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R K Stuart

Publications and source records attributed to R K Stuart.

At least 37 records · Page 2Linked to original sources
PubMed1991

Pharmacokinetics of leucovorin metabolites in human plasma as a function of dose administered orally and intravenously.

Studies have shown that conversion of leucovorin to the metabolite 5,10-methylenetetrahydrofolate (5,10-CH2FH4) is responsible for enhancement of the antitumor effects of fluorouracil given in combination with leucovorin, but the biochemical basis of this conversion in humans is not fully understood. To determine a possible sequence of metabolic steps, we studied the pharmacokinetics of leucovorin and its reduced folate metabolites in plasma in healthy volunteers. Groups of five subjects were given two equal doses of 10, 25, 125, 250, or 500 mg/m2 leucovorin, one orally and one intravenously at a 30-day interval. A sensitive radioenzymatic method that we developed previously was used to measure plasma concentrations of [S]5-formyltetrahydrofolate, 10-formyltetrahydrofolate (10-CHOFH4), 5-methyltetrahydrofolate (5-CH3FH4), and the combined 5,10-CH2FH4 plus tetrahydrofolate (FH4) pools. Intravenous administration of leucovorin resulted in dose-dependent accumulation of 5,10-CH2FH4 + FH4 exceeding 2 microM at peak levels. After oral and intravenous administration, 10-CHOFH4 and 5,10-CH2FH4 + FH4 exhibited peak levels earlier and were eliminated more rapidly than 5-CH3FH4. Accumulation of all metabolites after intravenous administration was linearly dose dependent, while oral administration appeared to result in saturation. We propose that the host activation of leucovorin suggested by these findings could be responsible for elevation of intratumor 5,10-CH2FH4 levels, thus enhancing the antitumor effects of fluorouracil. These results also suggest that 10-CHOFH4, 5,10-CH2FH4, and FH4 are intermediate metabolites and that 5-CH3FH4 is the terminal metabolite. In addition, our results indicate that attainment of high plasma levels of the metabolites active in modulation of the therapeutic effects of fluorouracil is best achieved through intravenous administration of high doses of leucovorin. Our future studies will address the proposed sequential conversion pathway and, thus, the mechanism by which pharmacologically relevant reduced folates accumulate in plasma after leucovorin administration.

Administration, Oral↗
PubMed1991

Role of glutathione in the in vitro synergism between 4-hydroperoxy-cyclophosphamide and cisplatin in leukemia cell lines.

To explain the sequence-dependent in vitro cytotoxic synergism between 4-hydroperoxycyclophosphamide (4-HC) and cisplatin in the K-562 human leukemia cell line, we have hypothesized that 4-HC decreases cellular glutathione (GSH) levels and that the resulting diminution of the cellular protective effect of GSH leads to the increased cytotoxicity of cisplatin. Exposure of K-562 cells to 4-HC resulted in a concentration- and time-dependent depletion of cellular GSH. To determine the effect of modulation of GSH levels on the toxicity of cisplatin, K-562 cells were exposed to buthionine sulfoximine (BSO) and/or GSH ethyl esters. Depletion of GSH to approximately 10% of control values by BSO potentiated the cytotoxicity of cisplatin, while rapid replenishment of GSH to within normal levels by GSH esters abolished the potentiation of BSO. Doubling cellular GSH by incubation with GSH esters protected against cisplatin cytotoxicity. Of importance, pretreatment of K-562 cells with BSO, in addition to increasing the cytotoxicity of 4-HC and cisplatin, abolished the synergism between the two drugs. The working hypothesis was also tested in two other cell lines in which the cytotoxic synergism between 4-HC and cisplatin was exhibited: the Raji cell line, a human lymphoblastic cell line, and the L1210-CPA cell line, a subclone of the murine L1210 leukemia with resistance to 4-HC. GSH levels in these two cell lines were not altered by incubation with concentrations of 4-HC at which the synergism was observed. In conclusion, the data for the K-562 cell line, indicating that (a) 4-HC depletes cellular GSH levels, (b) the lowering of cellular GSH levels enhances the toxicity of cisplatin, and (c) intact GSH stores are required for the synergism, strongly support the postulate that the cytotoxic synergism between 4-HC and cisplatin is modulated by GSH levels in this cell line. However, the lack of 4-HC-mediated depletion of GSH at concentrations of 4-HC resulting in cytotoxic synergism in the Raji and L1210-CPA cell line indicates that mechanisms other than modulation of GSH levels by 4-HC are responsible for the synergism in these cells.

