Ordering Patterns of Thrombophilia Testing and Its Appropriateness

Abstract Background Despite society recommendations to limit thrombophilia testing, this testing is often sent inappropriately. The results of thrombophilia testing frequently do not affect management. Additionally, interpretation of thrombophilia testing is confounded by acute thrombosis, anticoagulation (AC) therapy, and other medical comorbidities. Incorrect test selection is also a source of unnecessary testing [i.e. Factor V (FV) activity level instead of Factor V Leiden (FVL)]. The aim of our study was to assess ordering patterns for thrombophilia testing by qualifying the number of tests, identifying the requesting services, and assessing the appropriateness of testing. Methods This was a retrospective study of thrombophilia testing performed at LAC+USC Medical Center, Los Angeles, CA from January 1, 2019 to December 31, 2019. A laboratory query of thrombophilia testing was performed to identify eligible adult patients who received thrombophilia testing without a prior confirmed thrombophilia. Thrombophilia testing included: FVL and prothrombin 20210 gene mutations; activated protein C (APC) resistance; antithrombin, or protein C or S activity levels; and antiphospholipid syndrome (APS) evaluation with lupus anticoagulant, cardiolipin (CL) immunoglobulins IgM/G, and beta-2 glycoprotein (b2gp) IgM/IgG; and JAK2 V617F mutation. Homocysteine (HC) levels and methylenetetrahydrofolate reductase (MTHFR) gene mutation testing were considered to have limited clinical utility. FV activity and phosphatidylserine IgM/IgG were considered incorrect tests. The electronic medical record was reviewed for clinical history, indication for testing, requesting service, and appropriateness. The criteria for defining appropriateness were determined based on major society guidelines and literature review. The main criteria are summarized here. Testing was considered inappropriate for a provoked venous thromboembolism (VTE) or stroke/transient ischemic attack (TIA). For unprovoked VTE, testing was considered inappropriate for patients > 45 yo except for APS testing. For non-stroke arterial thrombosis, recurrent pregnancy loss or stillbirth, and diagnostic evaluation of suspected lupus, APS testing only was considered appropriate. Results 450 patients underwent thrombophilia testing with a mean age of 42 (range: 18-90); 76% were female and 81% were Hispanic. A total of 1698 thrombophilia tests were sent by 27 services. Testing was done in the following settings: inpatient (40%), outpatient (59%), and emergency department (1%). The mean tests per patient were 3.7 (range: 1-12). The most common requesting services were rheumatology (24%), obstetrics-gynecology (19%), and internal medicine/medicine-pediatrics (14%). Hematology requested 10% of tests. Common indications for testing were VTE (21%), rheumatology-related (25%), pregnancy-related (13%), ischemic stroke/TIA (9%), ocular-related (7%), non-stroke arterial thrombosis (3%), and dermatology-related (4%) (Table 1). 5% (84 tests) were sent for the evaluation of other non-thrombotic conditions. 8% (132 tests) were sent for > 1 indication, such as concurrent arterial and venous events. 840 tests (49%) were deemed inappropriate. Common reasons for inappropriate testing included provoked VTE events, stroke/TIA, APS testing after first pregnancy loss, current AC, and duplicate testing (Table 2). APS testing issues included testing for LAC while on AC, incomplete testing (both CL and b2GP not sent), incorrect tests (phosphatidylserine IgM/IgG), and repeat testing < 12 weeks from prior. Incorrect/redundant testing for FVL included: FV activity levels (36 total tests; 9 ordered in additional to FVL and 27 ordered instead of FVL) and APC resistance ordered simultaneously with FVL in 7 patients. Of note, 92 tests were sent for evaluation of non-thrombotic conditions. Conclusions Thrombophilia testing is often done inappropriately. Correct test selection is also a relatively common issue, particularly with APS and FVL testing. Given the large number of services ordering thrombophilia testing at our training hospital and this testing being sent for a variety of reasons, it is unlikely that physician education alone will lead to a substantial or sustained decrease in the number of inappropriate tests. Rather, it may be necessary to restrict at least some thrombophilia testing to certain services. Figure 1 Figure 1. Disclosures Piatek: Rigel: Consultancy, Research Funding; Alexion: Consultancy, Research Funding; Apellis: Research Funding; Dova: Consultancy, Speakers Bureau.

