Influence of laser power on atom probe tomographic analysis of boron distribution in silicon.

The relationship between the laser power and the three-dimensional distribution of boron (B) in silicon (Si) measured by laser-assisted atom probe tomography (APT) is investigated. The ultraviolet laser employed in this study has a fixed wavelength of 355nm. The measured distributions are almost uniform and homogeneous when using low laser power, while clear B accumulation at the low-index pole of single-crystalline Si and segregation along the grain boundaries in polycrystalline Si are observed when using high laser power (100pJ). These effects are thought to be caused by the surface migration of atoms, which is promoted by high laser power. Therefore, for ensuring a high-fidelity APT measurement of the B distribution in Si, high laser power is not recommended.

Dopant distributions in n-MOSFET structure observed by atom probe tomography.

The dopant distributions in an n-type metal-oxide-semiconductor field effect transistor (MOSFET) structure were analyzed by atom probe tomography. The dopant distributions of As, P, and B atoms in a MOSFET structure (gate, gate oxide, channel, source/drain extension, and halo) were obtained. P atoms were segregated at the interface between the poly-Si gate and the gate oxide, and on the grain boundaries of the poly-Si gate, which had an elongated grain structure along the gate height direction. The concentration of B atoms was enriched near the edge of the source/drain extension where the As atoms were implanted.

Reproductive status in GTD treated with etoposide.

OBJECTIVE: To investigate the mechanism and degree of ovarian dysfunction in gestational trophoblastic disease (GTD) patients treated with etoposide alone. STUDY DESIGN: Forty-seven patients with low-risk GTD were treated with etoposide alone, and pituitary-ovarian function was evaluated by measuring basal serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), estradiol and progesterone and by recording basal body temperature. Moreover, the responses of LH and FSH to the administration of LH-releasing hormone (LHRH) and the responses of prolactin to thyrotropin-releasing hormone (TRH) were analyzed after the completion of etoposide treatment. RESULTS: Increased basal LH and FSH levels were found in approximately 50% of patients, especially those over 40 years old. Although the LH and FSH responses to LHRH were exaggerated in patients with high basal FSH levels, the prolactin responses to TRH were normal. Ovulation resumed within 121 days after the cessation of chemotherapy in women under 39 years. However, five of nine patients over 40 years remained anovulatory during the follow-up period. CONCLUSION: Ovarian function was impaired in approximately 50% of patients treated with etoposide at the time of LHRH study, though pituitary function was preserved. This complication is age related but not related to the amount of etoposide exposure. Therefore, we must consider the possibility of permanent anovulation when we treat patients 40 years old and older.

[System for registering gestational trophoblastic disease in Chiba Prefecture in the past 20 years].

From 1974 to 1993, a total of 3,203 patients with hydatidiform mole (HM), were registered in Chiba Prefecture. The incidence of HM during the study period was 2.8 per 1,000 live births or one per 357 live births, but it decreased to 1.5 per 1,000 live births in these 3 years. In the past 20 years, the incidence of invasive mole (IM) and choriocarcinoma was 8.8% (282/3,203) and 0.91% (29/3,203), respectively. In addition, 1,599 patients with HM have been diagnosed and registered as complete mole (CM) and partial mole (PM) patients on the basis of macroscopic findings since 1981. Of these classified patients, the incidence of IM following CM and PM was 12.5% (141/1,130) and 1.5% (7/469), respectively. Moreover, 19 (1.7%) patients with CM and one (0.2%) patient with PM developed CC. In this study, it is clear that gestational trophoblastic disease after the evacuation of PM definitely occurred and the management of PM should be similar to that of CM so long as macroscopic criteria are employed.

[Studies on clinical usefulness of new tumor markers of ovarian cancer, CA54/61 and CA602--III. Measurement of serum samples from patients with various benign or malignant diseases].

Serum samples from patients with benign or malignant diseases were measured for the newly developed tumor markers CA54/61 and CA602 with the respective EIA kits for assessment of their utility as tumor markers. A total of 5236 patients were entered into the study, consisting of ovarian cancer patients, those with other cancers, pregnant women, and healthy volunteers. The CA54/61-positive rate with a cut-off value of 12 U/ml was 61.2% for ovarian cancers (50.4% with a cut-off value of 20 U/ml). A positive rate of 75.0% (64.4%) was achieved for mucinous cystadenocarcinoma, which was high, compared with that for CA125. On the other hand, the false-positive rate was 12.2%(5.9%) for benign ovarian tumors, and as low as 18.5%(8.7%) for endometriosis. The CA602-positive rate with a cut-off value of 63 U/ml was as high as 76.0% for ovarian cancers (69.8% with a cut-off value of 90 U/ml). On the other hand, the false-positive rate was relatively high at 21.9% (12.6%) for benign ovarian tumors, and 56.7% (40.0%) for endometriosis. These positive rates were therefore similar to those for CA 125. The levels of both CA54/61 and CA602 antigens well reflected the postoperative prognosis. These results suggest the utility of CA54/61 and CA 602 as tumor markers of ovarian cancers.

