Phase III randomised controlled trial of neoadjuvant chemotherapy plus radical surgery vs radical surgery alone for stages IB2, IIA2, and IIB cervical cancer: a Japan Clinical Oncology Group trial (JCOG 0102).

BACKGROUND: A phase III trial was conducted to determine whether neoadjuvant chemotherapy (NACT) before radical surgery (RS) improves overall survival. METHODS: Patients with stage IB2, IIA2, or IIB squamous cell carcinoma of the uterine cervix were randomly assigned to receive either BOMP (bleomycin 7 mg days 1-5, vincristine 0.7 mg m(-2) day 5, mitomycin 7 mg m(-2) day 5, cisplatin 14 mg m(-2) days 1-5, every 3 weeks for 2 to 4 cycles) plus RS (NACT group) or RS alone (RS group). Patients with pathological high-risk factors received postoperative radiotherapy (RT). The primary end point was overall survival. RESULTS: A total of 134 patients were randomly assigned to treatment. This study was prematurely terminated at the first planned interim analysis because overall survival in the NACT group was inferior to that in the RS group. Patients who received postoperative RT were significantly lower in the NACT group (58%) than in the RS group (80%; P=0.015). The 5-year overall survival was 70.0% in the NACT group and 74.4% in the RS group (P=0.85). CONCLUSION: Neoadjuvant chemotherapy with BOMP regimen before RS did not improve overall survival, but reduced the number of patients who received postoperative RT.

Low response rate of second-line chemotherapy for recurrent or refractory clear cell carcinoma of the ovary: a retrospective Japan Clear Cell Carcinoma Study.

Clear cell carcinoma (CCC) of the ovary has been recognized to show resistance to anticancer agents in the first-line chemotherapy. Our aim was to evaluate the effect of second-line chemotherapy in a retrospective study. A total of 75 patients diagnosed with CCC and treated between 1992 and 2002 in collaborating hospitals were reviewed. Criteria for the patients' enrollment were 1) diagnosis of pure-type CCC at the initial operation, 2) treatment after one systemic postoperative chemotherapy, 3) measurable recurrent or refractory tumor, 4) at least two cycles of second-line chemotherapy and assessable for the response, and 5) adequate clinical information. Regimens of first-line chemotherapy were conventional platinum-based therapy in 33 cases, paclitaxel plus platinum in 24 cases, irinotecan plus platinum in 9 cases, and irinotecan plus mitomycin C in 7 cases. Treatment-free periods were more than 6 months in 24 cases (group A) and less than 6 months in 51 cases (group B). In group A, response was observed in two cases (8%): one with conventional platinum therapy and another with irinotecan plus platinum. In group B, three cases (6%) responded: two with platinum plus etoposide and one case with irinotecan plus platinum. Median overall survival was 16 months in group A and 7 months in group B (P = 0.04). These findings suggest recurrent or resistant CCC is extremely chemoresistant, and there is only small benefit of long treatment-free period in CCC patients. Another strategy including molecular-targeting therapy is warranted for the treatment of recurrent or refractory CCC.

Sodium chloride enhances markedly the thermal stability of thermolysin as well as its catalytic activity.

Thermolysin, a thermophilic metalloproteinase, is markedly activated in the presence of high concentrations (1-5 M) of neutral salts. The activity increases in an exponential fashion with increasing salt concentration, and is enhanced 13-15 times with 4 M NaCl at pH 7.0 and 25 degreesC (K. Inouye, Effects of salts on thermolysin: activation of hydrolysis and synthesis of N-carbobenzoxy-l-aspartyl-l-phenylalanine methyl ester, and a unique change in the absorption spectrum of thermolysin, J. Biochem. 112 (1992) 335-340). In this study, the effect of NaCl on the thermal stability of thermolysin has been examined at 60-85 degreesC. The activation energy, Ea, for the thermal inactivation is 15 kcal/mol at 0 M NaCl, and increases up to 30-33 kcal/mol by the addition of 0. 5-1.5 M NaCl. Further increase in [NaCl] decreases the Ea value, and at 4 M NaCl it is almost the same as that at 0 M NaCl. Thermolysin at 0.5-1.5 M NaCl is twice as heat-stable as in the absence of NaCl. The NaCl dependence of the stability is different from that of the activity, suggesting that the effects of NaCl on activity and stability are independent. Thermolysin has been demonstrated to be not only a thermophilic enzyme but also a highly halophilic one.

Randomised study of immunotherapy with OK-432 in uterine cervical carcinoma.

