Phase I study of irinotecan by 24-h intravenous infusion in combination with 5-fluorouracil in metastatic colorectal cancer.

BACKGROUND: This study was intended to ascertain the feasibility of a combination therapy with irinotecan by 24-h intravenous infusion (24-h CPT-11) and 5-fluorouracil (5-FU) for patients with metastatic colorectal cancer, to estimate the dose-limiting toxicity (DLT) and the maximum tolerated dose (MTD), to determine the recommended dose (RD) for the Phase II study, and to evaluate the efficacy of the combination therapy. METHODS: The dosage regimen was as follows: CPT-11 was given by 24-h CPT-11 on day 1, followed by 24-h intravenous infusion of 5-FU on day 2. This regimen was repeated every 2 weeks. The dose of CPT-11 was escalated in five steps from 50 to 75, 100, 125, or 150 mg/m(2) (levels 1-5), whereas the dose of 5-FU was fixed at 800 mg/m(2). RESULTS: Twenty-six patients were recruited for this study, and 25 of the 26 patients were eligible for the assessment. The DLTs of 24-h CPT-11/5-FU therapy included grade 3 diarrhea in 1 patient treated at level 1, and grade 3 neutropenia in 1 patient and grade 4 neutropenia in 1 patient at level 4. In level 5, in 3 cases the next administration could not be done for 22 days or more as a consequence of anorexia. Thus, the level 5 was made a MTD and the level 4 was made a RD. The main side effects of grade 3 or higher, although nausea/vomiting occurred, were mild and tolerable in severity overall. The overall response rate was 24.0% (6PR/25). CONCLUSION: This study suggests that 24-h CPT-11/5-FU therapy is feasible and effective for treatment of metastatic colorectal cancer.

Activation mechanism of c-Jun amino-terminal kinase in the course of neural differentiation of P19 embryonic carcinoma cells.

P19 embryonic carcinoma cells, a model system for studying early development and differentiation, can differentiate into neurons and primitive endoderm-like cells depending on the culture conditions. We have previously reported that the activation of c-Jun amino-terminal kinase (JNK) is required for the retinoic acid-induced neural differentiation of P19 cells. However, the signaling pathway(s) responsible for the activation of JNK has not been known. In this study, we demonstrated that activities of MAPK kinase 4 (MKK4) and TAK1, one of the upstream kinases of MKK4, were enhanced in the neurally differentiating cells. Inhibition of the neural differentiation by an overexpression of protein phosphatase 2Cepsilon, an inactivator of TAK1, suggested a critical role of the TAK1 signaling pathway during the differentiation. Confocal microscopic analysis indicated that TAK1, phospho-MKK4, and phospho-JNK were colocalized with tubulin in the neurites and localized also in the nuclei of the differentiating cells. In contrast, two TAK1-binding proteins, TAB1 and TAB2, which are involved in the activation of TAK1, were localized in the neurites and the nuclei of the differentiating cells, respectively. These results suggest that two distinct TAK1-MKK4-JNK signaling pathways are independently activated at the different intracellular locations and may participate in the regulation of the neural differentiation of P19 cells.

PTEN gene mutation and high MIB-1 labeling index may contribute to dissemination in patients with glioblastoma.

Dissemination of glioblastomas is often observed, but the underlying mechanism is not well clarified especially from the genetic viewpoints. The present study examined whether PTEN gene mutations and MIB-1 labeling index (LI) correlate with the dissemination in 39 consecutive patients with glioblastomas. Dissemination was defined as leptomenigeal enhancement by magnetic resonance imaging performed in all patients every 2-3 months. We examined PTEN mutations in 26 patients using cDNA-based direct sequencing and MIB-1 LI in 38 patients. Median survival time of the 39 patients was 16.2 months. Dissemination was found in 17 of 39 patients (43.6%). PTEN mutation was significantly associated with dissemination (P=0.0140), and higher MIB-1 LI (> or =35%) resulted in earlier dissemination (P=0.0156). Kaplan-Meier survival plots showed a significantly poorer survival for patients with PTEN mutation (P=0.0012). The results indicate that the evaluation of PTEN mutation and MIB-1 LI are useful to predict dissemination and prognosis of glioblastomas.

Isolation of temperature-sensitive p53 mutations from a comprehensive missense mutation library.

