PubMed Health⌕ Search

Biomedical subjects

A Keating

Publications and source records attributed to A Keating.

At least 91 records · Page 5Linked to original sources
PubMed1995

Mini-BEAM as salvage therapy for relapsed or refractory Hodgkin's disease before intensive therapy and autologous bone marrow transplantation.

PURPOSE: To evaluate the efficacy of carmustine (BCNU), etoposide, cytarabine (Ara-C), and melphalan (mini-BEAM) as salvage therapy in patients with relapsed or refractory Hodgkin's disease who were potentially eligible to undergo intensive therapy and autologous bone marrow transplantation (ABMT). PATIENTS AND METHODS: Forty-four patients with refractory or relapsed Hodgkin's disease after front-line combination chemotherapy referred for consideration of ABMT were treated with mini-BEAM (BCNU 60 mg/m2 on day 1, etoposide 75 mg/m2 on days 2 to 5, Ara-C 100 mg/m2 twice per day on days 2 to 5, and melphalan 30 mg/m2 on day 6) to maximum response. Eleven patients were refractory to primary chemotherapy. Twenty-three patients were treated in first relapse and 10 in second or subsequent relapse; 21 received mini-BEAM as their first salvage regimen. Patients were restaged to determine disease status immediately before intensive therapy and transplant. RESULTS: The overall response rate was 84% (exact 95% confidence interval [CI], 70% to 92%), with a complete response (CR) rate of 32% (95% CI, 20% to 47%) and a partial response (PR) rate of 52%. No treatment-related deaths were observed. Myelosuppression was the major toxicity. Almost all patients required platelet transfusions. Eighty-four percent were given RBC transfusions, and 54% required intravenous antibiotics for fever while neutropenic. CONCLUSION: Mini-BEAM is a safe and effective regimen for treatment of refractory or relapsed Hodgkin's disease. Further studies are required to determine if responding patients have improved disease-free survival (DFS) after intensive therapy and ABMT.

Adolescent↗
PubMed1995

Outpatient bone marrow harvesting: an update.

We previously demonstrated in a pilot study that outpatient marrow harvesting is feasible and safe. Here, we provide an update and analysis of 149 outpatient marrow harvests and compare the results with 61 inpatient harvests. The mean patient age was 33 years. Mean outpatient harvest duration was 41 minutes, similar to inpatients. Average pre- and post-harvest hemoglobin values were 122 and 99 g/L, respectively, for outpatients and 119 and 93 g/L for inpatients. Fifty per cent of patients received an average of two units of homologous packed erythrocytes following the procedure versus 2.3 units for 57% of inpatients. Mean harvest volume was 1248 ml (versus 1341 ml for inpatients) and the mean number of mononucleated cells obtained was 2.73 x 10(8)/kg (3.23 x 10(8)/kg for inpatients). There were no serious complications in patients undergoing out-patient marrow harvesting. Sixteen outpatients (11%) had minor complications of hypotension, fever, nausea, vomiting or moderate pain. Thirteen patients (9%) required subsequent admission for a mean of 1.3 days. Eight inpatients (13%) had similar complications. All patients responded well to fluid replacement and analgesics. We confirm the safety of outpatient marrow harvesting for patients with hematologic malignancies.

Adult↗
PubMed1995

Microbial contamination of harvested bone marrow and peripheral blood.

The incidence and clinical consequences of microbiological contamination of autologous bone marrow and peripheral blood progenitor cells are not well documented. We therefore retrospectively analysed our experience with bacterial or fungal contamination of harvested bone marrow and/or peripheral blood. From January 1987 to the end of January 1994, 499 patients were harvested or which 301 were transplanted. A total of 3910 specimens obtained during three stages in the processing were assessed for microbial contamination: (1) in the operating room immediately after harvesting (1662 bags) with 2.1% culture positivity, (2) after processing for cryopreservation (1039 bags) with a further 1.1% cultures positive, and (3) after thawing at the time of reinfusion (1209 bags) of which 2.2% were culture positive. There were no culture positive specimens obtained from any peripheral blood progenitor cell products. The vast majority of the 85 culture positive specimens obtained from marrow were skin flora (89%) and 35% of all positive harvest specimens remained positive following processing and freezing. At least 36% of culture positive specimens were thought to have arisen as a result of exogenous contamination of blood culture bottles. Potentially pathogenic enteric organisms were present in nine (0.2%) specimens and infusion of these organisms occurred in four cases. A further seven patients were reinfused with marrow culture positive for skin organisms. No adverse clinical sequelae were noted following infusion of any contaminated products. However, clinical decision making continues to be influenced by culture results and multistage microbial culture continues to be of value in the management of our marrow recipients.