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Inherited Thrombophilia In Omani Women with Recurrent Pregnancy Loss

Blood ◽

10.1182/blood.v116.21.5124.5124 ◽

2010 ◽

Vol 116 (21) ◽

pp. 5124-5124

Author(s):

Khalil Al Farsi ◽

Shoaib Al Zadjali ◽

Karima Al Falahi ◽

Murtadha K. Al-Khabori ◽

Anil Pathare ◽

...

Keyword(s):

Research Funding ◽

Protein C ◽

Pregnancy Loss ◽

Recurrent Pregnancy Loss ◽

Median Number ◽

Prothrombin G20210a ◽

Control Group ◽

Molecular Testing ◽

Inherited Thrombophilia ◽

Thrombophilia Testing

Abstract Abstract 5124 Background: Recurrent pregnancy loss (RPL) is a common clinical problem. Inherited thrombophilia has been reported to be associated with RPL by different groups. Methods: We retrospectively analyzed the records of women who had thrombophilia testing for RPL, defined as 2 or more pregnancy losses, between the period of June 2006 and June 2010. The following thrombophilic disorders were included: protein C deficiency (PC), protein S deficiency (PS), anti-thrombin deficiency (AT), activated protein C resistance (APCR) and more recently, as molecular testing became available at our institution, factor V G1691A (FVL), prothrombin G20210A (PTG) and Methyl tetrahydrofolate reductase C677T (MTHRF) mutations. Women were excluded if testing was only done during pregnancy or in the immediate post-partum period. Results: A total of 136 women were identified. Median age was 32 (range: 18–44) with a median number of RPL of 3 (range: 2–11). Median number of first trimester losses was 2 (range 0–11). Two women had only second trimester losses and one had only third trimester losses. PS deficiency was identified in 8 women (5.8%), PC deficiency in 3 (2.2%), AT deficiency in 1 (0.7%) and APCR in 1 (0.7%). Of 30 women who had genetic analysis by PCR, 8 had abnormal results (MTHFR: 4 heterozygous, 1 homozygous; FVL: 2 heterozygous and PTG: 1 heterozygous). We did not find any correlation between the number of RPLs and the finding of a positive thrombophilia screen in the overall group or in the group of women who had molecular testing. Conclusion: Inherited thrombophilia is not as common in our patient population as described in other groups. However, a prospective study with a control group and a full panel of thrombophilia testing is needed to assess the prevalence and significance of such defects in women with RPL. Disclosures: Pathare: Sultan Qaboos University: Employment, Research Funding. Alkindi:Sultan Qaboos University: Employment, Research Funding.

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Protein C Global Assay in the Evaluation of Women with Idiopathic Pregnancy Loss

Thrombosis and Haemostasis ◽

10.1055/s-0037-1613149 ◽

2002 ◽

Vol 88 (07) ◽

pp. 32-36 ◽

Cited By ~ 15

Author(s):

Galit Sarig ◽

Naomi Lanir ◽

Ron Hoffman ◽

Benjamin Brenner

Keyword(s):

