Pharmacogenetic aspects of dabigatran administration for the prevention of venous thromboembolic complications in patients after orthopedic surgery: clinical cases

Relevance

Venous thromboembolic complications (VTE) remain one of the main threats to life and health after arthroplasty of large joints of the lower extremities. Early mobilization and drug therapy are mandatory measures for VTE prevention in this group of patients in the postoperative period. According to Russian clinical guidelines for the prevention, diagnosis and treatment of VTE, the use of low molecular weight heparins (LMWH) for 5 weeks is a priority [1]. The main direct oral anticoagulants (DOACs) currently used for the treatment of VTE are rivaroxaban, edoxaban and apixaban (direct factor Xa inhibitors) and dabigatran (direct thrombin inhibitor). Of these, apixaban, dabigatran and rivaroxaban are currently approved in the EU and USA for the treatment of VTE, as well as for the prevention and treatment of deep vein thrombosis (DVT) and pulmonary embolism (PE) in patients undergoing orthopedic surgery. Edoxaban is currently approved in Japan for the prevention of postoperative VTE and in the USA for the treatment and secondary prevention of VTE in the non-surgical population, and has also received a positive opinion from the European Committee for Medicinal Products. Compared to vitamin K antagonists (VKAs), DOACs provide rapid onset of action, have fixed dosing, no known food effects, fewer drug interactions, no need for routine monitoring of fixed doses, and a short offset of action [2].

Thrombin is an obvious target for anticoagulants because of its role as the final effector of blood coagulation [3].

When anticoagulant therapy for VTE prophylaxis is discontinued in patients after total knee arthroplasty (TKA), approximately 10% experience thrombosis recurrence within one year after the first event [4].

VTE is a life-threatening problem after TKA. However, the issue of optimal pharmacological prophylaxis remains controversial. The use of DOACs remains the most effective and proven method for VTE prevention after knee or hip arthroplasty. The convenience of their use is obvious both in the hospital and outpatient settings. The use of dabigatran etexilate significantly reduces the duration and costs of hospitalization, and the ease of drug administration ensures good treatment adherence. Nevertheless, the effectiveness of VTE prophylaxis with DOACs is not always 100% and has a number of specific complications. Great importance is attached to the search for new pharmacogenetic markers of thrombogenic risk (TR) and predictors of thromboembolic complications in patients with VTE.

Proven TR factors include advanced age, female gender, various diseases and pathological conditions such as arterial hypertension, coronary heart disease, varicose veins, obesity, diabetes mellitus, malignant tumors, and others.

Currently, a personalized approach to the choice of anticoagulant drug, effective and safe dose is relevant, since inadequate drug doses can lead to complications [5]. One of the methods for diagnosing coagulation hemostasis is the thrombodynamics test, which has been tested on samples of patients with various hemostasis pathologies, as well as cardiovascular diseases [6]. Among all thrombodynamics parameters, the clot growth rate is the most informative parameter in predicting VTE [7].

Research materials

The study included 60 patients (58 women and 2 men) aged 37 to 81 years. Patient information was entered into anonymized individual patient records. According to national clinical guidelines, all patients after total knee arthroplasty received VTE prophylaxis with dabigatran etexilate at a dose of 220 mg per day for 1 month.

To illustrate the pharmacogenetic approach to VTE prevention using the laboratory control method of thrombodynamics, 6 typical clinical cases were selected.

All patients underwent genotyping for carriage of the polymorphic variants rs1045642 and rs4148738 of the ABCB1 gene and rs2244613 of the CES1 gene using real-time polymerase chain reaction (Real-Time PCR). Peak and trough plasma concentrations of dabigatran were determined by high-performance liquid chromatography (HPLC). Determination of dabigatran etexilate concentration was performed in all patients on day 5 after surgery (TKA). Quantitative determination of dabigatran in blood plasma samples was performed by high-performance liquid chromatography with mass spectrometric detection using an internal standard [8]. An Agilent 6410 Triple Quadrupole LC/MS mass spectrometer was used. Sample preparation was performed by precipitation of blood plasma proteins. Deuterated dabigatran was used as an internal standard. To prepare solutions of dabigatran standards and its deuterated analogue, initial stock solutions of the compounds were used in a mixture of methanol and dimethyl sulfoxide (9:1 ratio). The concentrations of dabigatran and its deuterated standard in stock solutions were 10 μg/mL. The content of dabigatran in blood plasma samples was calculated by the ratio of the area of the chromatographic peak of the target compound to the area of the internal standard.

