Roy S. Weiner

Cryopreserved autologous marrow infusion following high dose cancer chemotherapy

Abstract Seventeen patients with advanced malignancies received escalating doses of chemotherapy with adriamycin and cyclophosphamide. Sixty-five per cent of the courses produced circulating granulocyte counts of 500 cells/mm3 or less. Febrile episodes occurred in only 15% of courses and were seen only with doses of chemotherapy that produced granulocyte nadirs of less than 200 cells/mm3. There were no episodes of septicaemia. Ten of the 17 patients received an intensive dose followed by reinfusion of cryopreserved, autologous bone marrow. In 5 of these patients, recovery to 500 granulocytes/mm3 was more rapid following the intensive, marrow-supported course by comparison with the prior, less intensive, unsupported course. We conclude that higher doses of adriamycin and cyclophosphamide than are conventionally used can be given without serious toxicity. Autologous bone marrow reinfusion may have a role in reducing the period of drug-induced granulocytopenia, but effective storage and recovery of human bone marrow remains a major problem.

Combination chemotherapy with adriamycin-cyclophosphamide for advanced ovarian carcinoma

Combination chemotherapy with Adriamycin‐cyclophosphamide was employed after surgical treatment in 60 women with Stage III‐IV ovarian adenocarcinoma. Of 53 evaluable patients, objective response was noted in 34 of 41 (83%) without prior cytotoxic therapy but in only two of 12 (17%) who had failed a single alkylating agent or radiotherapy (P < .005). Complete response was confirmed by a negative biopsy at the site(s) of prior disease in 12 patients. Eleven of the 12 biopsy‐confirmed complete responses were achieved in patients without pretreatment palpable tumor. Twenty‐four out of 41 patients with palpable masses responded but only one was confirmed as complete. Confirmed complete responses had a median duration of 24 months, whereas the median duration of all other responses was only seven months. The median survival for patients in whom Adriamycin‐cyclophosphamide was the initial chemotherapy was 24 months. The median survival in patients with palpable tumor exceeds that of historical controls matched for age, tumor cell type, and grade (P = .05); the median survival for the confirmed complete responders has not been reached. The toxicity of this regimen was acceptable at doses of Adriamycin and cyclophosphamide of 45 mg and 500 mg/M2 body surface area, respectively. Extensive excision of tumor followed by effective combination chemotherapy offers the best current approach toward improved patient survival in advanced ovarian cancer.

Prolonged cryopreservation of human bone marrow

We tested the viability of human bone marrow stored for 40 to 42 months in the vapor phase of liquid nitrogen. A median of 2 × 1010 nucleated cells obtained from eight patients were concentrated to 1.3 × 1010 using discontinuous centrifugation. These were stored in polyolefin bags in volumes of 100 to 500 ml using 10% dimethyl sulfoxide (DMSO) as a cryoprotectant. Cell number and granulocyte – monocyte colony – forming cell (CFU-c) plating efficiency were determined before freezing and after thawing, after dilution and removal of DMSO, and after 2 to 4 hr of additional incubation. The median difference in cell number and CFU-c plating efficiency after this prolonged storage was −9 and +2%, respectively. Dilution, washing, and a 2-hr incubation were associated with cell losses of 24, 24, and 19% and increases in CFU-c plating efficiency, ranging from 22 to 79%. The number of viable CFU-c was never significantly lower than the number of CFU-c stored or initially thawed. Vapor phase storage appears to be adequate for prolonged human bone marrow cryopreservation using CFU-c viability as a determinant.

Cryopreservation of lymphocytes for use in in vitro assays of cellular immunity

Human lymphocytes can be cryopreserved for the purpose of performing in vitro tests of cellular immunity. A systematic study of the conditions for freezing and recovering cells has shown that there is wide flexibility in cell concentrations of cryoprotective agent and the temperature and rate of dilution however, have definable optima. Cells frozen and thawed under optimal conditions retain their reactivity in MLC and can be used for sequential studies of immune responsiveness. Approximately 70% of viable and functional cells are recovered when the cells are frozen in 7.5--12.5% DMSO and no further cryoprotection is discernable when up to 50% serum is added to the freezing media. The temperature and rate of dilution are critical only in that cells diluted rapidly in THE COLD (10-fold in 2 min at 0 degrees C) are less responsive in MLC than are cells diluted slowly (10-fold in 10 min) in the cold or slowly or rapidly at room temperature.