing was performed standardly between 3 and 6 months following the index event after the evaluation of the patient by a hematologist taking into consideration age, risk factors, family history of VTE, co-morbidities and type of thromboembolism or pregnancy-related morbidity. A thrombophilia work-up was considered as “performed”, if one or more of the following thrombophilia parameters were tested: factor V Leiden (FVL) mutation status, prothrombin gene G20210A polymorphism status, protein C (PC) and S (PS) as well as antithrombin (AT) levels, lupus anticoagulant (LA), anticardiolipin antibodies, or anti-β2-glycoprotein I antibodies, which were usually ordered as a standard panel.
      Testing for PC (Protein C Berichrom®, Siemens; Protein C COAG, Siemens), PS (Free protein S, Asserachrom®, Diagnostica Stago from 2010 to 2015; Free Protein S Antigen, Innovance®, Siemens from 2015 to 2020), AT activity (LR Antithrombin, Coamatic®, Diapharma from 2010 to 2013; LRT Antithrombin, Biophen®, Endotell from 2013 to 2014 and Antithrombin Innovance®, Siemens from 2014 to 2020) was performed in the routine hemostasis laboratory (Bern University Hospital) and values below 70 %, 59 % and 69 % were considered as AT, PC and PS deficiencies, respectively. Antiphospholipid antibodies were tested using Varelisa diagnostic kits (Phadia®, ThermoFisher) from 2010 to 2014, fluorescence enzyme immunoassay (Phadia® 250, ThermoFisher) from 2014 to 2015 and automated chemiluminescence assay (Bio-flash®, Inova Diagnostics) from 2015 to 2020 and dilute Russell's viper venom time (Cryocheck®, Endotell). The diagnosis of an antiphospholipid antibody syndrome was established by persistent laboratory evidence of antiphospholipid antibodies at least 12 weeks later and presence of VTE, arterial thrombosis or criteria pregnancy-related morbidity [
      • Miyakis S.
      • Lockshin M.D.
      • Atsumi T.
      • Branch D.W.
      • Brey R.L.
      • Cervera R.
      • Derksen R.H.
      • PG D.E.G.
      • Koike T.
      • Meroni P.L.
      • Reber G.
      • Shoenfeld Y.
      • Tincani A.
      • Vlachoyiannopoulos P.G.
      • Krilis S.A.
      International consensus statement on an update of the classification criteria for definite antiphospholipid syndrome (APS.
      ]. Genetic mutations were detected by polymerase chain reaction method (FVL and Prothrombin, RealFast Assay®, Vienna Lab Diagnostics).