Risk Factor ◽

Protein C ◽

Pregnancy Loss ◽

Factor V Leiden ◽

Protein S ◽

Diagnostic Value ◽

Factor V ◽

Activation Time ◽

Apc Resistance ◽

Low Levels

SummarySince the majority of thrombophilic defects in women with pregnancy loss are in the protein C pathway, we have prospectively determined the diagnostic value of a protein C global assay in 60 consecutive women with pregnancy loss compared to 61 controls. Protein C activation time normalized ratio (PCAT-NR) in pregnancy loss women was significantly lower than controls (0.74 ± 0.16 vs. 0.99 ± 0.2; P <0.0001). PCAT-NR lower than cut off level of 0.8 were found in 42/60 (70%) of PL women compared to 7/61 (11%) of controls (OR = 18.0, 95% CI: 6.3-53.4, P < 0.0001).Cut-off level of 0.8 successfully identified all pregnancy loss women with abnormality in the protein C pathway (12 factor V Leiden, 7 APC-Resistance without factor V Leiden, 15 low levels of protein S and 1 of protein C). Moreover, PCAT-NR below 0.8 was documented in 15/29 (52%) of PL women without thrombophilic risk factor compared to 3/55 (5%) of controls (OR = 18.6, 95% CI: 4.2-95.2, P <0.0001).These results suggest that ProC Global may be useful as a screening test for protein C pathway abnormalities and may serve as a new thrombophilic risk factor in women with pregnancy loss.

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“Pseudo Homozygous” Activated Protein C Resistance due to Double Heterozygous Factor V Defects (Factor V Leiden Mutation and Type I Quantitative Factor V Defect) Associated with Thrombosis: Report of Two Cases Belonging to Two Unrelated Kindreds

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650290 ◽

1996 ◽

Vol 75 (03) ◽

pp. 422-426 ◽

Cited By ~ 53

Author(s):

Paolo Simioni ◽

Alberta Scudeller ◽

Paolo Radossi ◽

Sabrina Gavasso ◽

Bruno Girolami ◽

...

Keyword(s):

Protein C ◽

Dna Analysis ◽

Activated Protein C ◽

Factor V Leiden ◽

Type I ◽

Factor V ◽

Factor V Leiden Mutation ◽

Thrombotic Risk ◽

Apc Resistance ◽

Quantitative Factor

SummaryTwo unrelated patients belonging to two Italian kindreds with a history of thrombotic manifestations were found to have a double heterozygous defect of factor V (F. V), namely type I quantitative F. V defect and F. V Leiden mutation. Although DNA analysis confirmed the presence of a heterozygous F. V Leiden mutation, the measurement of the responsiveness of patients plasma to addition of activated protein C (APC) gave results similar to those found in homozygous defects. It has been recently reported in a preliminary form that the coinheritance of heterozygous F. V Leiden mutation and type I quantitative F. V deficiency in three individuals belonging to the same family resulted in the so-called pseudo homozygous APC resistance with APC sensitivity ratio (APC-SR) typical of homozygous F. V Leiden mutation. In this study we report two new cases of pseudo homozygous APC resistance. Both patients experienced thrombotic manifestations. It is likely that the absence of normal F. V, instead of protecting from thrombotic risk due to heterozygous F. V Leiden mutation, increased the predisposition to thrombosis since the patients became, in fact, pseudo-homozygotes for APC resistance. DNA-analysis is the only way to genotype a patient and is strongly recommended to confirm a diagnosis of homozygous F. V Leiden mutation also in patients with the lowest values of APC-SR. It is to be hoped that no patient gets a diagnosis of homozygous F. V Leiden mutation based on the APC-resi-stance test, especially when the basal clotting tests, i.e., PT and aPTT; are borderline or slightly prolonged.

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Elevated Levels of Prothrombin Activation Fragment 1+2 in Plasma from Patients with Heterozygous Arg506 to Gin Mutation in the Factor V Gene (APC-Resistance) and/or Inherited Protein S Deficiency

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650259 ◽

1996 ◽

Vol 75 (02) ◽

pp. 270-274 ◽

Cited By ~ 57

Author(s):

Benget Zöller ◽

Johan Holm ◽

Peter Svensson ◽

Björn Dahlbäck

Keyword(s):

