[A prospective study of adenosine triphosphate-tumor chemosensitivity assay directed chemotherapy in patients with recurrent ovarian cancer].

OBJECTIVE: To investigate the efficacy of adenosine triphosphate (ATP)-tumor chemosensitivity assay (TCA) directed chemotherapy in patients with recurrent epithelial ovarian cancer. METHODS: From August 2010 to June 2012, recurrent epithelial ovarian cancer patients were prospectively enrollmented in Cancer Hospital, Peking Union Medical College,Chinese Academy of Medical Sciences.The entry criteria are as follows: (1) Histologically proven to be epithelial ovarian cancer. (2) Patients of recurrent ovarian cancer with bidimensionally measurable tumor, or ascitic or pleural fluid for testing. (3) Karnofsky performance status > 60. (4) A life expectancy of at least more than 6 months.According to patients desires, they were assigned into two groups: assay-directed therapy group and physician's-choice therapy group, patients' clinical and pathological characteristics, response rate to chemotherapy and progression-free survival (PFS) were compared between two groups. RESULTS: A total of 113 patients with recurrent epithelial ovarian cancer were prospectively enrollmented to assay-directed chemotherapy (n = 56) or physician's-choice chemotherapy (n = 57).There was no difference in median age,types of recurrence, surgical-pathological stage, pathological type, tumor grade, times of recurrence, residual disease at secondary cytoreductive surgery between assay-directed group and physician's-choice group. The overall response rate (ORR) and median PFS in the ATP-TCA group was 66% (37/56) and 7 months, while the ORR in the control group was 46% (26/57, P = 0.037), the median PFS was 4 months (P = 0.040). For platinum-resistant patients, the ORR between ATP-TCA directed chemotherapy 59% (16/27) and control group 25% (7/28) were significantly different (P = 0.010), and the median PFS between two groups were also significantly different (5 months and 2 months, respectively, P = 0.003). CONCLUSION: ATP-TCA directed chemotherapy could improve ORR and PFS in patients with recurrent epithelial ovarian cancer, especially in platinum-resistant patients.

[The mRNA expression of BRCA1, ERCC1, TUBB3, PRR13 genes and their relationship with clinical chemosensitivity in primary epithelial ovarian cancer].

OBJECTIVE: To evaluate the expression of BRCA1, ERCC1, TUBB3 and PRR13 mRNA and their relationship with clinical chemosensitivity in primary ovarian cancer, and to assess the predictive value of joint detection of both BRCA1 and ERCC1 genes for the treatment of primary ovarian cancer. METHODS: Primary epithelial ovarian tumor samples were collected from 46 patients who underwent cytoreductive surgery. Real-time quantitative PCR was used to analyze the relative expression of BRCA1, ERCC1, TUBB3 and PRR13 mRNA in those cases. The correlation of clinical chemosensitivity and the test results was statistically analyzed. The efficacy of the joint prediction of clinical chemosensitivity by combining the two drug resistance gene detection was evaluated. RESULTS: The BRCA1 mRNA relative expression logarithm in the clinical-resistant group was 0.673±2.143, and clinical-sensitive group -1.436±2.594 (P=0.008). The ERCC1 mRNA relative expression logarithm in the clinical-resistant group was -0.529±1.982 and clinical-sensitive group -3.188±2.601 (P=0.001). BRCA1 and ERCC1 expression level is negatively correlated with platinum-based chemosensitivity. The PRR13 expressions in the two groups were not significantly different (P=0.074), and the TUBB3 expressions between the two groups were also not significantly different (P=0.619). When the intercept point value BRCA1 mRNA expression logarithm was -0.6, the predictive sensitivity, specificity, positive predictive value and negative predictive value were 73.3%, 75.0%, 84.6% and 60.0%, respectively, with the best comprehensive assessment. When the intercept point value of ERCC1 mRNA expression logarithm was -1, the predictive sensitivity, specificity, positive predictive value and negative predictive value were 80.0%, 68.8%, 82.8% and 64.7%, respectively, with the best comprehensive assessment. The combination detection of BRCA1 and ERCC1 can improve the chemotherapeutic sensitivity, specificity, positive predictive value and negative predictive value to 86.7%, 68.8%, 83.9% and 73.3%, respectively. CONCLUSIONS: BRCA1 and ERCC1 mRNA expression has a negative correlation with the clinical sensitivity of platinum-based chemotherapy. Combination detection of the two drug-resistance associated genes can improve the predictive efficacy of ovarian cancer chemosensitivity and beneficial to individual treatment of ovarian cancer.

