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Biomedical subjects

E M Areman

Publications and source records attributed to E M Areman.

18 recordsLinked to original sources
PubMed1999

Immunotherapy with interleukin-2 and alpha-interferon after IL-2-activated hematopoietic stem cell transplantation for breast cancer.

We previously demonstrated findings suggestive of autologous GVHD in patients receiving IL-2-activated peripheral blood stem cells (PBSC) with IL-2 after transplantation. A pilot study was designed to test tolerability, feasibility and frequency of autologous GVHD and engraftment using IL-2 and alpha-IFN post-transplantation. After cyclophosphamide (6 g/m2) and carboplatin (1800 mg/m2), patients with high-risk stage II or III breast cancer received chemotherapy and rhG-CSF mobilized autologous PBSC that had been cultured in IL-2 for 24 h. Subcutaneous administration of IL-2 began on day 0 at 6 x 10(5) IU/m2/day for 5 of 7 days each week and continued for 4 weeks. Once engraftment occurred, alpha-IFN was initiated at a dose of 1 x 10(6)/m2/day subcutaneously for 30 days. Thirty-four consecutive patients with stage II (n=20), IIIA (n=6) and IIIB (n=8) disease were treated. All patients were without evidence of disease at the time of transplantation. The average time required for the ANC to reach 500/mm3 was 10 days (range: 8-11 days) and for platelets to reach 20000/mm3 was 10.7 days (range: 6-21 days). Forty-seven percent of patients (n=16) completed the full course of immunotherapy; the remaining patients received attenuated doses due to patient's request (n=6), development of temperature >38 degrees C (n=3), development of neutropenia (n=3), serious infection (n=1) and miscellaneous reasons (n=5). Four patients experienced transient moderate toxicities (level 3) including elevated liver function tests, nausea, rash and capillary leak syndrome. Pathological findings suggestive of skin GVHD developed in 43% of patients (12/28 patients) when skin biopsies were evaluated in a blinded fashion. At 13 months post-transplant (median; range: 5-24 months), 28 patients (82%) remain disease-free. These results demonstrate the feasibility and toxicity of this regimen along with pathological findings compatible with autologous GVHD of the skin.

Adult↗
PubMed1999

Differential effects of IL-2 incubation on hematopoietic potential of autologous bone marrow and mobilized PBSC from patients with hematologic malignancies.

Culturing of hematopoietic progenitor cells for 24 h with IL-2 generates cytotoxic effector cells that mediate in vitro and possibly in vivo antitumor activity. We examined the effect of IL-2 incubation on progenitor cells from 24 patients with hematologic malignancies using paired autologous bone marrow (ABM) and PBSC to determine differences in hematopoietic potential. Cells were cryopreserved and stored in liquid nitrogen until conditioning therapy was completed. After thawing, cells were incubated with IL-2 for 24 h at 37 degrees C. Paired samples of ABM and PBSC from the same patient were analyzed for nucleated and mononuclear cell number, CD34 antigen expression, and colony-forming unit (CFU) activity before and after IL-2 incubation. There was a significant decrease in the average number of mononuclear cells (MNC) (x10(8)/kg) (<0.001) and CD34+ cells (x10(6)/kg) (0.006) from both ABM and PBSC after 24 h IL-2 culture (ABM MNC: 0.6+/-0.1 vs. 0.4+/-0.0, p = <0.001; PBSC MNC: 4.4+/-0.5 vs. 3.7+/-0.4, p = 0.03; ABM CD34+: 2.4+/-0.5 vs. 1.3+/-0.3, p = <0.001; PBSC CD34+: 6.6+/-1.8 vs. 5.0+/-1.2, p = 0.05). However, whereas ABM CFU/10(5) MNC plated (269.3+/-47.2 vs. 385.6+/-70.6) were significantly increased (p = 0.005), there was no change in PBSC CFU (271.0+/-47.2 vs. 257.3+/-48.5). The mean plating efficiency (%) of ABM CD34+ cells was markedly increased after IL-2 incubation (10.1+/-3.3 vs. 19.0+/-7.2, p = 0.04), although it was lower than that of PBSC CD34+ cells, which did not change significantly in culture (29.4+/-5.5 vs. 36.0+/-6.5). Additional work is in progress to determine the cause and significance of the enhanced plating efficiency of the ABM progenitor cells.

Adult↗
PubMed1997

Interleukin-2-activated hematopoietic stem cell transplantation for breast cancer: investigation of dose level with clinical correlates.