Bone Marrow↗
PubMed1995

Effect of antimicrobial prophylaxis on hematopoietic recovery following autologous bone marrow transplantation: ciprofloxacin versus co-trimoxazole.

We evaluated the effect of oral ciprofloxacin on neutrophil recovery in 20 consecutive patients undergoing autologous bone marrow transplantation (BMT) for malignant lymphoma and compared the results with a control group of 20 patients receiving co-trimoxazole and folinic acid. Both groups started the prophylactic antibiotic as well as granulocyte-macrophage colony-stimulating factor (GM-CSF) the day after marrow infusion and continued the former until the onset of febrile neutropenia (median duration of treatment 6 days for co-trimoxazole and 7 days for ciprofloxacin). The time of attain an absolute neutrophil count > or = 0.5 x 10(9)/L was significantly shorter in patients receiving ciprofloxacin (16 days vs 22 days; P = 0.006). There was no difference in time to attain a platelet count > or = 20 x 10(9)/L independent of transfusion or in time to the first febrile episode or incidence of bacteremia. We conclude that antibiotic prophylaxis with ciprofloxacin results in more rapid neutrophil recovery than prophylaxis with co-trimoxazole. This may result from a myelosuppressive effect of co-trimoxazole or an enhancement of neutrophil recovery by ciprofloxacin, or both.

Administration, Oral↗
PubMed1995

Optimizing the CD34+ and CD34+Thy-1+ stem cell content of peripheral blood collections.

We have previously described a sensitive and specific CD34 enumeration assay and report here a prospective analysis of 25 myeloma patients undergoing PBSC mobilization using this assay to determine the optimal days for collection of CD34+ and CD34+Thy-1+ cells after chemotherapy and growth factor mobilization. Correlations between frequency of peripheral blood CD34+ cells, circulating white blood cell (WBC) count, apheresis CD34+ cell count, nucleated cell count (NCC), and numbers of apheresis colony-forming units granulocyte/macrophage (CFU-GM) were determined. To assess levels of the more primitive subsets of CD34+ cells in the PBSC collections, coexpression of the Thy-1 antigen (CDw90) on CD34+ cells was also assessed. Marked heterogeneity was noted between patients with apheresis samples containing a median NCC of 4.2 x 10(8)/kg (range 1.3-8.1), median CFU-GM 17 x 10(4)/kg (range 0.15-32 x 10(4)/kg), and median CD34+ cell count of 1.39 x 10(6)/kg (range 0.02-6.6). The frequency of CD34+ cells in PBSC collections coexpressing Thy-1 was also heterogenous (6.2-50% of CD34+ cells), median 21.6%, mean 24.7 +/- 2%. The apheresis CD34+ cell count correlated with the peripheral blood CD34+ cell percentage (r = 0.71, p < 0.0001) but only weakly with the peripheral WBC. Apheresis CD34+Thy-1+ cell numbers correlated strongly with the circulating CD34+ cell numbers (r = 0.80), but no correlation was noted between these candidate stem cells and the peripheral WBC. In contrast, apheresis CFU-GM levels correlated most strongly with the peripheral WBC count (r = 0.61, p < 0.0001). The apheresis CD34+ cell count correlated with apheresis CFU-GM (r = 0.75, p < 0.0001) but not with the apheresis NCC. Apheresis CD34+Thy-1+ counts significantly correlated only with the apheresis CD34+ cell count and not with the apheresis CFU-GM or NCC. A higher percentage of circulating and apheresis CD34+ cells expressing Thy-1 were found on day 1 of collection, and the percentage of CD34+ cells expressing Thy-1 decreased on each subsequent day of measurement: median of 22% day 1 vs. 16.6% day 4, p = 0.04. This study therefore confirms that accurate quantitation of circulating CD34+ cells best predicts the optimal day for apheresis collection of CD34+ and CD34+Thy-1+ cells and is superior to the WBC count in this regard. Furthermore, the candidate stem cell (CD34+Thy-1+) subset is most prevalent during the earliest phases of CD34+ cell mobilization.

Blood Component Removal↗
PubMed1994

Variable transcription of BCR-ABL by Ph+ cells arising from hematopoietic progenitors in chronic myeloid leukemia.