Human cell clones, RSa and UVr-1, differing in their capability for UV-induced virus reactivation and phenotypic mutation.

UVr-1 is a human cell clone established as a variant with increased resistance to cell killing by ultraviolet light (UV, principally 254 nm wavelength) from a UV-sensitive cell clone, RSa. Both cells have been characterized to have much the same capacity of UV-induced DNA repair synthesis in whole cells, and the parent RSa cells were recently found to be hypermutable. In the present study UVr-1 cells were characterized in comparison RSa cells with respect to UV-induced virus reactivation and phenotypic mutation. Survival levels of UV-irradiated vaccinia virus and herpes simplex virus type 1 (HSV-1) were much the same in logarithmically proliferating UVr-1 and RSa cells. Correlated with these host cell reactivation levels, the same extent of UV-induced DNA repair replication synthesis was observed in isolated nuclei of the two cell clones. Enhancement of survival levels of UV-irradiated HSV-1 was detected when proliferating RSa cells were irradiated with UV prior to the virus infection. In contrast, this enhanced virus reactivation (EVR) was not detected in similarly irradiated and infected UVr-1 cells. As for phenotypic mutation frequencies assessed by the cloning efficiency of cells with increased resistance to ouabain cell killing (OuaR), OuaR mutants were not obtained from UVr-1 cells either with or without UV irradiation. When the proliferation of cells was synchronized, both EVR and OuaR mutations were detected in RSa cells irradiated with UV at any cell cycle phase, being greatest in the later half of the G1 phase. However, there was no detectable EVR or mutation in any phase of synchronous UVr-1 cells. The hypomutability of UVr-1 cells and hypermutability of RSa cells in a G1 cell cycle phase was also found even if 4-nitroquinoline 1-oxide was used as a mutagen or mutant cells with increased resistance to 6-thioguanine cell killing were estimated.

The application of immunotargeting into cancer chemotherapy with carboplatin: in vitro and in vivo studies.

Anti-tumor effects of the following 2 cis-diammin (1, 1-cyclobutandicarboxylato) platin II (carboplatin, Bristol-Myers-Squibb) conjugates were evaluated through both in vitro and in vivo experiments: (1) carboplatin coupled with anti-cytokeratin monoclonal antibody (MAb), TS1 via carboxymethyl dextran (carboplatin-carboxymethyl dextran-TS1), and (2) carboplatin-carboxymethyl dextran-avidin targeted to biotinylated TS1. Using 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinimide, carboplatin was conjugated to carboxymethyl dextran, TS1, or avidin, at high molar ratios. The staining positivity of carboplatin-carboxymethyl dextran-TS1 in indirect immunofluorescence was almost identical to that of the original MAb. The average dose of carboplatin given in each treatment was about 60% of a clinical dose. Regarding cytotoxicity, the free drug showed the strongest effect and the best dose-dependency in cell lines: HeLa and ZR-75-1. An in vivo study giving carboplatin-MAb conjugates or free drug to HeLa tumor bearing nude rats proved that the efficacy of carboplatin-carboxymethyl dextran-TS1 in HeLa tumor was not greater than that of the free carboplatin.

[A study of first and second line chemotherapies in gestational choriocarcinoma].

Twenty-eight patients with choriocarcinoma have received the three kinds of combination chemotherapy since 1983 at our department, i.e., MOA consisting of moderate dose methotrexate (MTX), actinomycin-D (Act-D) and vincristine, MEA (moderate dose MTX, Act-D and etoposide) and FA (high dose 5-Fluorouracil and Act-D). The clinical and laboratory data obtained in the 28 patients were summarized as follows; 1. The MOA regimen was administered to 4 patients primarily and to 2 secondarily. All of the 6 patients attained remission, but finally two (33.3%) developed relapse. 2. The MEA regimen was administered to 12 patients primarily and to 12 secondarily. Of the 24 patients, five (20.8%) were found to be resistant to the MEA regimen. Nineteen patients (79.2%) attained remission, but 2 (10.5%) developed recurrence. 3. The FA regimen was attempted in one patient primarily and in 6 secondarily. Although one patient died, the remaining 6 achieved remission and one relapse has been observed in the 6 cases. 4. By applying the above mentioned 3 combination chemotherapy regimens, the overall survival rate was pushed up from 64% to 90% in choriocarcinoma patients. 5. Three patients finally died of the disease but not from the side effects of the combination chemotherapies. The major adverse effects were alopecia, nausea, vomiting and myelosuppression. In particular, serious myelosuppression was caused by the MEA or FA regimen in 5-7% of all chemotherapy courses.

