The aim of this work was to study the evolution of neutrophil functions in non-neutropenic cancer patients. Thirty non-neutropenic patients, median age 35 years (range 19-52), with solid tumors (n = 21) or lymphomas (n = 9) entered a phase I study of five days of s.c. (n = 24) or i.v. bolus (n = 6) lenograstim, recombinant human glycosylated granulocyte colony-stimulating factor (rHuG-CSF Chugai-Rhône-Poulenc), with dose escalation from 1 to 40 micrograms/kg/day. Neutrophil functions were studied before lenograstim (D1) and 24 h after the last dose (D6). Granulocyte count rose in a significant way, and enzyme release, phagocytosis and bacterial killing were stimulated. All patients had improvement of at least one neutrophil function. Directed migration was depressed, although it was still in the normal range. These findings confirm that lenograstim is a potent activator of neutrophil functions in non-neutropenic cancer patients and may be useful as an adjunct to conventional antimicrobial therapy.

Colony stimulating factors and especially rHuG-GSF, the first available neutrophil growth factor, have led to considerable interest in the field of stem cell transplantation because of their ability to induce stem cell peripheralization either alone or in association with high-dose chemotherapy. Few data exist, however, on the impact of rHuG-CSF on large scale bone marrow collection and autologous bone marrow transplantation (ABMT). This phase I, non-randomized, dose escalation study of rHu-G-CSF (lenograstim) administered to 30 patients at doses ranging from 1 to 40 micrograms/kg/day for 5 days before bone marrow harvesting showed that priming with rHu-G-CSF in vivo increased the number of bone marrow cells and D14 myeloid restricted progenitors (CFU-GM) and led to a better neutrophil recovery after ABMT compared with a contemporary unprimed control population. Otherwise, this study established that 5 days of rHuG-CSF therapy, as a sole stimulus, induced a tenfold increase in the circulating CFU-GM amongst which immature progenitors, estimated by the Delta assay (secondary CFU-GM grown after 7 days of liquid culture/primary CFU-GM), are detected. These conclusions were valid for doses as low as 2 micrograms/kg/day which induced only mild neutrophilia up to the highest dose (40 micrograms/kg/day) and suggest that a short course of rHuG-CSF is beneficial in increasing the stem cell collection.

This trial studied the biodistribution, pharmacology, toxicity, immunogenicity, and biologic characteristics of a trace-labeled, anti-CD33, humanized monoclonal antibody M195 (Hu-M195) in patients with relapsed and refractory myeloid leukemia. Hu-M195 is a computer-modeled, "complementarity-determining region-grafted," IgG1, humanized version of M195. M195 is a murine monoclonal antibody that reacts with CD33, a 67-kD glycoprotein expressed on early myeloid progenitor cells and myeloid leukemia (acute myelogenous leukemia and chronic myelogenous leukemia) cells, but not normal stem cells. 131I-murine-M195 has already shown significant ability to cytoreduce patients with relapsed or refractory myeloid leukemias. Hu-M195 has higher avidity than the original mouse monoclonal antibody and, unlike murine M195, has the capability to mediate antibody-dependent cellular cytotoxicity against leukemia targets. Thirteen patients with relapsed or refractory myelogenous leukemia were treated with Hu-M195 at 4 levels of 0.5, 1.0, 3.0, and 10.0 mg/m2 in a phase I trial. Patients received a total of 6 doses per patient over 18 days. Two patients were retreated for a total of 12 doses. The first dose of Hu-M195 was trace-labeled with 131I to allow detailed pharmacokinetic and biodistribution studies by serial sampling of blood, radioimmunoassays of cells, and whole-body gamma-camera imaging. Cumulative total doses of up to 216 mg of Hu-M195 were administered safely. Reversible fever and rigors were observed after infusion at the highest dose levels. The entire bone marrow was specifically and clearly imaged within hours after infusion, with optimal biodistribution occurring at the 3 mg/m2 level. Adsorption of Hu-M195 onto targets in vivo was demonstrated by flow cytometry; near saturation of available sites occurred at the 3 mg/m2 dose level. Plasma and whole body half lives were 38 and 51 hours, respectively, which may reflect continual replenishment of target sites on new leukemia cells. 131I-Hu-M195 was rapidly internalized into the target cells in vivo within 1 hour. Human antihuman antibody responses were not observed. In conclusion, Hu-M195 can be administered safely in multiple doses, without significant toxicity or any evidence of immunogenicity, and can localize rapidly and efficiently to the bone marrow in patients with myeloid leukemias. Additional phase II trials with this agent alone or in combination with cytokines or isotopes are warranted at the optimal biologic dose.

