Masaji Yamauchi

Expression of carcinoembryonic antigen (CEA) and nonspecific crossreacting antigen (NCA) in gastrointestinal cancer; the correlation with degree of differentiation

In spite of its reputation as a tumour marker, little is known about the function of carcinoembryonic antigen (CEA). We examined the mRNA expression of CEA and NCA in 26 gastric and 14 colorectal cancers together with adjacent morphologically normal mucosae. There was no significant difference between the CEA mRNA levels of colorectal cancer and adjacent mucosa, whereas the CEA mRNA levels were significantly elevated in gastric cancer compared with normal gastric mucosa. The expression of NCA, on the other hand, was more cancer-specific and was significantly up-regulated in both gastric and colorectal cancers compared with the corresponding normal mucosae. No correlation was found between the mRNA level and plasma CEA value. No significant up-regulation was recognised in the node positive cancer, cancer with distant metastasis, or the metastatic tissues themselves. Marked diversity in the degree of differentiation in gastric cancer tissues, however, resulted in varied expression of the CEA gene family, compared with the constantly high expression found in colorectal cancer. Further analysis revealed significant up-regulation of NCA in well and moderately differentiated gastric cancers over poorly differentiated cancers, suggesting that NCA might have roles in differentiation.

Expression of nm23 H‐1 RNA levels in human gastric cancer tissues. A negative correlation with nodal metastasis

Background. Gene expression of nm23 has been investigated in number of tumors, including breast cancer, colon cancer, malignant melanoma, and hepatocellular carcinoma. Its down‐regulation has been shown to be associated with metastasis or disease progression in some of the tumors.

Pharmacokinetic Characteristics of 5‐Fluorouracil and Mitomycin C in Intraperitoneal Chemotherapy

Abstract— Eight patients with malignancies confined to the peritoneal space participated in this study. Five hundred milligrams 5‐fluorouracil or 10 mg mitomycin C was diluted in 1 L saline. The mixed solution was injected intraperitoneally through the semi‐permanent peritoneal catheter. Blood and peritoneal fluid were collected after injection. 5‐Fluorouracil concentrations in the peritoneal fluid were 1000 times those in serum, while mitomycin C concentrations were 100 times those in serum. Areas under the concentration vs time curve (AUC) were calculated by the trapezoidal method with extrapolation to infinity. The ratio of peritoneal fluid AUC to serum AUC was about 1400 for 5‐fluorouracil and 80 for mitomycin C. Patterns for the absorption and elimination from systemic circulation were similar for both compounds. Drug concentrations in the peritoneal fluid and serum were analysed according to the compartment model. The half‐life in the peritoneal fluid (t½p) and the rate constant from the peritoneal fluid to the systemic circulation (ka) were nearly equal for both 5‐fluorouracil and mitomycin C (t½p, 1·0 h for 5‐fluorouracil and 1·3 h for mitomycin C; ka 0·71 h−1 for 5‐fluorouracil and 0·68 h−1 for mitomycin C), although the apparent volume of distribution (Vds/F) and clearance in the peritoneal cavity (CLp) for mitomycin C (78 Lm−2 and 1.8 L h−1 m−2) were about twice the values for 5‐fluorouracil (149 L m−2 and 0·8 L h−1 m−2).

Expression of MDR1 and glutatione S transferase-π genes and chemosensitivities in human gastrointestinal cancer

The relationship was analyzed between drug resistance and MDR1 (with MDR signifying multiple drug resistance) and glutatione S transferase‐π (GST‐π) gene expression in four stomach and four colon cancer cell lines. Northern blot analysis by pmdr1 probe showed that stomach cancer cell lines had no detectable level of MDR1 mRNA expression. By contrast, some levels of MDR1 mRNA expression were found in two colon cancer cell lines, indicating doxorubicin resistance. To examine the MDR1 mRNA in each cell level, in situ hybridization was used. It was found that all colon cell lines and two stomach cell lines had more silver grains per cell than KB cells (a human KB kidney epidermoid carcinoma cell line). However, the number of silver grains in each cell was heterogeneous in the colon and stomach cell lines. Low‐level MDR1 mRNA expression could be detected even in cell lines without MDR1 mRNA expression by northern blot hybridization. These results suggest the possibility that all gastrointestinal cell lines can acquire multiple drug resistance. In addition, all examined gastrointestinal cell lines had high GST‐π mRNA expression. This GST‐π gene expression shows cisplatin resistance in the examined cell lines. Heterogeneity of GST‐π mRNA expression also was shown at the cellular level. Cancer 1992; 69:941–946.

