PB0134 - A novel method for measuring ex vivo intrinsic pathway-triggered thrombin generation in the development of REGN9933, a monoclonal antibody targeting factor XI

Sunday, June 23, 2024

13:45 – 14:45 ICT

Room: Exhibition Hall

Presenting Author(s)

KM

Karoline A. Meagher

Associate Director-Precision Medicine Strategy
Regeneron Pharmaceuticals, Inc
Tarrytown, New York, United States

Co-Author(s)

EM

Ethan Marin

Medical Director
Regeneron Pharmaceuticals, Inc
Tarrytown, New York, United States
DG

David E. Gutstein

Vice President, Strategic Program Direction – Hematology Translational & Clinical Sciences
Regeneron Pharmaceuticals, Inc.
Tarrytown, New York, United States
KL

KehDih Lai

Manager Cardiovascular Research
Regeneron Pharmaceuticals, Inc.
Tarrytown, New York, United States
RD

Robert J. Durso

Precision Medicine Biomarker Assay Quality Clinical Genetic Medicines and Experimental Sciences
Regeneron Pharmaceuticals, Inc
Tarrytown, New York, United States
HS

Holly Sorenson

Preclinical Operations Associate II
Regeneron Pharmaceuticals, Inc.
Tarrytown, New York, United States
MB

Michael E. Burczynski

Executive Director, Precision Medicine
Regeneron Pharmaceuticals, Inc
Tarrytown, New York, United States
VS

Venkata Sabbisetti

Director, Precision Medicine
Regeneron Pharmaceuticals, Inc
Tarrytown, New York, United States
JK

Jayaprakash Kotha

Vice President of Research
MLM Medical Labs, LLC
Memphis, Tennessee, United States
LJ

Lisa K. Jennings

Chief Scientific Officer
MLM Medical Labs, LLC
Memphis, Tennessee, United States
DC

Dan Chalothorn

Director in Cardiovascular and Renal Research
Regeneron Pharmaceuticals, Inc.
Tarrytown, New York, United States

Background: Utilizing thrombin generation as a clinical endpoint is challenging due to the lack of an automated and standardized assay. A novel protocol using a real-time, calibrated automated thrombogram (CAT) was explored during development of REGN9933, an investigational monoclonal antibody for thrombosis treatment that targets coagulation factor XI and blocks the intrinsic coagulation pathway.

Aims: To evaluate a CAT protocol for assessment of intrinsic pathway-triggered thrombin generation in clinical biomarker samples.

Methods: Thrombin generation was measured in citrated platelet-poor plasma from healthy donors after triggering the intrinsic pathway with ellagic acid (Thermo Scientific) in the presence of microparticles (Stago). Activated thrombin cleaves a synthetic substrate, producing a measurable fluorogenic product that reflects thrombin-generation kinetics. REGN9933/isotype control reagents and thrombin-generation methodology were developed and optimized using normal pooled plasma samples (Regeneron); all assays were performed on a CAT device with standard assay calibration. Validation of the fit-for-purpose clinical assay was conducted externally (MLM Medical Labs). The effects of intrinsic pathway blockade using REGN9933 versus isotype control were evaluated in naïve plasma from 20 healthy donors.

Results: In normal pooled plasma, ellagic acid induced rapid and robust increases in thrombin generation. Incubation with REGN9933 at 16–250 nM for 30 minutes inhibited intrinsic pathway-triggered thrombin generation in a dose-dependent manner at Regeneron laboratories (Figure 1A); isotype control had no effect. Results were replicated in normal pooled plasma at MLM Medical Labs (Figure 1B). Validation of the fit-for-purpose clinical assay in plasma from healthy donors also showed dose-dependent inhibition of thrombin generation with REGN9933, prior to analysis of clinical samples.

Conclusion(s): These data validate the CAT assay for reporting intrinsic pathway-triggered thrombin generation using ellagic acid. This assay has utility in clinical testing of anti-thrombotic compounds, including REGN9933, which demonstrated potent inhibition of intrinsic pathway-triggered thrombin generation.