Succinate Dehydrogenase Inhibition Test Research Articles

Molecular Factors Associated with Pemetrexed Sensitivity According to Histological Type in Non-small Cell Lung Cancer

This study was designed to investigate potential molecules that predict chemosensitivity to pemetrexed (Alimta®) in surgically resected non-small cell lung cancer (NSCLC). Chemosensitivity to ALM and other drugs was assessed by succinate dehydrogenase inhibition (SDI) test in 69 NSCLC samples (55 adenocarcinomas, and 14 squamous cell carcinomas). The mRNA expression levels of Alimta®-target enzymes [thymidylate synthase (TYMS); dihydrofolate reductase (DHFR) and glycinamide ribonucleotide formyltransferase (GARFT)], Alimta®-metabolizing enzymes [γ-glutamyl hydrase (GGH) and folylpolyglutamate synthase] and an Alimta® transporter [reduce folate carrier (RFC)] were measured and examined for potential correlations to chemosensitivity. The squamous cell carcinoma samples showed higher TYMS expression and lower RFC expression than did the adenocarcinoma samples. In the adenocarcinoma sample analyses, GGH expression was inversely correlated to sensitivity. The histology-dependent differences in chemosensitivity to Alimta® may be attributed to the histology-dependent differences in TYMS and RFC expression. In adenocarcinomas, GGH potentially represents a marker for chemosensitivity to Alimta®.

In Vitro Drug Sensitivity Tests to Predict Molecular Target Drug Responses in Surgically Resected Lung Cancer.

BackgroundEpidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) and anaplastic lymphoma kinase (ALK) inhibitors have dramatically changed the strategy of medical treatment of lung cancer. Patients should be screened for the presence of the EGFR mutation or echinoderm microtubule-associated protein-like 4 (EML4)-ALK fusion gene prior to chemotherapy to predict their clinical response. The succinate dehydrogenase inhibition (SDI) test and collagen gel droplet embedded culture drug sensitivity test (CD-DST) are established in vitro drug sensitivity tests, which may predict the sensitivity of patients to cytotoxic anticancer drugs. We applied in vitro drug sensitivity tests for cyclopedic prediction of clinical responses to different molecular targeting drugs.MethodsThe growth inhibitory effects of erlotinib and crizotinib were confirmed for lung cancer cell lines using SDI and CD-DST. The sensitivity of 35 cases of surgically resected lung cancer to erlotinib was examined using SDI or CD-DST, and compared with EGFR mutation status.ResultsHCC827 (Exon19: E746-A750 del) and H3122 (EML4-ALK) cells were inhibited by lower concentrations of erlotinib and crizotinib, respectively than A549, H460, and H1975 (L858R+T790M) cells were. The viability of the surgically resected lung cancer was 60.0 ± 9.8 and 86.8 ± 13.9% in EGFR-mutants vs. wild types in the SDI (p = 0.0003). The cell viability was 33.5 ± 21.2 and 79.0 ± 18.6% in EGFR mutants vs. wild-type cases (p = 0.026) in CD-DST.ConclusionsIn vitro drug sensitivity evaluated by either SDI or CD-DST correlated with EGFR gene status. Therefore, SDI and CD-DST may be useful predictors of potential clinical responses to the molecular anticancer drugs, cyclopedically.

Molecular mechanism of in vitro pemetrexed sensitivity according to histological type in non-small cell lung cancer.

