Warfarin-induced life-threatening bleeding associated with a CYP3A4 loss-of-function mutation in an acute limb ischemia patient: Case report and review of the literature

  • Authors:
    • Xiao‑Wei Ma
    • Chang‑Ning Hao
    • Zhi‑Chun Gu
    • Meng Ye
    • Min Li
    • Lan Zhang
  • View Affiliations

  • Published online on: June 14, 2017     https://doi.org/10.3892/etm.2017.4604
  • Pages: 1157-1162
Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )


Abstract

Patients with acute limb ischemia, deep venous thrombosis and pulmonary artery embolism may be treated with warfarin. The dose‑response interaction of warfarin is associated with numerous factors, depending on which an uncommon life‑threatening bleeding may occur. The present case study reported on a patient with acute limb ischemia and a history of warfarin-induced bleeding ten years previously and who again developed life threatening bleeding associated with warfarin treatment and received vascular surgery. In this patient, a cytochrome P450 3A4 loss‑of‑function mutation decreased the effective dose of warfarin. Although this was a rare case, clinicians should be alert to the bleeding risk associated with such rare genetic mutations.

Introduction

Warfarin therapy effectively reduces ischemic stroke and mortality amongst patients with various types of thromboembolism, such as deep venous thrombosis, as well as heart valve prosthesis, atrial fibrillation and stroke (1).

It is the most frequently used oral anti-coagulant. Bleeding is an important adverse drug response (ADR) of anti-coagulants due to its narrow therapeutic index and the wide variability in drug responses among individuals.

To date, the warfarin dose response has been associated with race, environmental, clinical status, and particularly genetic factors (2). Warfarin exerts its anti-coagulant effect by antagonizing vitamin K epoxide reductase complex (VKORC1), thereby reducing the activation of vitamin K-dependent clotting factors II, VII, IX and X. A series of genetic variations related with the pharmacodynamics and pharmacokinetics of warfarin have been reported, such as VKORC1, cytochrome P450 (CYP) 2C9 and CYP4F2 (3,4).

Only few reports on warfarin-induced side effects associated with CYP3A4 mutations are currently available, and the present study reported on warfarin-induced major bleeding in a patient with acute limb ischemia (ALI) associated with a CYP3A4 loss-of-function mutation, which may promote the development of warfarin research.

Case report

The present study was approved by the institutional review board (CWO) of Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University. Patients provided written informed consent. A 60-year-old man was admitted to the vascular surgery department due to ALI in the left lower extremity in May 2015. On reviewing his medical history, it was revealed that the patient's blood pressure, blood glucose, cholesterol, triglyceride, homocysteine, erythrocyte sedimentation rate, anti-thrombin-III as well as protein C and S were within normal ranges. His life style was healthy and he had never smoked. He had neither atrial fibrillation nor a ventricular aneurysm. According to the angiography results, his left profunda femoris was completely occluded, and his distal superficial femoral artery (SFA) to popliteal artery (PA) was filled with thrombus. A 4F/30 cm Uni-Fuse™ Thrombolytic catheter (AngioDynamics, Queensbury, NY, USA) was inserted and the patient was intra-arterially injected 1 million units urokinase by exact pump over the next 24 h. One day later, his SFA-PA-peroneal artery track was visualized and the thrombus was basically cleared. The occluded proximal anterior tibial artery (ATA) and strait distal superficial femoral artery were then re-canalized by implanting a XIENCE-PRIME 3.5×38 mm (Abbott Pharmaceutical Co. Ltd., Lake Bluff, IL, USA) and an Everflex 7×120 mm (Covidien/Medtronic, Dublin, Ireland). With the ATA and peroneal artery serving as below-the-knee (BTK) outflow tracks, his ischemic symptoms totally disappeared and the left ankle-brachial index was recovered from 0.43 to 1.14. The patient was discharged after 6 days and was prescribed warfarin (2.5 mg per day) and clopidogrel (75 mg per day).