OK-432, a streptococcal preparation, was administered to patients with stage Ib and II cervical carcinoma except for adeno- and adenosquamous carcinomas. To evaluate the efficacy of OK-432 precisely, 177 patients were stratified by clinical stage, radiotherapy, and lymph node metastasis after complete radical hysterectomy and pelvic lymphadenectomy. Within each stratum, patients were divided randomly into OK-432 and control groups. 85 patients received OK-432 and 92 patients did not. No significant difference was observed in overall 5-year disease free rates between the OK-432 and the control groups, although the mean diameter of erythema on SU-polysaccharide (SU-PS) skin test was larger in the OK-432 group than in the control group. In stage IIb, a significant difference was observed between the OK-432 and control groups. This difference, however, could be attributed in part to the different incidence of the lymph node metastasis. In stage II without lymph node metastasis, 5-year disease free rate was significantly higher in the OK-432 group.

Poly(ADP-ribose) synthesis in human cervical cancer cell-diagnostic cytological usefulness.

Poly(ADP-ribose) synthetase activity in human cervical cancer cell nuclei was found to be approximately twice that of normal cervical cell nuclei. Using antipoly(ADP-ribose) antibody, frozen section of normal tissue were stained by indirect-immunofluorescence and it was demonstrated that the fluorescence of nuclei in squamous cells decreased during maturation. Correspondingly, superficial cells appearing in smear specimens had less nuclear fluorescence compared with parabasal cells in the same smear. However, cancer cells in cervical smears had marked nuclear fluorescence using immunostaining and were easily distinguished from normal cells.

Conservative therapy for adenocarcinoma and atypical endometrial hyperplasia of the endometrium in young women: central pathologic review and treatment outcome.

Thirty-nine patients with endometrioid adenocarcinoma (EA) and atypical hyperplasia (AH) of the endometrium who received conservative treatment to preserve fertility were collected from member institutions of the Japan Gynecologic Oncology Study Group. Twenty-nine and ten were originally diagnosed with EA without myometrial invasion and AH, respectively. We performed a central pathological review to make definite diagnoses, and the diagnosis of EA in 29 cases was changed to AH in ten, complex hyperplasia in three and atypical polypoid adenomyoma in three, and AH in ten was changed to EA in one and simple hyperplasia in one. Nine of 12 women (75%) with EA and 15 of 18 women (83%) with AH had an initial response to medroxyprogesterone acetate (MPA) treatment. Two of nine responders with EA later developed relapse, and one of them had metastasis to the left obturator lymph node. Two became pregnant, and one delivered one full-term infant. One of the responders with AH had a relapse in the endometrium. Five became pregnant, and four delivered four normal infants. The young women with endometrial carcinoma localized in the endometrium who wish to preserve fertility may be treated as successfully with MPA as those with AH.

Effects of cobalt-substitution of the active zinc ion in thermolysin on its activity and active-site microenvironment.

Thermolysin is remarkably activated in the presence of high concentrations (1-5 M) of neutral salts [Inouye, K. (1992) J. Biochem. 112, 335-340]. The activity is enhanced 13-15 times with 4 M NaCl at pH 7.0 and 25 degrees C. Substitution of the active site zinc with other transition metals alters the activity of thermolysin [Holmquist, B. and Vallee, B.L. (1974) J. Biol. Chem. 249, 4601-4607]. Cobalt is the most effective among the transition metals and doubles the activity toward N-[3-(2-furyl)acryloyl]-glycyl-L-leucine amide. In this study, the effect of NaCl on the activity of cobalt-substituted thermolysin was examined. Cobalt-substituted thermolysin, with 2.8-fold increased activity compared with the native enzyme, is further activated by the addition of NaCl in an exponential fashion, and the activity is enhanced 13-15 times at 4 M NaCl. The effects of cobalt-substitution and the addition of salt are independent of each other. The activity of cobalt-substituted thermolysin, expressed as k(cat)/K(m), is pH-dependent and controlled by at least two ionizing residues with pK(a) values of 6.0 and 7.8, the acidic pK(a) being slightly higher compared to 5.6 of the native enzyme. These pK(a) values remain constant in the presence of 4 M NaCl, indicating that the electrostatic environment of cobalt-substituted thermolysin is more stable than that of the native enzyme, the acidic pK(a) of which shifts remarkably from 5.6 to 6.7 at 4 M NaCl. Zincov, a competitive inhibitor, binds more tightly to the cobalt-substituted than to native thermolysin at pH 4.9-9.0, probably because of its preference for cobalt in the fivefold coordination. The cobalt substitution has been shown to be a favorable tool with which to explore the active-site microenvironment of thermolysin.

A spectrophotometric study on the interaction of thermolysin with chloride and bromide ions, and the state of tryptophyl residue 115.