      2.3 Adherence to guidelines and accuracy

      Two persons working in the field of hematology evaluated the adherence to testing guidelines separately in all study participants in accordance to institutional recommendations, which were implemented at the center in 2014 and are fully or partially based on international guidance of British Society for Hematology [
      • Baglin T.
      • Gray E.
      • Greaves M.
      • Hunt B.J.
      • Keeling D.
      • Machin S.
      • Mackie I.
      • Makris M.
      • Nokes T.
      • Perry D.
      • Tait R.C.
      • Walker I.
      • Watson H.
      Clinical guidelines for testing for heritable thrombophilia.
      ,
      • Tait C.
      • Baglin T.
      • Watson H.
      • Laffan M.
      • Makris M.
      • Perry D.
      • Keeling D.
      Guidelines on the investigation and management of venous thrombosis at unusual sites.
      ,
      • Keeling D.
      • Mackie I.
      • Moore G.W.
      • Greer I.A.
      • Greaves M.
      • Haematology B.C.F.S.I.
      Guidelines on the investigation and management of antiphospholipid syndrome.
      ], American Society of Hematology [
      • Hicks L.K.
      • Bering H.
      • Carson K.R.
      • Kleinerman J.
      • Kukreti V.
      • Ma A.
      • Mueller B.U.
      • O'Brien S.H.
      • Pasquini M.
      • Sarode R.
      • Solberg Jr., L.
      • Haynes A.E.
      • Crowther M.A.
      The ASH choosing Wisely® campaign: five hematologic tests and treatments to question.
      ], International Consensus Statement [
      • Nicolaides A.N.
      • Fareed J.
      • Kakkar A.K.
      • Comerota A.J.
      • Goldhaber S.Z.
      • Hull R.
      • Myers K.
      • Samama M.
      • Fletcher J.
      • Kalodiki E.
      • Bergqvist D.
      • Bonnar J.
      • Caprini J.A.
      • Carter C.
      • Conard J.
      • Eklof B.
      • Elalamy I.
      • Gerotziafas G.
      • Geroulakos G.
      • Giannoukas A.
      • Greer I.
      • Griffin M.
      • Kakkos S.
      • Lassen M.R.
      • Lowe G.D.
      • Markel A.
      • Prandoni P.
      • Raskob G.
      • Spyropoulos A.C.
      • Turpie A.G.
      • Walenga J.M.
      • Warwick D.
      Prevention and treatment of venous thromboembolism–international consensus statement.
      ], The National Institute for Health and Care Excellence (NICE) [
      • Chong L.-Y.
      • Fenu E.
      • Stansby G.
      • Hodgkinson S.
      Management of venous thromboembolic diseases and the role of thrombophilia testing: summary of NICE guidance.
      ], Anticoagulation Forum of North America [
      • Stevens S.M.
      • Woller S.C.
      • Bauer K.A.
      • Kasthuri R.
      • Cushman M.
      • Streiff M.
      • Lim W.
      • Douketis J.D.
      Guidance for the evaluation and treatment of hereditary and acquired thrombophilia.
      ], European Society of Cardiology [
      • Konstantinides S.V.
      • Meyer G.
      • Becattini C.
      • Bueno H.
      • Geersing G.J.
      • Harjola V.P.
      • Huisman M.V.
      • Humbert M.
      • Jennings C.S.
      • Jiménez D.
      • Kucher N.
      • Lang I.M.
      • Lankeit M.
      • Lorusso R.
      • Mazzolai L.
      • Meneveau N.
      • Áinle F.N.
      • Prandoni P.
      • Pruszczyk P.
      • Righini M.
      • Torbicki A.
      • Van Belle E.
      • Zamorano J.L.
      2019 ESC guidelines for the diagnosis and management of acute pulmonary embolism developed in collaboration with the European Respiratory Society (ERS): the task force for the diagnosis and management of acute pulmonary embolism of the European Society of Cardiology (ESC).
      ] and American College of Obstetricians and Gynecologists (ACOG) [
      • ACOG Practice Bulletin No
      197: inherited thrombophilias in pregnancy: correction.
      ] (Table 1). The 2020 NICE guidelines for VTE [
      • McCormack T.
      • Harrisingh M.C.
      • Horner D.
      • Bewley S.
      Venous thromboembolism in adults: summary of updated NICE guidance on diagnosis, management, and thrombophilia testing.
      ] were not considered in determining the selection criteria, because they were published after initiation of the study. If study criteria in Table 1 were not met, thrombophilia testing was considered as not adherent to guidelines.
      Table 1Comparison of international thrombophilia testing guidelines and criteria used in this study.
      Table thumbnail fx1
      Red indicates no role of thrombophilia testing; green indicates unselective testing; blue indicates only selective testing; grey indicates no statement. Abbreviations: ACF, Anticoagulation Forum; ACOG, The American College of Obstetricians and Gynecologists; ASH, The American Society of Hematology; BCSH, The British Society of Hematology; ESC, The European Society of Cardiology; ISC, International Consensus Statement; NICE, The National Institute for Health and Care Excellence; y., years; VTE, venous thromboembolism.
      Measurement of PC and PS whilst on vitamin K antagonists or liver disease (Child-Pugh-Score B and C), PS in puerperium time or during estrogen-based treatment or thrombophilia work-up for PC, PS and AT in the acute phase (within 3 months of index event) was considered inaccurate [
      • Stevens S.M.
      • Woller S.C.
      • Bauer K.A.
      • Kasthuri R.
      • Cushman M.
      • Streiff M.
      • Lim W.
      • Douketis J.D.
      Guidance for the evaluation and treatment of hereditary and acquired thrombophilia.
      ] and classified as a negative result.

