Total body irradiation and granulocyte colony-stimulating factor-combined high-dose cytarabine as a conditioning regimen in allogeneic hematopoietic stem cell transplantation for advanced myelodysplastic syndrome: a single-institute experience.

In this study, we retrospectively evaluated the efficacy and safety of total body irradiation (TBI) and granulocyte colony-stimulating factor (G-CSF)-combined high-dose cytarabine as a conditioning regimen for allogeneic hematopoietic stem cell transplantation (HSCT) in patients with advanced myelodysplastic syndrome (MDS). We evaluated 22 patients with advanced MDS, including refractory anemia with excess blasts (RAEB; n=10), RAEB in transformation (n=2), acute myelogenous leukemia transformed from MDS (n=6) and chronic myelomonocytic leukemia (n=4). The conditioning regimen consisted of 12 Gy of TBI and high-dose cytarabine (3 g/m(2)) every 12 h for 4 days, and the cytarabine was combined with continuous administration of G-CSF. The stem cell sources were bone marrow or peripheral blood stem cells from human leukocyte antigen (HLA)-identical siblings (n=12) and bone marrow from HLA serologically matched unrelated donors (n=10). Three patients experienced disease relapse, two of whom died of disease progression. Of 22 patients, 16 are currently alive and disease-free. The 5-year estimated overall survival, disease-free survival, relapse and non-relapse mortality rates are 76.7, 72.2, 16.6 and 14.1%, respectively. These results suggest that G-CSF-combined high-dose cytarabine could be a promising component of the conditioning regimen of allogeneic HSCT for advanced MDS, providing a low incidence of both relapse and treatment-related mortality.

Tacrolimus and methotrexate for the prophylaxis of graft-versus-host disease after unrelated donor cord blood transplantation for adult patients with hematologic malignancies.

Eighteen patients with hematologic malignancies underwent cord blood transplantation (CBT) from unrelated donors after being conditioned with myeloablative or reduced-intensity regimens, and received tacrolimus and methotrexate (15 mg/m(2) on day 1, 10 mg/m(2) on days 3 and 6) as graft-versus-host disease (GVHD) prophylaxis. The median number of nucleated cells in infused cord blood was 2.66 x 10(7)/kg (range 1.90 to 4.15 x 10(7)/kg). Engraftment was achieved in 16 of 18 patients. The median time to absolute neutrophil count >0.5 x 10(9)/L was 21.5 days (range 17 to 32), and the median time to platelet count >2.0 x 10(9)/L was 36 days (range 26 to 57). Of the 16 evaluable patients, five and eight had grades I and II acute GVHD, respectively, and none had grades III/IV acute GVHD. The cumulative incidence of grade II acute GVHD was 44.4%. Chronic GVHD occurred in 7 of 15 evaluable patients: limited type in three patients, extensive type in four patients. Of the 18 patients, 14 were alive and disease-free between 173 and 1514 days after CBT (median 746 days). The probability of disease-free survival at 2 years was 79.1%. These results, although in a retrospective study, suggested that tacrolimus and short-term methotrexate effectively prevented the occurrence of severe acute GVHD after unrelated CBT, and may contribute to a high survival rate.

Establishment and characterization of two human mixed mesodermal tumor cell lines from the same patient.

A cell line designated HIRS -BM was established from fluid aspirated from the sternal bone marrow of a 16-year-old female. Another cell line ( HIRS -PB) was derived from the peripheral blood of the same patient. Both lines grew well, multilayering rapidly without contact inhibition, and 62 serial passages were successively done within 28 months. Both cultures contained spindle- or fibrous-shaped cells that revealed neoplastic and pleomorphic features, and these cells were characterized as possessing cross-striations in the cytoplasm. The cross-striations were detected by phosphotungstic acid hematoxylin stain. Some elongated cells were stained positively with anti-myoglobin by use of periodic acid-Schiff methods. The primary tumor in the uterus was diagnosed as a mixed mesodermal tumor composed of adenocarcinoma and rhabdomyosarcoma cells. The karyotype exhibited hyperploidy and large submetacentric marker chromosomes, and the modal chromosome number was 84. No difference was found between the 2 cell lines except for growth behavior and heterotransplantability . HIRS -BM cells grew more rapidly and were highly transplantable. The HIRS -BM cells were transplanted into the subcutis of BALB/c nude mice and produced mixed mesodermal tumors resembling the uterine tumor, while the HIRS -PB cells could not be transplanted. Due to the histogenesis of the mixed mesodermal tumor being's obscure with histologic observations only, this study was performed to obtain data by tissue culture of the tumor and resulted in support of the combination theory reported in the literature in regard to tumor.

