Synergistic effect of 4-hydroperoxycyclophosphamide and etoposide on a human promyelocytic leukemia cell line (HL-60) demonstrated by computer analysis.

Autologous stem cell transplantation using cryopreserved bone marrow offers the opportunity to rescue patients from hematopoietic toxicity caused by intensive chemotherapy. This approach is potentially useful for high-risk leukemias as well as for other cancers. The development of suitable methods for purging malignant cells from the bone marrow will offer a better chance of success for autologous stem cell transplantation. In this paper, we describe our efforts at purging myeloid cells. HL-60, a promyelocytic leukemia cell line, was used as a model. 4-Hydroperoxycyclophosphamide (4-HC) and VP-16-213 (VP-16) (either alone or in combination) were used to treat HL-60 cells and normal bone marrow. The cytotoxic effect of 4-HC (29.2 micrograms/ml; 100 microM) upon the HL-60 cell line was 99.8 +/- 0.12% (SE), and the colony-forming units-granulocyte, macrophage (CFU-cs) of normal bone marrow was inhibited by 82.5%. VP-16, at a concentration of 25 micrograms/ml (42.5 microM), can kill 99% of HL-60 cells and inhibit 72.7% of the CFU-cs. A drug mixture containing 4-HC (29.2 micrograms/ml) and VP-16 (10 micrograms/ml) (combination ratio, 1:0.342) reduces HL-60 cells to an undetectable number, and the CFU-cs were inhibited by 87.2%. The laboratory data were further analyzed for the synergistic effect of these two drugs by quantitative determination of the median effect plot and the multiple drug equation recently described by Chou and Talalay (Adv. Enz. Regul., 22: 27-55, 1984). Interactions of two drugs at different effect levels and at different combination ratios were then determined by computer simulation. At high effect levels, 4-HC and VP-16 in combination gave a synergistic cytocidal effect on HL-60 leukemic cells and gave an antagonistic inhibitory effect on normal bone marrow CFU-cs. This combination greatly increases the safety margin. Computer simulation of a dose effect relationship has also shown that the 4-HC:VP-16 combination ratio of 1:0.342 yields a better selective effect than a ratio of 1:0.856. This quantitative analysis suggests that the combination of these two drugs at the selected dose level offers a good method for purging nonlymphoblastic leukemia cells.

Autologous bone marrow transplantation for patients with poor-prognosis lymphoma.

Review of prognostic factors at Memorial Hospital in New York City has shown that adult patients with large-cell lymphoma (diffuse histiocytic lymphoma by Rappaport classification) who have high lactic dehydrogenase (LDH) and/or bulky mediastinal or abdominal disease are destined to do poorly with conventional combination chemotherapy, with a 2-year disease-free survival of about 20%. Patients who relapse after conventional combination chemotherapy have a similar poor prognosis. Thirty-one such patients with lymphoma were studied to evaluate the efficacy of intensive radiotherapy (hyperfractionated total body irradiation [TBI] [1,320 rad]), and cyclophosphamide (60 mg/kg/d for two days) followed by autologous bone marrow transplantation (ABMT). Our results show a disease-free survival advantage (P = .002) for 14 patients who underwent ABMT immediately after induction of remission with 79% surviving at a median follow-up 49.2+ months, compared with a median survival of 5.2 months for 17 patients administered ABMT while in relapse and/or after failing conventional treatment. Our results support the use of aggressive therapy as early treatment for patients with poor prognostic features.

Brassinolide is a selective ribosomal cistron regulator in onion leaf base tissue.

Outer epidermal cells from the basal, equatorial, near-apical, and apical regions of the third turgid onion (Allium cepa L. var. yellow, sweet Spanish) leaf base were treated (3 and 6 h in the dark = T3 and T6, respectively) with brassinolide (Br, a brassinosteroid plant growth regulator; effects on excised pieces compared with those in water controls: there were no statistical differences between the T3 and T6 results). Br induced increases in the volume and changes in morphologies of the major nucleoli to a greater extent than observed for major nucleoli in basal through near-apical controls. No major nucleoli were activated in control or Br-treated apical tissue. Minor nucleolar organizer regions in control and Br-treated tissue remained inactive in all locations. We propose that Br is a major ribosomal cistron regulator.

The role of light and growth regulators in the opening of the dentaria petiolar hook.

