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Diagnosis and Discussion: Case 1175

Further evaluation revealed normal recent PT and PTT results, suggesting that the patient’s prior hepatic coagulopathy had resolved. The factor VIII level had also normalized. Factor VIII is produced primarily by endothelial cells rather than hepatocytes and is often elevated in chronic liver disease and portal hypertension. Its normalization further supported resolution of the patient’s cirrhosis-associated coagulation abnormalities1. Review of the electronic medical record showed that the patient had undergone deceased donor liver transplant. This explained the normal APC-R result, as the donor liver was producing normal factor V.

Factor V Leiden is the most common inherited thrombophilia caused by a point mutation in the F5 gene, F5: c.1601G>A (p.Arg534GIn or R534Q); legacy nomenclature: c.1691G>A (p.Arg506GIn or R506Q), which produces an arginine-to-glutamine substitution at the protein C cleavage site in factor V. Under normal physiologic conditions, activated protein C, with protein S as a cofactor, acts as an anticoagulant by inactivating factors Va and VIIIa. In Factor V Leiden, the altered factor V molecule is resistant to cleavage by activated protein C, promoting a prothrombotic state2,3. Although Factor V Leiden accounts for most hereditary APC resistance, other rare F5 variants can alter factor V to reduce susceptibility to activated protein C4.

Factor V Leiden is more common in Caucasians (up to 5%) and rare in other ethnic groups. Heterozygosity for factor V Leiden is more common than homozygosity and is associated with a modest 3- to 7-fold increase in the risk of venous thrombosis compared to an approximately 80-fold increase in risk in homozygotes. The combination of factor V Leiden with other risk factors for venous thrombosis, such as antithrombin or protein S deficiency, further increase the relative risks of venous thrombosis 3,4.The APC-R assay is a functional screen that assesses whether the time to clot formation increases with added activated protein C. In normal plasma, activated protein C inactivates factors Va and VIIIa, prolonging the time to clot formation. In Factor V Leiden, the  R534Q (previously R506Q)variant makes factor V resistant to cleavage by activated protein C, so the prolongation in clotting time relative to baseline is reduced2.  The result is reported as the ratio of the clotting time with added activated protein C to the baseline clotting time. A ratio less than 2 is suggestive of a factor V variant. Current APC-R testing is performed using a second-generation assay in which patient plasma is mixed with factor V-deficient plasma. This corrects for elevations or deficiencies in other coagulation factors, such as in liver disease, warfarin therapy, or pregnancy. Rarely, APC-R assays can be falsely low in the setting of a lupus anticoagulant or falsely normal in the presence of direct thrombin inhibitors and factor Xa inhibitors2,5.

Genetic testing for Factor V Leiden directly evaluates the patient’s DNA for the pathogenic F5 variant, most commonly using peripheral blood leukocytes as the source of DNA. This is typically performed by quantitative PCR (qPCR)-based targeted mutation testing. APC-R testing reflects the functional phenotype of circulating factor V protein in plasma, whereas genetic testing reflects the genotype of the nucleated cells tested2,3.

Factor V is primarily synthesized in hepatocytes. After liver transplantation, the donor liver produces the circulating factor V. If the recipient has a germline Factor V Leiden variant but receives a donor liver that produces wildtype factor V, the circulating factor V will be functionally normal and the APC-R will also be normal. This explains the patient’s later APC-R result because the donor-derived hepatic factor V functionality is being evaluated, rather than the recipient’s original factor V. If genetic testing were repeated, the peripheral blood leukocytes would still demonstrate the Factor V Leiden mutation, as leukocytes are produced in the bone marrow. Prior case reports have shown that APC-R can normalize in patients with Factor V Leiden after orthotopic liver transplantation, while activated protein C resistance can also be acquired if the liver donor had a Factor V Leiden variant6,7.

Discrepant results in APC-R and germline Factor V Leiden testing may also be seen in a patient with Factor V Leiden who receives an allogenic stem cell transplant. Because the patient’s liver is still making  a variant factor V, the APC-R would be low. However, circulating leukocytes are made in the bone marrow and reflect the genotype of the stem cell donor. In such a patient, DNA from another source such as a buccal swab should be tested to confirm the diagnosis of Factor V Leiden8.

Ultimately, the normal APC-R assay demonstrated that the patient’s liver donor did not have Factor V Leiden and the liver is producing phenotypically normal factor V. This allowed the patient to consider discontinuation of anticoagulation.

References

  1. Pradhan-Sundd T, Gudapati S, Kaminski TW, Ragni MV. Exploring the Complex Role of Coagulation Factor VIII in Chronic Liver Disease. Cell Mol Gastroenterol Hepatol. 2021;12(3):1061-1072. doi:10.1016/j.jcmgh.2021.02.014
  2. Kadauke S, Khor B, Van Cott EM. Activated protein C resistance testing for factor V Leiden. Am J Hematol. 2014;89(12):1147-1150. doi:10.1002/ajh.23867
  3. Van Cott EM, Khor B, Zehnder JL. Factor V Leiden. Am J Hematol. 2016;91(1):46-49. doi:10.1002/ajh.24222
  4. Mohapatra AK, Todaro AM, Castoldi E. Factor V variants in bleeding and thrombosis. Res Pract Thromb Haemost. 2024;8(1):102330. Published 2024 Jan 26. doi:10.1016/j.rpth.2024.102330
  5. Moore GW, Van Cott EM, Cutler JA, Mitchell MJ, Adcock DM; subcommittee on plasma coagulation inhibitors. Recommendations for clinical laboratory testing of activated protein C resistance; communication from the SSC of the ISTH. J Thromb Haemost. 2019;17(9):1555-1561. doi:10.1111/jth.14532
  6. Loew A, Jacob D, Neuhaus P, Riess H. Resistance to activated protein C caused by factor V Leiden mutation and orthotopic liver transplantation. Transplantation. 2005;79(10):1422-1427. doi:10.1097/01.tp.0000158021.66004.19
  7. Hirshfield G, Collier JD, Brown K, et al. Donor factor V Leiden mutation and vascular thrombosis following liver transplantation. Liver Transpl Surg. 1998;4(1):58-61. doi:10.1002/lt.500040108
  8. Crookston KP, Henderson R, Chandler WL. False negative factor V Leiden assay following allogeneic stem cell transplant. Br J Haematol. 1998;100(3):600-602. doi:10.1046/j.1365-2141.1998.00585.x