Animals↗
PubMed1991

Case report: isoimmune inhibition of erythropoiesis following ABO-incompatible bone marrow transplantation.

A 26-year-old ABO-O positive patient with aplastic anemia received a bone marrow transplant from his genotypically HLA identical, but ABO-A positive, brother. Engraftment of myeloid and megakaryocytic lineages occurred within 4 weeks but pure red cell aplasia and transfusion dependent anemia persisted for 160 days. The authors postulated that the failure of erythropoiesis was due to a high titer of anti-A isohemagglutinins. They tested this hypothesis with clonal cell cultures and flow cytometric analysis of ABO antigen expression by colony forming cells in vitro. During the period of prolonged red cell aplasia, the patient had normal numbers (85 +/- 12 per 10(6) cells) of circulating donor derived, burst forming units-erythroid (BFU-E). Immunophenotypic analysis of erythroid burst colonies derived from culture of the patient's bone marrow cells showed that 91 +/- 5% of 274 nucleated red cells were A-antigen positive, confirming full donor engraftment. Autologous plasma and complement added on day 1 of culture did not affect the colony growth (82.5 +/- 15 per 10(6) cells). However, when the addition of complement was delayed until day 7 of culture, there was 90% inhibition of BFU-E (7.5 +/- 5 per 10(6) cells) compared to controls (p less than 0.0004). Based on this, the authors propose a model for expression of ABO antigens during erythropoiesis, in which BFU-E do not express ABO antigens but their progeny do. The data support the hypothesis that the mechanism of prolonged pure red cell aplasia after ABO-incompatible bone marrow transplantation is complement mediated immune destruction of erythroid progenitors past the stage of BFU-E in differentiation.

ABO Blood-Group System↗
PubMed1990

A phase II trial of high-dose cytarabine and cisplatin in previously untreated non-small cell carcinoma of the lung. A Piedmont Oncology Association Study.

Thirty-seven chemotherapy-naive patients with advanced non-small cell lung cancer (NSCLC) were treated with cytarabine (3 g/m2 intravenously [IV] during 3 hours) after IV bolus cisplatin (100 mg/m2 repeated every 3 weeks). Aside from nausea and vomiting, the principal toxicity was hematologic, with Grade IV myelosuppression in 32% and Grade III in 14%. Four patients died while on study. One complete and four partial responses were observed for an overall response rate of 14%. Responses were limited to lymph node and lung metastases and occurred in two of 17 adenocarcinomas, two of 12 squamous cell carcinomas, and one of eight large cell carcinomas. At this dose, the plasma level of cisplatin is only 3 micrograms/ml and the plasma level of cytarabine is 10 to 50 micrograms/ml, compared with the levels of 10 micrograms/ml and 1000 micrograms/ml, respectively, required for in vitro synergy. The severity of myelotoxicity observed indicates that, even at these levels, cisplatin enhances cytarabine activity. The combination may prove useful in malignancies that are sensitive to cytarabine, but is not of benefit in cytarabine-resistant malignancies such as NSCLC.

Adult↗
PubMed1990

Cellular glutathione as a protective agent against 4-hydroperoxycyclophosphamide cytotoxicity in K-562 cells.