      2.4 Influence of thrombophilia testing on treatment decisions

      For the entire study period, the thrombophilia center at the Bern University Hospital used a structured report form to document thrombophilia testing and subsequent treatment decisions including a description of diagnosis, thrombophilia test results, risk factors, therapy decision, and discussion. Based on this report, an experienced hematologist determined for each patient whether decision on the length of anticoagulation treatment and initiation or stopping the prophylactic or therapeutic anticoagulation were based on the thrombophilia test results or merely on clinical aspects. This decision was based on international guidelines, discussing both aspects of clinical-decision making [
      • Stevens S.M.
      • Woller S.C.
      • Bauer K.A.
      • Kasthuri R.
      • Cushman M.
      • Streiff M.
      • Lim W.
      • Douketis J.D.
      Guidance for the evaluation and treatment of hereditary and acquired thrombophilia.
      ,
      • May J.E.
      • Moll S.
      How I treat unexplained arterial thrombosis.
      ,
      • ACOG Practice Bulletin No
      197: inherited thrombophilias in pregnancy: correction.
      ,
      • Schünemann H.J.
      • Cushman M.
      • Burnett A.E.
      • Kahn S.R.
      • Beyer-Westendorf J.
      • Spencer F.A.
      • Rezende S.M.
      • Zakai N.A.
      • Bauer K.A.
      • Dentali F.
      • Lansing J.
      • Balduzzi S.
      • Darzi A.
      • Morgano G.P.
      • Neumann I.
      • Nieuwlaat R.
      • Yepes-Nuñez J.J.
      • Zhang Y.
      • Wiercioch W.
      American Society of Hematology 2018 guidelines for management of venous thromboembolism: prophylaxis for hospitalized and nonhospitalized medical patients.
      ] (Table 2). For the purpose of this study, exclusively thrombophilia testing-based treatment decisions regarding prophylactic and therapeutic anticoagulation treatment were considered, excluding other related advices, such as avoidance of estrogen-based treatment, the change of the type of anticoagulant or life style modification.
      Table 2Classification of thrombophilia result influence on treatment decisions.
      No influence on managementAnticoagulation therapy or prophylaxis should have been initiated or stopped, continued or discontinued irrespective of thrombophilia testing result based on:

      • Type of thromboembolic event (i.e., extended anticoagulation in unprovoked or recurrent venous thromboembolism, short-term anticoagulation in provoked venous thromboembolism).
      • Family or personal history for venous thromboembolism (i.e., prophylactic anticoagulation in risk situations).
      Influence on managementDecision to continue, stop or initiate prophylactic or therapeutic anticoagulation was based/should have been based on a thrombophilia testing result

      • Prolonged anticoagulation in provoked venous thromboembolism or arterial thrombosis due to high-risk thrombophilia.
      • Prophylactic anticoagulation in risk situations in asymptomatic family members or pregnancy-related morbidity merely due to a positive thrombophilia result.
      • Termination of anticoagulation in case of a negative thrombophilia work-up result

      2.5 Statistical analysis

      Continuous and categorical variables were compared using unpaired ANOVA test and χ2 test, respectively. The associations between clinical characteristics and treatment influence was evaluated using univariable and multivariable logistic regression models in the entire cohort and in subgroup analysis by indication for referral (i.e., patients with VTE, arterial thrombosis, or pregnancy-related morbidity, or asymptomatic persons with family history of VTE). We adjusted the logistic regression models for recurrent VTE and arterial thrombosis in the analysis including the entire study cohort; recurrent and unprovoked VTE in the analysis including only patients with VTE; age > 50 years and presence of ≥2 thrombotic risk factors in the analysis including only patients with arterial thrombosis; family history of VTE in a first-degree relative and female sex in the analysis including only asymptomatic patients; and family history of VTE in a first-degree relative and presence of ≥2 thrombotic risk factors in the analysis including only pregnancy-related morbidity. Only complete case analysis was performed, without an attempt to replace missing values with imputation methods. A p-value < 0.05 was considered statistically significant. All analyses were performed with R, version 4.1.1, and figures were edited with GraphPad Prism, version 9.1.2.

