Granulocytic Leukemia In Blast Crisis Research Articles

Synergistic action of tiazofurin with hypoxanthine and allopurinol in human neuroectodermal tumor cell lines

The activity of IMP dehydrogenase (EC 1.2.1.14), the key enzyme of de novo guanylate biosynthesis, was shown to be increased in tumor cells. Tiazofurin (TR), a potent and specific inhibitor of this enzyme, proved to be effective in the treatment of refractory granulocytic leukemia in blast crisis. We examined the effects of tiazofurin as a single agent and in combination with hypoxanthine and allopurinol in six different neuroectodermal tumor cell lines, the STA-BT-3 and 146-18 human glioblastoma cell lines, the SK-N-SH, LA-N-1 and LA-N-5 human neuroblastoma cell lines, and the STA-ET-1 Ewing tumor cell line. Tiazofurin inhibited tumor cell growth with ic 50 values between 2.2μM (LA-N-1 cell line) and 550 μM (LA-N-5 cells) and caused a significant decrease of intracellular GTP pools (GTP concentrations decreased to 39–79% of control). Incorporation of [8- 14C]guanine into GTP pools was determined as a measure of guanylate salvage activity; incubation with 100 μM hypoxanthine caused a 62–96% inhibition of the salvage pathway. Incubation with tiazofurin (100 μM) and hypoxanthine (100 μM) synergistically inhibited tumor cell growth, and the addition of allopurinol (100 μM) strengthened these effects. Therefore, this drug combination, inhibiting guanylate de novo and salvage pathways, may prove useful in the treatment of human neuroectodermal tumors.

Inhibition by tiazofurin of inosine 5′-phosphate dehydrogenase (IMP DH) activity in extracts of ovarian carcinomas

Cancer cells have an increased ability to synthesize GTP (guanosine triphosphate) because of increased activity of IMP DH (inosine 5′-phosphate dehydrogenase, EC 1.1.1.205). Because IMP DH activity is rate limiting for de novo biosynthesis of GTP, this enzyme was suggested as a sensitive target for chemotherapy. Tiazofurin (2-β- d-ribofuranosylthiazole-4-carboxamide) is converted in the cells into the active metabolite, TAD, (thiazole-4-carboxamide adenine dinucleotide) which potently inhibits IMP DH activity. By adding TAD to tissue extracts one can determine the extent of inhibition of IMP DH. We applied the IMP DH assay method to extracts of normal ovaries ( N = 11) and epithelial ovarian carcinomas ( N = 10). The IMP DH activity (mean ± SE) in ovarian carcinoma was 21.1 ± 5.8 which was markedly higher than that observed in normal ovaries (2.9 ± 0.7 nmol/hr/mg protein) ( P < 0.05%). The inhibition by TAD of IMP DH activity in ovarian carcinomas ( N = 4) was 81%. the results indicate that IMP DH activity is elevated sevenfold in ovarian carcinomas as compared to normal ovary and can be inhibited by exposure to tiazofurin (TAD). Similar high IMP DH activity and inhibition of the activity by TAD were observed in patients with chronic granulocytic leukemia in blast crisis among whom 70 to 80% remissions were reported. Since there is increased IMP DH activity in human ovarian carcinomas and in OVCAR-5 cells and tiazofurin and TAD inhibit IMP DH activity of these cells and the proliferation of human ovarian carcinoma xenografts in the mouse, tiazofurin may merit serious consideration for a Phase II trial for patients with recurrent/refractory epithelial ovarian carcinoma.

157. Tiazofurin (TR) treatment induced differentiation in patients with chronic granulocytic leukemia in blast crisis (CGL-BC) and was synergistic in HL-60 cells with ribavirin (RB), retinoic acid (RA) and gemcitabine (DFDC)

157. Tiazofurin (TR) treatment induced differentiation in patients with chronic granulocytic leukemia in blast crisis (CGL-BC) and was synergistic in HL-60 cells with ribavirin (RB), retinoic acid (RA) and gemcitabine (DFDC)

Schedule-dependent synergistic action of tiazofurin and dipyridamole on hepatoma 3924A cells.