Exposure of cells of the K-562 erythroleukemia cell line to 4-hydroperoxycyclophosphamide (4-HC), an analog of activated cyclophosphamide, causes a concentration-dependent inhibition of in vitro colony formation by these cells. For investigation of the role of glutathione (GSH) in the metabolism of 4-HC, GSH levels of K-562 cells were modulated by exposing the cells to buthionine sulfoximine (BSO), a specific inhibitor of GSH synthesis, and/or to GSH ethyl esters. Both the mono- and diethyl esters of GSH were synthesized in our laboratories and their identities were determined by chromatographic methods and fast-atom-bombardment mass spectrometry. An HPLC method including electrochemical detection used for thiol determination was applied for the measurement of GSH esters. Incubation of the cells with BSO depleted GSH levels to approximately 11% of control values and potentiated the cytotoxicity of 4-HC. By contrast, exposure to GSH esters approximately doubled GSH levels and protected the cells against the toxicity of 4-HC. Moreover, when cellular GSH levels were first depleted by BSO exposure and then replenished by incubation with GSH esters, the BSO-associated potentiation of 4-HC cytotoxicity was abolished. The work described herein extends the application of an HPLC method used for thiol determination to the measurement of GSH ethyl esters. In addition, it established that GSH acts as a competitive protecting agent against the in vitro toxicity of 4-HC in the K-562 cell line.

Antimetabolites↗
PubMed1990

Combinations of 4-hydroperoxycyclophosphamide (4-HC) and cisplatin for bone marrow purging in autologous marrow transplantation: an update.

We are studying the usefulness of combinations of 4-HC and cisplatin as a potential purging regimen for autologous bone marrow transplantation. In all of our studies, in vitro cytotoxicity was determined by clonogenic assay, and drug interaction was quantitated using the multiple drug-effect analysis method. The cells were incubated for one hour (4-HC) and/or 4 hours (cisplatin). We found that the drugs in combination had cytotoxic synergism against human leukemia cell lines (K-562 and Raji). The synergism was sequence-dependent (cells must be exposed to 4-HC first), was present at various molar ratios of the drugs, and most pronounced at high levels of cell kill. We also found that the drugs had cytotoxic synergism against normal human marrow progenitors (CFU-GM). However, the leukemic cells were approximately 55 times more sensitive to the combination than CFU-GM. In a murine system, the drugs were synergistic against L1210 leukemia cells and normal murine CFU-GM, but L1210 cells were at least 130 times more sensitive to the combination than CFU-GM. To determine the ability of L1210 cells to grow in vivo after exposure to the drugs, BDF1 mice were injected with 2 x 10(4) cells which had been incubated with 4-HC and/or cisplatin. The survival time of untreated controls was 13 +/- 2.8 days. For treated groups, the cure rates after 50 days of observation were 33% (4-HC, 40 uM), 0% (cisplatin, 8 uM), and 100% (4-HC + cisplatin). Finally, at concentrations resulting in equivalent toxicity to marrow CFU-GM, cisplatin seemed to be more toxic to murine spleen blast colony forming cells (CFC-BC) than 4-HC. The drugs in combination appeared to have at least additive toxicities against CFC-BC.

Animals↗
PubMed1989

In vitro synergism of 4-hydroperoxycyclophosphamide and cisplatin: relevance for bone marrow purging.