Blast-phase chronic myelogenous leukemia (CML) is the terminal phase in CML and is uniformly fatal. We treated 12 patients with blast-phase CML with a program of high-dose cytarabine 3.0 g/m2 and melphalan 140 mg/m2, followed by reinfusion of stem cells obtained from peripheral blood during the chronic phase. Seven patients achieved either a partial or complete hematologic remission, while five patients showed no response to therapy. One patient returned to chronic phase features with loss of a chromosomal abnormality acquired at blast phase, restoration of hematopoiesis, and a decrease in the amount of bone marrow blasts to less than 10%. Six patients cleared their peripheral blasts and showed recovery of their myeloid and platelet lineages, but all six required treatment for acceleration within 3 months. Of the five nonresponding patients, three died with aplastic bone marrow, one patient never cleared peripheral blasts after chemotherapy, and one patient had evidence of peripheral blasts 3 weeks after the autologous stem cell reinfusion. None of the patients returned to a normal karyotype. The ablative regimen was effective in eradicating bone marrow blasts to < 10% in 8 of 10 patients in whom interpretable bone marrow samples were performed following chemotherapy. Overall, the median survival for all patients from the time of stem cell reinfusion was 5.5 months. We conclude that autotransplants with peripheral blood can successfully be used to support hematopoiesis during high-dose therapy for CML blast crisis, however, has no role by itself in the curative therapy of blast crisis CML. A small number of patients can be restored to chronic phase features, and this may provide an opportunity to administer subsequent alternative treatments designed to eradicate the malignant stem cell population. Autotransplants with stem cells may also be used as therapy for patients without a histocompatible marrow donor. However, the autotransplant may be more effective when used during the chronic phase of CML, with the use of hematopoietic growth factors, and with reinfusion of stem cells depleted of the malignant Philadelphia chromosome-positive clone.

We analyzed the surface phenotype of CD34-positive (CD34+) cells in peripheral blood (PB) during the period of hematopoietic recovery following myelosuppressive chemotherapy. A significantly higher proportion of PB CD34+ cells coexpressed the CD13 and CD33 myeloid antigens (80.8%, 78.1%, respectively) than did BM CD34+ cells (45.8%, 37.8%, respectively) (both p < 0.01). In particular, the CD13 positivity of PB CD34+ cells harvested with granulocyte colony-stimulating factor (G-CSF) was significantly higher than that of those without G-CSF. Most of the PB CD34+ cells possessed the HLA-DR antigen, and less than 10% of the CD34+ cells coexpressed a mature cell antigen, such as CD14, GPA or Plt-1. The administration of G-CSF enhanced the appearance of significantly larger amounts of CD13+ 34+ and CD33+ 34+ cells (both p < 0.01). This G-CSF mobilization also resulted in an increased number of CD13- 34+ and CD33- 34+ cells and all types of colony-forming cells. On the other hand, macrophage colony-stimulating factor administration exerted little influence on the mobilization of PB CD34+ cells. Thus, G-CSF seemed to induce not only an expansion of circulating hematopoietic stem cells but also the myeloid differentiation of stem cells.

To evaluate the clinical value of growth factors (GFs) with peripheral-blood stem cells (PBSC) collected following mobilization with GFs, we randomized patients to receive or not to receive GFs following transplant.

We have conducted a 9-year multicenter trial of autologous bone marrow transplantation (ABMT) for acute myeloid leukemia (AML). Remission BM was purged in vitro using monoclonal antibodies (MoAbs; PM-81, AML-2-23) and complement targeting myeloid differentiation antigens (CD15, CD14). In 1988, the preparative regimen changed from 60 mg/kg/d cyclophosphamide x 2 and fractionated total body irradiation (TBI) total dose, 1,200 cGy (Cy/fTBI), to 4 mg/kg/d busulfan x 4 and 60 mg/kg/d Cy x 2 (Bu/Cy2). Recent analysis (October 1, 1993) shows that the Bu/Cy2 regimen along with the same MoAb purging method yields an improved outcome. Seven first complete-remission (CR) (CR1), 45 second- or third-CR (CR2/3), and 11 first-relapse (R1) patients were treated with chemotherapy and TBI or chemotherapy alone followed by ABMT with MoAb-purged BM. Median age at ABMT for those patients in CR 2/3 and R1 patients was 36 years. Twenty-nine CR 2/3 and R1 patients were conditioned with Cy/fTBI, and 27 CR2/3 and R1 patients were conditioned with Bu/CY. Using the Kaplan-Meier method, the CY/fTBI, CR2/3, and R1 patients have a 3-year disease-free survival (DFS) of 21%. On the other hand, the Bu/Cy2, CR2/3, and R1 patients have a 3-year DFS of 48%. Nineteen CR2/3 and R1 patients relapsed post-ABMT. On analysis by conditioning regimen, those treated with Cy/fTBI have a 3-year relapse rate (RR) of 58%, whereas the patients conditioned with Bu/Cy2 have a 39% 3-year RR. Long-term DFS can be achieved in about 50% of patients with advanced remissions and relapsed AML using Bu/Cy2 with MoAb-purged BM.

