Tissue plasminogen activator (t-PA) is a heart-attack clot-buster oftendescribed as "Genentech's blockbuster drug." Launched in 1987, therecombinant, fibrin-dissolving enzyme had by 1990 captured two-thirds of the $300-million-plus market for reopening blocked coronaryarteries in acute myocardial infarction. (See BioWorld Today, Sept. 7,1993, p. 1.)Coronaries aren't the only arteries dammed by life-threatening bloodclots. Thrombosis of vessels feeding the brain cause stroke; thosesupplying lungs produce pulmonary embolisms. Clots that occludearteries that nourish arms and legs usually require drastic surgery, oftenamputation.A Genentech-sponsored, 300-patient clinical trial to inject clot-threatened extremities with local doses of t-PA ended in failure lastyear, said the company's chief medical officer, "because of difficultiesplacing the catheter delivering the drug into the clot." (See BioWorldToday, Aug. 23, 1993, p. 3.)Now gene therapist Richard Mulligan, a member of the WhiteheadInstitute and professor of molecular biology at Massachussetts Instituteof Technology, has begun animal experiments to generate t-PAgenetically inside the vessel walls of limbs at thrombotic risk.What's more, his current inquiry into the effects of plasminogenactivator genes has led to a collaboration with Columbia neurobiologistEric Kandel, to test t-PA's possible role in memory and cognition.Mulligan began by creating a colony of mice in which he had knockedout either the gene for t-PA or that for u-PA (urokinase), or both.(Physicians have long used Abbott Laboratories' injectable urokinaseas a thrombolytic agent.) The report of his preliminary results, in thisweek's Nature (March 31), is titled "Physiological consequences ofloss of plasminogen activator gene function in mice."These consequences, Mulligan told BioWorld, are "very surprising,"because they are so mild. "Neither of these genes," he said, "playscritical roles in many of the processes in which they were thought toplay" _ notably, thrombosis, atherosclerosis, glomerulonephritis,acute respiratory distress syndrome, ovulation, and pathological cellinvasion in neoplasia and metastasis.Mulligan undertook these experiments because, despite thesesuppositions, "To date, genetic deficiencies of t-PA or u-PA have notbeen reported in man," his Nature paper noted.To be sure, mice deprived of both t-PA and u-PA gene functiondeveloped spontaneous fibrin deposits in liver, intestines, gonads andlungs, and non-healing skin and ear ulcers. Morover, they lysed bloodclots at a far lower rate than did animals lacking only one of the twogenes. Loss of only one had far less horrendous systemic effects ingeneral."This means," Mulligan observed, "that we need to think more aboutwhether or not other roles or other molecules are much more important.From Rescuing Hearts to Saving LimbsThe most telling result was the in vitro finding that circulating t-PA invery low concentrations "clearly plays a role in spontaneousdissolution of clots."Applying this tip-off to local limb thrombolysis, he and his group havea paper in the journal CIrculation, "that actually describes the generaltechnique where we modify endothelial cells [by engraftingplasminogen activator genes], then `paste' them back on the occludedvessels." He was able to show "that we can obtain attachment andpersistence of the cells where we put them, and that they do express thegenes."His animal model for these studies is the pig, which has blood vesselsclose in size and consistency to human coronary arteries. He envisagesusing this local gene therapy to repair the damage done to arterial wallsby such invasive procedures as angioplasty or arteriectomy, whichprovoke clot formation.Mulligan's group has already constructed retroviral vectors that expresst-PA, and are "in the midst of experiments where we are indeedresurfacing regions of the vasculature with transduced cells that expressits gene."A `New Twist' _ t-PA on the BrainWhat he describes as "a new twist we are now following up" involvesjoining forces with brain-researcher Eric Kandel, a senior investigatorof the Howard Hughes Medical Research Institute, and professor ofneurobiology at Columbia University.Kandel perceives a role for t-PA in the learning process called long-term potentiation (LTP). He told BioWorld that "LTP doubles synapticstrength in the hippocampus," and added, "so Richard Mulligan and hiscolleagues, and I and my colleagues, are exploring the consequences ofthe defect" in his knockout mice. Kandel is looking at how missing t-PA affects the animals' learning capability."We are quite far along," he said and observed, "The beauty aboutRichard's study is that he's also knocked out u-TPA, which doesnothing, as far as one knows, for learning or memory." n
-- David N. Leff Science Editor
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