Incubating hematopoietic stem cells with IL-2 in vitro for 24 h generates cytotoxic T cells. When infused into patients, these cells may stimulate a graft-versus-tumor (GVT) effect. This clinical trial was designed to assess the ability of IL-2 activated peripheral blood stem cells (PBSC) to reconstitute hematopoiesis, to investigate dose levels and dose-limiting toxicities of IL-2, and to evaluate clinical results and preliminary laboratory effects using a combination of IL-2-activated autologous PBSC followed by IL-2 after transplantation. Sixty-one women with stage II-IV breast cancer were treated. After the administration of carboplatin (200 mg/m2/day for 3 days) and cyclophosphamide (2 g/m2/day for 3 days), patients received autologous PBSC that were cultured in IL-2 for 24 h followed by parenteral administration of IL-2 beginning the day of transplantation. Three escalating doses of IL-2 were evaluated with increasing duration up to 4 weeks. Of the 57 patients receiving IL-2 after tranplantation, 19 patients (33.3%) were unable to complete the planned course of IL-2 therapy due to persistent fevers (n = 9), diarrhea (n = 2), pulmonary capillary leak syndrome (n = 3), development of a rash (n = 1), atrial fibrillation (n = 1), or patient's request (n = 3). One death occurred during hospitalization. Engraftment of neutrophils occurred on day 11.5 (mean; range 8-21 days) and platelets on day 11.7 (mean; range 7-33 days). The maximal tolerated dose of IL-2 was 6 x 10(5) IU/m2/day for 4 weeks. Disease-free survival rates for all stages were comparable to current reports in the literature. Preliminary laboratory evaluations include FACScan analysis of the IL-2 activated PBSC demonstrating an increased percentage of CD3+, CD25+, HLA-DR+ T cells. Phenotypically similar cells were present in peripheral blood samples of patients when tested 15 days after transplantation. This study demonstrates successful engraftment with IL-2-activated PBSC after high-dose chemotherapy for women with stage II-IV breast cancer. The regimen is feasible and, although toxicities are common, they are manageable and correlate with increasing dose and duration of IL-2.

Adult↗
PubMed1996

Hematopoietic potential of IL-2-cultured peripheral blood stem cells from breast cancer patients.

Patients receiving autologous transplants for various malignancies generally experience an increased incidence of relapse compared with patients receiving unmanipulated allogeneic transplants. We initiated a protocol for IL-2 activation of peripheral blood stem cells (PBSC) for induction of in vitro and in vivo autologous graft-versus-tumor (GVT) activity in patients with breast cancer. In this study we analyzed the effects of 24 h of IL-2 incubation on the hematopoietic potential of PBSC from these patients. Cells collected by leukapheresis were first cryopreserved and stored in liquid nitrogen, then thawed rapidly and incubated with IL-2 in a serum-free system for 24 h, with samples analyzed before and after incubation. Although there was a significant drop in mononuclear cells (MNC) (from 4.5 to 3.7 x 10(8)/kg) and CD34+ cells (from 12.3 to 7.5 x 10(6)/kg) after 24 h in culture, there was no significant change in colony-forming units (CFU) (from 12.5 to 11.5 x 10(5)/kg). Time to engraftment (neutrophils: < 0.5 x 10(9)/l; platelets: > 20 x 10(9)/l) was comparable to a cohort of similar patients receiving non-cultured PBSC transplants. These results indicate that mobilized frozen/thawed PBSC which have been cultured in IL-2 for 24 h retain adequate potential for hematopoietic reconsistution in this group of patients.

Breast Neoplasms↗
PubMed1994

The impact of harvest center on quality of marrows collected from unrelated donors.

The total number and distribution of nucleated cells in harvested bone marrow are potentially important determinants of patient outcome following bone marrow transplantation. In order to assess whether marrows collected from predominantly unrelated donors at Georgetown University Medical Center (GUMC) were different in cellular content from marrows collected at harvest centers outside of GUMC, we compared the nucleated cell counts and mononuclear cell subset distribution (CD34, CD3, CD4, CD8, CD19 antigen-positive cell content) of 10 consecutive marrows harvested at GUMC to 10 unrelated donor marrows from outside harvest centers. Significantly higher nucleated cell counts and CD34 antigen-positive cell content and significantly lower CD3 and CD4 antigen-positive T-cell numbers were demonstrated among the marrows harvested at GUMC. These results confirmed significant variability in marrow collection practices between GUMC and 10 different outside harvest centers and suggest that strict adherence to a specific collection procedure, involving small volume marrow aspirations and multiple puncture sites, results in a product with a high number of early hematopoietic progenitor cells and minimal contamination by peripheral blood. These data further suggest the need for careful monitoring of individual unrelated donor marrow collection centers' practices to optimize the quality of the harvested marrow.

Academic Medical Centers↗
PubMed1993

Use of a licensed electrolyte solution as an alternative to tissue culture medium for bone marrow collection.

