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Background: We conducted a prospective single-blind randomized study to assess whether a single 80,000 IU dose of human recombinant erythropoietin (HRE), given just 2 days before cardiac surgery, could be effective in reducing perioperative allogeneic red blood cell transfusion (aRBCt)., Study Design and Methods: Six-hundred patients presenting with preoperative hemoglobin (Hb) level of not more than 14.5 g/dL were randomly assigned to either HRE or control. The primary endpoint was the incidence of perioperative aRBCt. The secondary endpoints were mortality and the incidence of adverse events in the first 45 days after surgery, Hb level on Postoperative Day 4, and number of units of RBC transfusions in the first 4 days after surgery., Results: A total of 17% (HRE) versus 39% (control) required transfusion (relative risk, 0.436; p<0.0005). After baseline Hb was controlled for, there was no difference in the incidence of aRBCt between HRE (0%) and control (3.5%) among the patients with baseline Hb of 13.0 g/dL or more, which included the nonanemic fraction of the study population. The mean (range) Hb level on Postoperative Day 4 was 10.2 (9.9-10.6) g/dL (HRE) versus 8.7 (8.5-9.2) g/dL (control; p<0.0005). The distribution of number of units transfused was shifted toward fewer units in HRE (p<0.0005). The all-cause mortality at 45 days was 3.00% (HRE) versus 3.33% (control). The 45-day adverse event rate was 4.33% (HRE) versus 5.67% (control; both p=NS)., Conclusion: In anemic patients (Hb<13 g/dL), a single high dose of HRE administered 2 days before cardiac surgery is effective in reducing the incidence of aRBCt without increasing adverse events., (© 2015 AABB.)

Background: The peripheral blood progenitor cell (PBPC) mobilization capacity of EPO in association with either G-CSF or sequential GM-CSF/G-CSF was compared in a randomized fashion after epirubicin, paclitaxel, and cisplatin (ETP) chemotherapy., Study Design and Methods: Forty patients with stage IIIB, IIIC, or IV ovarian carcinoma were enrolled in this randomized comparison of mobilizing capacity and myelopoietic effects of G-CSF + EPO and GM-/G-CSF + EPO following the first ETP chemotherapy treatment. After ETP chemotherapy (Day 1), 20 patients received G-CSF 5 microg per kg per day from Day 2 to Day 13 and 20 patients received GM-CSF 5 microg per kg per day from Day 2 to Day 6 followed by G-CSF 5 microg per kg per day from Day 7 to Day 13. EPO (150 IU per kg) was given every other day from Day 2 to Day 13 to all patients in both arms of the study. Apheresis (two blood volumes) was performed during hematologic recovery., Results: The magnitude of CD34+ cell mobilization and the abrogation of patients' myelosuppression were comparable in both study arms; however, GM-/G-CSF + EPO patients had significantly higher CD34+ yields because of a higher CD34+ cell collection efficiency (57.5% for GM-/G-CSF + EPO and 46.3% for G-CSF + EPO patients; p = 0.0009). Identical doses of PBPCs mobilized by GM-/G-CSF + EPO and G-CSF + EPO drove comparable hematopoietic recovery after reinfusion in patients treated with identical high-dose chemotherapy., Conclusion: The sequential administration of GM-CSF and G-CSF in combination with EPO is feasible and improves the PBPC collection efficiency after platinum-based intensive polychemotherapy, associating high PBPC mobilization to high collection efficiency during apheresis.

Epoetin (Epo) is physiologically present in the human body, stimulating erythropoiesis from bone marrow. Anemia is observed in cancer patients submitted to high-dose chemotherapy (HDCT), mainly caused by myelosuppression. Epoetin alfa has been widely used to treat the anemia that develops in the HDCT setting. Controlled studies in patients with hematologic malignancies or solid tumors who received Epo following HDCT have shown a decreased red blood cell transfusion requirement at least in patients receiving allogeneic bone marrow transplantation (BMT), while results in patients receiving autologous BMT have been disappointing. The administration of Epo before HDCT, in a period when bone marrow is still responsive to growth factors, may represent a new strategy aimed at decreasing the degree of anemia in these patients. A combination of granulocyte-colony stimulating factor and Epo has proved to be effective in mobilizing stem cell and committed myeloid/erythroid precursors.