[Vertical transmission human T-cell lymphotropic virus type-I (HTLV-I)--genetic diagnosis and assessment of the probable routes of HTLV-I infection].

Human T-cell lymphotropic virus type-I (HTLV-I) provirus DNA from peripheral lymphocyte of 39 infants delivered by 26 pregnant carriers was detected by the nested double polymerase chain reaction (PCR) method to identify vertical transmission (VT) of HTLV-I. The 39 infants included 12 breast-fed and 27 bottle-fed infants. Particle agglutination (PA) assay and indirect immunofluorescence (IF) test with 467 cells were performed to detect anti-HTLV-I antibody. In breast-fed infants, two (16.7%) cases were both seropositive and PCR-positive and others were both negative, so there was perfect agreement between seropositivity and PCR-positivity. In bottle-fed infants, two (7.4%) cases were seropositive but PCR-negative. This seropositivity was supposed to be due to the transplacental maternal anti-HTLV-I antibody. In 25 seronegative bottle-fed infants, 4 (14.8%) cases were PCR-positive. No significant difference was found in the PCR-positivity rate between the breast-fed and bottle-fed groups. Our study showed the usefulness of the PCR method in identifying VT, the existence of silent carriers especially in bottle-fed infants and the possibility of transplacental or birth canal routes of HTLV-I infection.

The development of an enzyme immunoassay for placental tissue protein 17 (PP17) and its clinical significance.

We developed an enzyme immunoassay (EIA) for placental protein 17 (PP17) using avidin biotin binding, and measured the serum-PP17 levels of 37 healthy men, 103 nonpregnant women, 48 pregnant women, and 86 patients with gynecologic malignancies. The mean level was 12.8 ng/ml in healthy men and 44.2 ng/ml in nonpregnant women (p < 0.05). The calculated upper limit of normal was 97.8 ng/ml (mean + 2 sigma). The serum PP17 concentration was remarkedly reduced postmenopausally. Pregnant women showed a mean serum level of 19.2 ng/ml, which was significantly lower than that of nonpregnant women. Immunoserological results strongly suggest that PP17 is produced far more in the normal endometrium than in the placentae and decidua. Patients with gynecologic malignancies had obviously lower mean serum PP17 levels (8.3-19.9 ng/ml) than those found in healthy nonpregnant women. Measurement of the serum PP17 concentration might be useful in distinguishing gynecologic malignancies from various normal conditions.

[The effect of etoposide on ovarian function in patients with gestational trophoblastic disease].

Thirty-four patients with low risk gestational trophoblastic disease were treated with etoposide alone. Within 60 days after treatment, synthetic luteinizing hormone releasing hormone (LH-RH: 100 micrograms) was injected intravenously. The serum concentrations of LH, follicle-stimulating hormone (FSH) before and 30, 60, 120 minutes after LH-RH administration, estradiol (E2) and progesterone were measured. In addition, the basal body temperature (BBT) was taken continuously to evaluate ovulation. (1) The patients aged 40 or more in whom serum LH and FSH levels were higher before LH-RH administration, showed excessive response to LH-RH. These excessive responses were 45.5% and 7.1% in patients in their thirties and twenties, respectively. (2) Serum E2 and progesterone levels decreased with advanced age, which was not affected by total doses of etoposide. (3) Seven (77.8%) patients over 40 years were found to have anovulatory cycles, based on their BBT, and ovulation was confirmed in all the patients under 40 years. (4) Eleven pregnancies were confirmed. Nine delivered apparently healthy infants. The influence of etoposide on ovarian function was transient except for patients over 40 years old.

[Studies of clinical usefulness of new tumor markers of ovarian cancer, CA 54/61 and CA 602--CA 602 assay reagent kit, performance its normal value and correlations with other tumor markers].