Tiazofurin is an oncolytic nucleoside analog that has shown therapeutic activity in end-stage acute non-lymphocytic leukemia and in chronic granulocytic leukemia in blast crisis. Tiazofurin is anabolized to the active metabolite, TAD, which inhibits IMP dehydrogenase activity, leading to a reduction in guanylate pools and to the cessation of neoplastic cell proliferation. The drug exhibits potent cytostatic and cytotoxic activity against hepatoma 3924A cells in culture. In growth-inhibition and clonogenic assays, the 50% inhibitory concentration of tiazofurin was 3.8 and 4.2 microM, respectively. Dipyridamole, an inhibitor of nucleoside transport, curtails the salvage of nucleosides and bases for nucleotide biosynthesis. Dipyridamole exhibited cytotoxicity against hepatoma 3924A cells, with an LC50 of 24 microM and an IC50 of 29 microM being recorded. A combination of tiazofurin and dipyridamole provided synergistic cytotoxicity in hepatoma 3924A cells in culture. This synergistic activity was dependent on the order of addition of the drugs. Simultaneous addition of the two drugs produced antagonism, whereas preincubation of cells with tiazofurin or dipyridamole followed by addition of the second drug resulted in synergy. TAD concentrations were significantly higher (129% and 135%) in cells that had been pretreated with tiazofurin or dipyridamole before the addition of the second agent as compared with cells that had been treated simultaneously (113%). These studies indicate the importance of the order of the addition of drugs to obtain a synergistic response in combination chemotherapy and suggest the need for a careful selection of drug modulation in clinical trials of tiazofurin and dipyridamole.

Tiazofurin Down-Regulates Expression of c-Ki-ras Oncogene in a Leukemic Patient

The increased activity in cancer cells of inosine 5'-monophosphate dehydrogenase (IMP DH, EC 1.1.1.205), the rate-limiting enzyme of de novo GTP biosynthesis, was suggested as a sensitive target for chemotherapy. Tiazofurin (NSC 286193), through its conversion to the active metabolite, thiazole-4-carboxamide adenine dinucleotide (TAD), is a strong inhibitor of IMP DH. In our clinical trial, tiazofurin caused return to the chronic phase in patients with chronic granulocytic leukemia in blast crisis (Tricot, G.; Jayaram, H.N.; Weber, G.; Hoffman, R. Tiazofurin: Biological effects and clinical uses. Int. J. Cell Cloning 8:161-170; 1990). In K562 human leukemic cells, tiazofurin down-regulated the expression of c-Ki-ras and c-myc oncogenes, which was followed by induced differentiation. We now report down-regulation by tiazofurin of the c-Ki-ras oncogene in a patient with chronic granulocytic leukemia in blast crisis. A single tiazofurin infusion (2,200 mg/m2) on days one and two decreased IMP dehydrogenase activity (the apparent t1/2 was 30 min), GTP concentration (the apparent t1/2 was 6 hr), and expression of ras (the apparent t1/2 was 8 hr) and c-myc (the apparent t1/2 was 38.5 hr) oncogenes in the leukemic cells. No further tiazofurin was given, because on days three and four the chemotherapeutic impact became evident in a tumor-lysis syndrome and the blast cells were cleared from the periphery by day five. The decrease in IMP DH activity, GTP concentration, and expression of c-Ki-ras oncogene were early markers of the successful chemotherapeutic impact of tiazofurin in a patient with chronic granulocytic leukemia in blast crisis.

Regulation of de novo and salvage pathways in chemotherapy

An overview was presented of our approach of inhibition of de novo and salvage pathways in pyrimidine and purine metabolism. 1. 1. Combination of acivicin, an inhibitor of de novo biosynthesis, and dipyridamole, a transport inhibitor, provided synergistic cytotoxicity in hepatoma and colon carcinoma cells. 2. 2. AZT, a competitive inhibitor of the salvage enzyme, thymidine kinase, and 5-FU or MTX provided synergistic cytotoxicity in hepatoma 3924A. In human colon carcinoma HT-29 cells AZT and methotrexate yielded synergistic cytotoxicity and thymidine and hypoxanthine together provided protection from the action of these drugs. 3. 3. These observations are significant because in rat hepatoma 3924A and in human cell lines HT-29, HL-60 and K562 thymidine kinase activity was 16- to 67-fold higher than that of dTMP synthase. Therefore, inhibition of dTMP synthase activity alone may provide poor responses because the salvage pathways can circumvent this block. 4. 4. In leukemic patients treated with tiazofurin, an inhibitor of IMP dehydrogenase, the rate-limiting enzyme of GTP biosynthesis, and with allopurinol, which inhibits GPRT activity through raising plasma hypoxanthine levels, synergistic therapeutic results were obtained. The responses in sensitive patients entailed a decrease in IMP dehydrogenase activity and GTP concentration in leukemic cells and down-regulation of the ras and myc oncogenes. The down-regulation of the ras oncogene by tiazofurin through the decrease of GTP concentration has now been shown in K562, HL-60 and hepatoma cells and in patients with chronic granulocytic leukemia in blast crisis. Tiazofurin may be useful in studies on selective depression of the expression of the ras oncogene. 5. 5. In 27 consecutive patients 50% responded positively to tiazofurin treatment. From this group, 10 out of 12 patients (83%) with chronic granulocytic leukemia in blast crisis responded to tiazofurin treatment.