[Predicting clinical chemo-sensitivity of primary ovarian cancer using adenosine triphosphate-tumor chemosensitivity assay combined with detection of drug resistance genes].

OBJECTIVE: To predict clinical chemotherapy sensitivity of primary ovarian cancer by jointing adenosine triphosphate (ATP)-tumor chemo-sensitivity assay (TCA) method in vitro and detection of drug resistance genes, provide reference for clinical treatment. METHODS: Forty-seven primary epithelial ovarian tumor samples were collected from the patients who received cytoreductive surgery. Viable ovarian cancer cells obtained from malignant tissue were tested for their sensitivity to carboplatin (CBP), cisplatin (DDP), paclitaxel (PTX) and CBP + PTX using ATP-TCA method in vitro; at same time, real-time quantitative PCR was used to analysis BRCA1 and ERCC1 mRNA relative expression in forty-six specimens (1 frozen tumor samples mRNA were not detected due to serious degradation). The relationship between ATP-TCA test results, clinical indicators, and the effectiveness of the joint prediction on clinical chemo-sensitivity by combining these two methods were statistically analyzed using chi-square test. RESULTS: (1) The results showns that three programs of DDP, CBP and PTX + CBP were significantly related with clinical results (P < 0.05) in vitro, in which the compliance rate in PTX + CBP program was the highest 83% (39/47), and the predictive sensitivity, predictive specificity, positive predictive value, negative predictive value and predictive accurate rate were 90%, 71%, 84% and 80%, respectively. PTX + CBP combined in vitro test results was also related with residual tumor size and neoadjuvant chemotherapy, which was more prone to drug resistance with residual tumor larger than 2 cm (P = 0.023) and with neoadjuvant chemotherapy (P = 0.011). (2) BRCA1 mRNA expression levels in the clinical-resistant group and the clinical-sensitive group was 0.673 ± 2.143 and -1.436 ± 2.594 (P = 0.008), ERCC1 mRNA expression levels in the clinical-resistant group and the clinical-sensitive group was -0.529 ± 1.982 and -3.188 ± 2.601 (P = 0.001). There were also significant correlation among the expression levels of BRCA1, ERCC1 mRNA and clinical efficacy (P < 0.01). (3) ATP-TCA and detection of drug resistance genes combined to predict the clinical application of PTX + CBP resistance may occur in 8/9 cases. CONCLUSIONS: ATP-TCA may be an ideal method of in vitro drug sensitivity testing method, which could effectively predict clinical chemotherapy sensitivity. Combination of the drug-resistant associated genes detection method and the ATP-TCA method can increase the predictive effectiveness of ovarian cancer chemosensitivity and guide individual chemotherapy of ovarian cancer.

[Application of ATP-tumor chemosensitivity assay in primary epithelial ovarian cancer].

OBJECTIVE: To evaluate the predictive value of the adenosine triphosphate-tumor chemosensitivity assay (ATP-TCA) in the chemotherapy applied in primary epithelial ovarian cancer (PEOC), and to analyze if the neoadjuvant chemotherapy have any influence on the postoperative chemosensitivity. METHODS: ATP-TCA results from 61 PEOC specimens were analyzed retrospectively. Patients were divided into sensitive group and resistant group according to the ATP-TCA results. Sensitive index (SI) was applied to analyze the ATP-TCA results. The correlation between in vitro results and clinical outcome was assessed by univariate and multivariate analysis. RESULTS: SI set at > 250 had the highest test sensitivity, specificity, positive and negative predictive value of 91.6%, 73.9%, 84.6% and 85.0%, respectively. The ATP-TCA results had significant correlation with clinical outcome (chi(2) = 26.9, P < 0.001). Patients with tumors shown to be resistant had a higher risk of recurrence in comparison with those who were tested as sensitive (P = 0.030, OR = 0.033, 95%CI 0.002 approximately 0.724). The median progression-free survival (PFS) and overall survival (OS) of in vitro-sensitive patients were 26 months and 39 months, respectively, significantly longer than those in the in vitro drug-resistant group of patients (PFS 10 months and OS 25 months) (both P < 0.01). Neoadjuvant chemotherapy had a significant correlation with the clinical chemoresistance (chi(2) = 15.214, P < 0.001). CONCLUSION: ATP-TCA assay may effectively predict the chemosensitivity of primary ovarian cancer, and predict the early recurrence of the tumor.