We made a preclinical study of a newly developed tumor maker, CA 602, and its clinical study using serum samples available from 58 institutions located throughout Japan. In the preclinical study, a CA 602 assay kit was investigated for the reproducibility and precision of assay results; and in the clinical study, the kit was investigated for the normal value of the marker, for variations in assay result with age, menstrual cycle and term of pregnancy, and for correlations of assay results with those of other tumor markers. The tests of the kit for simultaneous reproducibility and interval reproducibility of assay results, and the results of analytical recovery and dilution tests were all favorable; the kit proved to be reliable in both precision and reproducibility. For the study, 2 cutoff levels were set: mean + 2 SD of healthy subjects, i.e., 63 U/ml, and the level which permits the maximal efficiency of differential diagnosis of benign from malignant ovarian tumors, i.e., 90 U/ml. The assay results showed that CA 602 levels were low in women aged 50 and over; the levels were high in the first half of pregnancy, and also high in the menstrual period to the early follicular phase. The assay results of CA 602 also proved to be intimately correlated with those of CA 125, which suggested that the 2 markers might be analogous to each other. CA 602 proving to be of high reproducibility even in the range of concentrations below the cutoff value, the measurement with the marker appeared to be of high precision, capable of detecting even the slightest variations in the antigen. CA 602 therefore appears of great value in the early detection of recurrent ovarian cancers.

Immunohistochemical and ultrastructural investigation of new membrane-associated placental tissue proteins (MP2 A, B, C, D, and E) in gynecologic neoplasms.

New membrane-associated placental tissue proteins (MP2 A, B, C, D, and E) were investigated immunohistochemically by avidin-biotin immunoperoxidase technique and immunoelectron microscopy in various gynecologic neoplasms and normal gynecologic tissues. MP2 A and MP2 B were not specific for malignant tumors. MP2 C was present in 67-100% of ovarian carcinomas, 100% of benign dermoid cysts, and 77% of endometrial carcinomas. Except for endocervical adenocarcinomas, MP2 D was hardly detectable in gynecologic malignancies. Although MP2 E was hardly detectable in benign gynecologic tumors, this protein was present in ovarian carcinomas, uterine squamous carcinomas, endocervical adenocarcinomas, and endometrial adenocarcinomas. These results suggest a possible clinical application of these MP2 proteins as a new tumor marker for gynecologic malignancies.

The immunohistochemical localization of new membrane-associated placental tissue proteins (MP2 A, B, C, D, and E) in human and cynomolgus monkey placentae.

New membrane-associated placental tissue proteins (MP2 A, B, C, D, and E) were investigated by avidin-biotin immunoperoxidase technique in the human and cynomolgus monkey placentae, decidua and umbilical cords. In human early placentae, MP2 A, B, C, and E were localized mainly in the membrane of villous syncytiotrophoblasts and cytotrophoblasts. Histiocytes in the villous stroma were positive for MP2 A, B, D, and E. In human term placentae, obvious positive staining for MP2 A, B, C, and E was observed in the membrane of villous syncytiotrophoblasts, in the amniotic epithelium, and in the umbilical cord sheath. Histiocytes in the villous stroma were positive for MP2 A, B, C, E, and especially for MP2 D. Importantly, MP2 A, C, and E were positive in polymorphonuclear neutrophils, since most of these common antigens are also carcinoma-associated, suggesting clinical usage of MP2 proteins as a new tumor marker. In the cynomolgus monkey placentae, similar immuno-staining results were obtained. The monkey can thus serve as a experimental model for the investigation of the placental proteins.

[A study of preclinical and clinical usefulness of new tumor markers of ovarian carcinoma, CA 54/61 and CA 602. 1. Evaluation of the diagnostic accuracy of CA 54/61, a study of the normal range of its values and its correlation with other tumor markers].

We made a preclinical study of CA 54/61, a recently developed marker of ovarian tumors, and also conducted a clinical study of it using serum samples collected from 58 institutions located throughout Japan. This paper describes the results of the preclinical study of the CA 54/61 marker that were obtained with a kit based on an enzyme immunoassay (EIA), and also the findings with the kit in the clinical study pertaining to the normal range of its values, its values relative to age, menstrual cycle, and pregnancy, and its correlations with other tumor markers. The tests for reproducibility of assay results, the analytical recovery test, and the dilution test all gave favorable results: the marker proved reliable in both precision and reproducibility. Two cut-off values were used: the mean + 2 S.D. of the mean for healthy subjects, or 20 U/ml; and the value for the maximum diagnostic efficacy, i.e., 12 U/ml. The assay results did not vary greatly with either age, menstrual cycle or pregnancy stage, which suggested that CA 54/61 might be a marker less liable to be affected by physiological conditions prevailing at the time of sample collection. The result correlated poorly with those of assays with other markers; thus CA 54/61 proved to differ in property from the previously recognized tumor markers.

