Scientists screened thousands of molecules to develop Hympavzi

Researchers spent years searching for a new treatment option for people with hemophilia A or B before arriving at the therapy that became Hympavzi (marstacimab).

Scientists at Pfizer, the company that markets Hympavzi, described the search in a study, “Discovery and optimization of marstacimab, a human monoclonal antibody targeting tissue factor pathway inhibitor for the treatment of hemophilia A and B,” published in mAbs.

The U.S. Food and Drug Administration’s (FDA) 2024 approval of Hympavzi marked the arrival of an alternative to traditional treatments that require burdensome infusions and, for some patients, lose effectiveness over time. Hympavzi is available as a once-weekly, under-the-skin injection, delivered via a prefilled syringe or pen, that can be administered quickly at home.

Hemophilia refers to a group of disorders in which a person’s blood cannot clot properly due to a missing or faulty clotting protein, specifically factor VIII (FVIII) in hemophilia A and factor IX (FIX) in hemophilia B. As a result, even minor injuries can lead to unusually prolonged bleeding episodes.

For decades, treatment largely relied on factor replacement therapies, which supply the body with the missing clotting factor either on demand to treat active bleeds or as prophylaxis to help prevent them. While these therapies can be highly effective, they require regular intravenous (into-the-vein) infusions, and they may lose effectiveness in some patients who develop inhibitors (antibodies that block the activity of the infused FVIII or FIX).

Recommended Reading

Jan. 8, 2025 Columns by Cazandra Campos-MacDonald

We must avoid being complacent with hemophilia treatment

Search for sustained protection begins with thousands of candidates

Unlike factor replacement therapies, Hympavzi is an antibody-based therapy that targets tissue factor pathway inhibitor (TFPI), a protein that acts as a natural brake on blood clotting. By blocking TFPI, the therapy helps increase thrombin production, a key protein involved in blood clot formation. This is intended to restore the blood’s ability to clot, thereby preventing or reducing bleeding episodes.

TFPI had long been considered an attractive treatment target. However, as the researchers noted, previously developed anti-TFPI antibodies, while “capable of potent TFPI neutralization,” showed a “short half-life in animal studies,” meaning they were cleared from the body relatively quickly.

The team sought an antibody that could strongly block TFPI to improve clotting while remaining active in the body long enough to provide sustained protection against bleeding with a weekly injection.

The search began with thousands of potential antibody candidates. After several rounds of screening, researchers narrowed the field to 288 unique antibodies that could bind to TFPI, ultimately identifying 48 that could effectively block TFPI’s activity.

Three lead candidates — TFPI-21, T.FPI-23, and TFPI-24 — were selected for further study, along with a fourth candidate, 4D8, identified through a separate screening approach. Further testing showed that 4D8 was the most potent TFPI-neutralizing antibody and bound most strongly to human TFPI, followed by TFPI-24, TFPI-23, and TFPI-21.

All four antibodies showed a similar ability to restore thrombin generation in human FVIII-deficient blood samples. A single dose of TFPI-21, TFPI-23, or TFPI-24 reduced bleeding after injury in mouse models of hemophilia A. 4D8 was not effective in this model because it did not bind mouse TFPI.

The researchers next turned to a challenge critical to a once-weekly treatment: finding an antibody that could effectively block TFPI while remaining in the bloodstream long enough to provide sustained protection against bleeding.

Studies in rabbits revealed that both 4D8 and TFPI-24 approached the limit of detection within four to six days after dosing, whereas TFPI-23 remained detectable for more than a week. Computer modeling further suggested that, despite binding to human TFPI less tightly than 4D8, TFPI-23’s longer persistence in the bloodstream could help maintain more consistent TFPI blockade between weekly doses.

Seeking the right balance between potency and durability, the researchers advanced TFPI-23 and TFPI-24 — the two antibodies with intermediate binding affinity — for further optimization. This process produced TFPI-106, derived from TFPI-23, and TFPI-118, derived from TFPI-24.

Both optimized candidates retained strong clot-promoting activity in human FVIII-deficient blood samples and reduced bleeding in mouse models of hemophilia A. They were then evaluated for additional characteristics important for long-term therapy, including stability, manufacturability, and low likelihood of triggering unwanted immune responses.

Based on the overall dataset, TFPI-106 was selected as the final clinical candidate. The molecule, later named marstacimab, would eventually be tested in the pivotal Phase 3 BASIS trial (NCT03938792), in which once-weekly treatment significantly reduced bleeding rates in people with hemophilia A or B, ultimately leading to its approval as Hympavzi.