      3. Results

      3.1 Study population and prevalence of thrombophilia

      A total of 5064 patients were screened for eligibility, of which we excluded 1356 patients due to lack of general consent and 22 persons because VTE, arterial thrombosis or pregnancy-related morbidity was not objectively confirmed, leaving a final study sample of 3686 participants.
      Patient characteristics at the time of thrombophilia work-up are shown in Table 3. Overall, 2218 patients (60 %) were women and median age was 44 years (standard deviation, ±16). Most patients were referred for VTE (2407 patients, 65 %), mainly DVT and/or PE (1840/2407 patients, 76 %), or unexplained arterial thrombosis (591 patients, 16 %), mainly stroke (446/591 patients, 75 %) (Supplementary Table 1). A total of 567 participants (15 %) had no prior thromboembolic event or pregnancy-related morbidity, but a positive family history for VTE in first-degree (341/567 persons, 60 %) or second-degree (165/567 persons, 29 %) family members. Few referrals were for pregnancy-related morbidity (121 patients, 3.3 %). Most patients (56 %) had no documented co-morbidity and about a third of patients no documented risk factor for thromboembolism (Table 3).
      Table 3Clinical characteristics of patients by adherence to thrombophilia testing guidelines.
      CharacteristicsTotal

      N = 3686
      No adherence to guidelines

      n = 2478 (67)
      Adherence to guidelines

      n = 1208 (33)
      P
      Age at consultation, mean ± SD (years)44 (16)47 (16)37 (13)<0.001
      Women, n (%)2218 (60)1443 (58)775 (64)<0.001
      Indication for consultation, n (%)<0.001
       VTE2407 (65)1611 (65)796 (66)
       Arterial thrombosis591 (16)449 (18)142 (12)
       Asymptomatic patients567 (15)334 (13)233 (19)
       Pregnancy-related morbidity121 (3.3)84 (3.4)37 (3.1)
      Provoking factors of VTE§, n (%)<0.001
       Unprovoked702 (19)415 (17)287 (24)
       Minor risk factor1267 (34)820 (33)447 (37)
       Major risk factor435 (12)373 (15)62 (5.1)
       Recurrent VTE§, n (%)589 (16)320 (13)269 (22)<0.001
       Family history of VTE in first-degree relative§, n (%)1159 (31)598 (24)561 (46)<0.001
      Number of co-morbidities*, n (%)<0.001
       02078 (56)1287 (52)791 (65)
       1841 (23)608 (25)233 (19)
       ≥2767 (21)583 (24)184 (15)
      Number of risk factors*, n (%)<0.001
       01323 (36)845 (34)478 (40)
       11219 (33)808 (33)411 (34)
       ≥21144 (31)825 (33)319 (26)
      Positive thrombophilia test result, n (%)1192 (32)768 (31)424 (35)<0.001
      Categorical values are compared by x2 test and continuous variables by ANOVA test. *At time of VTE, arterial thrombosis, pregnancy-related morbidity, or consultation in asymptomatic patients. Risk factors included smoking, immobilization >4 h, cancer, central intravenous catheter, infection, estrogen-based treatment, pregnancy, cancer, obesity, trauma, surgery, cancer, and its medication. Co-morbidities included diabetes, arterial hypertension, liver cirrhosis, kidney failure, rheumatic diseases, depression, dyslipidemia, lung diseases, neurological disorders, cardiovascular diseases, and chronic inflammatory diseases. §Values were missing for provoking factors of VTE (0.1 %), history of prior VTE at time of consultation (0.9 %), family history of VTE in first-degree relatives (1.3 %). Abbreviations: SD, standard deviation; VTE, venous thromboembolism.
      A total of 1756 patients (48 %) were referred to the thrombophilia center up to 2014 and 1930 (52 %) from 2014 to 2020. In 3550 patients (96 %) a partial or full thrombophilia work-up was performed (in 97 % of referred patients before 2014 and 96 % from 2014 to 2020). Female patients, asymptomatic patients, or patients with VTE provoked by a major risk factor were less likely to be tested (Supplementary Table 2). A total of 1260 thrombophilias were found in 1192 (34 %) patients. The most common type of hereditary thrombophilia was heterozygous FVL mutation (714 patients, 20 %), followed by heterozygous prothrombin G20210A mutation (193 patients, 5 %) and antiphospholipid antibody syndrome (119 patients, 3 %) (Table 4).
      Table 4Positive results of thrombophilia in adherent and not adherent work-ups.
      Type of thrombophilia, n (%)Total tested patients