Recent studies suggest that the BCR-ABL gene plays a critical role in the pathogenesis of Ph+ chronic myeloid leukemia (CML). We investigated the hematopoietic colonies derived from the marrows of 12 patients with Ph+ CML in chronic phase by reverse transcriptase-polymerase chain reaction (RT-PCR) amplification of BCR-ABL mRNA and by cytogenetics. Colonies were individually harvested and each colony divided into two portions, one for cytogenetics and the other for isolation of total RNA for PCR of BCR-ABL transcripts and for an RNA internal control. We found that 23% +/- 18% (mean +/- SD, range 0% to 60%) of Ph+ colonies did not transcribe the aberrant gene. In each case when BCR-ABL transcription was not detected, normal ABL mRNA was present. The data suggest that hitherto unknown mechanisms may regulate BCR-ABL expression in some Ph+ cells and indicate that caution should be exercised in the interpretation of results using RT-PCR analysis of hematopoietic colonies from clinical specimens and from experiments with antisense oligonucleotides directed at the BCR-ABL gene. These data also raise the notion of a transitional Ph+ precursor cell in which BCR-ABL may become upregulated and lead to a fully expressed phenotype. We conclude that further studies correlating the frequency of Ph+ PCR- progenitors with prognostic clinical variables are warranted.

Adult↗
PubMed1994

Hematopoietic cytokines enhance survival of SCID mice undergoing high-dose irradiation.

We have investigated the effect of hematopoietic cytokines on the survival of severe combined immune-deficient (SCID) mice that received a high dose of radiation. In this study, female SCID mice were irradiated at doses ranging from 500 to 600 cGy and then transplanted with 2 x 10(6) male Balb/c marrow cells. Groups of transplant recipients received stem cell factor (SCF), interleukin-1 (IL-1), and IL-3, alone or in combination, once daily for 5 days immediately after irradiation. Control posttransplant SCID recipients did not survive more than 2 weeks after irradiation with the dose over 500 cGy. SCF alone did not enhance survival, and treatment with IL-1 or IL-3 had very limited capacity to improve survival. IL-1 plus IL-3 has some radioprotective effect on SCID recipients, but the strongest synergistic radioprotective effect was observed in mice treated with a combination of SCF, IL-1, and IL-3. These mice survived for more than 4 months after an irradiation dose up to 600 cGy. We also examined the origin of hematopoietic stem cells in transplant recipients. Bone marrow cells were obtained from the SCID mice treated with a combination of cytokines at 2 and 4 months after transplant with male Balb/c marrow cells and irradiation with 600 cGy. These marrow cells were then transplanted into secondary lethally irradiated female Balb/c recipients. Twelve-day spleen colonies (CFU-S) were analyzed by amplification of the Y-chromosome sequence of the sex-determining region by polymerase chain reaction (PCR). All spleen colonies were of donor origin, indicating that the SCID recipients were fully reconstituted by donor cells. The results suggest that SCF, synergistic with IL-1 and IL-3, protects SCID mice from lethal doses of radiation and allows complete long-term engraftment of SCID recipients.

Animals↗
PubMed1994

Synergistic effect of stem cell factor with interleukin-3 or granulocyte-macrophage colony-stimulating factor on the proliferation of murine primitive hematopoietic progenitors.

We examined the effect of recombinant murine stem cell factor (SCF) on murine primitive hematopoietic stem cells in vivo. Marrow cells from 5-fluorouracil (5-FU)-treated male CBA/J mice were transplanted into lethally irradiated female littermates. Immediately after marrow transplant, the mice received SCF, interleukin-3 (IL-3), and granulocyte-macrophage colony-stimulating factor (GM-CSF) alone or in combination daily for 6 days. Day-12 colony-forming units-spleen (CFU-S) and marrow-derived colony-forming units-granulocyte/macrophage (CFU-GM) were assessed. Bone marrow cells from primary transplant recipients were transplanted into a secondary group of lethally irradiated mice, and the number of spleen colonies arising after 12 days' engraftment was determined as pre-CFU-S. SCF alone did not increase spleen colony formation in either primary or secondary recipients. In contrast, treatment of primary recipients with SCF and GM-CSF or IL-3 or with all three cytokines resulted in a synergistic increase of CFU-S in secondary recipients, indicating increased pre-CFU-S levels. The cytokine combinations also produced synergistic increases of CFU-GM in primary recipient marrow. Evaluation of spleen colonies in secondary recipients by PCR amplification of the Y-chromosome sex-determining region indicated that about 80% were of donor (male) origin. We conclude that SCF with IL-3 and/or GM-CSF increases pre-CFU-S proliferation.