e13090 Background: Pemetrexed (Alimta, ALM) is a new multitarget antifolate metabolite anticancer drug. Non-small cell lung cancer (NSCLC) is now treated with selective chemotherapy based on its classification as either non-squamous cell carcinoma or squamous cell carcinoma (SCC). This study was designed to investigate the relationship between in vitro chemosensitivity to ALM and both clinicopathological factors and the expressions of ALM-related enzymes and carrier molecule in NSCLC. Methods: Seventy NSCLC samples (56 adenocarcinoma and 14 SCC) were used. Chemosensitivity was assessed by the succinate dehydrogenase inhibition (SDI) test for three antimetabolic anticancer drugs, including ALM, 5-fluorouracil, gemcitabine. Samples showing more than the median of the succinate dehydrogenase (SD) activity, which reflects the cell viability, were defined as "Resistant" and samples showing less than the median were defined as "Sensitive." The mRNA expression levels of target enzymes of ALM (thymidylate synthase (TS), dihydrofolate reductase and glycinamide ribonucleotide formyltransferase), metabolic enzymes of ALM (γ-glutamyl hydrase (GGH) and folypolyglutamate synthase) and reduce folate carrier (RFC) were measured by reverse transcriptase polymerase chain reaction analysis to examine the influence on chemosensitivity to ALM. Results: Of 3 antimetabolic drugs, only ALM showed significant difference of chemosensitivity according to the histological type. SCC samples were more resistant to ALM than adenocarcinoma samples. In adenocarcinoma, no significant relationship between chemosensitivity to ALM and clinicopathological factors was observed. SCC samples showed higher expression of TS and lower expression of RFC than adenocarcinoma samples. In adenocarcinoma, only GGH was inversely correlated with chemosensitivity to ALM. Conclusions: The present study suggested that, in SCC, the expressions of TS and RFC were associated with the chemosensitivity to ALM. In adenocarcinoma, the expression of GGH was associated with the chemosensitivity to ALM.

Significance of FANCJ expression as a predictive marker of sensitivity to 5-fluorouracil in colorectal cancer.

10618 Background: Fanconi anemia protein, FANCJ, directly interacts with MLH1, a key protein involved in the DNA mismatch repair process. Deficient mismatch repair, or microsatellite instability, is a marker for the ineffectiveness of 5-fluorouracil (5-FU) and positive prognosis of colorectal cancer (CRC). We investigated the significance of FANCJ expression in CRC, focusing on the effects of 5-FU-based adjuvant chemotherapy. Methods: Clinicopathologic features and immunohistochemical expression of FANCJ and MLH1 were studied in 219 patients with CRC. We also analyzed 5-FU sensitivity in CRC cell lines with varying levels of FANCJ expression. Results: FANCJ expression was elevated in tumor tissues compared with normal epithelial tissue (p<0.001). High expression of FANCJ was significantly associated with 5-FU resistance measured by the succinate dehydrogenase inhibition test (p=0.02) and poor recurrence-free survival (RFS) (p=0.03). Among patients with stage II/III tumors who received 5-FU, patients with tumors exhibiting high FANCJ expression had significantly worse RFS than patients with tumors exhibiting low FANCJ expression (p=0.01). Among patients who did not receive adjuvant chemotherapy, FANCJ expression was not correlated with RFS (p=0.76). High FANCJ expression was correlated with 5-FU resistance in tumors with normal MLH1 expression (p=0.01), but not in tumors not expressing MLH1 (p=0.67). In vitro, FANCJ overexpression was correlated with 5-FU resistance in MLH1-proficient HCT116 3-6 cells, but not in MLH1-deficient HCT116 cells. Conclusions: FANCJ could be a useful biomarker to predict the response to 5-FU and prognosis of CRC, particularly in tumors with normal MLH1 expression.

Evaluation of ERCC1 Expression for Cisplatin Sensitivity in Human Hepatocellular Carcinoma

Excision repair cross-complementation group 1 (ERCC1) is one of the key enzymes in DNA repair. This study was designed to investigate the correlation between ERCC1 expression and chemosensitivity to cisplatin (CDDP) in patients with hepatocellular carcinoma (HCC). Eighty-seven HCC samples were analyzed by immunohistochemistry for ERCC1 and chemosensitivity was assessed by the succinate dehydrogenase inhibition (SDI) test for four anti-cancer agents, including CDDP. The ERCC1 expression was examined in HCC cell lines. ERCC1 siRNA was transfected to PLC/RPF/5 to investigate the correlation of ERCC1 expression and CDDP sensitivity. ERCC1 expression was observed in 33% of nuclei in immunohistochemical examination. Patients were divided into two groups as follows: ERCC1 high expression group (n=43): more than 33% of the nuclei were stained; ERCC1 low expression group (n=44): 33% or fewer of the nuclei were stained. Tumor size of low expression group was larger than that in the high expression group (p=0.02). The succinic dehydrogenase (SD) activity only for CDDP was significantly higher in the high expression group than that in the low expression group (p=0.02). An increased expression of ERCC1 was shown by immunohistochemical and Western blot analyses in PLC/RPF/5. ERCC1 expression was inhibited by ERCC1 siRNA transfection and the LC50 value (nM) of CDDP was reduced from 25.7 to 12.5 (p=0.01). Increased ERCC1 expression is associated with CDDP resistance in HCC specimens and cell lines. Therefore, immunohistochemical analysis for resected HCC tissues may be a useful predictor for the effectiveness of adjuvant chemotherapy, using CDDP.

