Researchers have discovered the mechanism behind the deaths and adverse events that occurred in patients receiving contaminated heparin.
In March, a team led by Ram Sasisekharan, PhD, of Massachusetts Institute of Technology in Cambridge, identified the contaminant responsible for the numerous adverse events and 81 deaths that have occurred since November 2007 in patients receiving heparin.
Now, Dr Sasisekharan and colleagues have identified the mechanism by which the contaminant, oversulfated chondroitin sulfate (OSCS), works. This finding was published in New England Journal of Medicine April 24.
The researchers found that OSCS activated the kinin-kallikrein pathway in human plasma, which can lead to the generation of the potent vasoactive mediator bradykinin. In addition, OSCS induced generation of C3a and C5a, which are potent anaphylatoxins derived from complement proteins.
Dr Sasisekharan’s team arrived at these conclusions by testing 29 lots of heparin obtained from the FDA. Thirteen of these lots had been associated with adverse events. A laboratory lot was also included to serve as a control.
In a blinded fashion, the researchers screened the heparin for the existence of OSCS. They then tested the effects heparin contaminated with 19.3% wt/wt OSCS had on human plasma.
At 2.5 µg/mL and 25 µg/mL, contaminated heparin showed activation of kallikrein, while the same doses of uncontaminated heparin did not.
At 250 µg/mL, the contaminated heparin did not demonstrate activation of kallikrein. Dr Sasisekharan and colleagues said this suggests that, at a high concentration, heparin may inhibit or cause the depletion of factor XII.
The researchers next examined the contaminated heparin for its ability to generate C3a and C5a. At 5 µg/mL and 50 µg/mL, contaminated heparin generated C5a, whereas the same doses of uncontaminated heparin did not. At 500 µg/mL, the contaminated heparin did not generate significant amounts of C5a.
Dr Sasisekharan and colleagues also found that activation of C3a and C5a were linked and dependent upon fluid-phase activation of factor XII.
To ensure the accuracy of these results, the researchers created synthetic OSCS via chemical sulfonation of chondroitin sulfate. This synthetic OSCS behaved in the same manner as the OSCS found in the contaminated lots of heparin— demonstrating activation of kallikrein and generating C3a and C5a.
In an attempt to better understand the effects of OSCS, the team tested their results on swine. Swine were chosen because their reactions to contaminated heparin were similar to those observed in humans.
Each pig received an infusion of 5mg of one of the following substances: control heparin, contaminated heparin, chondroitin sulfate A, or synthetic OSCS. The researchers monitored the pigs’ vital signs for an hour before the animals were euthanized. The team collected blood samples at baseline and 5, 10, 20, 40, and 60 minutes.
Six pigs received contaminated heparin. Of these, 2 experienced at least a 30% drop in blood pressure within the first 30 minutes after infusion. One pig experienced hypotension for more than 15 minutes.
In the pigs that received synthetic OSCS, adverse events were more severe. This was expected, as the dose of OSCS in this group was higher than that in the group of pigs receiving contaminated heparin. All pigs given synthetic OSCS experienced a profound drop in blood pressure and an increased heart rate. One pig had difficulty breathing.
None of the pigs given control heparin or chondroitin sulfate A experienced any adverse events.
Dr Sasisekharan and colleagues said the results of this study suggest that a simple in vitro bioassay could complement the tests currently used in the screening of heparin. This bioassay would uncover the presence of OSCS and other polysulfated contaminants that might cause patients harm.