Temperature-sensitive (ts) mutations have been used as a genetic and molecular tool to study the functions of many gene products. Each ts mutant protein may contain a temperature-dependent intramolecular mechanism such as ts conformational change. To identify key ts structural elements controlling the protein function, we screened ts p53 mutants from a comprehensive mutation library consisting of 2,314 p53 missense mutations for their sequence-specific transactivity through p53-binding sequences in Saccharomyces cerevisiae. We isolated 142 ts p53 mutants, including 131 unreported ts mutants. These mutants clustered in beta-strands in the DNA-binding domain, particularly in one of the two beta-sheets of the protein, and 15 residues (Thr155, Arg158, Met160, Ala161, Val172, His214, Ser215, Pro223, Thr231, Thr253, Ile254, Thr256, Ser269, Glu271, and Glu285) were ts hot spots. Among the 142 mutants, 54 were examined further in human osteosarcoma Saos-2 cells, and it was confirmed that 89% of the mutants were also ts in mammalian cells. The ts mutants represented distinct ts transactivities for the p53 binding sequences and a distinct epitope expression pattern for conformation-specific anti-p53 antibodies. These results indicated that the intramolecular beta-sheet in the core DNA-binding domain of p53 was a key structural element controlling the protein function and provided a clue for finding a molecular mechanism that enables the rescue of the mutant p53 function.

Biweekly administration regimen of docetaxel combined with CPT-11 in patients with inoperable or recurrent gastric cancer.

BACKGROUND: Both docetaxel (TXT) and irinotecan (CPT-11) are active chemotherapeutic agents for gastric cancer. We designed a biweekly administration regimen of TXT combined with CPT-11 for 4 weeks as one cycle in patients with inoperable or recurrent gastric cancer, and conducted a dose-escalation study. METHODS: Patients with histologically confirmed gastric cancer were treated with the regimen. The dosage levels of TXT and CPT-11 were as follows: level 1, 30 mg/m(2) and 50 mg/m(2); level 2, 35 and 50 mg/m(2); level 3, 40 and 50 mg/m(2); level 4, 40 and 60 mg/m(2); and level 5, 50 and 60 mg/m(2). The dose escalation was based on the dose-limiting toxicity (DLT) observed during the first cycle. RESULTS: Grade 4 neutropenia was observed at level 3, but no other DLT was observed at less than level 4 during the first cycle. However, three patients at level 3 could not continue treatment without a decrease in the dosage after the second cycle. Based on these results, level 2 was considered to be the clinically recommended dosages. CONCLUSION: Biweekly TXT and CPT-11 was well tolerated. The recommended dosages of TXT and CPT-11 for a phase II trial are 35 mg/m(2) and 50 mg/m(2), respectively.

Understanding the function-structure and function-mutation relationships of p53 tumor suppressor protein by high-resolution missense mutation analysis.

Inactivation of the tumor suppressor p53 by missense mutations is the most frequent genetic alteration in human cancers. The common missense mutations in the TP53 gene disrupt the ability of p53 to bind to DNA and consequently to transactivate downstream genes. However, it is still not fully understood how a large number of the remaining mutations affect p53 structure and function. Here, we used a comprehensive site-directed mutagenesis technique and a yeast-based functional assay to construct, express, and evaluate 2,314 p53 mutants representing all possible amino acid substitutions caused by a point mutation throughout the protein (5.9 substitutions per residue), and correlated p53 function with structure- and tumor-derived mutations. This high-resolution mutation analysis allows evaluation of previous predictions and hypotheses through interrelation of function, structure and mutation.

Evaluation of 18F-FDG PET in patients with advanced, metastatic, or recurrent gastric cancer.