      N = 3550
      No adherence to guidance

      N = 2448 (69)
      Adherence to guidance

      N = 1102 (31)
      P
      Factor V Leiden heterozygous mutation714 (20)455 (19)259 (23)<0.001
      Prothrombin G20210A heterozygous mutation193 (5.4)136 (5.6)57 (5.2)0.31
      Antiphospholipid antibody syndrome119 (3.3)77 (3.1)43 (3.6)0.41
      Protein S deficiency, <59 %101 (2.8)58 (2.4)43 (3.9)0.007
      Antithrombin deficiency, <70 %52 (1.5)35 (1.4)17 (1.5)0.70
      Factor V Leiden homozygous mutation48 (1.4)29 (1.2)19 (1.7)<0.001
      Protein C deficiency, <69 %28 (0.79)19 (0.78)9 (0.82)0.85
      Prothrombin G20210A homozygous mutation5 (0.14)2 (0.08)3 (0.27)0.31
      Categorical values are compared by x2 test. Testing was not performed or missing for factor V Leiden mutation (6 %), prothrombin G20210A mutation (13 %), antithrombin (20 %), protein C (30 %), protein S (29 %), antiphospholipid syndrome (11 %).

      3.2 Adherence to testing recommendations and accuracy of the work-up

      In 1208 patients (33 %) the indication for thrombophilia testing was adherent to guidelines (32 % from 2010 to 2014, 34 % from 2014 to 2020). Adherence was more likely in younger patients, women, patients without risk factors or co-morbidities, and those with recurrent VTE or a positive family history of VTE in first-degree relatives (Table 3). Non-adherence was more likely in patients with arterial thrombosis or VTE in uncommon locations (Supplementary Table 1). Overall, patients with guideline-adherent testing were more likely to be tested positive for any thrombophilia (Table 3). However, this association was limited to patients with heterozygous and homozygous FVL mutation and PS deficiency (Table 4). A total of 3325/3550 thrombophilia work-ups (94 %) were accurate, whereas 79 (2 %) were performed, whilst on VKA and 59 (2 %) on estrogen-based treatment or in pregnancy. Accuracy could not be determined in 87 patients (2 %).