Growth factor(s) produced by a human leukemic cell line growing in a protein-free chemically defined medium.

To examine whether human leukemic cells produce growth factor(s), a protein-free culture line of human erythroleukemic cells (K-562T1) has been established. This unique cell line has been continuously propagated in protein-free Ham's F-10 medium without any supplement for 5 yr. Growth-promoting activity was determined by measuring [3H]thymidine incorporation into DNA in serum-deprived chick embryo fibroblasts. The conditioned medium of K-562T1 contained the growth-promoting activity against chick embryo fibroblasts, mouse 3T3-L1 cells, and K-562 human leukemic cells. This leukemia-derived growth-promoting activity was heat and acid stable and trypsin sensitive. The activity was destroyed by dithiothreitol. Size exclusion chromatography revealed three peaks of activity, with apparent molecular weights of 13,500, 6,300, and 2,400, respectively.

Thermodynamic geometry and critical aspects of bifurcations.

This work presents an exploratory study of the critical aspects of some well-known bifurcations in the context of thermodynamic geometry. For each bifurcation its normal form is regarded as a geodesic equation of some model analogous to a thermodynamic system. From this hypothesis it is possible to calculate the corresponding metric and curvature and analyze the critical behavior of the bifurcation.

Treatment of Cushing's disease with reserpine and pituitary irradiation.

Eighteen patients with Cushing's disease were treated with reserpine and pituitary irradiation. Complete remission was obtained in 9 out of 18 patients after reserpine treatment of 1-2 mg per day for a mean period of 20.4 months, and pituitary irradiation with a mean of 5,865 rads. In another 9 patients, reserpine 0.8-2 mg per day for a mean period of 22.5 months, and pituitary irradiation with a mean of 6,650 rads, were employed. Of these 9 patients, an additional subtotal adrenalectomy was carried out in 6 patients who are now in complete remission. Because of severe psychic symptoms resulted from the original disease in 2 of the remaining 3 patients, subtotal adrenalectomy was performed first and pituitary irradiation and reserpine treatment followed. Remission was eventually obtained in these 2 cases. One patient refused the operation, and thus had little clinical remission. All of the 17 cases in remission were followed up for periods of 6 months to 10 yr. During this time, only one case which had responded to reserpine and pituitary irradiation relapsed, but regained remission following resumption of therapy. Another died of cerebral glioblastoma 4 yr after remission of the disease. It was noteworthy that endocrinologic data including: plasma levels of ACTH and 11-OHCS, suppressibility by dexamethasone, responses of plasma GH to arginine and to insulin loads, and diurnal rhythm of plasma 11-OHCS were nearly normal in a considerable number of the cases in remission. Effectiveness of the combined therapy with reserpine and pituitary irradiation for treating Cushing's disease may support a working hypothesis that reserpine acts through some as yet unknown mechanism to correct a presumed central nervous disorder, while suitable pituitary irradiation probably corrects the pituitary dysfunction directly.

Studies of cardiotonic agents. 8. Synthesis and biological activities of optically active 6-(4-(benzylamino)-7-quinazolinyl)-4,5-dihydro-5-methyl-3(2H)- pyridazinone (KF15232).