Plasma Concentration ◽

Protein C ◽

Protein S ◽

Factor V ◽

Protein S Deficiency ◽

Apc Resistance ◽

S Deficiency ◽

Increased Risk ◽

Factor V Gene ◽

V Gene

SummaryInherited resistance to activated protein C (APC-resistance), caused by a point mutation in the factor V gene leading to replacement of Arg(R)506 with a Gin (Q), and inherited protein S deficiency are associated with functional impairment of the protein C anticoagulant system, yielding lifelong hypercoagulability and increased risk of thrombosis. APC-resistance is often an additional genetic risk factor in thrombosis-prone protein S deficient families. The plasma concentration of prothrombin fragment 1+2 (F1+2), which is a marker of hyper-coagulable states, was measured in 205 members of 34 thrombosis-prone families harbouring the Arg506 to Gin mutation (APC-resistance) and/or inherited protein S deficiency. The plasma concentration of F1+2 was significantly higher both in 38 individuals carrying the FV:Q506 mutation in heterozygous state (1.7 ± 0.7 nM; mean ± SD) and in 48 protein S deficient cases (1.9 ± 0.9 nM), than in 100 unaffected relatives (1.3 ±0.5 nM). Warfarin therapy decreased the F1+2 levels, even in those four patients who had combined defects (0.5 ± 0.3 nM). Our results agree with the hypothesis that individuals with APC-resistance or protein S deficiency have an imbalance between pro- and anti-coagulant forces leading to increased thrombin generation and a hypercoagulable state.

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Molecular Characterization of a Type I Quantitative Factor V Deficiency in a Thrombosis Patient that Is “Pseudo Homozygous” for Activated Protein C Resistance

Thrombosis and Haemostasis ◽

10.1055/s-0038-1655948 ◽

1997 ◽

Vol 77 (02) ◽

pp. 252-257 ◽

Cited By ~ 32

Author(s):

Joan F Guasch ◽

Ruud P M Lensen ◽

Rogier M Bertina

Keyword(s):

Protein C ◽

Dna Analysis ◽

Activated Protein C ◽

Type I ◽

Factor V ◽

Sequencing Analysis ◽

Apc Resistance ◽

Quantitative Factor ◽

Factor V Deficiency ◽

Fv Leiden

SummaryResistance to activated protein C (APC), which is associated with the FV Leiden mutation in the large majority of the cases, is the most common genetic risk factor for thrombosis. Several laboratory tests have been developed to detect the APC-resistance phenotype. The result of the APC-resistance test (APC-sensitivity ratio, APC-SR) usually correlates well with the FV Leiden genotype, but recently some discrepancies have been reported. Some thrombosis patients that are heterozygous for FV Leiden show an APC-SR usually found only in homozygotes for the defect. Some of those patients proved to be compound heterozygotes for the FV Leiden mutation and for a type I quantitative factor V deficiency. We have investigated a thrombosis patient characterized by an APC-SR that would predict homozygosity for FV Leiden. DNA analysis showed that he was heterozygous for the mutation. Sequencing analysis of genomic DNA revealed that the patient also is heterozygous for a G5509→A substitution in exon 16 of the factor V gene. This mutation interferes with the correct splicing of intron 16 and leads to the presence of a null allele, which corresponds to the “non-FV Leiden” allele. The conjunction of these two defects in the patient apparently leads to the same phenotype as observed in homozygotes for the FV Leiden mutation.

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The A6936G polymorphism of the endothelial protein C receptor gene is associated with the risk of unexplained foetal loss in Mediterranean European couples

Thrombosis and Haemostasis ◽

10.1160/th-09-04-0224 ◽

2009 ◽

Vol 102 (10) ◽

pp. 656-667 ◽

Cited By ~ 9

Author(s):

Pascale Fabbro-Peray ◽

Pierre Marès ◽

Patrick Mismetti ◽

Géraldine Lissalde-Lavigne ◽

Éva Cochery-Nouvellon ◽

...

Keyword(s):

