Solving the Mystery: Heparin-induced Thrombocytopenia Without Heparin?

By Bryan Lin, PhD Candidate at Pryzdial Lab

Heparin is a natural anticoagulant that derives its name from the tissue it was first isolated from: hepar in Greek means liver. Listed in the WHO Model List of Essential Medicines, heparin has stood the test of time as one of the oldest drugs still employed in clinical treatment to reduce clotting. However, heparin use is not without consequence as its usage can result in an auto-immune clotting disorder known as heparin-induced thrombocytopenia (HIT). Interestingly, HIT can persist even after heparin has cleared from the body – a mystery that the CBR researchers aimed to solve.

Heparin can form a complex with another protein known as platelet factor 4 (PF4). PF4 is released by activated platelets in the blood and functions to neutralize heparin-like molecules, thus serving as a pro-coagulant. Some patients on heparin treatment can produce antibodies that recognize this PF4/Heparin complex. These antibodies are characteristic of HIT, with the resulting immune complex activating platelets and, in turn, contributing to the formation of a clot.

Unfortunately, the risk of thrombosis can persist for weeks after heparin therapy has ended even though heparin is rapidly cleared from the body. As the exact mechanisms that contribute to this recurrence of thrombosis had not been discovered, a collaborative effort by several investigators at renown institutions: Dr. Douglas Cines of the University of Pennsylvania Perelman School of Medicine and Drs. Hugh Kim and Ed Conway of the Centre for Blood Research, set out to explain this phenomenon.

A key segue to the findings, was that PF4 can bind to many manners of negatively-charged molecules. It is no secret that Dr. Conway’s laboratory is interested in the highly negatively-charged polyphosphates (polyPs), chains of phosphates of varying lengths that are found in all mammalian cells.

The collaborative effort, published in Blood Advances, found that HIT-inducing antibodies can bind to PF4 complexed with polyP. The PF4/polyP complex shares several characteristics with the classical PF4/heparin complex: a) both can form large complexes; b) both can be recognized by HIT-inducing antibodies; c) both can be taken up by macrophages; d) both activate complement; and e) in the presence of HIT-antibodies, both complexes can cause platelet activation and aggregation.

The recent publication describes the discovery that HIT-inducing antibodies can stabilize PF4/polyp complex, similarly to their activity with PF4/heparin, and thereby enhance complement protein deposition on these complexes. This feature potentially makes PF4/polyp complex act as a middleman to activate complement on the platelet surface and produce platelet activation.

Since human platelets are a rich source of polyPs, releasing them upon platelet activation, there was further interest in whether platelet polyPs can bind to PF4 to form complexes that could trigger HIT. Dr. Kim investigated this aspect using confocal microscopy and direct immunolocalization. It was found that polyP and PF4 were separate from one another in resting platelets but coalesced upon platelet activation.

PF4/polyP complex formation and binding to HIT-inducing antibodies can explain why patients that come off heparin can still experience complications of HIT. Now, researchers armed with this new evidence must still determine how PF4/polyP bind to platelets and the contribution of polyP to the development of HIT. This research has a large potential to open more strategies for combatting HIT.