Bone marrow for transplantation is traditionally collected into tissue culture medium with heparin. A licensed electrolyte solution (Plasma-Lyte A [PLA]) was used as a substitute for tissue culture medium in the harvesting of 28 bone marrows, 17 autologous and 11 allogeneic, which were subsequently transplanted. Data that were analyzed from the 25 evaluable patients consisted of the numbers of cells and colony-forming units in the transplanted marrow as well as the time to neutrophil and platelet engraftment. These results were compared with those in the 30 (26 evaluable) preceding transplanted marrows that were collected into a tissue culture medium (RPMI-1640 [RPMI]). The autologous marrow transplant patients in both the PLA and RPMI groups reached a neutrophil count of > or = 0.5 x 10(9) per L a mean of 19 days following transplantation. The patients who underwent transplantation with allogeneic bone marrow collected in RPMI achieved > or = 0.5 x 10(9) per L of neutrophils an average of 20 days following transplantation, while those who received marrow collected in PLA achieved engraftment of neutrophils to that level in a mean of 21 days. Because in vitro and in vivo results with RPMI and PLA are similar in this study, further work using a licensed solution for clinical bone marrow transplantation is indicated.

Adolescent↗
PubMed1993

A novel approach to immunomodulation of frozen human bone marrow with interleukin-2 for clinical application.

Interleukin-2 (IL-2) activation of fresh or frozen bone marrow (BM) in vitro generates killer cells with potent anti-tumor effect both in vitro and in vivo. The IL-2-activated BM (ABM) retains the capacity to reconstitute the hematopoietic system in an autologous bone marrow transplantation (ABMT) setting. The killer cells lose their cytotoxicity if the ABM undergoes the procedures of freezing and thawing. Therefore, for clinical application, the ABM has to be generated after thawing a frozen stock of BM before ABMT. The thawed BM cells are fragile and may undergo lysis, resulting in clump formation and cell loss. The frozen autograft also contains components of cryoprotectant mixture whose effects on the generation of ABM have not been defined. The present studies have been carried out to optimize a technique of handling the frozen BM for immunomodulation with IL-2 for 24 h at 37 degrees C prior to ABMT, with minimal loss of cells. IL-2-activation of BM was carried out in bags containing serum free medium which were designed to permit gaseous exchange. Addition of deoxyribonuclease (DNAse) (100 micrograms/ml of BM concentrate) immediately after thawing and the presence of heparin (20 units/ml) in the medium completely abrogated immediate or delayed clumping of cells. The presence of DNAse and/or heparin during in vitro culture did not affect the cell viability, cytotoxicity against tumor cells or the progenitor cell activity of the ABM; all these functions were well maintained even when BM was placed in culture immediately after thawing (without washing). There was no microbial contamination.(ABSTRACT TRUNCATED AT 250 WORDS)

Bone Marrow↗
PubMed1991

Bone marrow processing for transplantation.

As indications for BMT increase, so do variations in bone marrow processing and manipulation techniques. Many centers have their own unique methods of mononuclear cell purification, concentration and storage. This is particularly evident in the processing of bone marrow for autologous BMT to allow dose intensification as salvage therapy for malignant disease. Unique procedures have been developed to maximize yields, concentrate mononuclear cells necessary for engraftment, and reduce the likelihood of GVH disease. Graft rejection and disease relapse still remain a problem in some of these "manipulated" marrows. Newer procedures may allow titration of the optimum numbers of immune reconstituting cells; however, at this time, these techniques are not precise and the balance between preventing GVH disease at the expense of graft failure or relapse may still jeopardize disease-free survival. Innovative purging techniques that include pharmacologic and immunologic methods, continue to evolve, necessitating standards for bone marrow processing that are flexible yet practical. Quality control and viability assays are essential to verify the biologic proliferative potential of progenitor cells capable of marrow reconstitution. Although no standards are yet established, all centers should have criteria to monitor the quality of the processed marrow. Blood banks and transfusion services are well versed in regulations governing processing, labeling, storage, and quality control of blood components. Bone marrow is the ultimate blood component, and it stands to reason that methods outlined in this article be integrated into transfusion medicine.

Blood Component Transfusion↗
PubMed1991

Automated processing of human bone marrow can result in a population of mononuclear cells capable of achieving engraftment following transplantation.