Chemotherapy with a doxorubicin-containing regimen results in high overall response rates in the treatment of metastatic adenocarcinoma of the breast; the combination of doxorubicin, etoposide, and cyclophosphamide has produced a response rate of 84% in untreated patients with stage IV disease. Recent data suggest that selected groups of patients with metastatic disease treated with high-dose chemotherapy and stem cell support may enjoy unmaintained, long-term remission. This study was designed to establish the feasibility of administering the combination of high-dose infusional doxorubicin, etoposide, and cyclophosphamide followed by autologous stem cell rescue in patients with responsive metastatic, or high-risk primary breast cancer.

To examine the effect of granulocyte colony-stimulating factor (G-CSF) on hematopoietic recovery after high-dose chemotherapy and peripheral blood progenitor cell transplantation (PBPCT), 20 patients with hematologic malignancies were divided into two groups. One group was given G-CSF at a daily dose of 50 micrograms/m2 subcutaneously, the other received no G-CSF. Neutrophil recovery was accelerated in the G-CSF treated patients and exceeded 0.5 x 10(9)/l at a median of 10 days post-PBPCT compared with 14 days in the control group (p < 0.01). This reduction led to a decrease in antibiotic use and a trend toward fewer febrile days in the G-CSF treated group.

In advanced breast cancer high-dose consolidation chemotherapy with haematological rescue has resulted in prolonged disease free and overall survival in a small percentage of patients. Maximal reduction of the tumor burden by intensive induction treatment preceding the high-dose chemotherapy may favor that outcome. The aims of this study were to find a rapid highly effective induction regimen with acceptable toxicity and to examine the optimal time for peripheral blood progenitor cell (PBPC) collection for haematological rescue.

Between June 1989 and June 1992, 144 patients participated in sequential clinical trials using peripheral blood progenitor cells (PBC) as their sole source of hematopoietic rescue following high-dose chemotherapy. All patients had received prior extensive combination chemotherapy and had marrow defects that precluded autologous bone marrow transplantation (ABMT). PBC were collected according to a single apheresis protocol. The initial 86 patients (group 1) had PBC collected without mobilization. Beginning in April 1991, PBC were mobilized solely with recombinant human granulocyte-macrophage colony-stimulating factor (rHuGM-CSF). Thirty-four patients (group 2) received rHuGM-CSF at a dose of 125 micrograms/m2/d by continuous intravenous infusion, and 24 patients (group 3) received rHuGM-CSF at a dose of 250 micrograms/m2/d by continuous intravenous infusion. Patients underwent at least six aphereses and had a minimum of 6.5 x 10(8) mononuclear cells (MNC)/kg collected. Cytokines were not routinely administered immediately after transplantation. A median of nine aphereses were required to collect PBC in group 1 and seven aphereses for groups 2 and 3 (P = .03). The time required to recover 0.5 x 10(9)/L granulocytes after transplant was significantly shorter (P = .0004) for the mobilized groups; the median time to recovery was 26 days for group 1, 23 days for group 2, and 18 days for group 3. Transplantation of PBC mobilized with rHuGM-CSF resulted in a shorter time to platelet (P = .04) and red blood cell (P = .01) transfusion independence. Mobilization with rHuGM-CSF alone resulted in efficient collection of PBC, that provided rapid and sustained restoration of hematopoietic function following high-dose chemotherapy. Mobilization of PBC with rHuGM-CSF alone is an effective method for patients who have received prior chemotherapy and have bone marrow abnormalities.