      3.3 Influence of thrombophilia testing on treatment decisions

      A total of 293 test results (8.3 % of performed testing, 23 % of positive results for thrombophilia) influenced anticoagulation therapy (Supplemental Table 3). Test results influenced treatment in 56 of 1102 work-ups (5.1 %) that were adherent to guidelines, and in 237 of 2448 (9.7 %) non-adherent work-ups (absolute difference, 4.3 %; 95 % confidence interval, 2.9–6.3) (Fig. 1). Non-adherent testing was associated with influence on anticoagulation treatment compared to adherent testing (adjusted odds ratio 2.59, 95 % confidence interval, 1.89–3.56).
      Fig. 1
      Fig. 1Thrombophilia testing influence on therapy and adherence to guidance.
      A. Thrombophilia testing influence on therapy decision. B. Clinical characteristics of patients with non-adherent work-up with clinical utility. A total 12 work-ups could not be categorized due to unclear statement on treatment decision in the clinical report. Low-risk hereditary thrombophilia is defined by the presence of heterozygous factor V Leiden, heterozygous prothrombin G20210A mutation; high-risk hereditary thrombophilia comprises homozygous factor V Leiden, homozygous prothrombin G20210A mutation, antithrombin < 70 %, protein C < 69 %, protein S < 59 %, and compound thrombophilias. Abbreviations: ATE, arterial thrombosis; VTE, venous thromboembolism.
      Age < 50 years, female sex, absence of co-morbidities and risk factors, and referral for arterial thrombosis, pregnancy-related morbidity or asymptomatic persons with family history of VTE were associated with influence on anticoagulation therapy (Table 5). However, most of these associations were absent in subgroup analyses by indication for referral. When stratified by indication for referral, VTE provoked by a minor risk factor and age < 50 years were associated with influence on treatment decisions in patients referred for VTE. None of the evaluated clinical characteristics was associated with influence on anticoagulation therapy in patients with arterial thrombosis or pregnancy related-morbidity, or in asymptomatic persons with a family history of VTE (Table 5).
      Table 5Association between clinical characteristics and influence of thrombophilia testing on anticoagulation treatment in the full study cohort and by indication for consultation.
      Clinical characteristicCrude OR (95 % CI)Adjusted OR (95 % CI)
      Full cohort (n = 3686)
       Age < 50 years2.20 (1.64–2.95)2.20 (1.64–2.95)
       Women1.92 (1.47–2.51)1.92 (1.47–2.51)
       Indication for consultation
      VTE1 (ref)1 (ref)
      Asymptomatic patients7.09 (5.11–9.84)7.12 (5.12–9.90)
      Arterial thrombosis6.60 (4.76–9.17)6.60 (4.75–9.16)
      Pregnancy-related morbidity15.73 (9.95–24.86)15.80 (9.98–24.99)
       Number of co-morbidities
      ≥21 (ref)1 (ref)
      11.33 (0.89–1.98)1.32 (0.89–1.97)
      01.61 (1.15–2.26)1.61 (1.14–2.26)
       Number of risk factors for thromboembolism
      ≥21 (ref)1 (ref)
      12.10 (1.49–2.96)2.10 (1.49–2.96)
      02.42 (1.74–3.38)2.42 (1.73–3.37)
      Patients with VTE (n = 2407)