Experission of glutathione-S-transferases α and π in gastric cancer: a correlation with cisplatin resistance

Glutathione-S-transferase (GST) in one of several factors that are proposed to affect tumor sensitivity to anticancer drugs, including cisplatin (CDDP). Attempts are made herein to evaluate the significance of the enzymes in resistance to CDDP in clinical samples of gastric cancer. A total of 22 gastric cancer specimens, 16 of which were obtained with matching normal mucosae, underwent immunoblotting with polyclonal antibodies against GST-α and GST-π. At the same time, the chemosensitivity of 15 gastric cancer specimens to CDDP was evaluated by the succinic dehydrogenase inhibition (SDI) test. The expression of GST-π was detected in all the specimens, and its content in the neoplasms exhibited a significant positive correlation with that in the matched normal mucosae. The expression of GST-α was detected in 18 of 22 cancer specimens (82%), but its content in the neoplasms did not correlate with that in the matched mucosae. A comparison of the drug-sensitivity findings with the results of immunoblotting revealed a weak but interesting correlation between the protein levels of GST-α and CDDP resistance. The cellular content of GST-α correlated weakly with CDDP resistance in gastric cancer, and its quantification could contribute to prediction of the clinical effects of CDDP in patients with gastric cancer.

Prediction of doxorubicin resistance in gastrointestinal cancer by P-glycoprotein staining

Feasibility of immunohistochemical staining of P-glycoprotein for the prediction of doxorubicin resistance in gastrointestinal cancers was examined. Among 10 cancer cell lines which consist of two gastric cancer cell lines and eight colon cancer cell lines, seven cell lines were stained positively by the monoclonal antibody to P-glycoprotein, C219. In consequence of the evaluation on the effect of doxorubicin on these tumour cells by means of succinic dehydrogenase inhibition test (SDI test), zero out of seven cell lines stained positively by C219 was sensitive to doxorubicin, but two out of three cell lines stained negatively were sensitive. Among 23 fresh surgical specimens of gastrointestinal cancers which consisted of 15 gastric cancers and eight colon cancers, seven tumour tissues were stained positively by C219. All P-glycoprotein positive tumours were resistant to doxorubicin. On the other hand, four of 16 P-glycoprotein tumours were sensitive to doxorubicin. These data indicate that positively stained cancer cells by C219 are resistant to doxorubicin.

Establishment of Drug Resistance in Human Gastric and Colon Carcinoma Xenograft Lines

We established multidrug‐resistant human gastric and colon xenograft lines by means of intratu moral injections of four agents, doxorubicin (DXR), cisplatin (CDDP), 5‐fluorouracil (5‐FU) and mitomycin C (MMC), into subcutaneous SC1NU and SW480 tumors once a week or less. Such intermittent drug exposure is commonly used in clinical chemotherapeutic protocols. All xenograft lines acquired resistance to the injected drugs as evaluated by in vivo drug‐resistance tests. Many of the drug‐resistant lines showed various patterns of cross resistance to other drugs. In order to analyze the mechanism of resistance in vivo, we investigated the expression of drug resistance gene, which has been extensively studied in vitro. We used four complementary DNAs (cDNAs) for multidrug resistance (MDR1), glutathione S‐transferase‐ (GST‐), thymidylate synthase (TS) and dehydrofolate reductase (DHFR), as probes. We observed GST‐, DHFR and TS mRNA expression at various levels, but MDR1 mRNA expression was found only in SW480/DXR by the method of poly (A+) RNA selection. Four resistant SW480 lines had higher TS mRNA expressions. Six resistant lines had stronger GST‐ mRNA expression. Five resistant lines had higher DHFR mRNA expression. Drug resistance genes related to the treated drug were also expressed in this in vivo model; MDR1 in SW480/DXR, GST‐ in SW480/CDDP and in SC1NU/CDDP and TS in SW480/5‐FU. In contrast to in vitro resistant lines which have been reported as models of drug resistance, the expression of drug resistance genes in vivo was not always correlated to the acquisition of cross resistance. These resistant xenograft lines and the methods developed to induce drug resistance in vivo should be useful for studies on the mechanism of drug resistance in the clinical setting.

