Lymphoma cell contamination of PBSC: a murine model

Abstract

Summary:In P-388 bearing BDF 1 mice stem cell mobilisation was tested on the survival of lethally irradiated isologous recipients. Contamination of the graft with lymphoma cells was evaluated by the number of clonogenic lymphoma cells (CFU-L) and the ability of the graft to induce lymphoma in non-irradiated recipients. A mobilisation protocol (200 mg/kg cyclophosphamide (CY) i.p. or 200 mg/kg CY followed by 125 g/kg G-CSF administered every 12 h for 3 consecutive days, starting 14 days after lymphoma initiation) that resulted in a substantial stem cell mobilisation in normal mice, mobilised too few CFU-L to induce lymphoma in the recipients: 50 l of blood obtained after mobilisation protected lethally irradiated mice but did not induce lymphoma in normal recipients. A minimum graft of bone marrow (2 × 10 5 cells, with 5580 CFU-L) from untreated P-388 bearing donors killed irradiated recipients presumably by lymphoma and not bone marrow failure. The mobilisation protocol reduced CFU-L so much that no lymphoma-associated death occurred even after the transplantation of 10 × 10 5 bone marrow cells. These data suggest that, although the PBSC mobilisation protocol may also mobilise some clonogenic lymphoma cells, with the minimum graft size no lymphoma transfer can be observed.

G-CSF Alone vs cyclophosphamide plus G-CSF in PBPC mobilization of patients with lymphoma: results depend on degree of previous pretreatment

“…in many patients, 24,25 and they can be mobilized by current mobilization strategies; 24 however, the seeding efficiency of these cells and hence their role in determining relapse after autologous transplantation is not clear. 26,27 Tumour cell contamination of apheretic harvests obtained after 'cyclophosphamide plus G-CSF' or after 'G-CSF alone' has been studied in patients with breast cancer and in patients with lymphomas and no differences have been detected. 11,15,28 'G-CSF alone', compared to 'cyclophosphamide plus G-CSF', results in apheretic harvests with a higher content of nucleated cells and lymphocytes; furthermore, a fast immune recovery has been reported after autologous transplantation of PBPC harvested using 'G-CSF alone'.…”

Leukaemogenic potency of WEHI-3B cells grown in vitro or in leukaemic mice

“…Following transplantation of 0.5-2.0 × 10 6 nucleated blood cells from leukaemic mice 3 weeks after inoculation of WEHI-3B cells (2 × 10 6 WBC are equivalent to the minimum graft size needed to protect supralethally irradiated recipients, 17 but we wished to see the effect of lower cell doses, too), leukaemia-associated death occurred in all but one of the 21 unconditioned recipients. When applying a mobilisation protocol mimicking the clinical situation for obtaining PBSC for autologous transplantation (CY or CY followed by G-CSF administration) 5 none of the 18 recipients died of leukaemia (P = 0.003 at 0.5 × 10 6 or 1 × 10 6 cells and P = 0.002 at 2 × 10 6 cells, Fisher's test, Table 3a).…”

“…17 Although we were aware of the fact that CML cells show growth factor dependent growth, in order to be able to quantify contaminating leukaemia-inducing cells in WBC or bone marrow, we tried to establish a system where quantitative assay of in vitro colony-forming leukaemia cells (CFU-L) and survival rate and/or survival time of mice transplanted with WEHI-3B cells of different origin (ie from suspension culture or from WEHI-3B bearing mice) could be correlated.…”

“…and G-CSF (recombinant human G-CSF, Neupogen; Hoffman-La Roche, Basel, Switzerland) was injected s.c. in a daily dose of 250 g, divided into two equal doses on days 3, 4, 5 and 6 after CY administration. Three hours after the last injection, mice were exsanguinated from the axillary vessels, circulating CFU-GM and CFU-L were assayed and various amounts of blood cells up to the minimum graft size for stem cell transplantation 17 were injected into unirradiated recipients.…”

“…This idea supports the use of animal models where colony-forming leukaemia cells in vitro (CFU-L) and cells inducing leukaemia in vivo (putative leukaemia stem cells) can be studied in parallel. In our previous experiments we studied the lymphoma cell contamination of PBSC in a murine model 17 and found that, due to the relatively little contamination, no lymphoma-associated death could be observed when PBSC were transplanted after mobilisation with cyclophosphamide (CY) or a combination of CY and G-CSF. Quite recently a murine leukaemia model 18 was offered to follow the contamination of bone marrow with leukaemia cells transduced with a retroviral vector encoding human nerve growth factor receptor.…”