According to clinical guidelines, the Caprini score was chosen for individual VTE prediction in patients, as it is the most validated model worldwide [1, 9--10]. To predict bleeding risk, the RIETE score proposed in the clinical guidelines was used, as it is the most convenient and appropriate for the study objectives [1].

Description of clinical cases

Clinical case 1

A 58-year-old woman with a high risk of VTE — 11 points on the Caprini score — developed acute occlusive DVT of the right lower leg on day 2 after TKA, which required comprehensive medical and surgical treatment. There was a high risk of PE. According to molecular genetic testing (MGT), the patient was found to have the CC genotype of the rs2244613 polymorphism of the CES1 gene and the TT genotype of the rs4148738 polymorphism of the ABCB1 gene (Fig. 1).

Fig. 1. Association of the rs4148738 polymorphism of the ABCB1 gene with C~max~,ss of dabigatran

Clinical case 2

A 59-year-old patient (heterozygous genotype for ABCB1 and CES1) with a high risk of VTE — 10 points on the Caprini score — showed a high clot formation rate of 47.8 μm/min according to the thrombodynamics test. 4 weeks after TKA, she developed acute thrombosis of the popliteal vein, which could be associated with low activity after surgery due to intense pain in the contralateral joint. The patient underwent conservative thrombolytic therapy according to clinical guidelines for 2 months. According to control ultrasound examination of the veins of the lower extremities, resolution through thrombus recanalization occurred after 2 months. Against the background of increased glomerular filtration rate (GFR), the patient's peak dabigatran concentration was below the average level obtained in our study.

Clinical case 3

A 71-year-old patient (with the CC genotype of the rs1045642 polymorphism and TT genotype of the rs4148738 polymorphism of the ABCB1 gene, and the AA genotype of the rs2244613 polymorphism of the CES1 gene), with an extremely high risk of thrombosis according to the Caprini score — 13 points, and the highest clot formation rate according to the thrombodynamics test — 58.7 μm/min. Nevertheless, the patient did not experience thrombosis, minor or major bleeding, and the postoperative period was uneventful. Thus, despite the individual very high risk of thrombosis, high efficacy of dabigatran was noted — the clot formation rate after taking the drug decreased to 31.4 μm/min (46.7% of the initial value), and according to the coagulogram, the diluted prothrombin time (PTT) increased. High safety of the drug was also noted — no bleeding was recorded (Fig. 2).

Fig. 2. Association of the rs4148738 polymorphism of the ABCB1 gene with C~max~,ss of dabigatran

Clinical case 4

A 45-year-old patient with an extremely high risk of thrombosis (12 points on the Caprini score). According to MGT — heterozygous for all three studied polymorphisms: rs1045642 and rs4148738 of the ABCB1 gene, and rs2244613 of the CES1 gene.

Nevertheless, the patient developed a minor hematoma, which could be associated with a technically complex (more traumatic and prolonged) surgical intervention. In our opinion, the combination of heterozygous carriage for all three studied polymorphisms is the most unpredictable option in terms of the development of thrombosis and/or bleeding.

Clinical case 5

A 64-year-old patient with the TT genotype for the C3435T polymorphism of the ABCB1 gene in combination with the CC genotype of the rs4148738 polymorphism of the ABCB1 gene, and the AA genotype of the rs2244613 polymorphism of the CES1 gene, had high baseline and peak dabigatran concentrations, which is associated with an increased risk of bleeding (Fig. 3).

Fig. 3. Association of the rs1045642 polymorphism of the ABCB1 gene with C~max~,ss of dabigatran

Clinical case 6

A 61-year-old patient recorded the maximum concentration of dabigatran — 800 ng/mL. The patient was receiving concomitant therapy with a combined antihypertensive and lipid-lowering drug (amlodipine + lisinopril + rosuvastatin). In this case, there was an interaction between the drugs and dabigatran. Also, the patient once received an injection of fondaparinux (pharmacodynamic interaction). 2 days after surgery, macrohematuria occurred.

In this case, dabigatran discontinuation and administration of tranexamic acid to stop the bleeding were required. On the second day, dabigatran was prescribed with a 50% dose reduction for the entire treatment period; no thrombosis and/or bleeding was recorded during follow-up. This patient was excluded from the study and analysis.

Discussion

In our study, no statistically significant dependence of dabigatran concentration on gender, age, body mass index, or renal function was found.