Unexpectedly, at 9 days after discharge, the patient presented to the emergency room with a sudden onset of hemoptysis. Laboratory analysis revealed that the international normalized ratio (INR) of the prothrombin time was 5.02 R. After intravenous infusion of 40 mg vitamin K1, his INR was decreased to 1.06R (reference value: 0.8–1.5). The patient's prescription was therefore modified by eliminating warfarin and retaining clopidogrel when he was discharged 24 h later. The patient's clinical characteristics on second admission are listed in Table I.

Table I.

Patient characteristics at the time-point of second admission (sudden onset of hemoptysis after warfarin + clopidogrel treatment for 9 days).

Table I.

Patient characteristics at the time-point of second admission (sudden onset of hemoptysis after warfarin + clopidogrel treatment for 9 days).

VariableResultReference value
EthnicityChinese-Han
SexMale
Height175 cm
Weight64 kg
FeverNo
SmokingNo
Alcohol consumptionNo
Diabetes mellitusNo
HypertensionNo
Hepatic diseaseNo
Chronic kidney diseaseNo
Protein C/S deficiencyNo
Age (years)60
Alanine aminotransferase (IU/l)19.513–69
Aspartate aminotransferase (U/l)26.015–46
Blood urea nitrogen (µmol/l)2.802.5–7.1
Creatinine (µmol/l)51.053–115
White blood cells (×109/l)8.993.97–9.15
Platelets (×109/l)17685–303

Fifty days later, the patient presented with another left ALI with typical 6P symptoms. The angiogram revealed that a thrombus had formed in the upper part of the left SFA, which was more proximal to BTK arteries than the previous time. Mechanical thrombectomy using an Angioget system (Boston Scientific Corp., Marlborough, MA, USA) and traditional balloon angioplasty to re-establish the SFA-PA-ATA/peroneal artery tracks. The patient was prescribed aspirin (100 mg per day) and cilostazol (100 mg per day) to inhibit platelet aggregation, as well as rivaroxaban (10 mg per day) to neutralize coagulation factor X. To date, the patient has remained in remission. The treatment of the patient is illustrated in a flow diagram in Fig. 1.

The genetic information was also analyzed. DNA extraction was performed using a QIAamp DNA Blood Mini kit (Syngen, Inc., Sacramento, CA, USA) according to the manufacturer's protocol. The concentration of isolated DNA was measured using a NanoDrop spectrophotometer (NanoDrop; Thermo Fisher Scientific, Inc., Pittsburgh, PA, USA) according to the manufacturer's protocol. The associated genetic variation in the pharmacogenomics knowledge base (https://www.pharmgkb.org/) was obtained by whole-genome sequencing (Illumina Hiseq2500; Illumina Inc. San Diego, CA, USA; sequencing depth, ×100). Rare genetic variations [minor allele frequency (MAF) <1%] were analyzed in this patient. There was no rare mutation in CYP2C9 or VKORC1, but a splicing mutation in CYP3A4 (rs55808838; MAF=0.04%) was identified by the Sanger method (Fig. 2).

Discussion

As is known, the dose-response association of warfarin is influenced by several factors, including ethnicity, environmental factors, drug interactions, clinical status and genetic factors. With regard to bleeding complications, as many aspects as possible were considered for the present case:

i) Clinical characteristics. It has been reported that abnormal kidney/liver function and several complications, such as fever, hypertension and diabetes mellitus affect the effective dose of warfarin (57). Recently, the white blood cell count was also found to affect inter-patient variations in the response to warfarin (8).

ii) Drug interactions. A multitude of drugs have been associated with the effective dose of warfarin, such as amiodarone, fluconazole and antibiotics, according to the warfarin product monograph. The patient of the present study was only prescribed clopidogrel (75 mg/day) besides warfarin. To the best of our knowledge, there is no evidence that clopidogrel affects the dose of warfarin and it was not a major factor associated with the bleeding.