The activity of thermolysin is greatly enhanced in the presence of high concentrations of neutral salts [Holmquist, B. and Vallee, B.L. (1976) Biochemistry 15, 101-107; Inouye, K. (1992) J. Biochem. 112, 335-340]. NaBr and NaCl are the most effective for the activation. An absorption difference spectrum with a peak around 293 nm, which is characteristic of the red-shift of a tryptophyl residue caused by charge effects, was observed on mixing of thermolysin with NaCl. As the peak disappeared in the presence of competitive inhibitors of the enzyme (phosphoramidon and zincov), it was considered to be derived from a tryptophyl residue (Trp 115) located in the active site of the enzyme. On the other hand, this peak was not observed on the mixing of thermolysin and NaBr, indicating that the slight difference in size between chloride and bromide ions is critical for the interaction with the tryptophyl residue. NaCl and NaBr exhibit comparable effects on the activation of thermolysin regardless of the considerable discrepancy in their effects on the absorptivity difference around 293 nm. This suggests that the interaction of salts with Trp 115 is not necessarily correlated with the activation of thermolysin.

Effect of salts on the solubility of thermolysin: a remarkable increase in the solubility as well as the activity by the addition of salts without aggregation or dispersion of thermolysin.

Thermolysin is remarkably activated in the presence of high concentrations (1-5 M) of neutral salts [Inouye, K. (1992) J. Biochem. 112, 335-340]. The activity is enhanced 13-15 times with 4 M NaCl at pH 7.0 and 25 degrees C. In this study, the effect of neutral salts on the solubility of thermolysin has been examined. Although the solubility was only 1.0-1.2 mg/ml in 40 mM Tris-HCl buffer, pH 7.5, in the temperature range between 0 and 60 degrees C, it was increased greatly by the addition of salts. With NaCl, the solubility showed a bell-shaped behavior with increasing NaCl concentration, and the maximum solubility (10 mg/ml) was at 2.0-2.5 M NaCl. With LiCl and NaI, it increased progressively to 20-50 mg/ml with increasing salt concentration up to 5 M. The solubility observed in the presence of salts decreased with increasing temperature from 0 to 60 degrees C, and also with the order of chaotropic anion effect. The molecular weight of thermolysin was estimated to be 33.0(+/-2.5) x 10(3) in the presence of 0-3 M NaCl, suggesting that thermolysin exists as a monomer in the presence or absence of 3 M NaCl. The possibility that aggregation and/or dispersion of thermolysin might be related to the remarkable activation by salt was ruled out.

Molecular diagnostic detection of free cancer cells in the peritoneal cavity of patients with gastrointestinal and gynecologic malignancies.

Free cancer cells exfoliated from cancer-invaded serosa contribute to peritoneal dissemination, the most frequent pattern of recurrence in patients with gastric and ovarian cancers. This study was designed to evaluate the prognostic significance of free cancer cells in peritoneal washes detected using the reverse transcriptase-polymerase chain reaction (RT-PCR) and cytology. RT-PCR analysis with primers specific for the carcinoembryonic antigen (CEA) gene was found to be more sensitive than cytology for detection of free tumor cells in the peritoneal washes, collected at laparotomy from 199 gastric carcinoma patients, with higher detection rates for each of the T-categories in the TNM classification. Six patients with synchronous and 5 with recurrent peritoneal dissemination were found among 25 advanced cancer patients with positive PCR and negative cytology results. Positive PCR results were significantly associated with poor survival of curatively resected advanced gastric carcinoma patients (P < 0.001). A rapid method for detecting CEA mRNA using the LightCycler and the dsDNA binding dye SYBR green I was also developed. The results obtained using this technique were essentially the same as those obtained using the conventional RT-PCR method. Furthermore, RT-PCR analysis with primers specific for MUC1 epithelial mucin were performed on peritoneal washes from patients with ovarian cancer. Peritoneal washes from 21 of 25 ovarian carcinoma patients, including all 17 with positive cytology results, were positive for MUC1 mRNA, again indicating a higher sensitivity using this method than conventional cytology. Highly sensitive and rapid detection of free cancer cells in peritoneal washes, most reliably by RT-PCR, is a powerful technique to predict peritoneal dissemination in patients with gastric and ovarian cancers.

A newly synthesized bifunctional inhibitor, CKA1083, enhances adriamycin activity against human ovarian carcinoma cells.