Animals↗
PubMed1994

Sensitive detection and enumeration of CD34+ cells in peripheral and cord blood by flow cytometry.

Peripheral blood stem cell autografts are increasingly used to reconstitute hematopoiesis after intensive, potentially marrow-ablative therapy. Assessment of autograft adequacy by enumeration of hematopoietic progenitors in colony-forming assays is handicapped by lack of reproducibility and prolonged assay time. Alternative approaches of graft assessment by flow-cytometric enumeration of stem/progenitor cells bearing the CD34 antigen can be hampered by low specificity and sensitivity. Here, we report a rapid and reliable multiparameter flow-cytometric approach to accurately enumerate CD34+ cells in peripheral blood (PB) mononuclear cells (MNCs). Total nucleated white blood cells (WBCs) are quantified by staining with fluorescein isothiocyanate (FITC)-conjugated CD45 antibody. Simultaneous staining by phycoerythrin (PE)-conjugated CD34 antibody defines an approximate number for the CD34+ progenitor/stem cell subfraction. When starting CD34+ cell numbers are low (0.01-0.5%), other nonspecifically stained leukocytes make accurate enumeration impossible. However, when the CD34+ fraction is analyzed for CD45 expression vs. side scatter (granularity), true CD34+ blast cells form a discrete cluster exhibiting low-density CD45 expression and low side-scatter characteristics. Cells within this "blast region" can be readily distinguished from lymphocytes, monocytes, granulocytes, and other events that can contaminate the CD34+ population. Here, we used this sensitive procedure to enumerate CD34+ cells in steady-state PB samples (0.03-0.09%), normal bone marrow (BM) aspirates, and umbilical cord blood collections (0.33-1.98%). This approach thus provides a means to analyze CD34+ cells in specimens from patients who have been extensively treated with chemotherapy and those undergoing PB stem cell mobilization with cytokines. Additionally, it is useful for assessment of CD34+ cells in a variety of clinical samples exhibiting perturbations of the hematopoietic progenitor/stem cell compartments.

Antigens, CD↗
PubMed1994

Peripheral neuropathy following high-dose etoposide and autologous bone marrow transplantation.

We investigated the incidence of peripheral neuropathy in 142 consecutive patients receiving high-dose etoposide between August 1988 and December 1991. A retrospective chart review was conducted. Six patients with hematologic malignancies presented with a new sensory polyneuropathy after receiving an intensive therapy regimen consisting of etoposide 60 mg/kg i.v. and melphalan 140-160 mg/m2 i.v. followed by autologous bone marrow transplantation. Neurologic symptoms began 2-8 weeks after transplant, were grade 2-3 in severity and pursued a chronic course with slow improvement over months. Electromyographic studies in three of the patients tested confirmed distal sensory polyneuropathy. All 6 patients had previously received vincristine 1-60 months prior to autotransplant. We conclude that peripheral neuropathy of moderate severity is a potential complication of high-dose etoposide therapy but appears to be self-limited.

Adolescent↗
PubMed1993

Structure-function analysis of the C-terminal segment of human interleukin-6.

It has been hypothesized that interleukin-6 (IL-6) and granulocyte-colony-stimulating factor (G-CSF) may fold as four-alpha-helix bundle proteins. To probe the functional role of the putative fourth helical segment of IL-6 (D-helix), a chimeric IL-6/G-CSF analog containing the predicted D-helix of G-CSF as well as a panel of IL-6 D-helix point mutants were analyzed for their respective secondary structure, antigenicity, and receptor binding and biological activities. The putative D-helix of IL-6 could not be replaced by its G-CSF counterpart in spite of their high degree of similarity and thus is indispensable for the antigenic and functional integrity of the IL-6 receptor binding site. Conversely, the grafting of the G-CSF D-helix did not confer any G-CSF activity to IL-6. A synthetic helical peptide containing the IL-6 D-helix was inactive, even when mixed with or linked to a peptide from the A-helix known to be involved in the active site. However, the conserved residues F173, R179, and R182 found in the D-helices of both IL-6 and G-CSF critically contribute to the architecture of the IL-6 active site. Indeed, mutation of F173 or R179 markedly affected IL-6 receptor binding and biological activities, but not the conformation of a major neutralization epitope. Furthermore, substitution of R182 resulted in a significant unfolding of the D-helix accompanied by a drastic loss in IL-6 antigenicity and functional activities. Nevertheless, residues other than F173, R179, and R182 also contribute to IL-6 specificity.