Autologous bone marrow transplantation with 4-hydroperoxycyclophosphamide (4-HC)-purged bone marrow gives long-term remission in almost half of relapsed acute nonlymphocytic leukemia and non-Hodgkin's lymphoma patients, but relapse of disease is the main cause of failure, suggesting ineffective purging in some cases. Cisplatin (CP) has activity against a variety of human tumors and is not commonly used for initial therapy of leukemia and lymphoma. Using established human leukemia cell lines, combinations of 4-HC and CP were investigated as a potential regimen for improving the ex vivo removal of leukemia cells from bone marrow. The cell lines (K-562 and Raji) were incubated for 1 (4-HC) or 4 h (CP), washed, and assayed for inhibition of colony formation in semisolid media. In both cell lines, CP (4h) was more potent than 4-HC (1 h). Combinations of the drugs in various molar ratios were studied after the cells were sequentially incubated with 4-HC and CP. The effects of the drugs were analyzed using the multiple drug-effect analysis of Chou and Talalay. Analysis of data on in vitro inhibition of colony formation suggested that all combinations studied were synergistic in both cell lines, with the greatest synergism being found in the Raji cell line. In addition, for K-562 cells we could detect at least a 4.6 log reduction in cloning with the CP:4-HC combination (1:10 molar ratio). We conclude that CP is a potential candidate in drug combinations for ex vivo bone marrow purging because of its high potency against human leukemia cell lines, its synergistic activity in combination with 4-HC, and its ability to reduce a high tumor burden when combined with 4-HC.

Bone Marrow↗
PubMed1989

Tumor-promoting phorbol esters support the in vitro proliferation of murine pluripotent hematopoietic stem cells.

The effect of tumor-promoting phorbol esters on the in vitro proliferation of mouse pluripotent hematopoietic stem cells (CFU-S) was examined using a short-term in vitro culture system and an 11-d spleen colony assay. Phorbol myristate acetate (PMA, 10(-7) M), but not the inert compound phorbol, supported the in vitro survival of day 11 CFU-S for 72 h in a manner similar to IL 3. PMA also enhanced the effect of IL 3 on the in vitro survival of day 11 CFU-S and as little as 1 h of exposure to PMA was sufficient for this purpose. The effect of PMA on CFU-S survival in vitro was not mediated by prostaglandins, did not require an established adherent cell population, and was observed at a concentration of 10(-9) M. PMA alone did not enhance the in vitro survival of day 11 CFU-S at very low concentrations of FCS but was still able to potentiate the effect of IL 3 on these cells. PMA also enhanced the in vitro survival of day 11 CFU-S from mice treated with 5-fluorouracil or from marrow cells exposed to merocyanine 540 and light. The interaction of PMA with day 11 CFU-S was not inhibited by a neutralizing antiserum to IL 3 but was inhibited by the protein kinase inhibitor H-7. Together, the data indicate that tumor-promoting phorbol esters interact with pluripotent hematopoietic stem cells. Like IL 3, their effect appears to be permissive and involves stem cells with marrow repopulating ability.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1989

Breast cancer.

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Breast Neoplasms↗
PubMed1988

The occurrence of a peripheral T-cell lymphoma in a chronically immunosuppressed renal transplant patient.

A 32-year-old man received a cadavaric renal transplant in 1975 for end-stage renal disease and, thereafter, was treated with azathioprine and methylprednisolone for chronic immunosuppression. In 1985, he presented with fever and pancytopenia that persisted despite withdrawal of the immunosuppressive agents. Lymph node and liver biopsies demonstrated malignant lymphoma within the sinuses of the node and the sinusoids of the liver. A splenectomy was performed for persistent pancytopenia, and the spleen demonstrated malignant lymphoma of the diffuse mixed large and small cell type exclusively within the cords of the red pulp. The immunophenotype of the tumor cells was obtained by frozen section immunoperoxidase staining with monoclonal antibodies and flow cytometric analysis. The tumor cells were positive for the Pan T cell markers CD3 and CD2, but were negative for the subset markers CD4 and CD8. A DNA hybridization study conducted on the splenic tissue conclusively identified the clonal nature of the malignant T cells by demonstrating rearrangement of the T cell receptor beta gene. In spite of multiple chemotherapeutic regimens, the patient developed increasing peripheral blood involvement and died with disseminated lymphoma. This case appears to be unique in that it is the first report of a chronically immunosuppressed transplant recipient to develop a malignant lymphoma of the mature T cell type, and several of the pathologic features of the tumor have not been observed previously.

Adult↗
PubMed1988

Consultation letters as a method for assessing in-training performance in a department of medicine.

