Børge Thing Mortensen

Changing bone marrow micro‐environment during development of acute myeloid leukaemia in rats

The Brown Norwegian rat transplanted with promyelocytic leukaemic cells (BNML) has been used as a model for human acute myeloid leukaemia. We have previously shown that both the blood supply to the bone marrow and the metabolic rate decrease in relation to the leukaemic development in these rats. Here we have investigated how the development and progression of this leukaemia affect oxygenation, pH and proliferation of normal and leukaemic cells in vivo. Bone marrow pH was measured by a needle electrode. Nitroimidazol‐theophylline (NITP) was used to identify hypoxic cells, and we applied bromodeoxyuridine (BrdUrd) to identify DNA replicating cells.  The leukaemia progressed slowly until day 27 after which a rapid deterioration could be observed leading to severe changes over the following 5 d. In whole blood there was evidence of progressing metabolic acidosis. In bone marrow the fraction of leukaemic cells increased to > 90% and the pH dropped to about 6.5. The fraction of NITP+ cells increased to > 80% in bone marrow and to about 40% in blood. The fraction of BrdUrd+ cells was unchanged in blood, but decreased in bone marrow both for normal cells (from about 20% to 5%), and for leukaemic cells (from about 45% to 25%), evidently as a result of the severely changed micro‐ environment. In this study we have demonstrated in vivo the development of an acidic and hypoxic bone marrow hampering normal haemopoiesis during leukaemic growth. Our data support the notion of BNML as a valuable tool for studying leukaemogenesis.

Comparative pharmacokinetics of single-dose administration of mammalian and bacterially-derived recombinant human granulocyte-macrophage colony-stimulating factor.

Abstract: Pharmacokinetics of recombinant human non‐glycosylated bacterially‐synthesized (E. coli) granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) were studied following single intravenous (i.v.) and subcutaneous (s.c.) bolus injection, and compared to equivalent doses of glycosylated mammalian‐derived CHO‐GM‐CSF. Each route of administration gave a different GM‐CSF concentration‐time profile. The highest peak serum concentrations (Cmax) were observed following i.v. bolus injection. After i.v. administration, a two‐phase decline in concentration was noted for both types of GM‐CSF with a significantly shorter ***t1/2 α of 7.8 minutes for the E. coli GM‐CSF versus 20.0 min for the CHO‐GM‐CSF, while no significant difference was observed for the terminal phase. Following s.c. administration of equivalent doses, a higher peak serum concentration was observed in the E. coli‐treated patients and, again, a faster elimination where pretreatment serum levels were reached after 16–20 h, versus more than 48 h after administration of CHO‐GM‐CSF. Although the non‐glycosylated E. coli GM‐CSF thus seems to undergo a faster elimination that the glycosylated CHO‐GM‐CSF no significant difference could be demonstrated in the in vivo effect of corresponding doses of the two compounds with respect to stimulation of granulopoiesis — with reservation for small patient numbers and a large individual variations in response.

Recombinant human interleukin-3: pharmacokinetics after intravenous and subcutaneous bolus injection and effects on granulocyte kinetics

The pharmacokinetics of E. coli derived recombinant human interleukin‐3 (rhIL‐3) was studied following intravenous (i.v.) and subcutaneous (s.c.) bolus injection of rhIL‐3. After i.v. bolus injection in eight patients, serum peak levels of 34.5‐135.0 ng/ml were reached, followed by a rapid decline with a t1/2α of 17 ± 2 min and a t1/2β of 59 ± 7 min. After s.c. bolus injection in five patients, the absorption was more prolonged with peak serum levels reached at 2.8 ± 0.4h. Elimination was also more protracted, and serum base‐line levels were reached at 14‐24 h.

1α(OH)D3 (ETALPHA)® treatment and receptor studies in 16 patients with chronic and myeloproliferative disorders

10 patients with CLL and 2 with CML were treated with gradually increasing doses of 1α(OH)D3, up to 4 μg daily during 6 wk. 3 patients with preleukemia and 1 with myelofibrosis were treated with 2 μg daily of 1α(OH)D3 for a prolonged period up to 17 wk. The treatment with 1α(OH)D3 did not result in changes of disease parameters in any of the patients under study. Receptor studies for 1,25(OH)2D3 were performed in 8 CLL patients and revealed only 1 patient with increased specific receptor binding capacity. The maximum tolerable dose of 1α(OH)D3 varied individually, but was in the range of 2–4 μg daily.

Clinical experience with bacteraemia in patients with leukaemia and allied neoplastic diseases.

134 bacteraemic episodes in 112 patients with leukaemia and allied disorders over a 5-year period were reviewed. The patients were divided, with respect to neoplastic disease, into 3 prognostic groups according to estimated periods of survival of years (group 1), months (group II) or weeks (group III). High mortality was correlated to group III, gram-negative rod bacteraemia and septic shock. Low leucocyte levels and intensive antineoplastic treatment did not adversely influence the prognosis of the bacteraemic patients. Appropriate antibiotic therapy was correlated with a significantly lower mortality, and it is concluded that aggressive antibiotic therapy is indicated when bacteraemia is suspected from clinical judgement.

