Distinct Gene Expression Pattern of CD34+ Stem and Progenitor Cells Mobilized by Pegfilgrastim after Cytotoxic Therapy in Multiple Myeloma in Comparison to G-CSF.

Background: Current regimens for peripheral blood stem cell (PBSC) mobilization in patients with multiple myeloma are based on daily subcutaneous injections of G-CSF starting shortly after cytotoxic therapy. Recently a polyethylenglycole (PEG)-conjugated G-CSF (pegfilgrastim) has been introduced which has a substantially longer half-life than the original formula. Here, we compared the molecular phenotypes of CD34+ stem and progenitor cells mobilized by G-CSF with those mobilized by pegfilgrastim.

Study design and Methods: We examined immunomagnetically enriched CD34+ cells from leukapheresis products of 8 patients who received G-CSF and of 8 patients who were given pegfilgrastim using Affymetrix HG Focus GeneChips covering 8793 genes. The statistical scripting language ‘R’ was used for data analysis. Significantly differentially expressed genes were identified with the Significance Analysis of Microarrays (SAM) algorithm.

Results: Comparing CD34+ cells mobilized by G-CSF with pegfilgrastim-mobilized CD34+ cells 108 genes were differentially expressed (fold change 1.25 – 14.0, q- value 2.45–14.44%). 38 genes had a higher and 70 genes had a lower expression in CD34+ cells mobilized by G-CSF.

We found upregulation of genes characteristic for erythropoietic differentiation including haemoglobin chains and Erythroid Kruppel-like factor in G-CSF-mobilized CD34+ cells. Utilizing clonogenic assays we were able to functionally corroborate this finding as G-CSF-mobilized cells gave rise to a significantly higher number of burst-forming units erythroid (BFU-E) as compared to colony forming units granulocyte-macrophage (CFU-GM) (p=0.016). Cell cycle regulating genes were differentially expressed as well. Genes encoding for proteins that cause cell cycle arrest including human HTm4 were upregulated in G-CSF-mobilized cells, as opposed to an upregulation of cell cycle-promoting genes including Cyclin D2 and Hepatocyte Leukemia Factor (HLF) in pegfilgrastim-mobilized cells.

Moreover in pegfilgrastim-mobilized CD34+ cells we saw an upregulation of multiple genes involved in cellular immunogenicity like MHC class I and II antigens and genes encoding for proteins playing a role in antigen presentation.

Conclusion: Unconjugated G-CSF seems to be associated with an increased mobilisation of erythroid progenitors or an induction of erythropoiesis. Pegfilgrastim might result in mobilization of more immunogenic CD34+ cells. Unconjugated G-CSF and pegfilgrastim both seem to have an effect on cell cycle. Unconjugated G-CSF might rather induce cell cycle arrest and pegfilgrastim seems to lead to an increase of the cell cycle activity. This may be due to potentially different effects of continuously high serum levels of G-CSF maintained by pegfilgrastim and the pulsatile daily G-CSF injections on CD34+ cells.