BACKGROUND: A newly synthesized drug, CKA1083 ((S)-N-[2-(4-benzyloxy-carbonylpiperazinyl)-1-(P-methoxybenzyl) ethyl]-N-methyl-N(5-isoquinolinesulfonamide)), has the same glutathione-S-transferase (GST)-binding site structure as W-77, a bifunctional inhibitor that enhances the cytotoxicity of Adriamycin for human ovarian carcinoma cells. We examined the effects of CKA1083 on the cytotoxicity of Adriamycin and the resistance of human ovarian carcinoma cells to this drug. MATERIALS AND METHODS: We used GST-pi transfected cells and Adriamycin-sensitive or -resistant cells of human ovarian carcinoma. GST-pi activity, the intracellular Adriamycin content, and the cytotoxicity of Adriamycin in these cell lines in the presence or absence of CKA1083 were measured and compared to the findings obtained with W-77 or verapamil. RESULTS: CKA1083 inhibited GST-pi activity in an uncompetitive manner and more strongly than W-77. It enhanced the cytotoxicity of Adriamycin for GST-pi transfected cells by about 3-times. Further, CKA1083 increased the intracellular Adriamycin content about 3-fold in two Adriamycin-resistant cell lines (NOS2AR and NOS3AR). CKA1083 (10 microM) reduced the IC50 of Adriamycin to 1/38 in NOS2AR cells and 1/21 in NOS3AR cells, and overcame Adriamycin resistance more effectively than both W-77 and verapamil. CONCLUSIONS: CKA1083 enhanced the antitumor effect of Adriamycin more than W-77 by inhibiting both GST activity and P-glycoprotein.

The significance of tumor size in clinical stage IB cervical cancer: Can a cut-off figure be determined?

The purpose of this study was to investigate the influence of tumor size on pathologic variables and the prognosis of patients diagnosed as clinical stage IB cervical cancer. Five hundred sixty six patients with clinical stage IB cervical cancer treated surgically at the Aichi Cancer Center between 1976 and 1995 were studied. The incidence of pathologic variables that increased as tumors enlarged was unchanged beyond 4.0 cm. Although univariate analysis revealed that the prognosis worsened as a tumor enlarged, there was no significant difference in prognoses between 3.1-4.0 cm and 4.1-5.0 cm tumors. While multivariate analysis showed tumor size as an independent prognostic variable, there was no difference between the odd ratios of 3.1-4.0 cm and 4.1-5.0 cm tumors. Tumor size was an independently significant risk factor for the prognosis of clinical stage IB cervical cancer. While the definition of 4.0 cm as a cut-off point was useful as far as determining an association with pathologic variables, it may be an insufficient indicator of poor prognosis. The "bulky" tumors should be defined as clinical lesions greater than 5.0 cm, though few patients would have tumors that meet that criterion.

Associations between tumor diameter and prognostic variables of epithelial ovarian cancer.

PURPOSE: Associations of tumor diameter in epithelial ovarian cancer with clinical and pathological prognostic variables were investigated. METHODS: The clinical and pathological records of 233 patients diagnosed with epithelial ovarian cancer and treated at Aichi Cancer Center were studied. RESULTS: Tumor diameters of 44 patients (18.9%) were < 5 cm, 90 (38.6%) were 5-10 cm, and 99 (42.5%) were > 10 cm. While 90.9% (40/44) of < 5 cm tumors presented with FIGO stage III-IV, 40.4% (40/99) of > 10 cm tumors were advanced. Intra-abdominal disease was also significantly associated with tumor diameter, although differences among lymph-node status were not significant. The incidence of serous and endometrioid adenocarcinoma in < 5 cm tumors were 75.0% (33/44) and 11.4% (5/44), respectively, while those of > 10 cm tumor were 32.3% (32/99) and 17.2% (17/99). Multivariate analysis revealed that tumor diameter was not an independent prognostic variable. CONCLUSION: Tumor diameter of ovarian cancer is associated closely with histological subtypes and stage of disease, especially intra-abdominal disease.

[The efficacy of mass screening for uterine cancer].

Mass screening for uterine cancer, mainly by cytological examination, has been conducted since 1962. The total number of examinees has been increasing and has remained at the same level for the last decade. The coverage rate of screening is only 16%, much lower than the target rate of screening, at present. According to the increase in the number of examinees, the mortality has fallen gradually, because the patients with cancer detected early on are increased and treated successfully with better QOL. However, there are some problems, as follows. 1) Although the coverage rate among the first-time examinee, women in their 30s and 70s are lower than the other groups, the rate of dysplasia and uterine cancer detected among them, are higher. 2) False, negative rate of cytology is relatively high. It is suggested the systemic mass screening is effective to reduce mortality from uterine cancer. It is necessary, however, to improve and certify the further efficacy of screening in reduction of mortality of uterine cancer, referring to procedures of cytology, especially for early detection of adenocarcinoma, and registration system from the point of cost-effectiveness.