Enhanced expression of DNA topoisomerase II genes in human medulloblastoma and its possible association with etoposide sensitivity

Medulloblastoma (MB) is the most common malignant neuroepithelial tumor of childhood. The DNA topoisomerase II (Topo II) inhibitor etoposide has been widely used for the treatment of MBs; however, it remains unknown whether MB cells are more sensitive to etoposide than other malignant neuroepithelial tumor cells. In this study, we tested the chemosensitivities of malignant neuroepithelial tumors (26 glioblastomas, 9 anaplastic astrocytomas, and 5 MBs) to etoposide and vincristine using the succinate dehydrogenase inhibition test and found that MB cells are more sensitive to etoposide and more resistant to vincristine than other tumor cells. We performed quantitative reverse-transcription polymerase chain reaction to evaluate the expression of genes related to etoposide sensitivity, and found co-overexpression of DNA topoisomerase II (Topo II) alpha and beta mRNA in MBs. In addition, the levels of Topo IIalpha and beta mRNA in these tumors correlated with etoposide sensitivity. Immunohistochemical studies using surgical samples of these tumors demonstrated that the percentages of Topo IIalpha immunopositive cells (Topo IIalpha labeling index) correlated with those of Ki-67 immunopositive cells (MIB-1 labeling index); however, neither the Topo IIalpha nor the MIB-1 labeling index correlated with the levels of Topo IIalpha mRNA or etoposide sensitivity. Based on these observations, Topo IIalpha and beta mRNA expression, but not the Topo IIalpha labeling index, might be a useful marker for sensitivity to etoposide in human malignant neuroepithelial tumors.

Increased expression of multidrug resistance-associated proteins in bladder cancer during clinical course and drug resistance to doxorubicin.

Overexpression of the P-glycoprotein/multidrug resistance 1 (MDR1) and multidrug resistance protein 1 (MRP1) gene is closely associated with the clinical outcome of various malignancies, and it is involved in responses to some anticancer chemotherapeutic agents including doxorubicin. Six human MRP subfamily members (MRP2-7) with structural similarities to MRP1 have been identified. Recently, the relationships between MRP2 and MRP3 expression levels of some cancer cells and drug sensitivity to doxorubicin have been reported, but the relationship between the clinical samples and drug sensitivity remains unclear. We determined the expressions of the MDR1, MRP1, MRP2 and MRP3 gene in bladder cancer during the clinical course and sought to learn whether the expression was correlated with drug responses to doxorubicin. Doxorubicin, used in chemotherapeutic treatment including intravesical and systemic chemotherapy, is an important anticancer agent for the treatment of bladder cancer. We used quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis for our study, and the sensitivity to doxorubicin in bladder cancer was determined using the in vitro succinate dehydrogenase inhibition test. Using 47 clinical samples of bladder cancer, we confirmed the significant correlation of MDR1, MRP1 and MRP3 mRNA levels with resistance to doxorubicin. We showed that the expression of MDR1, MRP1, MRP2 and MRP3 in recurrent tumors and residual tumors after chemotherapeutic treatment was higher than that in untreated primary tumors. In particular, the MDR1 expression in residual tumors was 5.7-fold higher than that in untreated primary tumors.

SDI法による皮膚悪性腫瘍の抗癌剤感受性の検討

SDI法による皮膚悪性腫瘍の抗癌剤感受性の検討

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

Background. 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.