Objective: A fusion protein made of human granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (EPO), referred to as MEN 11303, has been tested for biologic activity using mobilized CD34(+) cells., Methods and Results: MEN 11303 and a combination of GM-CSF/EPO produced the same amount of colony-forming unit granulocyte-macrophage (CFU-GM), of burst-forming unit erythroid (BFU-E), and of multipotent CFU-mixed. After 15 days, liquid cultures of CD34(+) cells exposed to MEN 11303 yielded a total cell number larger than that obtained with an equimolar mixture of GM-CSF and EPO, with a clear prevalence of cells exhibiting an erythroid phenotype. A colony-forming cell assay established from CD34(+) cells precultured with MEN 11303 for 7 days yielded a greater amount of BFU-E than GM-CSF/EPO combination. Exposing CD34(+) cells to MEN 11303 for 7 days in liquid culture resulted in higher recoveries of cells expressing a comparatively less differentiated hematopoietic phenotype and of long-term culture initiating cells. A cell-based binding-competition assay using the human EPO-receptor (EPO-R) transfected murine Ba/F3EPOR cell line showed that MEN 11303 bound to EPO-R with a sixfold lower affinity but induced a more sustained receptor phosphorylation. MEN 11303 supported the growth of Ba/F3EPOR cells more efficiently than EPO and remained detectable in the spent culture medium for a longer time., Conclusions: MEN 11303 and the combination of GM-CSF/EPO are equally potent in recruiting hematopoietic progenitors into cycle, but the fusion protein is superior in promoting the expansion of committed erythroid percursors. Primitive hematopoiesis is less affected by MEN110303 than GM-CSF/EPO combination. Part of these effects may reflect the peculiar interaction of the EPO moiety of MEN 11303 with the EPO-R.

Purpose and Methods: The ability of granulocyte colony-stimulating factor (G-CSF) plus erythropoietin (EPO) treatment was compared in a randomized fashion with that of G-CSF treatment alone in promoting hematologic recovery and peripheral-blood progenitor-cell (PBPC) mobilization in previously untreated patients with advanced ovarian cancer who underwent their first course of epirubicin, paclitaxel, and cisplatin (ETP) chemotherapy during a phase II study of intensive outpatient ETP chemotherapy followed by high-dose carboplatin, etoposide, and melphalan (CEM) late intensification with PBPC support., Results: Comparative analysis of hematologic recovery of 50 randomized patients, after ETP chemotherapy, showed that life-threatening neutropenia occurred in 88% of the patients treated with G-CSF alone, whereas it occurred in only 4% of patients treated with G-CSF + EPO. Significantly different WBC and polymorphonuclear leukocyte (PMN) counts were observed in the two distinct arms on the day of WBC nadir (P <.0001 and P <.0001, respectively). Moreover, the addition of EPO to G-CSF increased PBPC mobilization and collection as compared with that in G-CSF-treated patients (P =.0009 and P =.0026, respectively), who required a significantly higher number of leukaphereses than G-CSF + EPO-treated patients (P =.0076) to obtain the planned minimum dose of PBPCs. Qualitative analysis by cloning assay of PBPCs collected in both arms revealed that G-CSF- and G-CSF + EPO-mobilized PBPCs have comparable in vitro functional properties., Conclusion: This randomized comparison revealed that EPO significantly increases most of the hematologic effect produced by G-CSF administration after chemotherapy. This biologic property of EPO translated in vivo into a global improvement of patients' hematologic status.

Ex vivo stroma-free static liquid cultures of granulocyte colony-stimulating factor (G-CSF)/chemotherapy-mobilized CD34+ cells were established from patients with epithelial solid tumors. Different culture conditions were generated by adding G-CSF, granulocyte-macrophage colony-stimulating factor (GM-CSF), Flt3 ligand (Flt3), megakaryocyte growth and development factor (Peg-rHuMGDF), GM-CSF/erythropoietin (EPO) hybrid protein (MEN11303), and interleukin-15 (IL-15) to the basic stem cell factor (SCF) + interleukin-3 (IL-3) + EPO combination. This study showed that, among the nine different combinations tested in our 5% autologous plasma-containing cultures, only those containing IL-3/SCF/Flt3/MEN11303 and IL-3/SCF/Flt3/MEN11303/IL-15 significantly expanded colony-forming unit granulocyte-macrophage (CFU-GM), burst-forming unit erythroid (BFU-E), long-term culture-initiating cells (LTC-IC), CD34+, and CD34+/CD38- cells after 14 days of culture. Particularly, the addition of IL-15 to IL-3/SCF/Flt3/MEN11303 combination produced a significant increase of LTC-IC, with an average 26-fold amplification as compared to input cells, without any detrimental effect on CFU-GM and BFU-E expansion. This combination also produced a statistically significant 3.6-fold expansion of primitive CD34+/CD38- cells. Moreover, this study confirms the previously described erythropoietic effect of MEN11303, which, in our experience, was the only factor capable of expanding BFU-E. Compared to equimolar concentrations of GM-CSF and EPO, MEN11303 hybrid protein showed a significantly higher capacity of expanding CFU-GM, BFU-E, LTC-IC, CD34+, and CD34+/CD38- cells when these cytokines were tested in combination with IL-3/SCF/Flt3. These cultures indicated that Peg-rHuMGDF addition to IL-3/SCF/EPO/Flt3 does not affect CFU-GM and BFU-E expansion but, unlike G-CSF or GM-CSF, it does not decrease the ability of Flt3 to expand primitive LTC-IC. These studies indicate that, starting from G-CSF/chemotherapy-mobilized CD34+ cells, concomitant expansion of primitive LTC-IC, CFU-GM, BFU-E, CD34+, and CD34+/CD38- cells is feasible in simple stroma-free static liquid cultures, provided IL-3/SCF/Flt3/MEN11303/IL-15 combination is used as expanding cocktail in the presence of 5% autologous plasma.