       Age < 50 years1.93 (1.12–3.33)1.73 (1.01–3.01)
       Female1.84 (1.08–3.12)1.60 (0.93–2.76)
       Risk factors for VTE
      Unprovoked1 (ref)1 (ref)
      Minor risk factor2.90 (1.46–5.75)2.94 (1.48–5.84)
      Major risk factor0.97 (0.35–2.69)0.97 (0.35–2.69)
      Family history of VTE in first-degree relative1.14 (0.67–1.92)1.15 (0.68–1.94)
      Recurrent VTE0.45 (0.22–0.91)0.49 (0.24–0.99)
       Number of co-morbidities
      ≥21 (ref)1 (ref)
      11.02 (0.43–2.43)1.07 (0.45–2.54)
      01.78 (0.89–3.56)1.86 (0.93–3.72)
      Patients with arterial thrombosis (n = 591)
       Age < 50 years1.04 (0.66–1.63)1.01 (0.64–1.60)
       Female1.17 (0.75–1.82)1.31 (0.83–2.07)
       Family history of VTE in first-degree relative0.79 (0.44–1.44)0.81 (0.44–1.48)
       Recurrent arterial thrombosis0.90 (0.50–1.61)0.94 (0.51–1.73)
       Number of risk factors for thromboembolism
      ≥21 (ref)1 (ref)
      12.39 (1.33–4.29)1.41 (0.83–2.40)
      01.69 (0.87–3.28)1.23 (0.71–2.13)
       Number of co-morbidities
      ≥21 (ref)1 (ref)
      11.34 (0.79–2.29)1.24 (0.71–2.16)
      01.14 (0.66–1.97)1.01 (0.56–1.80)
       Asymptomatic patients (n = 567)
      Age < 35 years1.77 (1.04–3.02)1.67 (0.93–2.99)
      Women2.06 (1.03–4.12)1.81 (0.85–3.86)
       Family history of VTE in first-degree relative0.06 (0.03–0.11)0.06 (0.03–0.11)
       Number of risk factors for thromboembolism
      ≥21 (ref)1 (ref)
      12.25 (0.64–7.92)1.38 (0.35–5.43)
      02.80 (0.84–9.34)2.02 (0.55–7.47)
       Number of co-morbidities
      ≥21 (ref)1 (ref)
      15.97 (0.76–47.13)6.79 (0.81–57.05)
       05.46 (0.73–40.81)6.16 (0.78–48.88)
      Women with pregnancy-related morbidity (n = 121)
       Age < 30 years0.75 (0.34–1.65)0.76 (0.32–1.76)
       Family history of VTE in first-degree relative0.12 (0.03–0.52)0.13 (0.03–0.61)
       Number of risk factors for thromboembolism
      ≥21 (ref)1 (ref)
      10.38 (0.06–2.29)0.38 (0.06–2.29)
      01.56 (0.39–6.32)1.56 (0.39–6.32)
       Number of co-morbidities
      ≥21 (ref)1 (ref)
      10.23 (0.02–2.37)0.89 (0.13–6.27)
      00.75 (0.12–4.71)0.21 (0.02–2.43)
      Influence of clinical characteristics was calculated by logistic regression model. Models were adjusted for recurrent venous and arterial thromboembolism in the entire cohort; recurrent and unprovoked VTE in patients with VTE; age > 50 years and presence of ≥2 risk factors in patients with arterial thrombosis; family history for VTE in a first-degree relative and female sex in asymptomatic patients; family history of VTE in a first-degree relative and presence of ≥2 risk factors in pregnancy-related morbidity. Risk factors included smoking, immobilization >4 h, cancer, central intravenous catheter, infection, estrogen-based treatment, pregnancy, cancer, obesity, trauma, surgery, cancer, and its medication. Co-morbidities included diabetes, arterial hypertension, liver cirrhosis, kidney failure, rheumatic diseases, depression, dyslipidemia, lung diseases, neurological disorders, cardiovascular diseases, and chronic inflammatory diseases. Abbreviations: CI, confidence interval; OR, odds ratio; VTE, venous thromboembolism.