Possible cytogenetic distinction between lymphoid and myeloid blast crisis in chronic granulocytic leukemia.

This study consists of 25 patients with chronic granulocytic leukemia in blast crisis (BC) or with acute leukemia who had a Ph1 chromosome and one or more other chromosome abnormalities and who were investigated by cytochemistry and immunocytochemistry techniques to determine whether the predominant blasts were myeloid or lymphoid. The disorder was myeloid in 15 patients, lymphoid in 8, and mixed in 2. Among the 15 patients with myeloid disorders, 13 (86.6%) had an additional Ph1 chromosome, i(17q), +8, +19, or some combination of these abnormalities. None of the eight patients with a lymphoid disorder had +8, +19, or i(17q), but one had an additional Ph1 chromosome. Among the eight patients with lymphoid disorders, two had structural abnormalities of chromosome 7 and two were monosomy 7. None of the patients with myeloid disease had a structurally abnormal chromosome 7, but one was monosomy 7. Our findings suggest that the number of chromosomes in an abnormal clone may be unreliable for distinguishing between lymphoid and myeloid BC. Most patients with myeloid disease had only abnormal metaphases, whereas many patients with lymphoid disorders had both normal and abnormal metaphases. This finding may partially explain why many patients with lymphoid BC respond better to treatment than do those with myeloid BC.

Myeloid lineage specificity and high sensitivity of monoclonal antibody GM 58/8 proves its usefulness as a diagnostic reagent in acute leukemia

Monoclonal antibody (MoAb) GM 58/8 was earlier reported to be directed against an antigen expressed by myeloid progenitors (CFU-GM), myeloid precursors, granulocytes, and monocytes. Immunophenotyping of 216 cases of acute leukemia [acute myeloblastic leukemia (AML) = 147 and acute lymphoblastic leukemia (ALL) = 69] and 18 cases of chronic granulocytic leukemia in blast crisis (CGLBC) with this antibody showed that GM 58/8 reacted with 92% of AML cases (M1-M5) and 100% of myeloblastic crisis in CGL cases. All cases of ALL, lymphoblastic crisis in CGL, erythroleukemia, and erythroblastic crisis in CGL were unreactive with GM 58/8. The antibody revealed the myeloid phenotype in an additional 15 cases of otherwise unclassifiable acute leukemia and six cases of CGLBC. Eleven cases of acute "mixed lineage" leukemia were also diagnosed with the help of GM 58/8. The high specificity (100%) and sensitivity (92%) of MoAb GM 58/8 for myeloblastic leukemia is unmatched by almost all previously described myeloid MoAb and proves its usefulness as a single diagnostic reagent for AML and myeloblastic crisis in CGL.

Correlation between chromosomal abnormalities and blast phenotype in the blast crisis of Ph-positive CGL

We carried out cytogenetic analysis in 23 patients with Ph-positive chronic granulocytic leukemia in blast crisis. In all cases the type of blast cell was characterized by cytochemistry, immunologic markers, and ultrastructural studies. Twelve cases were classified as myeloid transformation, six as lymphoid, two as mixed (lymphoid and myeloid), and two were unclassifiable. Duplication of Ph was the most frequent abnormality in the whole series. Trisomy 8, i(17q) and trisomy 19 were seen only in patients with myeloid blast crisis (53%, 30%, and 23%, respectively). Our findings suggest that the nature of additional chromosome abnormalities arising in blasts with features of myeloid differentiation are different from those in blasts showing lymphoid differentiation.

Histomorphologic study of bone marrow in acute leukemia following chemotherapy and autologous bone marrow transplantation.

Twenty-six patients with acute myeloid leukemia, acute lymphoid leukemia and chronic granulocytic leukemia in blast crisis were studied by means of multiple biopsies during a polychemotherapeutic or autologous bone marrow transplant protocol. Following chemotherapy, 3 main phases were observed: leukemic cellular depletion, stromal bone marrow reconstruction, and bone marrow hemopoietic restoration. Following intensive chemotherapy (in 2 patients after cyclophosphamide and total body irradiation) and autologous bone marrow transplantation, the 3 phases appeared to be shorter. A focal or diffuse increase in marrow fibrosis was a common finding in leukemia. An effective antileukemic therapy resulted in a decrease in fibrosis, whereas in some cases a further increase was a precocious sign of leukemia relapse.