iii) Vitamin K supplementation. Another common concern regarding the use of warfarin is the interaction with food rich in vitamin K (9). It has been reported that vitamin K intake was responsible for ~3% of dosage variations in Chinese patients (10), while certain studies found a negative correlation with vitamin K intake (11). The available evidence does not support the modification of dietary vitamin K intake when starting therapy with warfarin. The patient of the present study was on a regular Chinese diet and it was not the major factor associated with the bleeding.

iv) Genetic factors. Given that the clinicopathological findings were associated with genetic factors, written informed consent was obtained from the patient to perform genetic testing. Five genetic variations associated with a required increase in the warfarin dose, namely CYP4F2, γ glutamyl carboxylase, protease, serine 53 and NAD(P)H quinone dehydrogenase 1 were identified, besides the common mutation in VKORC1 (rs7294) in Chinese individuals (Table II) (1217).

Table II.

Genetic variations.

Table II.

Genetic variations.

GeneSingle nucleotide polymorphismEvidence levelGenotypeEffect on dose
CYP2C9rs17998531ACC
CYP2C9rs10579101AAA
VKORC1rs99232311AAADecrease
VKORC1rs72941BCC
VKORC1rs99344381BGG
CYP4F2rs21086221BCTIncrease
CYP2C9rs79001942AGG
CYP2C9rs49176392AAA
CYP2C9rs561654522ATT
CYP2C9rs283716862ACC
VKORC1rs23596122AAA
VKORC1rs80508942ACC
VKORC1rs177084722AGG
VKORC1rs28847372AAA
VKORC1rs617422452ACC
CALUrs3390972BAA
CALUrs127778232BGG
CALUrs71961612BGG
NR1I3rs25018733CC
EPHX1rs18777243CC
GGCXrs25925513GAIncrease
GGCXrs6996643CTIncrease
rs127141453CC
CYP2C9rs93320963CC
CYP2C9rs70895803AA
CYP2C9rs93321313AA
CYP2C9rs105096803GG
CYP2C9rs283716853CC
CYP2C9rs10579103AA
STX4rs108714543CC
PRSS53rs111506064CCIncrease
VKORC1rs72007494GG
PRSS53, VKORC1rs178861994AA
VKORC1rs99344384GG
VKORC1rs178808874GG
VKORC1rs611620434AA
NQO1rs105174AA
NQO1rs18005664GAIncrease
rs21897844GG
THBDrs10425804TT
HNF4Ars32121984
VKORC1rs1048945424AA
rs104894541 TT
VKORC1rs1048945404AA
VKORC1rs1048945394CC

[i] CYP, cytochrome P450; VKORC1, vitamin K epoxide reductase complex; CALU, calumenin; NR1I3, nuclear receptor subfamily 1 group I member 3; EPHX1, epoxide hydrolase 1; GGCX, γ glutamyl carboxylase; STX4, syntaxin 4; PRSS53, protease, Serine 53; NQO1, NAD(P)H quinone dehydrogenase 1; THBD, thrombomodulin; HNF4a, hepatocyte nuclear factor 4α.

The patient's warfarin dose was adjusted using five built-in pharmacogenetics-based warfarin dosing algorithms for the Chinese population and one by the International Warfarin Pharmacogenetics Consortium algorithm, as the equations based on Caucasian populations may not be suitable for predicting the dose of warfarin in Chinese patients (Table III).

Table III.

Six published pharmacogenetics-based warfarin dosing algorithms.

Table III.

Six published pharmacogenetics-based warfarin dosing algorithms.