      4. Discussion

      In this large single-center, cross-sectional cohort study of 3686 patients, the indication for thrombophilia testing was in accordance with guidelines in only 33 % of patients, which did not improve after implementation of internal guidelines. Adherence to guidelines partially improved the diagnostic, but not the therapeutic yield of testing. In contrast, test results in patients in whom testing was adherent to guidelines were less likely to influence treatment decisions than those in patients with non-adherent testing (absolute difference, 5.4 %). Whilst age < 50 years, female sex, absence of co-morbidities and risk factors, and referral for arterial thrombosis, pregnancy-related morbidity, or asymptomatic persons with family history of VTE were associated with influence on treatment decision in the entire cohort, these associations disappeared in subgroup analysis by indication for referral. The only clinical characteristics that were associated with treatment influence in subgroup analysis by indication for referral was VTE provoked by a minor risk factor and age < 50 years in patients that were referred for VTE.
      Our cohort comprised mostly younger patients without comorbidities and more women than men, probably due to frequent referral for family planning and primary infertility. Most studies on adherence to thrombophilia guidelines were conducted in a primary care setting and more frequently included patients with pre-existing co-morbidities or major risk factors than our study [
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      ] and was found to be poor. To our knowledge, this is the first study reporting data from a tertiary hematology center comprising all types of indications for consultation. Our study showed similar results with adherence to guidelines in only one-third of work-ups. Considering that 96 % of cohort patients were tested, our center did not carefully select patients that were referred by general practitioners or other specialists. Further, no significant effect on adherence was found after implementing the institutional selection criteria in 2014. Requests of patients themselves to be tested and expectations of referring physician may have likely contributed to this finding, in addition to physicians' preference and knowledge. Although better established local guidelines and implementation of clear selection criteria have been shown to improve adherence at other centers [
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      Because testing was performed in 96 % of patients, we were able to assess the influence of testing on treatment decision in a rather unselected population. Overall, only 8 % of thrombophilia tests and 25 % of positive results were relevant for determining further anticoagulation treatment. Surprisingly, the proportion of test with influence on treatment was higher in patients in whom testing was not adherent to current guidelines (absolute difference, 5.4 %), which highlights the limited clinical utility of current selection criteria, especially in women with pregnancy-related morbidity, asymptomatic patients and patients with arterial thrombosis, where only a very restricted testing is suggested [
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      ]. In our study, only age < 50 years and presence of a minor risk factor in patients with VTE was associated with influence on treatment decision, which highlights the discordance between the selection criteria aiming at diagnostic or therapeutic yield of thrombophilia work-up. This finding corresponds with only few guidelines [
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      • Griffin M.
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      • Spyropoulos A.C.
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      Prevention and treatment of venous thromboembolism–international consensus statement.
      ], suggesting testing in young patients with weakly provoked VTE, if discontinuation of anticoagulation at 3–6 months is planned, and highlights the importance of the therapy-related testing.
      Our study has also limitations. First, we retrospectively determined treatment influence, which may be prone to information bias and possible misclassification of the influence of thrombophilia testing on treatment decisions. However, the structured reporting system and testing pattern during the entire study period and cross-validation of the data by two individuals likely limited missing values and random misclassification. Second, due to the inhomogeneous selection criteria worldwide and a large study time span, internal selection criteria were used as a reference to the status of adherence, limiting the generalizability of the study results. However, due to a large study population, comparable work-up rate and adherence over the years and clearly defined selection criteria, which are largely consistent with published guidelines, the study gives a comprehensive insight regarding the adherence to thrombophilia testing and its therapeutic yield. Third, we did not consider any other impact on treatment than anticoagulation therapy, such as the avoidance of estrogen-related medications, possible indication to substitution therapy in case of PC and AT deficiency, higher motivation to life style modification, and the change of type of anticoagulant, which would have resulted in higher impact and usefulness of thrombophilia testing. Nevertheless, the accumulated experience through this systematic analysis reflects most of the general influence and constitutes a solid basis of the next testing strategies to be discussed. Fourth, a very small proportion of patients with PS type II deficiency might have been missed, because no systematic measurement of PS activity was performed [
      • Marlar R.A.
      • Gausman J.N.
      • Tsuda H.
      • Rollins-Raval M.A.
      • Brinkman H.J.M.
      Recommendations for clinical laboratory testing for protein S deficiency: communication from the SSC committee plasma coagulation inhibitors of the ISTH.
      ]. Moreover, some AT type II defects may be detected only by genetic testing (AT Dublin, AT Wibble, and AT Rouen VI) or detected more accurately with anti-IIa-based activity assays (AT Cambridge II, AT Denver, and AT Stockholm) [
      • Van Cott E.M.
      • Orlando C.
      • Moore G.W.
      • Cooper P.C.
      • Meijer P.
      • Marlar R.
      Recommendations for clinical laboratory testing for antithrombin deficiency; communication from the SSC of the ISTH.
      ], which was not performed in our center. However, the prevalence of these types of thrombophilias is generally very low, reducing the impact of this limitation.
      In conclusion, adherence to thrombophilia testing guidelines was poor in our tertiary thrombophilia center. Since thrombophilia testing was less likely to influence the treatment decisions in patients in whom testing adhered to guidelines than in those in whom testing would not have been indicated, better criteria to improve the therapeutic yield of thrombophilia work-up are needed. A more comprehensive testing in patients <50 years old and in weakly provoked VTE should be further explored.

      CRediT authorship contribution statement

      K.V.B. and A.A.S.: Designed the protocol and the analysis plan. K.V.B., T.T. and A.A.S.: Interpreted the data and drafted the manuscript. A.H.: Performed the statistical analysis. H.B and F.S.: Under the supervision of K.V.B. collected the data. D.A, A.R, J.A.K.H., J.S.S. and K.A.J.: Intellectually reviewed the manuscript.

      Declaration of competing interest

      The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: T.T. is an investigator of the CanVECTOR Network; the Network receives grant funding from the Canadian Institutes of Health Research (CDT-142654). The remaining authors declare no competing financial interests. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

      Appendix A. Supplementary data

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