Increased expression of an ATP-binding cassette superfamily transporter, multidrug resistance protein 2, in human colorectal carcinomas.

The expression of ATP-binding cassette superfamily transporter genes, such as P-glycoprotein/multidrug resistance (MDR) 1 and MDR protein (MRP) 1, is often up-regulated in various tumor types and is involved in responses to some anticancer chemotherapeutic agents. Five human MRP subfamily members (MRP2-6) with structural similarities to MRP1 have been identified. The relationships between MRP2-6 mRNA levels and drug resistance are not well understood. Data on 45 patients with colorectal cancer were analyzed. Of the ATP-binding cassette superfamily genes, we asked whether mRNA levels of MDR1, MRP1, MRP2, and MRP3 correlated with drug resistance to anticancer agents. For this analysis, we used quantitative reverse transcription-PCR, and the sensitivity to anticancer agents in surgically resected colon carcinomas was determined using the in vitro succinate dehydrogenase inhibition test. MDR1, MRP1, and MRP3 were highly expressed in normal colorectal mucosa, and the relative mRNA levels of MDR1, MRP1, and MRP3 in cancerous tissues compared with noncancerous tissues were decreased or unchanged. By contrast, MRP2 mRNA expression was low in normal colorectal mucosa and specifically increased in cancer regions compared with noncancerous regions. Of the anticancer agents prescribed for patients with colorectal cancers, including doxorubicin, mitomycin C, cisplatin, 5-fluorouracil, etoposide, and a camptothecin derivative, mRNA expression of MRP2 was significantly associated with resistance to cisplatin. MRP2 may be important for resistance to cisplatin treatment in colorectal cancer.

Establishment of a cisplatin-resistant gastric carcinoma cell line OCUM-2M/DDP.

A cis-diamminedichloroplatinum (CDDP)-resistant scirrhous gastric cancer cell line, OCUM-2M/DDP, was established by chronic exposure of cells of the parent scirrhous gastric cancer cell line, OCUM-2M, to CDDP at progressively increasing concentrations. The OCUM-2M/DDP cell line had an 11.3-fold higher level of resistance relative to its parent cell line as determined by a succinate dehydrogenase inhibition test. The biological and biochemical characteristics of the resistant and parent cell line were compared. There were differences in the modal chromosome number and DNA index, suggesting that some alterations of the DNA in the CDDP-resistant cells had occurred. Neither the parent nor resistant cell line expressed mdr-1 mRNA. After exposure to CDDP for 4 h, the intracellular platinum content of OCUM-2M cells was significantly higher than that of OCUM-2M/DDP cells (51.9 +/- 1.8 vs 16.4 plus 1.0 ng/mg protein, mean +/- SD, respectively). The GSH levels in OCUM-2M cells and OCUM-2M/DDP cells were 3.5 +/- 1.0 micrograms/mg protein and 16.8 +/- 1.2 micrograms/mg protein, respectively. These levels were also significantly different. These findings suggest that the possible mechanisms of acquired resistance to CDDP in OCUM-2M/DDP cells may be a decrease in intracellular CDDP accumulation and detoxication by GSH. This OCUM-2M/DDP cell line could be used in further investigations of the mechanism of CDDP resistance in gastric cancer.

Chemosensitivity test for repeated arterial infusion chemotherapy by reservoir for unresectable hepatocellular carcinoma.