UNLABELLED: PET with 18F-FDG has been widely used in oncology, but its application for stomach neoplasms has been limited. The aim of this study was to evaluate the visual diagnostic accuracy of (18)F-FDG PET for advanced, metastatic, or recurrent gastric cancer and to generate semiquantitative values for lesions. METHODS: 18F-FDG PET scans were obtained on 42 patients (29 men, 13 women; age, 27-78 y; median age, 63 y): 20 patients with a PT931/04 scanner and 22 patients with a SET2400W scanner. The PT931/04 has a spatial resolution of 6.0 mm at full width at half maximum (FWHM) and covers 15 cm above and below the targeted lesion, and the SET2400W has a spatial resolution of 3.9 mm at FWHM and images the entire body. All PET images were interpreted visually, and tracer uptakes were quantitated as standardized uptake values (SUVs) on SET2400W images. RESULTS: The sensitivity, specificity, and accuracy as a whole were as follows: 71%, 74%, and 73%, respectively, with the SET2400W scanner and 47%, 79%, and 62%, respectively, with the PT931/04 scanner. Values were high for primary lesions, liver, lymph node, and lung metastases, but were low for bone metastases, ascites, peritonitis, and pleuritis carcinomatoses. SUVs were 8.9 +/- 4.2 (primary lesions, 19 patients/19 lesions), 6.5 +/- 2.2 (liver, 9/55), 6.1 +/- 2.5 (lymph nodes, 14/38), 6.5 +/- 1.8 (abdominal wall, 4/7), 3.9 +/- 2.0 (bone, 3/27), and 4.7 +/- 2.6 (lung, 2/3). Comparing SUVs and histologic findings for 17 untreated patients, values for well-differentiated and moderately differentiated adenocarcinomas versus poorly differentiated adenocarcinomas and signet ring cell carcinomas were 13.2 +/- 6.3 (4/4) versus 7.7 +/- 2.6 (13/13) (P < 0.05) for the primary lesions, 7.0 +/- 2.4 (5/39) versus 5.6 +/- 2.8 (2/2) for the liver, and 5.5 +/- 1.9 (9/28) versus 8.8 +/- 3.3 (3/8) (P < 0.05) for the lymph nodes. CONCLUSION: Our results indicate that 18F-FDG PET is a useful diagnostic modality for advanced, metastatic, or recurrent gastric cancer but not for detecting bone metastases, peritonitis, or pleuritis carcinomatoses. 18F-FDG uptake by gastric cancers is relatively high but does not parallel histopathologic features of malignancy.

Evaluation of the diagnostic accuracy of the stop codon (SC) assay for identifying protein-truncating mutations in the BRCA1and BRCA2genes in familial breast cancer.

Screening for protein-truncating mutations of the BRCA1 and BRCA2 genes is useful in genetic testing for familial breast cancer because, first, the methods are usually simple and not expensive, and second, the detected mutations indicate pathogenic mutations in general. We evaluated the diagnostic accuracy of the stop codon (SC) assay for detecting protein-truncating mutations in the BRCA1 and BRCA2 genes by comparing the results with DNA sequencing in samples from 29 patients with breast cancer from 24 Japanese families with a history of breast cancer. Protein-truncating mutations were detected in 5 of the 24 families (20.8%; two in the BRCA1 gene and three in the BRCA2 gene). Among the 176 DNA fragments examined using the SC assay, the existence of three protein-truncating mutations (one in the BRCA1 gene and two in the BRCA2gene) was predicted correctly by the assay. Only one reverse transcriptase-polymerase chain reaction fragment was positive for the SC assay but was negative using DNA sequencing. Our study showed clearly that the SC assay is sensitive (3 of 3, 100%) and specific (172 of 173, 99%) for detecting pathogenic protein-truncating mutations in the BRCA1 and BRCA2 genes, and that it could be useful for screening larger populations.

Analysis of the human APC mutation spectrum in a saccharomyces cerevisiae strain with a mismatch repair defect.

Somatic APC mutations in colorectal tumors with an RER phenotype reflect excessive frameshift mutations, especially in simple repetition tracts within the coding sequence. Because this type of mutation is characteristic of cells with a deficient DNA MMR system, the APC mutation signature of RER tumors may be attributable to a defect in the MMR system. However, there is little experimental evidence to prove that the spectrum of mutations and the APC gene distribution are directly influenced by MMR system defects. We therefore examined the mutation spectrum of the MCR of the APC gene after transfection into both MMR-proficient and MMR-deficient yeast strains and compared it with a previously reported human APC mutation database. Small insertions or deletions in mono- or dinucleotide repeats were more common in the MMR-deficient than in the MMR-proficient strain (91.2% vs. 38.1%, Fisher's exact test p < 0.0001). Furthermore, the 2 mutation hot spots, 4385-4394(AG)(5) and 4661-4666(A)(6), found in the yeast system corresponded with those in human tumors. Combining our data with those from human tumors, there appears to be hypermutable mutations in specific simple repetitive sequences within the MCR, which are more prevalent in MMR-deficient cells and RER tumors than in MMR-proficient cells and non-RER tumors. We therefore consider that the differences in the spectra of RER and non-RER tumors are attributable at least in part to the MMR system of the host cells.