This study examined the feasibility of using outpatient consultation letters as a means of assessing the clinical performance of clerks and house staff in a large teaching hospital. A total of 222 assessments were obtained based on samples of consultation problems from two clinical services. The method involved a two-stage rating scale based on the performance areas of Data Base, Problem Formulation, Diagnostic Tests, Management and Overall Assessment. The technique was investigated in terms of the objectivity, reliability, validity and efficiency of the performance ratings. Product-moment coefficients were used to assess inter- and intrarater agreement in scoring the consultation letters. These analyses identified a consistent difficulty in assessing the category of Data Base. For the remaining performance categories, interrater coefficients in the range .19-.76 and intrarater coefficients in the range .23-.96 were obtained. Other preliminary findings indicated good potential for a reliable and valid assessment. The method is technically feasible, and is based on a novel and reasonably unobtrusive approach to assessing trainee performance in a clinical setting.

Canada↗
PubMed1987

Nutritional support of bone marrow transplant recipients: a prospective, randomized clinical trial comparing total parenteral nutrition to an enteral feeding program.

Although standard supportive care for bone marrow transplant (BMT) recipients includes total parenteral nutrition (TPN), it has not been shown that this is the most appropriate method of nutritional support. To determine whether current BMT recipients require TPN during the early recovery period, we conducted a prospective, randomized clinical trial comparing TPN and an individualized enteral feeding program (counseling, high protein snacks and/or tube feeding). Nutritional assessment included measurement of serum proteins, anthropometry, and body composition analysis. For the latter, total body water and extracellular fluid were measured by standard radioisotope dilution techniques and used to quantitate body cell mass and body fat plus extracellular solids (FAT + ECS). In 27 TPN patients, body composition 28 days after BMT, expressed as a percentage of baseline, was body cell mass, 100%, extracellular fluid, 108%, FAT + ECS, 108%, and in 30 enteral feeding program patients, was body cell mass, 93%, extracellular fluid, 104%, and FAT + ECS, 94%. Only the difference in FAT + ECS was statistically significant (p less than 0.01). Compared to the enteral feeding program, TPN was associated with more days of diuretic use, more frequent hyperglycemia, and more frequent catheter removal (prompted by catheter-related complications), but less frequent hypomagnesemia. There were no significant differences in the rate of hematopoietic recovery, length of hospitalization, or survival, but nutrition-related costs were 2.3 times greater in the TPN group. We conclude that TPN is not clearly superior to individualized enteral feeding and recommend that TPN be reserved for BMT patients who demonstrate intolerance to enteral feeding.

Adolescent↗
PubMed1987

Dye-mediated photolysis of normal and neoplastic hematopoietic cells.

The purpose of this study was to determine the sensitivity to merocyanine 540 (MC 540)-mediated photolysis of normal human hematopoietic progenitor cells and four leukemia cell lines (Daudi, Raji, K562 and HL-60). Late erythroid progenitors were the most sensitive normal cells. Early erythroid progenitors were of intermediate sensitivity. Granulocyte/macrophage progenitors and multipotent progenitors were the least sensitive normal marrow cells. A combination of dye concentration, serum concentration, and illumination that eliminated 50% of multipotent progenitor cells reduced the concentration of leukemic cells by greater than or equal to 4.5 log. It is conceivable that this difference in photosensitivity can be exploited for the extracorporeal purging of autologous remission marrow grafts.

Bone Marrow Cells↗
PubMed1986

Autologous bone marrow transplantation in patients with acute nonlymphocytic leukemia, using ex vivo marrow treatment with 4-hydroperoxycyclophosphamide.

