The use of haemoregulatory peptides (pEEDCK monomer and dimer) for reduction of cytostatic drug induced haemopoietic damage

Abstract

We have previously shown that the stem cell inhibitory peptide pGlu-Glu-Asp-Cys-Lys (pEEDCK monomer) leads to a good tolerance of otherwise lethal multiple ara-C doses and an increased survival of ara-C + peptide treated mice. This effect was due to the prevention of drug-induced CFU-S proliferation, thus keeping stem cells in a quiescent state insensitive to ara-C. Here we show that the pEEDCK monomer also inhibits stem cell proliferation after clinically relevant (non-lethal) ara-C doses. This leads to a sustained (100%) stem cell number in the femoral bone marrow, which was greatly reduced without protective peptide treatment (27%). We have measured the kinetics of influx of CFU-S into the empty S-phase (after two consecutive ara-C injections). This influx reached peak levels of 60–70%; pEEDCK treatment reduced it to 25–30%. Due to its cysteine content the pEEDCK monomer is easily oxidized and forms a symmetric disulfide-bonded dimer (pEEDCK) 2. This dimer is a potent stimulator of haemopoiesis. Various modes of protective peptide treatment (monomer and dimer) were investigated in conjunction with a standardized protocol of 2 × 300 mg/kg ara-C given 12 h apart. 1. (a) ara-monomer-ara: The administration of pEEDCK-monomer 2 h before the second ara-C injection retarded the onset of neutropenia, shortened its duration and improved recovery after depression. The degree of short-term neutropenia was not changed. 2. (b) ara-ara-HN2-dimer: Post chemotherapy infusion of the stimulatory (pEEDCK) 2 dimer led to considerable increases of progenitor levels (6.8 CFU-GM/1000 bone marrow cells vs. 1.2 CFU-GM/1000 in normal mice) 2 days after cytostatic treatment when CFU-GM were not detectable in unprotected mice. This increase was followed by greatly elevated granulocyte counts (8000 PMN/mm 3 vs. 750 PMN/mm 3 in normal mice). In the dimer-treated mice, up to 75% of the peripheral leukocytes were mature PMN (normal, 10%). 3. (c) ara-monomer-ara-dimer: ara-C and monomer treatment as above (a) followed by dimer infusion led to complete protection of haemopoiesis. Mice treated with the protective pEEDCK monomer plus stimulatory dimer did not develop the leukocyte depression seen in unprotected animals. Our results show that the haemoregulatory peptide monomer and dimer can be used to improve the haemotological status of mice treated with clinically relevant doses of cytostatic drugs (anti-metabolite and alkylating, alone and in combination). The pEEDCK monomer and dimer are equally active also on human haemopoietic cells. They are highly specific for haemopoiesis and lack toxic side-effects. Both peptides can be obtained easily by chemical synthesis. They are thus highly promising candidates for use as haemoprotectors in cancer chemotherapy.