[Application of ATP-tumor chemosensitivity assay in recurrent epithelial ovarian cancer].

OBJECTIVE: To explore the value of adenosine triphosphate-tumor chemosensitivity assay (ATP-TCA) in individualized treatment of recurrent epithelial ovarian cancer (REOC), and to evaluate the correlation between the in vitro chemosensitivity assay and clinical drug sensitivity. METHODS: Sixty-nine REOC specimens were tested by ATP-TCA assay retrospectively. The patients were divided into strong sensitive, moderate sensitively and resistant groups according to the ATP-TCA assay results. The clinical results were evaluated according to imaging and serum CA125 analysis. The correlation between in vitro ATP-TCA assay and clinical outcome was statistically analyzed by χ(2) test. The progression free survival (PFS) and overall survival (OS) of each group were analyzed using Kaplan-Meier method. RESULTS: The results of ATP-TCA assay had significant correlation with clinical outcome. The clinical chemotherapy outcome became better with increased drug sensitivity in vitro (χ(2) = 9.066, P = 0.004). The sensitivity, specificity, positive predictive value, negative predictive value and accuracy rate for ATP-TCA method to predict the clinical chemotherapy sensitivity of REOC were 87.5%, 45.9%, 58.3%, 80.9% and 65.2%, respectively. The mean PFS of strong sensitive group, moderately sensitive group and resistant group were 187.1 days, 195.0 days and 60.3 days, respectively. The mean OS were 476.7, 335.7 and 237.5 days, respectively, following the start of TCA-directed therapy. The PFS and OS of the two sensitivity groups in vitro were significantly longer than that of the in vitro-resistant group (P < 0.01). CONCLUSION: The results of ATP-TCA assay are well correlated with clinical treatment responses. The assay may be an important and useful method for individualized chemotherapy for recurrent ovarian cancer.

[The effect of siRNA-Her2/neu on the drug sensitivity of Her2/neu-overexpressing lung adenocarcinoma cell line].

OBJECTIVE: To explore the effect of synthesized siRNA targeting Her2/neu oncogene on the drug sensitivity of Her2/neu-overexpressing lung adenocarcinoma cell line. METHODS: The experiments consisted of four groups including an untreated control group, an empty vector group, an unrelated siRNA group, and a Her2/neu siRNA group. Every experiment was repeated five times. Lung adenocarcinoma cell line Calu-3 was transfected with siRNAs formulated LipofectAMINE 2000, and Her2/neu protein and P-gp of each group were determined by flow cytometry (FCM). The chemosensitivity of transfected cells to cisplatin (DDP) was measured by MTT. Cell apoptosis detection kit (Annexin V method) was used to examine the drug induced apoptosis rate. RESULTS: The Her2/neu protein and P-gp positive expression rate in the Her2/neu siRNA group, the untreated control group, the empty vector group and the unrelated siRNA group were [(25.04 +/- 1.56)%, (4.24 +/- 1.01)%], [(98.24 +/- 2.23)%, (5.11 +/- 2.98)%], [(95.67 +/- 1.98)%, (6.98 +/- 2.47)%] or [(94.79 +/- 0.87)%, (5.59 +/- 3.66)%], respectively. Introduction of the sequence specific siRNA into Her2/neu positive Calu-3 cells in vitro greatly reduced the cell surface expression of the Her2/neu protein, but had no effect on P-gp level. Consequently the inhibitory rate of DDP in combination with siRNA targeting Her2/neu was (67.1 +/- 2.3)%, but it was (48.1 +/- 3.5)%, (46.3 +/- 5.9)% and (50.2 +/- 2.9)% in the untreated control, the empty vector and the unrelated siRNA groups, respectively. There was a significant difference between Her2/neu siRNA group and other three groups (P < 0.01). The FCM results showed the apoptosis rate of DDP combined with siRNA-Her2/neu was elevated as compared with the unrelated siRNA group, the empty vector group or the untreated control group. CONCLUSION: Sequence specific siRNA-Her2/neu was capable of enhancing the chemosensitivity of Calu-3 cells to cisplatin.