S-Phase Induction by Interleukin-6 Followed by Chemotherapy in Patients with Chronic Lymphocytic Leukemia and Non-Hodgkin'S Lymphoma

Interleukin-6 (IL-6) has in vitro demonstrated growth regulatory effects on tumor cells from patients with chronic lymphocytic leukemia (CLL) and lymphoma. The proliferation rate of these cells is usually very low and this is thought to be one of the reasons for the lack of a curative potential of cytostatic chemotherapy in CLL and low grade NHL. Recombinant human (rh) IL-6 might increase the in vivo proliferation rate leading to a higher sensitivity for chemotherapy. We tested this hypothesis by administering rhIL-6 to 9 CLL patients and 3 NHL patients in doses of 2.5 micrograms/kg, 5 micrograms/kg and 10 micrograms/kg s.c. daily for 5 days followed by CHOP chemotherapy on the last day of rhIL-6 injection. Six patients had two treatment cycles. The proportion of cells in S-phase was determined by the bromodeoxyuridine labeling index (LI). Three patients achieved a partial remission, one patient had progressive disease and the remaining patients demonstrated no change. Two patients, who received 10 micrograms/kg/day rhIL-6, demonstrated a significant increase in LI, one of these was first observed in the second treatment cycle. A significant decrease was seen in two patients receiving 2.5 micrograms/kg and 5 micrograms/kg respectively. Immunophenotypic assessment demonstrated that rhIL-6 increased the expression of CD20 in all CLL patients with a reversal after cessation of rhIL-6. We conclude that rhIL-6, in the dosage and schedule used in this study, did not increase the proportion of the cells in S-phase and that the growth stimulatory effects of rhIL-6 in CLL in vivo probably are insignificant. However, the role of rhIL-6 in CLL as inducer of increased CD20 expression prior to anti-CD20 antibody treatment remains to be determined.

Bone marrow culture and haemopoietic growth factors: Recent developments and current status

The use of semi‐solid cultures of bone marrow cells is briefly reviewed. In the last two decades this technique has greatly improved our understanding of the hierarchical structure of cell production and its regulatory factors. It is now well established that relatively few multipotent stem cells, with self‐renewing capacity, are feeding cells into the differentiating pool of all the different lineages of mature cells found in blood. The proliferation and differentiation in this system are controlled by specific haemopoeitic growth factors, which also regulate the function of the mature cells. Several of these factors are now produced in recombinant form, allowing further studies of their biological significance. Clinical trials with two of the factors have supported the experimental evidence for their in vivo function, and they can be administered without serious side effects. The clinical access to these factors is promising for future treatment of cancer patients and may possibly completely change the outcome of several haematological and infectious diseases.

S-phase induction by interleukin-6 followed by chemotherapy in patients with refractory multiple myeloma

The plasma cell labeling index (PCLI) in patients with multiple myeloma (MM) is relatively low and this has been associated with the low rate of remission following chemotherapy. Interleukin-6 (IL-6) has been demonstrated to be a major growth factor of myeloma cells. In order to increase the S-phase proportion of myeloma cells, which might increase the sensitivity to chemotherapy, we gave rhIL-6 followed by chemotherapy to 15 myeloma patients with refractory disease. A total of 25 treatment cycles were administered since ten patients had two cycles. The rhIL-6 dose was 2.5 (n = 3), 5.0 (n = 6) and 10.0 microg/kg (n = 6) by subcutaneous injection once daily for 5 days and chemotherapy was administered on the last day of rhIL-6 injection. The effect of rhIL-6 treatment on labeling index (LI) was heterogeneous, but no statistically significant change was noted for this particular group as a whole. In two patients an increase (mean 7.7%) in LI of mononuclear bone marrow cells during the rhIL-6 treatment was demonstrated and in one patient a decrease of 2.8% was seen. Assessment of PCLI demonstrated an increase of 2.9% in one out of six patients and a decrease of 1.9% in one out of six patients. None of the 15 patients achieved remission according to standard criteria. During the rhIL-6 treatment, 14 of the 15 patients developed mild constitutional adverse events (AE) well known in patients treated with IL-6, and none of the AE in the subsequent chemotherapy phase were related to IL-6. In conclusion, our study demonstrated that rhIL-6 can be administered safely to patients with refractory MM, but the cell cycle recruitment approach was not sufficiently effective to be of clinical value.

Pharmacokinetic and pharmacodynamic studies of subcutaneously administered recombinant human interleukin-3 following chemotherapy for non-Hodgkin's lymphoma

Summary. The pharmacokinetics and the pharmacodynamic profile of subcutaneously administered recombinant human non‐glycosylated interleukin‐3 (rhIL‐3) was studied in lymphoma patients after standard CHOP chemotherapy. 30 patients received 0.5, 1.0, 5.0, 7.5 and 10/ig/kg (six patients at each dose level) of rhIL‐3 for 14 d. Serum rhIL‐3 samples were obtained regularly during the treatment and serially over a 24 h period on the first (cycle day 2) and the last (cycle day 15) day of rhIL‐3 treatment for pharmacokinetic evaluation. Following s.c. injection on cycle day 2, the maximum rhIL‐3 serum concentration ranged from 289pg/ml (0‐5/ig/kg) to 4690pg/ml (10/xg/kg). Both the maximum serum concentration (R= 0.90, P< 0.0001) and the area under the serum concentration‐time curve (R= 0.95, P< 0.0001) were related to dose.