Clot penetration and retention by plasminogen activators promote fibrinolysis

Tissue-type plasminogen activator (tPA) remains the sole thrombolytic approved by the FDA for the treatment of pulmonary embolism (PE). tPA has not been replaced by third generation plasminogen activators, e.g. Reteplase (Ret) and Tenecteplase (TNK) that circulate with longer life-spans and in theory should have more extended potency in vivo. One reason for this paradox is the inability to assign units of activity to plasminogen activators based on specific biologically relevant standards, which impairs objective comparison. Here, we compare clot permeation, retention and fibrinolytic activities of tPA, TNK and Ret in vitro and clot composition over time with outcome in a mouse model of disseminated pulmonary microembolism (ME). When clots were incubated in the continuous presence of drug, tPA, TNK and Ret lysed fibrin clots identically in the absence of PA inhibitor-1 (e.g. PAI-1). Ret, which has lower fibrin affinity and greater susceptibility to inhibition by PAI-1 than tPA, was less effective in lysing plasma clots, while TNK was less effective when the fibrin content of the clots was enhanced. However, when clots were afforded only brief exposure to drug, as occurs in vivo, Ret showed more extensive clot permeation, greater retention and lysis than tPA or TNK. These results were reproduced in vivo in a mouse model of ME. These studies indicate the need for more relevant tests of plasminogen activator activity in vitro and in vivo and they show that clot permeation and retention are important potential predictors of clinical utility.