Clinical case No. 1 demonstrated that carriage of the CC genotype of the rs2244613 polymorphism of the CES1 gene is associated with a lower concentration of dabigatran, which increased the risk of VTE. According to the thrombodynamics test, a high clot growth rate was noted before and after dabigatran administration. In our 2018 study, we found an association between the CC genotype of the rs2244613 polymorphism of the CES1 gene and a lower peak concentration of dabigatran, with a median of 116.3 ng/mL [11]. No statistical significance was obtained (*p*=0.27) due to the small sample size, but later, in a meta-analysis conducted in 2020, it was demonstrated that the frequency of bleeding was 26.8% lower in the group of carriers of the C allele of the rs2244613 polymorphism of the CES1 gene (AC and CC genotypes), compared to non-carriers (AA genotype) (OR 0.732, 95% CI 0.629--0.851; *p* <0.001; heterogeneity: Q=2.183; *p*=0.535) among patients taking dabigatran as indicated. Carriage of rs2244613 was significantly associated with a reduced incidence of bleeding during dabigatran etexilate therapy [12].

Analysis of clinical case No. 2 demonstrated that heterozygous carriage for the C3435T polymorphisms of the ABCB1 gene and rs2244613 of the CES1 gene, combined with normal or increased GFR, led to a decrease in dabigatran concentration. According to the thrombodynamics test, a high clot formation rate persisted before and after dabigatran administration. Thus, in this case, increasing the duration of anticoagulant therapy should be considered.

Clinical case No. 3 demonstrated the most favorable variant of genetic polymorphism: the CC genotype of the rs1045642 polymorphism and the TT genotype of the rs4148738 polymorphism of the ABCB1 gene, and the AA genotype of the rs2244613 polymorphism of the CES1 gene, in which, despite the initially high TR, high efficacy and safety of dabigatran were demonstrated. We consider this combination of genetic polymorphisms as the most favorable option.

Clinical case No. 4 presents a patient with heterozygous carriage for all three studied polymorphisms; in this case, it is the most difficult to predict the risk of thrombosis and bleeding in terms of prognosis.

In clinical case No. 5, in our opinion, the patient with the TT genotype for the C3435T polymorphism of the ABCB1 gene in combination with the CC genotype of the rs4148738 polymorphism of the ABCB1 gene, and the AA genotype of the rs2244613 polymorphism of the CES1 gene, had the highest risk of bleeding. The need to consider treatment options with dose reduction of anticoagulants was noted. For optimal safety control, determination of the drug concentration in patients with this genotype is necessary.

Analysis of clinical case No. 6 revealed pharmacokinetic and pharmacodynamic drug interactions: amlodipine, lisinopril, rosuvastatin, and dabigatran. Statins, including rosuvastatin, are modulators of P-glycoprotein, which encodes the efflux of statins, and their pharmacokinetics depends on the C3435T polymorphism of the ABCB1 gene [13--15]. Amlodipine is a substrate of P-glycoprotein, and its concentration also depends on the C3435T polymorphism of the ABCB1 gene [16]. Lisinopril, in turn, is a substrate of P-glycoprotein [17]. Thus, the drug interaction led to the development of a serious adverse reaction (AR) — renal bleeding.

Conclusions

1. When carrying out VTE prophylaxis in patients after orthopedic interventions, the pharmacogenetic characteristics of the patient that affect the metabolism of anticoagulants are important.

2. The pharmacokinetics of dabigatran are influenced by polymorphisms of genes encoding P-glycoprotein and carboxylesterase: ABCB1 and CES1.

3. The TT genotype for the C3435T polymorphism of the ABCB1 gene is associated with a higher concentration of dabigatran and a high risk of bleeding.

4. The CC genotype of the rs2244613 polymorphism of the CES1 gene correlates with a lower concentration and an increased risk of thrombosis.

5. The most favorable combination of gene polymorphisms was demonstrated: the CC genotype of the rs1045642 polymorphism and the TT genotype of the rs4148738 polymorphism of the ABCB1 gene; the AA genotype of the rs2244613 polymorphism of the CES1 gene. High efficacy and safety of dabigatran were observed, despite unfavorable thrombosis risk factors.

6. Heterozygous carriage for all three studied polymorphisms rs1045642 and rs4148738 of the ABCB1 gene and rs2244613 of the CES1 gene has the most unpredictable effect on the risk of bleeding and/or thrombosis.

7. A case of interaction between dabigatran and other drugs was demonstrated, which led to the development of severe bleeding that required discontinuation of dabigatran.

8. The thrombodynamics test can be considered as a promising laboratory test for diagnosing hypercoagulation and monitoring the effectiveness of dabigatran therapy.

Thus, wider introduction of genetic testing among patients after orthopedic operations receiving dabigatran for VTE prophylaxis will significantly improve the effectiveness and safety of anticoagulant therapy in the future.