Single-strand conformational polymorphism analysis of the M(r) 170,000 isozyme of DNA topoisomerase II in human tumor cells

Five cell lines selected for resistance to the cytotoxicity of inhibitors of DNA topoisomerase II have point mutations in the gene that codes for the M(r) 170,000 form of this enzyme. In each case, the mutation results in an amino acid change in or near an ATP binding sequence of the M(r) 170,000 isozyme of topoisomerase II. We used single-strand conformational polymorphism analysis to screen for similar mutations in other drug-resistant cell lines or in leukemic cells from patients previously treated with etoposide or teniposide. We also analyzed the region of the gene that codes for amino acids adjacent to the tyrosine at position 804 of topoisomerase II which binds covalently to DNA. CEM/VM-1, CEM/VM-1-5, and HL-60/AMSA human leukemic cell lines were used as controls; 3 of 3 known mutations were detected by migration differences of polymerase chain reaction products from the RNA extracted from these three lines. A previously unknown mutation was found in the tyrosine 804 region of the M(r) 170,000 topoisomerase II expressed by CEM/VM-1 and CEM/VM-1-5 cells. Sequence analysis showed that substitution of a T for a C at nucleotide 2404 resulted in an amino acid change of a serine for a proline at amino acid 802. No mutations in any of the ATP binding sequences or in the tyrosine 804 region were detected in polymerase chain reaction products from RNA extracted from human leukemia HL-60/MX2 or CEM/MX1 cells (both cell lines selected for resistance to mitoxantrone) or in human myeloma 8226/Dox1V cells (selected for resistance by simultaneous exposure to doxorubicin and verapamil). No mutations were detected in polymerase chain reaction products from RNA extracted from blasts of 15 patients with relapsed acute lymphocytic leukemia, previously treated with etoposide or teniposide. We conclude that: (a) single-strand conformational polymorphism analysis is useful for screening for mutations in topoisomerase II; (b) resistance to the cytotoxicity of inhibitors of DNA topoisomerase II is not always associated with mutations in ATP binding sequences or the active site tyrosine region of M(r) 170,000 topoisomerase II; and (c) mutations similar to those detected in drug resistant cells selected in culture have not been identified in blast cells from patients with relapsed acute lymphocytic leukemia, previously treated with etoposide or teniposide.     

Percutaneous transhepatic drainage of the nondilated biliary system

Although reticulocyte counts can be reliably performed for up to 48 h after storage in EDTA, it is unclear whether this is applicable to the pediatric age group. In order to evaluate this, manual reticulocyte counts were performed on 20 specimens from pediatric patients stored at 4 degrees C for up to 24 h post collection. Samples were evaluated at 1-3, 6, 12, 18, and 24 h after storage in EDTA vacutainer tubes at 4 degrees C. The age of the subjects ranged from 1 day to 9 years with a median age of 3 years. Patients' reticulocyte counts ranged from 0 to 27% (5.89 +/- 7.21). No clinically significant changes were evident in the reticulocyte count over 24 h after specimen collection. The mean of the 20 specimens at 1-3 h was 5.50 and at 24 h was 5.40 (P > .05). The standard deviation of the mean values ranged from 7.03 to 7.26 (P > .05). The results indicate that reticulocyte counts may be performed on previously drawn blood held at 4 degrees C for up to 24 h post collection in a pediatric population without significant difference from baseline values.     

An investigation of seated postural stability

Results of tests undertaken to identify which measures of seated posture control are most effective in two areas, distinguishing differences in the x and y direction control strategies for a given task and distinguishing differences in overall control strategies for pairs of different tasks, are presented. The test platform, calibration tests, test protocol, and data analysis method are described. The results of statistical analyses performed on the data are summarized     

Oncogene rearrangement in non-Hodgkin's lymphoma with a 14q+ chromosome of unknown origin

Southern blot analysis was performed with a panel of DNA probes to detect rearrangements of c-myc, bcl-1, bcl-2 and bcl-3 in 14 cases of B-cell non-Hodgkin's lymphoma (NHL) with a clonal cytogenetic rearrangement involving the chromosome 14q32 locus and no known donor chromosome [t(14;?)(q32;?)]. In our experience, 21% of all chromosomal abnormalities involving the 14q32 locus in B-cell NHL are of this type. We found oncogene rearrangements in five of the 14 cases: bcl-1 rearrangement on one mantle zone lymphoma, bcl-2 rearrangements in two follicular lymphomas, and c-myc rearrangements in two small noncleaved cell lymphomas. We conclude that a 14q32+ abnormality of unknown origin is a relatively frequent karyotypic finding in B-cell NHL. In one third of the cases, known oncogenes that have been previously described in reciprocal translocations involving the immunoglobulin heavy chain locus were shown to be involved in the 14q32+ abnormality. The translocations in the other cases are likely to have involved one of the above oncogenes with breakpoints not revealed by the probes employed, other known oncogenes, or oncogenes that have not yet been identified.     