Suppression of primary breast, colon, gastric and bladder cancers cell growth in vitro by CKBM, a natural product.

CKBM is a product composed of natural ingredients and had been shown to possess certain anti-cancer effects in vitro and in vivo. The aim of the present study is to analyze the chemosensitivity in the treatment of primary colon, breast, gastric and bladder cancer cells by CKBM. A total of 77 patients with cancers of breast, colon, stomach or bladder were included in the present study. Primary cancer cells were isolated from the surgical removed tumors and treated with various dosages of CKBM for 5 days. ATP is then extracted and measured by luminescence assay. CKBM treatment inhibited primary colon, breast, gastric and bladder cancer growth dose-dependently. The IC values were smaller from tumor cells at early stages, when compared with the ones at later stages. The present study strongly indicated that CKBM exerted cytotoxic effect on primary cancer cells.

[Correlation between ATP bioluminescence tumor chemosensitivity assay and clinical response in ovarian cancer].

OBJECTIVE: To determine the correlation between results of ATP bioluminescence tumor chemosensitivity assay (ATP-TCA) of human ovarian cancer specimens in vitro and clinical chemo-therapeutic responses of patients. METHODS: Thirty-four freshly taken ovarian cancer specimens (28 cases) and ascites (6 cases) and 9 chemotherapeutic drugs were tested in vitro for cancer chemosensitivity by ATP-TCA. RESULTS: Among the 34 ovarian cancer cases, the efficacy of ATP-TCA is 94.0%, the sensitivity, the specificity, the positive and negative predicting values, and an overall predicting value in vitro and vivo were 90.0%, 91.7%, 94.7%, 84.6% and 90.6%, respectively. CONCLUSION: The results of ATP-TCA assay are correlated well with clinical treatment responses. The assay may be an important and useful method for individual-based chemotherapy of cancers.

[Expression of retinoic acid receptor-beta mRNA and p16, p53, Ki67 proteins in esophageal carcinoma and its precursor lesions].

OBJECTIVE: To study the expression of retinoic acid receptor-beta (RAR-beta) mRNA and p16, p53, Ki67 proteins in squamous-cell carcinoma of the esophagus and its precursor lesions in a high risk population. METHODS: A total of 397 tissue specimens were collected from individuals with normal mucosa (NM, n = 25), mild dysplasia (MiD, n = 69), moderate dysplasia (MoD, n = 106), severe dysplasia (SD, n = 51), carcinoma in situ (CIS, n = 78), and squamous-cell carcinoma (SC, n = 68). Expression of RAR-beta mRNA was detected by in situ hybridization, and that of p16, p53 and Ki67 proteins by immunohistochemistry. RESULTS: The frequencies of RAR-beta mRNA expression in NM, MiD, MoD, SD, CIS and SC were 96.0%, 89.9%, 67.9%, 68.6%, 62.8%, and 63.2%, respectively. The frequencies of p16 expression were 88.0%, 71.0%, 64.2%, 51.0%, 53.8% and 52.9%; those of p53 expression were 4.0%, 39.1%, 57.5%, 52.9%, 67.9% and 69.1%; those of Ki67 expression were 0, 40.6%, 61.3%, 58.8%, 59.0% and 75.0%, respectively. CONCLUSION: There are no significant differences in four biomarkers expression between carcinoma of the esophagus and its precursor lesions.

[Expression of a plant associated human cancer antigen in breast cancer].

OBJECTIVE: To study the expression of a glycoprotein of plant origin in normal, benign and malignant breast tissues. METHODS: Expression of a plant glycoprotein was examined in 5 samples of normal breast tissues, 20 fibro-adenoma and 136 breast cancer by SABC immunohistochemical staining and the results were analyzed by SPSS statistics software. RESULTS: No positive staining was detected in normal breast tissues (0/5). Weak staining was observed in 4 of 20 (20.0%) breast fibro-adenoma. Positive staining was demonstrated in 116 out of 136 (85.3%) breast cancer specimens. The differences were statistically significant. The expression of plant-associated human cancer antigen was related to pathological grade (P < 0.05), tissue invasiveness (P < 0.01) and recurrence (P < 0.05), but not to patients' age, tumor size and c-erbB-2 expression. CONCLUSION: The plant glycoprotein studied may be a human cancer-associated antigen which might be a potential marker of breast cancer.

