Targeting protein disulfide isomerase with the flavonoid isoquercetin to improve hypercoagulability in advanced cancer

Abstract

Background: Protein disulfide isomerase (PDI) is a thiol isomerase secreted by vascular cells that is required for thrombus formation. Quercetin flavonoids inhibit PDI activity and block platelet accumulation and fibrin generation at the site of a vascular injury in mouse models, but the clinical effect of targeting extracellular PDI in humans has not been studied.

Methods: We conducted a multicenter phase II trial of sequential dosing cohorts to evaluate the efficacy of targeting PDI with isoquercetin to reduce hypercoagulability in cancer patients at high risk for thrombosis. Patients received isoquercetin at 500 mg (cohort A, n = 28) or 1000 mg (cohort B, n = 29) daily for 56 days, with laboratory assays performed at baseline and the end of the study, along with bilateral lower extremity compression ultrasound. The primary efficacy endpoint was a reduction in D-dimer, and the primary clinical endpoint included pulmonary embolism or proximal deep vein thrombosis.

Results: The administration of 1000 mg isoquercetin decreased D-dimer plasma concentrations by a median of -21.9% (P = 0.0002). There were no primary VTE events or major hemorrhages observed in either cohort. Isoquercetin increased PDI inhibitory activity in plasma (37.0% in cohort A, n = 25, P < 0.001; 73.3% in cohort B, n = 22, P < 0.001, respectively). Corroborating the antithrombotic efficacy, we also observed a significant decrease in platelet-dependent thrombin generation (cohort A median decrease -31.1%, P = 0.007; cohort B median decrease -57.2%, P = 0.004) and circulating soluble P selectin at the 1000 mg isoquercetin dose (median decrease -57.9%, P < 0.0001).

Conclusions: Isoquercetin targets extracellular PDI and improves markers of coagulation in advanced cancer patients.

Trial registration: Clinicaltrials.gov NCT02195232.

Funding: Quercegen Pharmaceuticals; National Heart, Lung, and Blood Institute (NHLBI; U54HL112302, R35HL135775, and T32HL007917); and NHLBI Consortium Linking Oncology and Thrombosis (U01HL143365).

Keywords: Clinical Trials; Coagulation; Hematology; Thrombosis.

Conflict of interest statement

Conflict of interest: JIZ serves as a consultant for Parexel and served on advisory boards for Bayer, Seattle Genetics, and Incyte. TW receives research support and serves on advisory boards for Janssen and Pfizer. RF is a consultant for PlateletDiagnostics. JDS is an employee of Rubius Therapeutics. DN serves as a trustee with stock ownership in Madrigal Pharmaceuticals and receives research funding from Pharmacyclics. HAL reports spousal consulting income from Alexion. MH discloses equity in Champions Oncology and research support and/or honoraria from Celgene, Pfizer, Bioline, Asana, Erytech, Bicycle, Berg, Roche, EMD, Oncomatryx, MedImmune, Bayer, Novartis, Targovax, MSD, and Bioncotech.

Figures

Figure 1. Flow diagram of patients according to isoquercetin treatment allocation.

Figure 2. Measurement of D-dimer following administration of isoquercetin.

Waterfall plot showing baseline versus end-of-study comparisons of D-dimer values for each patient according to the dose of isoquercetin administered. (A) Median change in D-dimer was +9.9% (paired t test, P = 0.92) with 500 mg isoquercetin. (B) Median decrease in D-dimer was –21.9% with 1000 mg isoquercetin (P = 0.0002).

Figure 3. Cumulative incidence of venous thromboembolism.

Venous thromboembolisms (VTE) were monitored clinically and by lower extremity ultrasound at completion of the 2-month study. Shown is the proportion of patients remaining free of VTE through the course of the study. There were no primary VTE in either the 500-mg isoquercetin cohort (A) or the 1000-mg isoquercetin cohort (B). The cumulative incidence of all secondary VTE endpoints (i.e., superficial thrombosis and distal thrombosis) shown in blue for both the 500-mg cohort (C) and the 1000-mg cohort (D).

Figure 4. Measurement of plasma PDI inhibitory activity following isoquercetin administration.

Waterfall plot showing baseline versus end-of-study comparisons for each cancer patient. (A) Median change in PDI inhibitory activity was +37.0% (paired t test, P < 0.001) with 500 mg isoquercetin. (B) Median change in PDI inhibitory activity was +73.3% with 1000 mg isoquercetin (P < 0.001).

Figure 5. Measurement of platelet-dependent thrombin generation activity following isoquercetin administration.

Waterfall plot showing baseline and follow-up platelet-dependent thrombin generation following isoquercetin administration. Change (%) for each patient shown in a waterfall plot for 500 mg isoquercetin (A) and 1000 mg isoquercetin (B).

Figure 6. Measurement of plasma P selectin levels following isoquercetin administration.

Waterfall plot showing baseline and follow-up soluble P selectin values in plasma following isoquercetin administration. Change (%) for each patient shown in a waterfall plot for 500 mg isoquercetin (A) and 1000 mg isoquercetin (B).