Author/(Ref.), yearEthnicity Pharmacogenetics-based warfarin algorithmCalculated dose (mg/day)
IWPC; Klein et al (12), 2009Mixed√Weekly dose = 5.6044–0.2614 (age in decades) +0.0087 (height in cm) +0.0128 (weight in kg) −0.8677 (VKORC1 A/G) −1.6974 (VKORC1 A/A)-0.9357 (CYP2C9*1/*3) −0.1092 (Asian)a3
Miao et al (13), 2007ChineseDaily dose = 6.22–0.011 (age in years) +0.017 (weight in kg) −0.775 (CYP*3)-3.397 (VKORC1 × 1) −4.803 (VKORC1 × 2)a2.02
Huang et al (14), 2009ChineseDaily dose = exp [0.727–0.007 (age in years)+0.384 (BSA in square meters)+0.403 (VKORC1 6484TC) −0.482 (CYP2C9*1/*3)-1.583 (CYP2C9*3/*3)]2.67
Zhong et al (15), 2012Chinese√Daily dose = 1.68143–0.0029 (age, years) +0.30784 (BSA, m2) −0.2633 (VKORC1 g.3588G.A) −0.19114 (CYP2C9*3) +0.14735(CYP4F2 c.1297G>A) −0.1797 (amiodarone) −0.4138 (fluconazole) -(0.1888) diltiazem2.79
Tan et al (16), 2012Chinese√Daily dose = 2.140–0.370 (VKORC1-1639 G>A)-0.332 (CYP2C9*3)+0.324 (BSA, m2)-0.004 (age, years) −0.231 (no. of INR-increasing drugs) +0.105 (smoking habit) −0.135 (pre-operative stroke history) −0.108 (hypertension)2.99
Chen et al (17), 2014ChineseDaily dose = 0.135+1.7816 (rs7294) −1.2146 (rs1057910) +1.2886 (BSA, m2) −0.0196 (age, years) +0.7086 (target INR) +0.1596 (rs2108622) +0.3736 (diabetes mellitus) −0.5816 (amiodarone) −0.2526 (digoxin)2.84

a Genotyped for VKORC1-1639G>A (rs9923231) and CYP2C9*3 (rs1057910). IWPC, international Warfarin Pharmacogenetics Consortium; INR, international normalized ratio; BSA, body surface area.

Even if no other genetic variations were included in the dosing algorithms, not all of the dosing algorithms listed may be suitable for predicting the dose of warfarin in the present Chinese patient. In addition, the patient had a similar response to warfarin treatment ~10 years previously.

Compared to the normal Chinese population, the patient was unusually sensitive to warfarin. It was suggested that he had a rare genetic mutation leading to the high sensitivity to warfarin, as no other decrease-dose factors were found in this patient.

To the best of our knowledge, the present study was the first to report on warfarin-induced life threatening bleeding associated with a CYP3A4 loss-of-function mutation. S-warfarin is metabolized via CYP3A4 and the splicing mutation of CYP3A4 may therefore affect the effective dose of warfarin.

Only few studies have previously assessed CYP3A4 polymorphisms (18,19), and to the best of our knowledge, the rs55808838 polymorphism has not been previously reported. These previous studies on pharmacogenetic polymorphisms reported that other CYP3A4 polymorphisms may be associated with drug-induced thrombosis. CYP3A4*1G is the most frequent mutant allele of CYP3A4 in Asians and may be associated with a lower rate of clopidogrel resistance, which was, however, not the case in the patient of the present study.

The major limitation of the present study was that the effect of this rare mutation on the effect of warfarin in a single case was hard to verify. Therefore, as many aspects of this case as possible were reviewed, such as the patient's clinicopathological characteristics, drug interactions and vitamin K supplementation. All known genetic variations were considered and included into the pharmacogenetics-based warfarin dosing algorithms. However, all not all factors reviewed explain for the sensitivity to the warfarin in this patient. However, it is likely that this rare genetic mutation of CYP3A4 was associated with the recurrent warfarin-induced bleeding.

The present case report illustrated the importance of considering genetic mutations for assessing unusual warfarin-induced bleeding and presented the methods used for this. Although this was a rare case, clinicians should be alert to the bleeding risk associated with such rare genetic mutations.

Acknowledgements

The present study was supported by the China National Natural Science Foundation (nos. 81500372, 81322025, 81171623, 81371875 and 81421001) and the Shanghai Pujiang Program (no. 15PJ1405000).