Alpha-FP normalization as a prognostic factor for mediastinal origin embryonal carcinoma: report of five cases.

It is well known that a subgroup of germ cell tumors, embryonal carcinoma of extra-gonadal origin have a poor prognosis. We have encountered five cases of mediastinal embryonal carcinoma treated with high-dose chemotherapy (HD-CT) supported by auto-PBSCT in four, and resection in three. Our cases indicated that normalization of the alpha-FP tumor marker level during standard chemotherapy is a very important factor for cure, and the resection of the residual mass after chemotherapy is indicated due to the great risk of remnant malignant cells despite HD-CT.

Phase II study of paclitaxel with 3-h infusion in patients with advanced gastric cancer.

BACKGROUND: To increase the options for agents for gastric cancer chemotherapy, we performed a phase II clinical trial on the use of a 3-h infusion of paclitaxel to confirm its efficacy and the feasibility of its use in patients with advanced gastric cancer. METHODS: Thirty-two (32) patients with measurable metastatic gastric cancer were enrolled in this study. Seventeen patients (53%) had received prior chemotherapy for metastatic disease, 4 patients (13%) had adjuvant chemotherapy alone, and 11 patients (34%) were chemotherapy-naive. Paclitaxel was intravenously infused for 3 h, at a dose of 210 mg/m(2), once every 3 weeks. To prevent hypersensitivity reactions, standard premedication was administered to all patients. RESULTS: Nine (28%; 9/32 ) objective partial responses (PRs) were observed (95% confidence interval [CI], 14%-47%), and the remaining 23 patients showed stable (12 patients; 37.5%) and progressive disease (11 patients; 34.4%). The median time to response was 20 days (range, 14-38 days). The median response duration was 87 days (range, 50-103 days). The median survival of all patients was 234 days (range, 13-646+ days). The major adverse reactions were myelosuppression (grade 3/4 leukopenia and neutropenia were observed in 59% and 88% of the patients, respectively), alopecia, and peripheral neuropathy. Peripheral neuropathy was observed in 19 patients, however, most of the patients recovered after the completion of treatment. CONCLUSION: A 3-h infusion of paclitaxel is an effective therapy for advanced gastric cancer and is clinically well tolerated by the patients.

Phase I studies of nogitecan hydrochloride for Japanese.

BACKGROUND: SmithKline Beecham synthesized camptothecin analogs and identified nogitecan hydrochloride (topotecan) with a broad spectrum of antitumor activity and less toxicity than camptothecin. Because preclinical and overseas clinical data indicated the antitumor effect of nogitecan hydrochloride with a 5-day repeat-dose schedule, we carried out phase I studies in Japan to determine the maximum tolerated dose (MTD), pharmacokinetics, and antitumor effect of nogitecan hydrochloride. METHODS: Phase I studies of nogitecan hydrochloride given by single and 5-day repeat dosing were carried out in patients with various solid tumors at 15 medical institutions in Japan. Pharmacokinetic evaluations were performed for both single and 5-day repeated dosing. RESULTS: The dose-limiting factor (DLF) was reversible leucopenia, and the maximum tolerated dose (MTD) was higher than 22.5 mg/m2 in the single-dose study. In the 5-day repeat-dose study, the DLF was also reversible leucopenia, and the MTD was estimated to be 1.5 mg/m2 per day. The plasma concentration of nogitecan hydrochloride increased with increasing dose, and the half-life after single dosing ranged from 3 to 5h. There was no evidence of accumulation or delayed excretion during 5-day repeat dosing. CONCLUSION: Based on these results and the finding that there were responders among patients treated at 1.5 mg/m2 per day by 5-day repeat dosing in overseas studies, 5-day repeat dosing of 1.2mg/m2 per day, one dose level lower than the MTD, was selected for phase II studies in Japan.