The effect of a novel, small non-peptidyl molecule butyzamide on human thrombopoietin receptor and megakaryopoiesis.
BACKGROUND
Thrombocytopenia is a common problem in the management of patients with cancer and other conditions that affect hematopoietic cells. In previous clinical trials, the polyethylene-glycol-conjugated recombinant human megakaryocyte growth and development factor increased platelet counts in patients with idiopathic thrombocytopenic purpura and solid tumors undergoing chemotherapy. However, antibodies to polyethylene-glycol-conjugated recombinant human megakaryocyte growth and development factor develop in healthy volunteers and patients undergoing chemotherapy and cross-react with endogenous thrombopoietin. As a result, clinical development of polyethylene-glycol-conjugated recombinant human megakaryocyte growth and development factor was discontinued in 1998. The aim of this study was to identify an orally bioavailable human Mpl activator that does not develop autoantibodies against endogenous thrombopoietin.


METHODS
We screened our chemical library and created a novel non-peptidyl thrombopoietin receptor, Mpl activator named butyzamide. We evaluated the effect of butyzamide on megakaryopoiesis in vitro using Ba/F3 cells expressing Mpl and human hematopoietic stem cells. For the evaluation of its in vivo effect, we administered butyzamide orally to immunodeficient NOD/Shi-scid,IL-2R gamma(null) (NOG) mice transplanted with human fetal liver-derived CD34(+) cells and investigated the production of human platelets.


RESULTS
Butyzamide specifically reacted with human Mpl and activated the same signal transduction pathway as thrombopoietin. However, unlike thrombopoietin, butyzamide did not react with murine Mpl and was shown to require the histidine residue in the transmembrane domain of Mpl for its agonistic activity. Butyzamide induced colony-forming unit-megakaryocytes and polyploid megakaryocytes from human CD34(+) hematopoietic progenitor cells, and its effects were comparable to those of thrombopoietin. When butyzamide was administered orally at the doses of 10 and 50 mg/kg for 20 days to NOG mice transplanted with human fetal liver-derived CD34(+) cells, the human platelet count increased by 6.2- and 22.9-fold, respectively.


CONCLUSIONS
Butyzamide is an orally bioavailable human Mpl activator, and appears to have potential for clinical development as a therapeutic agent for patients with thrombocytopenia.
Nogami
Wataru
W
Division of Hematology, Department of Internal Medicine, Keio University School of Medicine, Tokyo, Japan.
Yoshida
Hiroshi
H
Koizumi
Kenzo
K
Yamada
Hajime
H
Abe
Kenji
K
Arimura
Akinori
A
Yamane
Noriko
N
Takahashi
Koji
K
Yamane
Akiko
A
Oda
Atsushi
A
Tanaka
Yoshikazu
Y
Takemoto
Hiroshi
H
Ohnishi
Yasuyuki
Y
Ikeda
Yasuo
Y
Miyakawa
Yoshitaka
Y
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D001792:Blood Platelets / Q000166:cytology / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008533:Megakaryocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008689:Methacrylates; D051379:Mice; D015394:Molecular Structure; D010455:Peptides / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects; D053628:Receptors, Thrombopoietin / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D013844:Thiazoles / Q000737:chemistry / Q000494:pharmacology*; D013926:Thrombopoietin / Q000819:agonists / Q000378:metabolism
D018952:Antigens, CD34; D008689:Methacrylates; D010455:Peptides; D053628:Receptors, Thrombopoietin; D013844:Thiazoles; C534742:butyzamide; C082218:MPL protein, human; D013926:Thrombopoietin