We previously reported that (+/-)-6-(4-(benzylamino)-7-quinazolinyl)-4,5- dihydro-5-methyl-3(2H)-pyridazinone (+/-)-1, KF15232) showed potent cardiotonic activity with a strong myofibrillar Ca(2+)-sensitizing effect. As an extension of our work, we attempted to synthesize optically active 1. (+/-)-4-(4-(Benzylamino)-7-quinazolinyl)-3-methyl-4-oxobutyric acid (-)-menthyl ester (6) was separated into both diastereoisomers, and each was converted to optically pure 1 (> 99% ee) in an enantioselective manner. In order to determine the absolute configuration of the isomers, an alternative synthesis of optically active 1 was employed. The precursor of (-)-1 ((+)-9) was obtained by enantioselective synthesis from (R)-D-alanine. Consequently, we concluded that the absolute configuration of (-)-1 at the 5-position of the pyridazinone ring was R. The cardiotonic effects and inhibitory activities to PDE III and V of racemic 1 and (-)-1 were more potent than those of (+)-1. These compounds also demonstrated greater vasorelaxant effects in guinea pig aorta. In contrast, (+)-1 showed only weak cardiotonic and vasodilating effects, although the compound displayed potent Ca(2+)-sensitizing activity. Racemic and (-)-1 attracted our interest for the treatment of congestive heart failure.

Interconversion and stability of duocarmycins, a new family of antitumor antibiotics: correlation to their cytotoxic and antimicrobial activities in vitro.

Stability and interconversion of duocarmycins were studied in relation to their cytotoxicities and antimicrobial activities. The compounds studied included duocarmycin A and SA, which have a spirocyclopropylhexadienone moiety, and four halogenated seco-compounds of duocarmycin A: duocarmycin B1, B2, C1 and C2, from which the cyclopropane ring structure is absent. Duocarmycins were potent cytotoxic compounds to cells. The cytotoxic activity seen on Balb 3T3/H-ras cells after 72 h drug exposure was in the following order (IC50 (nM): concentration for 50% growth inhibition); SA (0.05) > A (0.3) > B2 (1.5) > B1 (3.0) > C2 (20) > C1 (40). Average minimum inhibitory concentrations (MICs) of duocarmycins against microorganisms showed essentially the same ranking order as that of cytotoxicity. There was a large difference between SA and A in their stability in aqueous solvents. For halogenated seco-compounds, a good correlation was found between their cytotoxicities in vitro and their conversion rate to duocarmycin A, suggesting that halogenated seco-compounds undergo closure to the spirocyclopropylhexadienone structure, the pertinent active form, in cells.

Hydroxyakalone, a novel xanthine oxidase inhibitor produced by a marine bacterium, Agrobacterium aurantiacum.

A new xanthine oxidase inhibitor named hydroxyakalone was isolated from the culture broth of a marine bacterium Agrobacterium aurantiacum N-81106. Structure of hydroxyakalone was determined to be 4-amino-1H-pyrazolo[3,4-d]pyrimidine-3-one-6-ol by the spectral studies of hydroxyakalone and its permethyl derivative. The concentration to induce 50% inhibition (IC50) was 4.6 microM against xanthine oxidase.

MS-282a and MS-282b, new inhibitors of calmodulin-activated myosin light chain kinase from Streptomyces tauricus ATCC 27470.

MS-282a and MS-282b were isolated from the culture broth of Streptomyces tauricus ATCC 27470 as inhibitors of smooth muscle myosin light chain kinase (MLCK). MS-282a and MS-282b inhibited the activity of chicken gizzard MLCK with IC50 values of 3.8 microM and 5.2 microM, respectively. Cyclic AMP-dependent protein kinase, cyclic GMP-dependent protein kinase and protein kinase C were not inhibited by 150 microM MS-282a at all. It is likely that MS-282a blocks MLCK activity by antagonizing calmodulin since 1) the compound inhibited calmodulin-dependent but not calmodulin-independent activity of MLCK; 2) the inhibition of MLCK was antagonized by increasing concentrations of calmodulin, and 3) the compound inhibited calmodulin-dependent cyclic nucleotide phosphodiesterase.