[Phase II study of 254-S (cis-diammine glycolato platinum) for gynecological cancer].

A phase II study of 254-S was conducted in 134 patients with gynecological malignancies by the gynecology section of the 254-S cooperative study group. The drug was administered at least twice at a dose of 100 mg/m2 by intravenous infusion at 4 week intervals. Forty-two of the 102 evaluable patients responded, including 8 CRs and 34 PRs, with a response rate of 41.2%. The response rate was 37.7% for ovarian cancer and 46.3% for cervical cancer. The response rate of 46.3% for cervical cancer was the highest reported for any single anticancer agent available in Japan. The major side effect was hematotoxicity, in particular thrombocytopenia and leukopenia, while nephrotoxicity was rarely observed. These results suggest that 254-S is an active cisplatin analogue with reduced nephrotoxicity and is a very promising anticancer agent for the treatment of ovarian and cervical cancer.

CA54/61 as a marker for epithelial ovarian cancer.

Using a new one-step, double-determinant enzyme immunoassay, we performed quantitative measurements of a mucin-type glycoprotein antigen (CA54/61) that we recently detected in sera of ovarian carcinoma patients. When the cutoff value was set at 12 units/ml, at which a high diagnostic efficiency was demonstrated [or at 20 units/ml (mean + 3 SD of healthy females)], the positive rates of ovarian serous, mucinous, clear cell, and endometrioid carcinomas were 76% (or 63%), 63% (or 55%), 57% (or 52%), and 50% (or 38%), respectively. Even in mucinous cystadenocarcinoma, more than one-half of the cases were positive, indicating the potential utility of the assay in the diagnosis of mucinous tumors. In sera from patients with benign ovarian tumors, only 9% (or 4%) of the cases were positive, indicating the quite high specificity of this test for ovarian carcinomas. To make a comparison between CA54/61 and CA125, we set the cutoff level of CA125 at 110 units/ml, at which value a high diagnostic efficiency was demonstrated [or at 35 units/ml (mean + 3 SD of healthy females)]. When both CA54/61 and CA125 were assessed in sera from 36 patients with mucinous cystadenocarcinoma, the positive rates of CA54/61 and CA125 were 64% (or 56%) and 36% (or 56%), respectively, suggesting that CA54/61 is of clinical value as a new tumor marker for ovarian cancers, including mucinous tumors.

Expression of a human multidrug resistance gene in human ovarian carcinoma cell lines.

To investigate the possible role of the multidrug resistance phenotype to chemoresistance in human ovarian carcinoma, we have analyzed human multidrug resistance gene (mdr 1) expression in 8 human ovarian adenocarcinoma cell lines. An increase in P-glycoprotein level specific to multidrug-resistant tumor cells was not apparently associated with the increase in resistance to vincristine (VCR) or doxorubicin (Adriamycin). Mdr 1 transcripts (4.5 kilobases) were observed in the RNA preparation obtained from only one cell line (SHIN-3) that showed the highest resistance to both drugs in vitro and in vivo. No cell lines showed mdr 1 DNA amplification. These results suggest that the insensitivity of human ovarian carcinoma to chemotherapy could be partly explained by the expression of mdr 1.

[Changes in various differentiation-related markers after differentiation induction of a pluripotent human embryonal carcinoma (EC) cell line by hexamethylene bisacetamide].

A pluripotent human EC cell line (NEC14) could be induced to morphologically differentiate by treatment with 10(-2) M HMBA for 3 days in vitro. The changes in various differentiation-related markers (cell surface antigens, lectin binding sites, intermediate filaments, secreted products and extracellular matrix proteins) after induction of differentiation were examined in order to clarify the differentiation lineage. The results were as follows: 1) The most conspicuous changes in cell surface antigens after differentiation were the expression of major human histocompatibility antigens (HLA-A,B,C) and the changes in stage specific embryonic antigens (SSEA-1-/SSEA-3(+)----SSEA-1+/SSEA-3-). 2) Vimentin, mesenchymal intermediate filament, was only detected after the differentiation. 3) Tenascin, an extracellular matrix protein produced in mesenchymal cells, was produced after the differentiation. These results indicate that HMBA can induce NEC14 cells to differentiate into mesenchymal elements of embryonal mesoderm.