The phenomenon of the etiolated hook is not restricted to the hypocotyl of the dicotyledenous plant (e.g., Phaseolus) but appears to serve a similar, adaptive function in the petioles of certain rhizomatous plants. The commonly employed regulants of hypocotyl hook opening were tested for their effect on the petiolar hook of Dentaria diphylla. The hook was found to require both light (red light promoted, far red inhibited) and the intact leaf for opening. The leaf requirement was fully replaced by gibberellic acid (0.04% in lanolin) but only in light; cobalt chloride (0.1-1.0 mm) promoted a partial opening in dark with or without leaf; and coumarin (1 mm), indoleacetic acid (1-4% in lanolin), and ethylene 10 microliter per liter all inhibited opening of hooks with or without lamina. The absolute requirement for light and leaf tissue and the replacement of proximal tissue by GA(3) alone represent marked differences in the physiology of hypocotyl and petiolar hooks. These differences are believed to indicate the necessity for concomitant leaf maturation in petiolar hook opening.

Gamma chain-associated cytokine receptors signal through distinct transducing factors.

The IL-2, IL-4, and IL-7 signaling pathways have been shown to utilize shared components. The receptors for these cytokines are composed of ligand-specific binding chains that associate with a shared signaling subunit, the common gamma (gamma c) chain. In addition, IL-2, IL-4, and IL-7 induce activation of a common set of nonreceptor tyrosine kinases, Jak-1 and Jak-3. We have further investigated the signaling events induced by these cytokines and find that the gamma c-associated receptors activate distinct signal transducing factors (STFs). In addition, we show that a 94-kDa STAT-related protein (p94) is activated in response to IL-2 and IL-7, but not IL-4. These data indicate that IL-2, IL-4, and IL-7 activate distinct signaling molecules which might be differentially recruited to the receptor complex by the ligand-specific units of the IL-2, IL-4, and IL-7 receptors.

Ion metabolism in a halophilic blue-green alga, Aphanothece halophytica.

The intracellular ion content of the halophilic blue-green alga, Aphanthece halophytica was studied as a function of age, external sodium and external potassium concentration. Intracellular Na+ was found to be about 0.38 millimoles/g dry mass. Intracellular K+ concentrations were as high as 1 M and varied directly with external salinity. Intracellular Ca++ and Mg++ were in the range previously reported for fresh water blue-green algae despite their extremely high extracellular concentrations. Average cell size is consistent at room temperature with two exceptions. When the outside K+ is lower than 6.5 mM the cells tend to be smaller with less intracellular K+ and high Ca++. In stationary phase cultrues the cells are larger with high intracellular Mg++ and low K+.

Comparative regimens for the ex vivo chemopurification of B cell lymphoma-contaminated marrow.

The success of autologous bone marrow transplantation for B cell lymphoma may depend on the efficacy of in vitro purification of patients' tumor cell-contaminated marrow. In this study, we tested the toxicity of seven different chemotherapeutic agents against two B cell lymphoma lines (LY-16 and SK-DHL-2) as compared to normal human bone marrow granulocyte-macrophage progenitor cells (CFU-GM). 4-Hydroperoxycyclophosphamide (4-HC), VP-16-213 (VP-16), nitrogen mustard, and vincristine showed a highly selective toxicity against cultured lymphoma cells; i.e., at doses sufficient to induce a 4-log clonogenic tumor cell reduction (4-HC 21 micrograms/ml, VP-16 50 micrograms/ml, nitrogen mustard and vincristine 5 micrograms/ml), 10.0 +/- 6.7, 3.0 +/- 3.2, 23.2 +/- 22.7 and 24.0 +/- 17.0% (mean +/- 1 SD), respectively, of normal bone marrow CFU-GM were preserved. The differential sensitivity of tumor cells and normal hematopoietic precursors was less prominent after exposure of cells to cis-diamminechloroplatinum II (cis-platinum); thus, at a drug dose of 100 micrograms/ml, all detectable lymphoma cells could be eradicated (i.e., greater than or equal to 4 log reduction) while a CFU-GM recovery of only 0.2 +/- 0.2% was observed. In contrast, adriamycin and bleomycin, at the highest tumoricidal concentrations tested (5 and 100 micrograms/ml, respectively) did not exhibit a selective toxicity toward lymphoma cell lines. In summary, our results suggest that nitrogen mustard and vincristine, as well as 4-HC and VP-16, may be useful agents for the ex vivo treatment of bone marrow grafts form B cell lymphoma patients.