Hematopoietic cell transplantation using plasma and DMSO without HES, with non-programmed freezing by immersion in a methanol bath: results in 213 cases

Abstract

Summary:A simplified cryopreservation method for bone marrow (BM) and peripheral blood progenitor cells (PBPC) was utilized in hematopoietic cell transplantation of 213 patients with hematological or solid neoplasms after ablative chemotherapy (187 with peripheral blood progenitor cells and 26 with bone marrow). Cells were cryopreserved, after addition of autologous fresh plasma with DMSO, without HES, by freezing to ؊80؇C in a methanol bath and non-programmed freezer. For the patients autotransplanted with PBPC, the median period necessary for recovery of more than 0.5 × 10 9 /l granulocytes was 11 days (range 6-44), and 15 (8-204) days were required to obtain more than 20 × 10 9 /l platelets. For the patients autotransplanted with BM, the median period necessary to recover Ͼ0.5 × 10 9 /l granulocytes was 12 days (range 9-33), and 24 (12-57) days to obtain more than 20 × 10 9 /l platelets. These results support this method as being very effective in achieving high-quality cryopreservation. The procedure, which uses a non-programmed freezer, simplifies and reduces enormously the cost of the technical measures currently in use, enabling its adoption in almost any clinical oncological institution. Keywords: hematopoietic cell transplantation; cryopreservation; plasma and DMSO; non-programmed freezing; methanolThe transplantation of hematopoietic progenitor cells, derived both from bone marrow and peripheral blood, is an ever more frequent procedure in the treatment of numerous neoplastic diseases. The technology for this type of therapeutic procedure requires, in the majority of cases, the freezing and storage of these cells for variable periods of time. The freezing procedure is traditionally carried out using machinery which incorporates pre-established, comCorrespondence: Dr F Hernández-Navarro, Servicio Hematología y Hemoterapia, Hospital La Paz, Paseo de la Castellana 261, 28046, Madrid, Spain Received 7 July 1997; accepted 21 October 1997 puterized freezing programs. 1-4 Experience using direct or non-programmed freezing systems for hemotopoietic progenitor cells and their transplant is less widespread. [5][6][7][8][9][10][11] In this article, we summarize our experience using a simple, non-programmed freezing method for the transplant of hematopoietic cells in patients with neoplastic disease. Two different in vitro experiments were designed to analyze the reproducibility and the effect of the methanol bath on freezing rates. Finally, a limited comparative study of in vitro cell viability following programmed and non-programmed freezing is also presented. Materials and methods Cooling ratesIn order to analyze the reproducibility of the cooling rates with the methanol system and the impact of this technique on the standardization of the freezing, two different in vitro experiments were performed.In the first (Figure 1), four different bags containing 160 ml of hematopoietic cells in 10% DMSO and autologous plasma were cryopreserved using a mechanical freezer at Ϫ80°C in a methanol bath as previously describe...

Long‐term hematologic reconstitution after autologous peripheral blood progenitor cell transplantation: a comparison between controlled‐rate freezing and uncontrolled‐rate freezing at 80°C

“…PBPCs cryopreservation is usually performed using the CRF method, followed by storage in either a liquid or a vapor phase of nitrogen. Considering the costs and complexity of this technique, several groups are currently investigating a faster procedure of CRF at –80°C for PBPCs or marrow freezing, 6‐16 and recently consistent clinical and biologic data have been reported in large groups of patients 8,14 …”

“…Some studies indicate that either uncontrolled‐rate freezing (URF) at –80°C or CRF are both characterized by similar HR, 10,11 but the clinical impact of URF, particularly long‐term HR, has not been sufficiently studied 8 …”

“…Alternative methods have been proposed: In 1987, Stiff et al 5 described a simple method of marrow URF at -80ЊC using a mechanical freezer; this technique of URF was applied by the same transplant centers and in some cases it was followed by storage at -80ЊC in the same mechanical freezer. [6][7][8][9][10][11][12][13][14][15][16] In clinical studies of URF, different concentrations (ranging from 5 to 10%) of DMSO were used for marrow or PBPC cryopreservation.…”

The occurrence of adverse events during the infusion of autologous peripheral blood stem cells is related to the number of granulocytes in the leukapheresis product

“…For cryopreservation, the apheresis product was diluted with an equal volume of cryoprotectant solution containing 5-10% DMSO and autologous plasma from the excess previously removed. The cells were frozen in methanol at À80 1C, as described previously, 7 and then stored in the vapour phase of liquid nitrogen. Cryopreserved cells were infused at least 48 h after the administration of the last dose of chemotherapy.…”

A single apheresis to achieve a high number of peripheral blood CD34+ cells in a lithium-treated patient with acute myeloid leukaemia

“…In a single apheresis, performed with a cell separator COBE Spectra (Cobe Laboratories, Lakewood, CO, USA), the percentage of CD34 ϩ cells was 0.43% and the number of CD34 ϩ cells was 4.66 ϫ 10 6 /kg, determined by methods previously described. 3 At the time of mobilisation, the dose of lithium carbonate was 1200 mg/day and the serum lithium concentration was 0.86 mmol/l. No side-effects of lithium therapy were observed while lithium carbonate was administered.…”