A concentrate of mononuclear bone marrow cells is often desired for ex vivo treatment with pharmacologic agents, monoclonal antibodies, cytokines, and other agents prior to transplantation. A method has been developed for automated separation of mononuclear cells from large volumes of harvested bone marrow. A programmable instrument originally designed for clinical ex vivo cell separation and the plasma-pheresis of patients and blood donors was adapted to permit rapid preparation, in a closed sterile system, of a bone marrow product enriched with mononuclear cells. A mean (+/- SEM) of 53 +/- 30 percent of the original mononuclear cells was recovered in a volume of 125 +/- 42 mL containing 82 +/- 12 percent mononuclear cells. This technique removed 95 +/- 9 percent of the red cells in the original marrow. No density gradient materials or sedimenting agents were employed in this process. Of 36 marrows processed by this technique, 19 autologous (6 of which were purged with 4-hydroperoxycyclophosphamide) and 7 allogeneic marrows have been transplanted, with all evaluable patients achieving a neutrophil count of 0.5 x 10(9) per L in a mean (+/- SEM) of 21 +/- 6 days.

Bone Marrow Cells↗
PubMed1990

Processing and storage of human bone marrow: a survey of current practices in North America.

We conducted a two-step survey to question 110 transplant centers in the United States and Canada regarding marrow processing and storage policies and procedures. Approximately 65% of the centers surveyed responded to the questionnaires. Major differences with respect to patient diagnoses, amount of marrow harvested, purging method applied, freezing procedure, storage bag, cell concentrations, storage duration, interval until transplantation, cell counting, viability determination and so forth were reported. Among those centers responding 13% stored not only autologous but also allogeneic marrow. There was no consensus regarding patient consent for duration of storage, coverage of cost for cryopreservation or utilization of stored marrow after a patient's death. Additional studies will be necessary to correlate in vitro methods of marrow storage with clinical transplantation results, and to determine the cost/benefit ratio of this approach to various diagnoses. This should provide the basis for the establishment of standards and should facilitate the approach to various ethical questions.

Bone Marrow↗
PubMed1990

Automated isolation of mononuclear cells using the Fenwal CS3000 blood cell separator.

We describe a method for in vitro isolation of mononuclear cells from peripheral blood or bone marrow using a Fenwal CS3000 Apheresis device without employing density gradients or sedimenting agents. The automatic processing program requires minimal operator intervention and no subjective operator decisions. A mean of 67% of starting mononuclear cells were recovered in a 100 ml product having 95% mononuclear cells and less than 1% of the original red blood cells. The average processing time was 35 minutes.

Bone Marrow Cells↗
PubMed1990

Cryopreservation and storage of human bone marrow: a survey of current practices.

As bone marrow transplantation is being used with increasing frequency, problems of storage space and cost, inventory control and disposal have arisen. Issues such as maximum storage time and acquisition of consent for marrow disposal need to be addressed before a large inventory is accumulated. Consideration should also be given to using non-infused marrows for research purposes. Eighty-three bone marrow transplant centers were surveyed in an attempt to establish a data base with regard to guidelines for storage of cryopreserved human bone marrow. Fifty-two centers (62.7%) responded to the questionnaire, 5 of which did not have an active cryopreservation program. The remaining 47 centers freeze and store autologous marrow from patients with leukemia, lymphoma, neuroblastoma and a large diversity of other conditions including solid tumors. Twelve centers (25.5%) specify maximum storage times of up to 5 years, but only 9 centers (19.1%) require the donor to sign a specific consent form for marrow disposal if it is not used for transplantation within a given time. Eighty-five percent of the responding centers reinfuse at least half of the marrows they freeze within 12 months of harvesting. It appears that at least 90% of marrows that are being reinfused have been stored for three years or less. However, the storage time of non-infused marrows extends even further, and autologous marrow has been reinfused successfully as long as eight years after storage.

Bone Marrow Transplantation↗
PubMedDate not supplied

Bulk cryopreservation of lymphocytes in glycerol.

The authors describe a method for freezing large amounts of peripheral blood lymphocytes (PBL) in a 20 percent glycerol solution. Between 0.6 and 4.3 X 10(9) cells in autologous plasma were frozen in polyethylene freezing bags in a final volume of 50 ml. The recovery after thawing averaged 89 +/- 14 percent with a mean viability by trypan blue dye exclusion of 80 +/- 7 percent (n = 11). In aliquots of fresh and frozen-thawed PBL from the same subjects radiolabeled with 111In, the radiolabeling efficiency for both fresh and thawed cells was 49 +/- 15 percent (p = 0.98, n = 5). The mitogen mean stimulation indices for glycerol-frozen cells (471 with phytohemagglutinin-M, 176 with pokeweed mitogen, and 380 with concanavalin A) were superior to those of cells frozen by a standard technique with dimethylsulfoxide (DMSO) (141, 47, and 123; p less than 0.05) and comparable to those of fresh PBL (403, 75, and 147). In a mixed lymphocyte culture, glycerol-frozen PBL showed significantly greater responsiveness to a pool of stimulator cells than did PBL frozen in DMSO (p = 0.03). Thawed cells are viable and functional as demonstrated by their response to mitogens and their ability to stimulate and respond in mixed lymphocyte culture.

Blood Preservation↗