References

1 

Lam MP and Cheung BM: The pharmacogenetics of the response to warfarin in Chinese. Br J Clin Pharmacol. 73:340–347. 2012. View Article : Google Scholar : PubMed/NCBI

2 

Takahashi H and Echizen H: Pharmacogenetics of warfarin elimination and its clinical implications. Clin Pharmacokinet. 40:587–603. 2001. View Article : Google Scholar : PubMed/NCBI

3 

Vecsler M, Loebstein R, Almog S, Kurnik D, Goldman B, Halkin H and Gak E: Combined genetic profiles of components and regulators of the vitamin K-dependent gamma-carboxylation system affect individual sensitivity to warfarin. Thromb Haemost. 95:205–211. 2006.PubMed/NCBI

4 

Li S, Zou Y, Wang X, Huang X, Sun Y, Wang Y, Dong L and Jiang H: Warfarin dosage response related pharmacogenetics in Chinese population. PLoS One. 10:e01164632015. View Article : Google Scholar : PubMed/NCBI

5 

Gage BF, Yan Y, Milligan PE, Waterman AD, Culverhouse R, Rich MW and Radford MJ: Clinical classification schemes for predicting hemorrhage: Results from the National Registry of Atrial Fibrillation (NRAF). Am Heart J. 151:713–719. 2006. View Article : Google Scholar : PubMed/NCBI

6 

Pisters R, Lane DA, Nieuwlaat R, de Vos CB, Crijns HJ and Lip GY: A novel user-friendly score (HAS-BLED) to assess 1-year risk of major bleeding in patients with atrial fibrillation: The euro heart survey. Chest. 138:1093–1100. 2010. View Article : Google Scholar : PubMed/NCBI

7 

Self TH, Oliphant CS, Reaves AB, Richardson AM and Sands CW: Fever as a risk factor for increased response to vitamin K antagonists: A review of the evidence and potential mechanisms. Thromb Res. 135:5–8. 2015. View Article : Google Scholar : PubMed/NCBI

8 

Harada T, Ariyoshi N, Shimura H, Sato Y, Yokoyama I, Takahashi K, Yamagata S, Imamaki M, Kobayashi Y, Ishii I, et al: Application of Akaike information criterion to evaluate warfarin dosing algorithm. Thromb Res. 126:183–190. 2010. View Article : Google Scholar : PubMed/NCBI

9 

Holmes MV, Hunt BJ and Shearer MJ: The role of dietary vitamin K in the management of oral vitamin K antagonists. Blood Rev. 26:1–14. 2012. View Article : Google Scholar : PubMed/NCBI

10 

You JH, Wong RS, Waye MM, Mu Y, Lim CK, Choi KC and Cheng G: Warfarin dosing algorithm using clinical, demographic and pharmacogenetic data from Chinese patients. J Thromb Thrombolysis. 31:113–118. 2011. View Article : Google Scholar : PubMed/NCBI

11 

Kakkar AK, Mueller I, Bassand JP, Fitzmaurice DA, Goldhaber SZ, Goto S, Haas S, Hacke W, Lip GY, Mantovani LG, et al: Risk profiles and antithrombotic treatment of patients newly diagnosed with atrial fibrillation at risk of stroke: Perspectives from the international, observational, prospective GARFIELD registry. PLoS One. 8:e634792013. View Article : Google Scholar : PubMed/NCBI

12 

International Warfarin Pharmacogenetics Consortium, ; Klein TE, Altman RB, Eriksson N, Gage BF, Kimmel SE, Lee MT, Limdi NA, Page D, Roden DM, et al: Estimation of the warfarin dose with clinical and pharmacogenetic data. N Engl J Med. 360:753–764. 2009. View Article : Google Scholar : PubMed/NCBI

13 

Miao L, Yang J, Huang C and Shen Z: Contribution of age, body weight, and CYP2C9 and VKORC1 genotype to the anticoagulant response to warfarin: Proposal for a new dosing regimen in Chinese patients. Eur J Clin Pharmacol. 63:1135–1141. 2007. View Article : Google Scholar : PubMed/NCBI