The experimental and clinical usefulness of a chemosensitivity test (Nuclear Damage Assay) was studied. Karyologic degenerative changes were observed as an indicator of drug sensitivity, in repeated arterial infusion chemotherapy (RAIC) using a reservoir for advanced hepatocellular carcinoma (HCC). In the experimental study, this sensitivity test was performed using five liver cell lines against 15 drugs. At the same time, the succinate dehydrogenase inhibition (SDI) test was also performed. Comparison of the results between these two tests gave a high consistency rate of 81%. Clinically, the karyologic sensitivity test was carried out in 135 patients with unresectable HCC. Drug sensitivity could be evaluated in as many as 89% of the total 135 patients. Of the patients, 43 received RAIC on an outpatient basis via a subcutaneously implanted reservoir. The objective response of RAIC on tumours of the 43 patients was evaluated as complete response, partial response, in 3 (9%) and 8 (23%) in 35 patients treated with positive drugs (positive group), and as 0 (0%) and 0 (0%) of 8 patients treated with negative drugs (negative group), respectively. As regards the prognosis, 1 year and 1.5 year survival rates were 70 and 45% in the positive group, and 42 and 0% in the negative group, respectively. As objective response in the positive group tended to be better than that in the negative group, and prognosis in the positive group was significantly better than that in the negative group, this sensitivity test appears to contribute to the improvement of therapeutic results if used to select drugs suitable for RAIC for advanced HCC.

Glutathione S transferase Pi is a powerful indicator in chemotherapy of human lung squamous-cell carcinoma.

We immunohistochemically investigated the expression of glutathione S transferase pi (GST- pi) and clarified the correlation between GST-pi and the results of chemosensitivity testing on the tissue of primary human squamous-cell carcinoma of the lung. The expression of GST-pi was evaluated in 105 cases and their level of chemosensitivity was estimated by the in vitro succinate dehydrogenase inhibition test for cisplatin, Adriamycin, cyclophosphamide, mitomycin C, vindesine and fluorouracil. Tumors in which 25% and more of the cells stained for GST-pi were classified as having a high GST-pi expression, while those tumors demonstrating less than 25% of the cells staining for GST-pi were considered to have a low expression GST-pi. The percentage of high GST-pi was 52% (53 of 105) while that of low GST-pi was 48% (52 of 105). No significant correlation between the expression of GST-pi and clinicopathologic factors was observed, while no significant difference in the survival of the two groups was found either. An increase in succinate dehydrogenase activity was recognized in the high-GST-pi group compared with the low-GST-pi group for each anticancer drug; however, no statistical significance was seen except for cisplatin. In the cases with adjuvant combination chemotherapy using cisplatin after a complete resection, all cases demonstrating a relapse were associated with a high GST-pi. These findings thus indicate that an overexpression of GST-pi is related to the resistance to cisplatin in human lung squamous-cell carcinoma. It is therefore important to select carefully the optimal anticancer drug for high-GST-pi cases.

Expression of glutathione S-transferase-pi and sensitivity of human gastric cancer cells to cisplatin.

The authors examined the correlation between the expression of glutathione S-transferase-pi (GST pi) and the sensitivity of gastric cancer to anticancer agents. In 62 human gastric carcinomas, the expression of GST pi was immunohistochemically evaluated, and sensitivity to the anticancer drugs, cisplatin (CDDP), doxorubicin (DXR) aclacinomycin A (aclarubicin), (ACR), 5-fluorouracil (5-FU), mitomycin C (MMC) and carboquone (carbazilquinon) (CQ) was examined using the in vitro succinate dehydrogenase inhibition test. The authors used the Chinese hamster ovary (CHO) cell line and its variant CDDP-resistant cell line C/CDP-2, and they analyzed the relationship among CDDP-sensitivity, mRNA expression, and immunohistochemical staining. Immunohistochemically detected GST-pi-positive tumors were noted in 35 of 62 excised tumors. There was no significant correlation between GST-pi expression and clinicopathologic features or prognosis. The succinate dehydrogenase activity of each drug for tumors, with regard to negative or positive GST-pi, was 34.9 plus or minus 13.7% and 46.0 plus or minus 22.0% for CDDP, with a significant statistical difference (P < 0.05). However, there was no statistical difference for the other drugs tested. C/CDP-2 cells showed a lower sensitivity to CDDP, a higher expression of mRNA for GST pi and a stronger immunohistochemical staining than CHO cells. The overexpression of GST-pi is significantly related to the sensitivity of gastric cancer to CDDP.

Expression of P-glycoprotein influences resistance against anthracyclines in clinical gastric carcinomas.

