A Low‐Molecular‐Weight Factor Extracted from Human Placenta Inhibits the Entry into Cell Cycle of Murine Pluripotent Stem Cells

In 1977, Frindel et al.' reported the presence in fetal calf bone marrow of a factor of low molecular weight capable of inhibiting in vivo the recruitment of hematopoietic murine pluripotent stem cells (CFU-S) into DNA synthesis. The semipurified factor increased the survival of mice that had received high doses of cytosine arabinoside (ara-C), a phase-specific drug.* The inhibitory molecule has been identified) as the tetrapeptide Ac-N-Ser-Asp-Lys-Pro (AcSDKP); it has recently been synthesized and is now produced by Beaufour Laboratories. Considering the potential clinical importance of CFU-S proliferation inhibitors for the protection of bone marrow during anticancer chemotherapy and the possibility of monitoring the inhibitor by immunoanalytical methods, we thought it necessary to determine if a similar inhibitor is present in man. Due to the difficulties in obtaining large amounts of normal human bone marrow, investigations were carried out on human term placenta, which is known to contain hematopoietic growth factors. The CFU-S inhibitory fraction was extracted from a pool of 5 kg of frozen human term placenta by the procedure of Lenfant et aL4 The biological activity assay was that previously describedS5 Briefly, 6 hours after injection of cytosine arabinoside (ara-C), mice received 100-200 mg/mouse of the placental fractions or saline (controls); 6 hours later their bone marrow was analyzed for CFU-S content and percentage in DNA synthesis of these cells. A competitive enzyme immunoassay (EIA) using a rabbit anti-AcSDKP antibody* was performed on HPLC fractions according to the technique of Pradelles et aL6 After chromatography on Biogel and Sep-Pak C 18 cartridges, two fractions, A', and B',, were selected because their retention time was identical to that of AcSDKP. They were tested for their inhibitory activity on murine CFU-S entry into cycle after treatment of mice with 20 mg of ara-C. As shown in TABLE 1, a significant inhibitory activity was found in peak A , but not in peak B14. When the HPLC fractions of semipurified placental extract were checked for the presence of AcSDKP activity in