14 

Huang SW, Chen HS, Wang XQ, Huang L, Xu DL, Hu XJ, Huang ZH, He Y, Chen KM, Xiang DK, et al: Validation of VKORC1 and CYP2C9 genotypes on interindividual warfarin maintenance dose: A prospective study in Chinese patients. Pharmacogenet Genomics. 19:226–234. 2009. View Article : Google Scholar : PubMed/NCBI

15 

Zhong SL, Yu XY, Liu Y, Xu D, Mai LP, Tan HH, Lin QX, Yang M and Lin SG: Integrating interacting drugs and genetic variations to improve the predictability of warfarin maintenance dose in Chinese patients. Pharmacogenet Genomics. 22:176–182. 2012. View Article : Google Scholar : PubMed/NCBI

16 

Tan SL, Li Z, Song GB, Liu LM, Zhang W, Peng J, Zhang T, Jia FF, Zhou G, Zhou HH and Zhou XM: Development and comparison of a new personalized warfarin stable dose prediction algorithm in Chinese patients undergoing heart valve replacement. Pharmazie. 67:930–937. 2012.PubMed/NCBI

17 

Chen J, Shao L, Gong L, Luo F, Wang J, Shi Y, Tan Y, Chen Q, Zhang Y, Hui R and Wang Y: A pharmacogenetics-based warfarin maintenance dosing algorithm from Northern Chinese patients. PLoS One. 9:e1052502014. View Article : Google Scholar : PubMed/NCBI

18 

Blanco F, Muriel C, Labrador J, Gonzalez-Porras JR, Gonzalez-Sarmiento R and Lozano FS: Influence of UGT2B7, CYP3A4, and OPRM1 gene polymorphisms on transdermal buprenorphine pain control in patients with critical lower limb ischemia awaiting revascularization. Pain Pract. 16:842–849. 2016. View Article : Google Scholar : PubMed/NCBI

19 

Liu R, Zhou ZY, Chen YB, Li JL, Yu WB, Chen XM, Zhao M, Zhao YQ, Cai YF, Jin J and Huang M: Associations of CYP3A4, NR1I2, CYP2C19 and P2RY12 polymorphisms with clopidogrel resistance in Chinese patients with ischemic stroke. Acta Pharmacol Sin. 37:882–888. 2016. View Article : Google Scholar : PubMed/NCBI

Related Articles

Journal Cover

August-2017
Volume 14 Issue 2

Print ISSN: 1792-0981
Online ISSN:1792-1015

Sign up for eToc alerts

Recommend to Library

Copy and paste a formatted citation
x
Spandidos Publications style
Ma XW, Hao CN, Gu ZC, Ye M, Li M and Zhang L: Warfarin-induced life-threatening bleeding associated with a CYP3A4 loss-of-function mutation in an acute limb ischemia patient: Case report and review of the literature. Exp Ther Med 14: 1157-1162, 2017
APA
Ma, X., Hao, C., Gu, Z., Ye, M., Li, M., & Zhang, L. (2017). Warfarin-induced life-threatening bleeding associated with a CYP3A4 loss-of-function mutation in an acute limb ischemia patient: Case report and review of the literature. Experimental and Therapeutic Medicine, 14, 1157-1162. https://doi.org/10.3892/etm.2017.4604
MLA
Ma, X., Hao, C., Gu, Z., Ye, M., Li, M., Zhang, L."Warfarin-induced life-threatening bleeding associated with a CYP3A4 loss-of-function mutation in an acute limb ischemia patient: Case report and review of the literature". Experimental and Therapeutic Medicine 14.2 (2017): 1157-1162.
Chicago
Ma, X., Hao, C., Gu, Z., Ye, M., Li, M., Zhang, L."Warfarin-induced life-threatening bleeding associated with a CYP3A4 loss-of-function mutation in an acute limb ischemia patient: Case report and review of the literature". Experimental and Therapeutic Medicine 14, no. 2 (2017): 1157-1162. https://doi.org/10.3892/etm.2017.4604