In 58 human gastric cancers, the expression of P-glycoprotein (P-gp) was evaluated immunohistochemically and chemosensitivity was determined using the in vitro succinate dehydrogenase inhibition (SDI) test. Tumors which contained over 75% stained cells were scored as positive, and 14 of 58 cases (24%) were positive. There was no significant correlation between P-gp expression and clinicopathologic features. The succinate dehydrogenase (SD) activity for each drug of P-gp positive and negative tumors was as follows: 81.8 +/- 15.2% vs. 66.3 +/- 16.1% for Adriamycin (ADM), 75.5 +/- 14.2% vs. 59.1 +/- 17.6% for aclacinomycin A (ACR), 71.7 +/- 15.0% vs. 61.1 +/- 14.0% for mitomycin C (MMC), and 57.5 +/- 18.4% vs. 47.0 +/- 16.7% for cisplatin (CDDP). The increase in SD activity was evident in P-gp positive tumors compared with negative ones in cases of ADM (P = 0.0044), ACR (P = 0.0105), and MMC (P = 0.0353). We suggested that P-gp expression is closely related to chemosensitivities of human gastric cancers to anthracyclines.

SYNERGISM OF CYTOTOXICITY BETWEEN CIS-DIAMINEDICHLORO-PLATINUM-(II) AND CIS-DIAMINE(1,1-CYCLOBUTANEDICARBOXYLATE)-PLATINUM-(II)

In an attempts to increase the antitumor effect and to reduce normal tissue toxicity, the combined cytotoxic effect of cis-Diamminedichloroplatinum (II) (CDDP) and cis-diammine(1,1-cyclobutane dicarboxylate) platinum (II) (CBDCA) was investigated using HeLa and colon 26 cell lines and the combination index (CI). Cytotoxicity of the combination of CDDP and CBDCA on 27 surgically resected specimens of human gastric and colorectal adenocarcinomas was also evaluated using the in vitro succinate dehydrogenase inhibition (SDI) test. The CI values varied with the dose ratio examined (1:1-1:6) of CDDP and CBDCA, with findings that CI<1, synergy, was obtained at fraction affected (Fa)>0.75 for HeLa cells and at Fa<0.9 for colon 26 cells in cases of a dose ratio of 1:1 to 1:2. Of all 27 clinical human adenocarcinomas, the succinate dehydrogenase (SD) activity was significantly lower in cancer cells concomitantly exposed to both CDDP and CBDCA than in those exposed to either drug alone. These positive effects of a combination of two platinum analogues on human malignant tissues have heretofore not been reported, which would warrant the clinical application of this combination for human malignant tumors.

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

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.

Expression of glutathione-S-transferases alpha and pi 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-alpha and GST-pi. 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-pi 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-alpha 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-alpha and CDDP resistance. The cellular content of GST-alpha 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.

Use of the succinate dehydrogenase inhibition test in assessing the heat sensitivity of tumor cells.

The in vitro succinate dehydrogenase (SD) inhibition (SDI) test was used to determine the heat sensitivity of tumor cell lines and human tumor cells. The tumor cells were exposed to heat in vitro and the decrease in SD activity was assayed using a colorimetric assay, the SDI test. With respect to survival curves of HeLa cells, the SDI test correlated well with the clonogenic assay and the dye exclusion assay. Decrease in the SD activity of HeLa cells after the heat treatment (41-44 degrees C, 10 min to 5 h) depended on both the temperature and the duration of heat exposure. S- and G2/M-phase-rich HeLa cells were more sensitive to heat than were the G1-phase-rich cells. The SDI test exhibited a wide variation in the heat sensitivity among four cell lines (HeLa, B-16, V-79, and a human GT-1 squamous cell carcinoma). Variation in heat sensitivity was also detected among individual tumor tissues obtained from clinical specimens of gastric, esophageal, and colorectal cancers. Gastric cancer tissues were more sensitive to heat than were esophageal and colorectal tissues. We recommended that if the SDI test is used to assess the heat sensitivity of clinical tumor tissues in vitro, appropriate therapy for individual patients can be designed.

ＳＤＩを用いた温熱感受性試験の確立に関する研究

ＳＤＩを用いた温熱感受性試験の確立に関する研究