Polyoxyethylene‐modified Superoxide Dismutase Reduces Side Effects of Adriamycin and Mitomycin C

Polyoxyethylene‐modified superoxide dismutase (SOD‐POE) is a newly developed long‐acting superoxide dismutase. Adriamycin (ADR) and mitomycin C (MMC) generate superoxide, which contributes to their cytocidal effects or side effects. We examined whether SOD‐POE could prevent the side effects induced by superoxide generated by antitumor agents, and the following results were obtained. SOD‐POE did not influence the antitumor effects of ADR and MMC either in vitro or in vivo, but prevented the toxic death of BALB/c, nu/nu male mice caused by overdoses of ADR or MMC. As for its effective sites, SOD‐POE prevented a decrease in the specific activity of rotenone‐sensitive NADH‐ubiquinone oxidoreductase (complex I) in heart muscle mitochondrial respiratory chain function in BALB/c male mice administered 10 mg/kg ADR, and prevented damage to the sarcoplasmic reticulum and mitochondria of mouse heart muscle by ADR as observed by electron microscopy. Furthermore, SOD‐POE prevented bone marrow suppression induced by MMC in Donryu rats. The above results suggest that combination chemotherapy with SOD‐POE would make it possible to increase the maximum permissible doses of antitumor agents, improving the efficacy of these agents.

Expression of π-glutathione S-transferase gene (GSTP1) in gastric cancer: lack of correlation with resistance against cis-diamminedichloroplatinum(II)

Abstract Class π-glutathione S-transferase (GSTP-1) is one of several factors proposed to affect drug sensitivity to cisdiamminedichloroplatinum (II) (CDDP). It has also been investigated as a potential marker for the serodiagnosis of various types of cancers. In this study, attempts were made to quantify mRNA levels of the enzyme in healthy and cancerous gastric mucosa specimens, and to evaluate their significance in inherent drug resistance to CDDP. Thirty gastric cancer specimens were analysed by northern blotting with radiolabelled GSTP1 cDNA. Of these, the chemosensitivities of 22 specimens were evaluated by the succinic dehydrogenase inhibition (SDI) test. GSTP-1 mRNA was detected in all the specimens, with slightly increased, but non-significant expression in the neoplasms. Comparison of these drug sensitivities with results of northern blotting analysis showed no inverse correlation, as was expected from the widely investigated role of the enzyme in drug resistance.

Evaluation of usefulness of in-vitro drug sensitivity testing for adjuvant chemotherapy of stomach cancer

AbstractBackground. The selection of effective drugs is the key for tumor chemotherapy. The succinic dehydrogenase inhibition (SDI) method, the simplest chemosensitivity test, is widely used at many institutions in Japan. As adequate proof of the usefulness of this test in the clinical setting has not yet been established, a prospective randomized study was undertaken. Methods. An adjuvant chemotherapy study incorporating the SDI test was performed in patients with gastrectomized stomach cancer. Fifteen test centers were set up throughout Japan, and the tests were performed on test specimens collected from neighboring institutions. In order to insure homogeneity of the test techniques, both intra- and inter-institutionally, simulation tests in tumor-bearing nude mice were carried out simultaneously. Three hundred and thirty-three eligible patients were categorized as stratum I (curative resection) or stratum II (non-curative resection) and were randomly allocated to three chemotherapy groups group A (mitomycin C), group B (doxorubicin), and group C (cisplatin), Correlations between survival rates and the inhibition index (II; a percentage value calculated from the optical density of tumor cell suspersions caltured with the anticancer drugs), which was arbitrarily divided into three categories: ≥60%; 59%–30%; and ≤29%, were analyzed in the three groups. Results. Significant differences in survival rates, in terms of the II, were shown in the stratum I group C (cisplatin), whereas no difference in terms of II was found in groups A and B. Conclusion. The SDI sensitivity test was found to be useful for cisplatin; however, the test failed to show usefulness for mitomycin C and doxorubicin.