Thrombopoietin Raises Platelet Counts in Animals

NASHVILLE--In the last year, at least four groups have reportedcloning proteins that appear to be the long-elusive megakaryocytecolony-stimulating growth factor thrombopoietin, which acts viathe mpl cell surface receptor.

Laboratory studies have shown that thrombopoietin appears to haveboth early- and late-acting effects on the mega-karyocytic lineage.It results in increases in the number, size, and ploidy of mega-karyocytecolonies in bone marrow and has a potent effect on megakaryocytematuration, leading to increased numbers of circulating platelets.

In a study presented at the scientific sessions of the AmericanSociety of Hematology (ASH) meeting, use of a recombinant mousethrombopoietin developed by ZymoGenetics, Inc. (Seattle) was shownto stimulate recovery of platelets and, unexpectedly, red bloodcells in myelosuppressed mice.

"These results indicate that cells able to respond to thegrowth factor are present in the marrow after cytoreductive therapy. . . and suggest that thrombopoietin may be useful clinicallyin speeding the recovery of both megakaryocytic and erythroidlineages in states of marrow failure," Katherine Sprugel,PhD, director of preclinical studies at ZymoGenetics, said inher presentation.

Initial studies to determine effective doses found that 25,000units/day produced circulating platelet levels in normal micemore than fourfold greater than baseline, but the effect droppedoff at higher doses, suggesting that the dose-response curve maybe bell-shaped, Dr. Sprugel said. After treatment was stopped,platelet counts returned to normal within 7 to 10 days.

Despite the high circulating platelet counts, no overt toxicitywas seen in any of the mice, and histopathology evaluation atthe end of the study was unremarkable, she said.

To test the effectiveness of the agent, a myelosuppressed rodentmodel was required, and Dr. Sprugel's group used a model developedat Genetics Institute (Cambridge, Mass). These mice were exposedto single doses of sublethal total body irradiation and a singleinjection of carboplatin (Paraplatin). Starting 24 hours later,the mice received daily intraperitoneal injections of vehicleor thrombopoietin (25,000 or 75,000 units/day) for 18 days.

The mice receiving thrombopoietin showed a dramatic improvementin recovery of circulating platelets, reaching baseline levels10 to 14 days earlier than the vehicle-treated group. There wasno significant difference in effect between the two doses. Inaddition, Dr. Sprugel said, the fact that treatment with thrombopoietinalone was effective suggests that additional exogenous cytokineswill not be required for effective therapy.

An unexpected finding, Dr. Sprugel said, was the dramatic effectof thrombopoietin on erythrocyte recovery, indicating that theagent may not be lineage specific. Therapy with thrombopoietinreduced the red blood cell nadir and improved and sped the recoveryof red blood cell counts.

"This effect appears to be due to an increased red bloodcell production rather than decreased bleeding," she saidin her presentation. In contrast, thrombopoietin had little orno effect on the recovery of the myeloid lineage.

A questioner from the audience noted that the mouse studies maynot be transferable to the clinic, since transplant patients willhave much lower platelet counts than could be produced in themice. Dr. Sprugel responded that, obviously, patients would notneed transfusion at the platelet counts produced in the mice,but "it is very difficult to keep rodents alive at the sortof platelet levels that would be clinically relevant."