Protein C ◽

Pregnancy Loss ◽

Receptor Gene ◽

Coagulation Factor ◽

Factor V ◽

Endothelial Protein C Receptor ◽

Foetal Loss ◽

Factor Ii ◽

Embryonic Loss ◽

Factor V Gene

SummaryThe endothelial protein C receptor (EPCR) is expressed by trophoblast cells. Mid-gestation pregnancy loss is described in animals with a haemochorial placenta lacking EPCR. The A6936G allele of the EPCR gene (PROCR) may be associated with lower EPCR densities on trophoblasts, but data are lacking for its effect on the risk of pregnancy loss in humans. A 1:2 case-control study on unexplained pregnancy loss was nested in the NOHA First cohort: 3,218 case couples and 6,436 control couples were studied for PROCR A6936G, coagulation factor V gene (F5) G1691A and coagulation factor II gene (F2) G20210A polymorphisms. Ethnicity and time of pregnancy loss defined through biometry-based gestational ages (embryonic loss < 10th week ≥ foetal loss) were analysed. The PROCR A6936G allele, in mothers and fathers, was associated only with foetal loss in both Europeans and non-Europeans. Increasing probability levels of carrying a homozygous child were increasingly associated with the risk of foetal demise. The F5 G1691A and F2 G20210A alleles, only in mothers, were only and independently associated with foetal loss in Europeans. In our population, the PROCR A6936G allele describes women, but also men and thus couples, at risk for first unexplained foetal loss. This risk is independent of the foetal loss risk conferred to our local Mediterranean European women by the F5 G1691A and F2 G20210A alleles. Data confirm that the relationship between thrombophilias and pregnancy loss varies according to ethnicity and loss type.

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A Factor V Genetic Component Differing From Factor V R506Q Contributes to the Activated Protein C Resistance Phenotype

Blood ◽

10.1182/blood.v90.4.1552 ◽

1997 ◽

Vol 90 (4) ◽

pp. 1552-1557 ◽

Cited By ~ 127

Author(s):

F. Bernardi ◽

E.M. Faioni ◽

E. Castoldi ◽

B. Lunghi ◽

G. Castaman ◽

...

Keyword(s):

Venous Thromboembolism ◽

Protein C ◽

Activated Protein C ◽

Functional Assay ◽

Factor V ◽

Resistance Phenotype ◽

Apc Resistance ◽

Genetic Components ◽

Factor V Gene ◽

V Gene

AbstractFactor V gene polymorphisms were investigated to detect components that may contribute to the activated protein C (APC) resistance phenotype in patients with venous thromboembolism. A specific factor V gene haplotype (HR2) was defined by six polymorphisms and its frequency was found to be similar in normal subjects coming from Italy (0.08), India (0.1), and Somalia (0.08), indicating that it was originated by ancestral mutational events. The relationship between the distribution of normalized APC ratios obtained with the functional assay and haplotype frequency was analyzed in patients heterozygous for factor V R506Q (factor V Leiden). The HR2 haplotype was significantly more frequent in patients with ratios below the 15th percentile than in those with higher ratios or in normal controls. Moreover, the study of 10 patients with APC resistance in the absence of the factor V R506Q mutation showed a 50-fold higher frequency of HR2 homozygotes. The HR2 haplotype was associated with significantly lower APC ratios both in patients with venous thromboembolism and in age- and sex-matched controls. However, the two groups showed similar HR2 haplotype frequencies. Plasma mixing experiments showed that an artificially created double heterozygote for the factor V R506Q mutation and the HR2 haplotype had an APC ratio lower than that expected for a simple R506Q heterozygote. Time-course experiments evaluating the decay of factor V in plasma showed the normal stability of the molecule encoded by the factor V gene marked by the HR2 haplotype, which ruled out the presence of a pseudo-homozygous APC resistance mechanism. Our results provide new insights into the presence of factor V genetic components other than the factor V R506Q that are able to contribute to the APC resistance phenotype in patients with venous thromboembolism.

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Thrombophilia And Arterial Ischemic Stroke

Advances in Bioscience and Clinical Medicine ◽

10.7575/aiac.abcmed.ca1.45 ◽

2017 ◽

pp. 45

Author(s):

A.A. Abrishamizadeh

Keyword(s):