We studied 25 patients with acute nonlymphocytic leukemia in second remission (20 patients) or third remission (5 patients) in whom autologous bone marrow transplantation was performed with use of marrow incubated ex vivo with the alkylating agent 4-hydroperoxycyclophosphamide. Patients received intensive cytoreductive therapy with busulfan and cyclophosphamide or cyclophosphamide and total body irradiation, followed by an infusion of marrow that had been collected in remission, treated with 4-hydroperoxycyclophosphamide, and cryopreserved. Four patients died from bacterial or fungal sepsis within the first month after transplantation, and one patient with persistent marrow hypoplasia died from gram-negative sepsis 155 days after infusion with autologous marrow. In the remaining patients, peripheral-blood levels of neutrophils in excess of 0.5 X 10(9) per liter and platelet counts over 50 X 10(9) per liter were attained at median intervals of 29 and 57 days after transplantation, respectively. Nine patients had leukemic relapses at 73 to 316 days (median, 182 days) after infusion of autologous marrow, for an actuarial relapse rate of 46 percent. Eleven patients (eight in second remission and three in third) remained in remission at a median of more than 400 days (range, greater than 230 to greater than 1653 days) after transplantation. The observed disease-free survival after transplantation with autologous marrow treated with 4-hydroperoxycyclophosphamide compares favorably with the results of syngeneic or allogeneic transplantation in similar groups of patients.

Acute Disease↗
PubMed1986

Hematologic and cytogenetic remission of 5q-refractory anemia after syngeneic bone marrow transplantation.

Refractory macrocytic anemia with hypolobulated megakaryocytic nuclei and partial deletion of the long arm of chromosome 5 has been termed the 5q- syndrome. Although long survival has been reported in a few cases of 5q- refractory anemia, accumulating evidence suggests that this syndrome is a preleukemic state with risk of transformation to acute nonlymphocytic leukemia as well as complications of bone marrow failure. This report describes the first apparently successful therapy for this disorder in a young man who originally presented with a clinical picture consistent with pure red cell aplasia and normal marrow chromosomes but with hypolobulated megakaryocytic nuclei. He was treated with vitamins, androgens, and sequential trials of immunosuppressive therapy, all without response. Two years after diagnosis, repeated marrow cytogenetic studies showed a 5q- abnormality in 70 percent and later in 100 percent of marrow metaphases. Because of transfusion-induced hemosiderosis and the availability of a cytogenetically normal monozygotic twin, bone marrow transplantation was undertaken. In light of the clonal (and suspected preleukemic) nature of the 5q- syndrome, the patient's marrow was ablated with a busulfan plus cyclophosphamide regimen used for patients with nonlymphocytic leukemia. Sustained engraftment of cytogenetically normal marrow ensued. Two years after transplantation, and following six months of regular phlebotomy, the patient was hematologically normal with a normal serum ferritin level.

Adult↗
PubMed1986

Antigenic analysis of hematopoiesis. V. Characterization of My-10 antigen expression by normal lymphohematopoietic progenitor cells.

The My-10 glycoprotein is an hematopoietic cell surface antigen expressed specifically by undifferentiated (blast) cells, constituting 1%-4% of normal adult bone marrow leukocytes. We used several immunological and in vitro culture methods to analyze the expression of this unique antigen on a variety of lymphohematopoietic progenitor cells. Colony-forming cells (CFC) for granulocyte-monocyte colonies (CFC-GM) and erythroid colonies (BFU-E) were predominantly My-10 positive. CFC with higher proliferative potential were more strongly My-10 positive than CFC with lower proliferative potential, and those for mixed-lineage and blast cell colonies were even more uniformly My-10 positive. Cells maintaining CFC-GM number in short-term marrow culture (pre-CFC) were found to be My-10 positive, as were lymphoid precursors defined by their content of intranuclear terminal deoxynucleotidyl transferase. More mature erythroid precursors (CFU-E) were heterogeneous for antigen expression and lost My-10 antigen progressively, in parallel with advancing maturational stage. The My-10 antigen permits rapid identification and purification of hematopoietic progenitor cells for further study or potential clinical application. The disappearance of the My-10 antigen, moreover, may be a probe for differentiation-linked cellular events.

Antigens, Surface↗