A combination of erythropoietin (EPO) plus stem cell factor (SCF) drove purified unfractionated granulocyte colony stimulating factor (G-CSF)/chemotherapy mobilized peripheral blood CD34+ cells to selective erythroid differentiation in liquid culture with an average 28-fold increase in the total cell number after 21 d. From day 6 of culture cytologic and cytofluorimetric characterization revealed that cultured cells belonged to the erythroid lineage with a gradual wave of maturation along the erythroid pathway to terminal cells. A similar pattern of erythroid differentiation was observed when the same peripheral blood CD34+ cells were culture with EPO plus SCF in serum-free medium. This cytokine combination produced selective erythroid differentiation with the complete exhaustion of the clonogenic potential on day 21. In parallel experiments the same circulating CD34+ cells underwent granulocytic/ monocytic differentiation in liquid culture in response to granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin-3 (IL-3) and SCF, demonstrating that these CD34+ progenitors had intact pluripotent differentiating potential. Conversely, bone marrow CD34+ cells isolated from bone marrow allografts were unable to selectively differentiate along the erythroid pathway when they were exposed to EPO plus SCF combination. However, these cells maintained a greater number of colony forming cells on day 21 of culture compared to mobilized peripheral blood CD34+ cells. This model is a simple and reliable way to obtain selective erythroid differentiation of peripheral blood G-CSF/ chemotherapy mobilized CD34+ progenitor cells in liquid culture. The absence of cytokines such as GM-CSF and IL-3 in the culture medium permits studies on in vitro erythropoiesis without disturbance of prevalent myelopoiesis.

In order to investigate the effects of erythropoietin (EPO) plus granulocyte colony-stimulating factor (G-CSF) administration after peripheral blood progenitor cell transplantation (PBPCT) we performed a phase I/II study in patients with high-risk cancer. 15 consecutive patients were treated wit recombinant human G-CSF (rhG-CSF) at the dose of 5 micrograms/kg subcutaneously (s.c.) every 24 h until day + 12 and with recombinant human EPO (rhEPO) at the dose of 150 IU/kg s.c. every 48 h until day + 11 following PBPCT. Their haemopoietic recovery was compared to that obtained in eight historic and control patients who did not receive any cytokines after PBPCT. No side-effects were observed during EPO plus G-CSF treatment and the treatment was not discontinued in any of the patients before completion of the treatment plan. The administration of EPO plus G-CSF after PBPCT produced a significant increase in the rate of white blood cell (WBC) (P = 0.0005), polymorphonuclear leucocyte (PMN) (P = 0.0005) and platelet (PLT) (P = 0.0105) recovery compared to the control group. The acceleration in haemopoietic recovery observed in the EPO plus G-CSF-treated patients produced a significant reduction of the days with WBC < 1 x 10(9)/l (P = 0.0009), PMN < 0.2 x 10(9)/l (P = 0.0030) and PMN < 0.5 x 10(9)/l (P = 0.0006). EPO plus G-CSF-treated patients required a significantly lower number of single donor PLT transfusions (P = 0.0142) and did not experience neutropenic fever, but historic control patients experienced fever > 38 degrees C for a median period of 4 d (0-12) with a medial period of parenteral antibiotic administration of 7.5 d (0-17). The length of the hospital stay was significantly shorter in the study group than in the historic control group (P = 0.0264). In conclusion, we can confirm that EPO plus G-CSF treatment is feasible and potentiates the haemopoietic recovery after PBPCT, thus simplifying the clinical management of cancer patients who undergo high-dose chemotherapy.

We tested in vitro the effect of recombinant human erythropoietin (rhEPO) plus recombinant human G-CSF (rhG-CSF) on purified human CD34+ haemopoietic progenitors (HP) and in vivo in patients who had undergone anti-cancer chemotherapy for advanced ovarian cancer. In this preliminary experience we found that, in vitro, rhEPO potentiates the effect of rhG-CSF on HP growth and differentiation toward the granulocyte-macrophage lineage. rhEPO plus rhG-CSF produced in vitro a proliferative stimulus of HP which represents 26% of the maximum stimulation obtained using IL-1, IL-3, IL-6, G-CSF, GM-CSF and stem cell factor in combination. In the patients treated with rhEPO plus rhG-CSF after chemotherapy, we observed a favourable trend for platelet and neutrophil recoveries compared with a control group treated with rhG-CSF alone and a significantly higher haematocrit nadir was observed in the rhEPO plus rhG-CSF series. In the patients treated with rhEPO plus rhG-CSF we observed a significant increase of circulating colony-forming unit granulocyte-macrophage (CFU-GM) and burst forming unit-erythroid (BFU-e) compared with the rhG-CSF series. Our results, in vitro and in vivo, encourage the in vivo use of rhEPO plus rhG-CSF to improve blood cell recoveries of patients who have undergone conventional or high-dose chemotherapy. Moreover, rhEPO plus rhG-CSF was demonstrated to be a good HP mobilising treatment for blood stem cell collection after chemotherapy.