DESCRIPTION (provided by applicant): Chronic thrombo-embolic pulmonary hypertension (CTEPH) is characterized by progressive right ventricular failure and, if left untreated, death. It is preceded by acute pulmonary thrombo-embolism. Unlike other patients with acute pulmonary embolism, however, patients with CTEPH do not lyse the thrombotic material within the pulmonary arteries. Instead, the thrombi are remodeled into intravascular scars that obstruct flow, often with disastrous consequences. Our goal is to understand the mechanisms responsible for the failure to resolve the acute thrombo-emboli in CTEPH. Our previous studies had disclosed no defect within fibrinolytic enzymes in these patients. However, we demonstrated that fibrinogen purified from CTEPH patients is somewhat resistant to lysis by exogenous plasmin. For this reason, we plan to search within the CTEPH patient population for genetic variants of fibrinogen and post-translational modifications which might make it resistant to fibrinolysis. Fibrinogen variants will be searched for in CTEPH patients using three complementary methods. First, purified fibrinogen will be polymerized and cross-linked to simulate normal thrombosis, then exposed to plasmin under controlled conditions to determine the profile of peptide fragments yielded during fibrinolysis, as well as the kinetics of their release. Next, genetic variants in targeted portions of fibrinogen will be looked for using the polymerase chain reaction (PCR) and DMA sequencing. Finally, primary amino acid sequence and post translational modifications will be searched for in fibrinogen from CTEPH, using purified fibrinogen samples fragmented with proteases. The fragments will be subjected to liquid chromatography-mass spectroscopy with data-dependent MS-2 data acquisition, followed by computer-assisted sequence analysis.