The stability of LaNi5 in a Cu matrix

The reaction of LaNi₅ with Cu powder during sintering at high temperature was examined in order to obtain better conductivity and ductility. Pressurized and forged copper-LaNi₅ pellets were investigated by optical metallography, DTA, XRD and TEM. The need for heat treatment after pressing and/or forging is vital to obtain good diffusion bonding between the copper particles and to improve the material strength. No interaction between La, Ni and Cu were observed after heating at 1173 K but a very stable La₂O₃ phase was created around the LaNi₅ particle. Rolling up to 85% reduction showed good ductility.     

Low-temperature interface reaction in aluminium-silicon carbide particulate composites produced by mechanical alloying

An experimental investigation has been carried out on the reaction that takes place between 3 and 20 μm SiC particles and the aluminium alloy 1050 matrix of composite materials prepared by a mechanical alloying process. The work is different from that undertaken by other researchers in that the SiC-Al interface reaction has been studied in the temperature range 853–933 K, i.e., with the matrix initially in the solid state. Differential thermal analysis, X-ray diffraction and scanning electron microscopy all show that the SiC-Al reaction initiates in the solid state at temperatures as low as 883 K. The reaction produces Al4C3 and Si, the latter entering into solid solution in the aluminium matrix. At temperatures exceeding 903 K, the reaction produces a liquid phase and at this stage the speed of the interface reaction increases significantly. The results are discussed in terms of Al-Si-C metastable equilibrium and the kinetics of the reaction. This revised version was published online in November 2006 with corrections to the Cover Date.     

Efficient generation of a reshaped human mAb specific for the {alpha} toxin of Clostridium perfringens

We have used the technique of antibody reshaping to producea humanized antibody specific for the a toxin of Clostridiumperfringens. The starting antibody was from a mouse hybridomafrom which variable (V) region nucleo-tide sequences were determined.The complementarity-determining regions (CDRs) from these Vregions were then inserted into human heavy and light chainV region genes with human constant region gene fragments subsequentlyadded. The insertion of CDRs alone into human frameworks didnot produce a functional reshaped antibody and modificationsto the V region framework were required. With minor frameworkmodifications, the affinity of the original murine mAb was restoredand even exceeded. Where affinity was increased, an alteredbinding profile to overlapping peptides was observed. Computermodelling of the reshaped heavy chain V regions suggested thatamino acids adjacent to CDRs can either contribute to, or distort,CDR loop conformation and must be adjusted to achieve high bindingaffinity.     

Young children''s understanding of pretense expressions of independent agency

Data from the Cancer Registry of Slovenia were used in a cohort study to determine whether the incidence of second primary cancers in patients with first primary breast cancer differs from the incidence expected in the general population. Special interest was given to long-term survivors. The expected numbers of second primary cancers were calculated by multiplying the number of appropriate person-years at risk by the corresponding age- and calendar-period-specific cancer incidence rates for women in Slovenia. The risk of a second primary cancer was expressed as the standardized incidence ratio (SIR). Of the 8,917 patients newly diagnosed in the period 1961-85 and followed-up to the end of 1994, 547 (6.2 percent) developed second primary cancers, whereas 410 (4.7 percent) were expected (SIR = 1.3, 95 percent confidence interval [CI] = 1.2-1.4). The risk was higher among younger patients. In long-term survivors, the risk was increased significantly for second primary cancer of the breast (SIR = 1.4, CI = 1.1-1.7), lung cancer (SIR = 1.6, CI = 1.1-2.3), melanoma (SIR = 2.7, CI = 1.5-4.4) and non-melanoma skin cancers(SIR = 2.0, CI = 1.6-2.4), corpus uteri cancer(SIR = 1.6, CI = 1.2-2.1), ovarian cancer(SIR = 2.3, CI = 1.7-3.0), and thyroid cancer (SIR = 2.5, CI = 1.2-4.6). Our results confirm the findings of several cohort studies carried out in Europe, the United States, and Japan, indicating that breast cancer patients should be monitored carefully for the occurrence of second primary cancers.     

Formation of titanium and cobalt germanides on Si (100) using rapid thermal processing

Titanium and cobalt germanides have been formed on Si (100) substrates using rapid thermal processing. Germanium was deposited by rapid thermal chemical vapor deposition prior to metal evaporation. Solid phase reactions were then performed using rapid thermal annealing in either Ar or N₂ ambients. Germanide formation has been found to occur in a manner similar to the formation of corresponding silicides. The sheet resistance was found to be dependent on annealing ambient (Ar or N₂) for titanium germanide formation, but not for cobalt germanide formation. The resistivities of titanium and cobalt germanides were found to be 20 μΩ-cm and 35.3μΩ-cm, corresponding to TiGe₂ and Co₂Ge, respectively. During solid phase reactions of Ti with Ge, we have found that the Ti₆Ge₅ phase forms prior to TiGe₂. The TiGe₂ phase was found to form approximately at 800° C. Cobalt germanide formation was found to occur at relatively low temperatures (425° C); however, the stability of the material is poor at elevated temperatures.