Ischemic Stroke ◽

Vitamin K ◽

Arterial Thrombosis ◽

Protein C ◽

Antithrombin Iii ◽

Factor V Leiden ◽

Protein S ◽

Factor V ◽

Factor V Leiden Mutation ◽

C Protein

Ischemic stroke (IS) is a common cause of morbidity and mortality with significant socioeconomic impact especially when it affects young patients. Compared to the older adults, the incidence, risk factors, and etiology are distinctly different in younger IS. Hypercoagulable states are relatively more commonly detected in younger IS patients.Thrombophilic states are disorders of hemostatic mechanisms that result in a predisposition to thrombosis .Thrombophilia is an established cause of venous thrombosis. Therefore, it is tempting to assume that these disorders might have a similar relationship with arterial thrombosis. Despite this fact that 1-4 % of ischemic strokes are attributed to Thrombophillia, this alone rarely causes arterial occlusions .Even in individuals with a positive thrombophilia screen and arterial thrombosis, the former might not be the primary etiological factor.Thrombophilic disorders can be broadly divided into inherited or acquired conditions. Inherited thrombophilic states include deficiencies of natural anticoagulants such as protein C, protein S, and antithrombin III (AT III) deficiency, polymorphisms causing resistance to activated protein C(Factor V Leiden mutation), and disturbance in the clotting balance (prothrombin gene 20210G/A variant). Of all the inherited thrombophilic disorders, Factor V Leiden mutation is perhaps the commonest cause. On the contrary, acquired thrombophilic disorders are more common and include conditions such as the antiphospholipid syndrome, associated with lupus anticoagulant and anticardiolipin antibodies.The more useful and practical approach of ordering various diagnostic tests for the uncommon thrombophilic states tests should be determined by a detailed clinical history, physical examination, imaging studies and evaluating whether an underlying hypercoagulable state appears more likely.The laboratory thrombophilia screening should be comprehensive and avoid missing the coexisting defect and It is important that a diagnostic search protocol includes tests for both inherited and acquired thrombophilic disorders.Since the therapeutic approach (anticoagulation and thrombolytic therapy) determines the clinical outcomes, early diagnosis of the thrombophilic disorders plays an important role. Furthermore, the timing of test performance of some of the thrombophilic defects (like protein C, protein S, antithrombin III and fibrinogen levels) is often critical since these proteins can behave as acute phase reactants and erroneously elevated levels of these factors may be observed in patients with acute thrombotic events. On the other hand, the plasma levels of vitamin K-dependent proteins (protein C, protein S and APC resistance) may not be reliable in patients taking vitamin K antagonists. Therefore, it is suggested that plasma-based assays for these disorders should be repeated3 to 6 months after the initial thrombotic episode to avoid false-positive results and avoid unnecessary prolonged anticoagulation therapy. The assays for these disorders are recommended after discontinuation of oral anticoagulant treatment or heparin for at least 2 weeks.

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Factor V Cambridge: A New Mutation (Arg306→Thr) Associated With Resistance to Activated Protein C

Blood ◽

10.1182/blood.v91.4.1140 ◽

1998 ◽

Vol 91 (4) ◽

pp. 1140-1144 ◽

Cited By ~ 165

Author(s):

David Williamson ◽

Karen Brown ◽

Roger Luddington ◽

Caroline Baglin ◽

Trevor Baglin

Keyword(s):

Venous Thrombosis ◽

Cleavage Site ◽

Protein C ◽

Activated Protein C ◽

Factor V Leiden ◽

Factor V ◽

Degree Relative ◽

Prothrombinase Complex ◽

Apc Resistance ◽

The Common

AbstractA new factor V mutation associated with resistance to activated protein C and thrombosis (factor V Cambridge, Arg306→Thr) was found in one patient from a carefully selected group of 17 patients with venous thrombosis and confirmed APC resistance in the absence of the common Gln506 mutation. The Arg306 mutation was also present in a first degree relative who also had APC resistance. Other potential causes of APC resistance, such as a mutation at the Arg679 site and the factor V HR2 haplotype, were excluded. Subsequent screening of 585 patients with venous thromboembolism and 226 blood donors did not show any other individual with this mutation. Factor VThr306 is the first description of a mutation affecting the Arg306 APC cleavage site and is the only mutation, other than factor V Leiden (Arg506→Gln), that has been found in association with APC resistance. This finding confirms the physiologic importance of the Arg306 APC-cleavage site in the regulation of the prothrombinase complex. It also supports the concept that APC resistance and venous thrombosis can result from a variety of genetic mutations affecting critical sites in the factor V cofactor.

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