Amino Acid Sequence↗
PubMed1993

Role of stromal cells and macrophages in fibronectin biosynthesis and matrix assembly in human long-term marrow cultures.

Fibronectin is a major component of the extracellular matrix of adherent layers of human long-term marrow cultures where it may stabilize the extracellular matrix network and provide adhesion sites for primitive hemopoietic cells. This study was devised to analyze the role of adherent cell populations in fibronectin synthesis, matrix assembly, and degradation. In cultures performed under the conditions described by Gartner and Kaplan, immunoprecipitation after metabolic labeling showed that adherent cells synthesized a fibronectin variant comprising the EDa domain and lacking the EDb one. Vascular smooth muscle-like stromal cells were the cell subset responsible for this synthesis. Once synthesized by stromal cells, EDa+fibronectin was secreted into the supernatant and incorporated into the extracellular matrix. The cumulation in the extracellular matrix was predominant by weeks 5 and 6 of culture, when a decrease in the stromal cell intracytoplasmic content of fibronectin was observed. Stromal cells from a transformed cell line, L2Ori-, were also able to synthesize the EDa+fibronectin variant, although for these cells the assembly into the extracellular matrix was partly impaired. Besides stromal cells, other cell types participated in fibronectin synthesis: early-adhering granulomonocytic cells and macrophages appearing later in culture were able to synthesize an EDa-, EDb- fibronectin variant, clearly distinct from the EDa+ variant produced by stromal cells. Studies on cultures in which macrophage growth was stimulated at the expense of stromal cells by adding granulocyte-macrophage colony-stimulating factor (50 ng/mL) to the culture medium showed a striking decrease in amounts of fibronectin measured in the adherent layer. This decrease was caused by a lack of incorporation of fibronectin in the extracellular matrix, disclosing a major difference between stromal cells and macrophages in terms of matrix assembly. This study confirms the similarity between stromal cells and vascular smooth muscle cells, because in vivo subendothelial intimal aortic smooth muscle cells and cultured smooth muscle cells from the aortic media express the EDa+, EDb- fibronectin variant. Furthermore, our results suggest that the level of fibronectin in adherent layers is regulated by stromal cells and macrophages. The balance between these two cell populations may therefore be crucial for the local control of hemopoiesis by regulating the extracellular fibronectin available for the adhesion of hematopoietic cells. Our data indicate that it may be essential to study the adhesion of stem cells to EDa+, EDb- fibronectin instead of EDa-, EDb- soluble fibronectin, as found in human plasma.

Bone Marrow↗
PubMed1993

Androgen-induced inhibition of cell proliferation in an androgen-insensitive prostate cancer cell line (PC-3) transfected with a human androgen receptor complementary DNA.

A full length human androgen receptor complementary DNA was introduced into androgen receptor-negative PC-3 cells to determine if androgen sensitivity could be established in this cell line and to assess what influence, if any, androgen exposure would have on the growth of these cells. The androgen receptor complementary DNA was inserted into pSG5 in the region controlled by the SV40 promoter. This construct was cotransfected with pSR1neo into PC-3 cells and stably transfected cells were selected and screened for the expression of the androgen receptor. Active expression of the receptor was demonstrated by Western blotting using a rabbit anti-androgen receptor antiserum and by [3H]methyltrienolone binding to cytosol extracts. Saturation ligand-binding analysis revealed the presence of a single class, high affinity (Kd = 0.122 nM) androgen-binding site in cytosol extracts of transfected cells but not in extracts from mock-transfected cells. In cells expressing the transfected androgen receptor, androgen decreased the proliferation rate and cloning efficiency and induced a more differentiated phenotype. These results demonstrate that PC-3 cells have retained the mechanisms required to respond to the activated androgen receptor and that the loss of androgen sensitivity in these cells is due to the lack of functional androgen receptor. This also provides a technique for determining whether androgen-resistant tumor cells contain functional androgen receptors or whether androgen sensitivity is due to abnormalities in downstream signaling pathways. The apparent androgen-induced decreased malignant state of these transfected cells suggests new directions for the treatment of prostate cancer.

Animals↗