Intravenous vincristine infusion: phase I trial.

In an attempt to sustain potentially cytotoxic concentrations of vincristine in man, a five-day continuous infusion of vincristine after an initial intravenous bolus injection was administered to 30 patients with refractory malignancies. Three dosage levels were explored (0.5 mg/m2, 0.75 mg/m2, and 1.0 mg/m2 daily for five days). Neurologic and hematologic toxicity were severe at the high dose level, whereas mild to moderate toxicity occurred at the 0.5 and 0.75 mg/m2 dose levels. Objective responses were noted in 11 patients (37%) with the following malignancies: non-Hodgkin's lymphoma (4), acute non-lymphoblastic leukemia (2), chronic granulocytic leukemia in blast crisis (1), carcinoma of the breast (3), and small cell carcinoma of the lung (1). Responses were observed at each infusion dose level. Nine of the 11 responders had previously progressed while receiving conventional intravenous bolus injection of vincristine. These data suggest that the clinical usefulness of vincristine may be enhanced by the use of infusion techniques. A maximum daily dose of 0.5 mg/m2 given for five days is recommended for future trials of intravenous vincristine infusion.

The light microscopic demonstration of hydroperoxidase-positive Phi bodies and rods in leukocytes in acute myeloid leukemia

Unique fusiform or spindle-shaped particles (Phi bodies) and rods with hydroperoxidase (catalase and/or peroxidase) activity are present in human granulocyte precursors only in acute myelogenous leukemia (AML). These newly recognized particles are much more numerous and prominent than Auer rods. They may be rapidly and readily identified using the microscope in marrow or peripheral blood films when the procedures recommended in this paper for fixation, incubation for hydroperoxidase demonstration in 3,3'-diaminobenzidine (DAB)/H2O2 medium, copper salt treatment and counterstaining (optional) with the Papanicolaou method are employed. Films prepared in the same manner but treated with benzidine/H2O2 medium for myeloperoxidase did not reveal these particles. We believe that Phi bodies are pathognomonic of AML since they are almost invariably present in AML patients with active disease. Their presence serves to distinguish AML from acute lymphocytic leukemia and from chronic granulocytic leukemia in blast crisis. Since the particles disappear in disease remission and reappear upon relapse, the recommended procedure is not only useful in diagnosis but in guiding therapy. When a very rapid diagnosis is needed, it is not necessary to counterstain the preparations, but the nuclei, cytoplasm and plasmalemma can readily be observed in the granulocyte precursors when they are counterstained by the Papanicolaou method. This treatment does not diminish the clarity of the Phi bodies and rods which stain by virtue of their peroxidatic activity. This cytochemical diagnostic procedure should be considered for adoption by hematology laboratories.

Unusual Cytogenetic Changes in Chronic Granulocytic Leukemia in Blast Crisis: Hyperdiploid and Two Ph′ Chromosomes.

s1 May 1966Unusual Cytogenetic Changes in Chronic Granulocytic Leukemia in Blast Crisis: Hyperdiploid and Two Ph′ Chromosomes.William H. KnospeWilliam H. KnospeSearch for more papers by this authorAuthor, Article, and Disclosure Informationhttps://doi.org/10.7326/0003-4819-64-5-1165_2 SectionsAboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissions ShareFacebookTwitterLinkedInRedditEmail ExcerptThe only consistent cytogenetic abnormality heretofore described among the human leukemias has been the Ph′ chromosome of chronic granulocytic leukemia. Several previous reports have described hyperdiploid cell lines with two Ph1 chromosomes.Case 1, a 25-year-old man, developed chronic granulocytic leukemia after a 5-month illness. He responded poorly to busulfan, and his course was complicated by massive painful lymphadenopathy and blast crisis. A lymph node biopsy section was initially interpreted as reticulum cell sarcoma but later revised to infiltration by primitive granulocytes. The granulocytes had a clonal line of 52 chromosomes including two Ph′ chromosomes. Lymphocytes from blood and lymph... This content is PDF only. To continue reading please click on the PDF icon. Author, Article, and Disclosure InformationAffiliations: MC Washington, D. C. PreviousarticleNextarticle Advertisement FiguresReferencesRelatedDetails Metrics 1 May 1966Volume 64, Issue 5Page: 1165-1165KeywordsBiopsyBloodChromosomesCytogeneticsGranulocytesLymphLymph nodesLymphocytesSarcoma Issue Published: 1 May 1966 PDF DownloadLoading ...