RES-1149-1 and -2, novel non-peptidic endothelin type B receptor antagonists produced by Aspergillus sp. III. Biochemical properties of RES-1149-1, -2 and structure-activity relationships.

RES-1149-1 and -2, produced by Aspergillus sp. RE-1149, were found to be non-peptidic antagonists for endothelin type B receptor (ET(B) receptor). RES-1149-1 and -2 selectively inhibited the endothelin-1 (ET-1) binding to ET(B) receptor in a competitive manner with IC50 values of 1.5 microM and 20 microM, respectively. RES-1149-1 inhibited the increase in intracellular Ca2+ concentration elicited by 1 nM ET-1 in COS-7 cells expressing human ET(B) receptor, but not in the case of cells expressing ET(A) receptor. In addition, some structure-activity relationships are described.

RP-1551s, a family of azaphilones produced by Penicillium sp., inhibit the binding of PDGF to the extracellular domain of its receptor.

Nine azaphilones designated RP-1551-1, -2, -3, -4, -5, -6, -7, -M1, and -M2 were isolated from the culture broth of Penicillium sp. SPC-21609 as inhibitors of PDGF binding to its receptor. RP-1551s inhibit the binding of PDGF AA to the extracellular domain of PDGF alpha-receptor with IC50 values ranging from 0.1 to 2 microM without affecting PDGF BB binding to the extracellular domain of PDGF beta-receptor. PDGF binding was not restored after the PDGF alpha-receptor extracellular domain was washed in an attempt to remove the RP-1551-1 bound to the receptor. This result suggests that RP-1551-1 may irreversibly interact with the PDGF alpha-receptor. Since many azaphilone compounds possess high reactivity with an amino group, RP-1551-1 may prevent PDGF AA binding by reacting with amino groups on the alpha-receptor extracellular domain.

Interindividual variation of maximal blood levels of tacrolimus after its oral administration in hematopoietic cell transplant recipients.

We investigated the pharmacokinetics of oral tacrolimus in 31 hematopoietic cell transplant recipients, identifying 2 subgroups based on the differences between C(0) (trough) and C(max) (maximal) levels: group A (n = 21; 68%) with a C(max)-C(0) value of <10 ng/mL, and group B (n = 10; 32%) with a C(max)-C(0) value of >or=10 ng/mL. Although the C(0) and C(12) values were not significantly different between the 2 groups, the mean area under the concentration curve for 12 hours (AUC(0-12)) was significantly greater in group B than group A (200.9 +/- 36.3 vs 155.1 +/- 43.1 ng.h/mL; P < .05), and the mean half-life was significantly shorter in group B than group A (13.55 +/- 6.70 vs 18.17 +/- 6.30 hours; P < .05). Thus after the oral administration of tacrolimus, we observed a notably high AUC due to high peak level, which we were unable to predict simply by measuring the trough level. A pharmacokinetic analysis of each patient was essential to optimize the oral tacrolimus dose.

Detection of minimal residual disease in cerebro-spinal fluid of a patient with acute myelogenous leukemia with t(16;21)(p11;q22) translocation by reverse transcriptase-polymerase chain reaction.

We describe a patient with acute myelogenous leukemia (AML) with t(16;21)(p11;q22) translocation, whose minimal residual disease (MRD) both in cerebrospinal fluid (CSF) and bone marrow (BM) was monitored by reverse transcriptase-polymerase chain reaction (RT-PCR). A TLS/ERG-FUS fusion transcript, which is known to be expressed by t(16;21)(p11;q22) translocation, was detectable by RT-PCR both in BM and CSF cells in the first complete remission, suggesting the existence of MRD. The disease relapsed 6 months after its onset and allogeneic peripheral blood stem cell transplantation (PBSCT) was undergone. A TLS/ERG-FUS fusion transcript became rapidly below the detection level after PBSCT. These findings suggest the usefulness of RT-PCR for the detection of MRD in CSF, which contains a limited number of cells, as well as BM.