Incompatibility of blood group antigens in bone marrow transplantation (BMT) requires the depletion of red blood cells from donor bone marrow in order to avoid fatal transfusion reactions. We purified 72 human bone marrows prior to transplantation from red blood cells by a modified method using simple centrifugation and sedimentation in 6% dextran. Small-volume samples (n = 4) were prediluted with compatible homologous red blood cells to reduce the loss of stem cells during the preparation steps. 12 of 30 allogeneic bone marrows were ABO major incompatible with the patient's blood group. Although we did not observe any transfusion reactions nor graft rejections, in ABO incompatible transplantations, donor reticulocytes appeared 10 days later in the peripheral blood than in patients who received an ABO compatible bone marrow. Concordantly, incompatible patients needed 1.7 times more red blood cell units after BMT.

The effect of recombinant human erythropoietin (rhEPO) and interleukin 3 (IL3) on circulating haematopoietic progenitors consisting mainly of immature burst-forming-units-erythrocytes (BFU-E), was investigated in ten paediatric patients treated by regular haemodialysis. During a 30-week study rhEPO treatment resulted in a rise of median haemoglobin levels from 6.7 g/dl to > 10 g/dl in all patients. Before initiating rhEPO treatment the number of circulating BFU-E in chronic renal failure patients responded to grading doses of rhEPO in vitro similar to that in control children; however, the dose-response curves were not predictive for the in vivo response to rhEPO. After an initial rise in five patients BFU-E numbers declined by week 30 of rhEPO treatment. BFU-E numbers decreased to 35% of pretreatment values. The number of granulocyte-macrophage colony forming cells (GM-CFC) also decreased during rhEPO treatment. Addition of IL3 to the culture medium containing saturating concentrations of granulocyte-macrophage colony stimulating factor did not stimulate BFU-E numbers of patients before rhEPO treatment or those of controls. However, 2 weeks after start of rhEPO treatment IL3 increased the growth of patient's BFU-E in vitro to 220% of pretreatment levels, followed by a gradual decrease of stimulation until the end of observation. These findings indicate that: (1) long-term recruitment of circulating haematopoietic progenitors during rhEPO treatment is low in children with renal anaemia; (2) rhEPO sensitivity of circulating BFU-E is not predictive for the in vivo response; (3) rhEPO treatment results in enhanced sensitivity of BFU-E to IL3.

A total of three topoisomerase I inhibitors, including topotecan, CPT-11 (irinotecan), and intoplicine, have been studied in both preclinical and clinical/clinical pharmacology studies. In in vitro testing against human tumor colony-forming units, all three compounds were significantly more effective when tested as a continuous exposure as compared with a 1-h exposure. The dose-limiting toxicities were different for all three of the agents, with neutropenia and thrombocytopenia being dose-limiting for topotecan; diarrhea, for CPT-11; and hepatotoxicity, for intoplicine. In these phase I studies a number of marginal responses were noted with topotecan; partial and marginal responses, with CPT-11 (particularly in patients with colon cancer); and no response, with intoplicine. The detailed pharmacology of all three agents documented a very short half-life for topotecan, an intermediate half-life for CPT-11, and a prolonged half-life for intoplicine. Based on our experience to date, these compounds (particularly CPT-11) have promise as useful additions to our tremendous therapeutic armamentarium.

The quick reduction of differentiated myeloma cells by VAD chemotherapy (vincristine, adriamycin, dexamethasone) causes, according to the investigation by Bell et al., the acceleration of the proliferation of myeloma stem cells. In 1990 Bell demonstrated that this proliferation could be stopped by administering 500 mg of cyclophosphamide in 1-week intervals. We therefore modified the classical VAD scheme to the following "C-VAD" scheme:vincristine 0.5 mg/day in continuous infusion on the first to the 4th day, adriamycin 9 mg/m2/day in continual infusion on the 1st to the 4th day, dexamethasone 40 mg p.o. or i.v. always on 4 days starting with the 1st, 10th and 20th days, cyclophosphamide 600 mg i.v. on the 5th, 10th and 20th days). A further cycle follows on the 28th day. In the present paper the effect and the tolerance of this C-VAD scheme is evaluated: In the group of 21 patients with refractory myeloma 9 remissions were achieved, 5 partial remissions, in 6 patients the disease progressed, 1 patient died after the 2nd cycle without the possibility of evaluating the therapeutic response. The mean remission length was 10.2 months. The tolerance of chemotherapy was satisfactory, C-VAD chemotherapy did not cause any serious drop in the number of leucocytes and thrombocytes. Echocardiographically lower adriamycin cardiotoxicity was demonstrated in continual administration in comparison with the bolus administration. The C-VAD scheme is considered to be suitable for comparison with the VAD chemotherapy in a randomized study.