[Antitumor effect and apoptosis induction in human cancer cell lines by BRM-SJS].

BACKGROUND & OBJECTIVE: It had been observed that BRM-SJS had antitumor effect in our clinical practice. This study was designed to investigate the antitumor activity of BRM-SJS, and mechanism of its action. METHODS: In vitro antitumor experiments with MTT method, meanwhile cell morphology, flow cytometry, and agarose gel electrophoresis were performed for determining apoptosis in several tumor cell lines. RESULTS: BRM-SJS had antitumor effects on human Suzhou human glioma (SHG-44), breast carcinoma (MCF-7), and human pancreas carcinoma (PANC1) in vitro, the IC50 values of BRM-SJS were 0. 299 mg/ml, 1.853 mg/ml and 9.416 mg/ml respectively. At the 2. 5 mg of BRM-SJS on SHG-44 and MCF-7, marked morphological changes, including cell shrinkage and condensation of chromosomes, were observed with electric microscope. The increase of apoptosis in SHG-44 and MCF-7 cells treated with BRM-SJS extracts 0.625 -2.5 mg for 14 -48 h was observed by Annexin-V/PI flow cytometry analysis. Agarose gel electrophoresis of DNA from SHG-44 and MCF-7 cells treated with BRM-SJS extracts 1.25 -5 mg for 24 h or 48 h showed marked DNA Ladder pattern. CONCLUSION: Antitumor activity of BRM-SJS may be related with inducement of apoptosis of tumor cells.

[Heterogeneity of human breast cancer cells and their biological behavior].

OBJECTIVE: To investigate the heterogeneity of human breast cancer cells, their influence on biological behavior of tumor cells and clinical implications. METHODS: The subpopulations of MCF-7 breast cancer cells were isolated by Percoll gradient centrifugation. DNA content and cell cycle distribution were detected with flow cytometry. Tumor chemosensitivity analysis was performed with MTT assay. RESULTS: Heterogeneity was observed in DNA content and cell cycle distribution among four subpopulations of breast cancer cells, which were related to their proliferation ability and chemosensitivity results. CONCLUSION: Hereditary instability and intrinsic characteristics of most tumor cells, not only lead to tumor progression and heterogeneity but also cause the loss of monoclonality and the generation of subclones. Further study on some profiles of tumor heterogeneity such as DNA content, cell cycle distribution and their influence on tumor proliferation and chemosensitivity may very well improve the clinical treatment.

Expression of a plant-associated human cancer antigen in normal, premalignant and malignant esophageal tissues.

AIM: To study the relationship between the expression profiles of a plant-associated human cancer antigen and carcinogenesis of esophagus and its significance. METHODS: We analyzed expression of a plant-associated human cancer antigen in biopsy specimens of normal (n=29), mildly hyperplastic (n=29), mildly (n=30), moderately (n=27) and severely dysplastic (n=29) and malignant esophageal (n=30) tissues by immunohistochemistry. RESULTS: The plant-associated human cancer antigen was mainly confined to the cytoplasm and showed diffuse type of staining. Positive staining was absent or weak in normal (0/30) and mildly hyperplastic tissue samples (2/29), while strong staining was observed in severe dysplasia (23/29) and carcinoma in situ (24/30). There was significant difference of its expression between normal mucosa and severely dysplastic tissues (P<0.001) or carcinoma in situ (P<0.001). Significant difference was also observed between mild dysplasia and severe dysplasia (P<0.001) or carcinoma in situ (P<0.001). An overall trend toward increased staining intensity with increasing grade of dysplasia was found. There was a linear correlation between grade of lesions and staining intensity (r=0.794, P<0.001). Samples from esophageal cancer showed no higher levels of expression than those in severely dysplastic lesions (P>0.05). CONCLUSION: The abnormal expression of this plant-associated human cancer antigen in esophageal lesions is a frequent and early finding in the normal-dysplasia-carcinoma sequence in esophageal carcinogenesis. It might contribute to the carcinogenesis of esophageal cancer. The abnormal expression of this plant-associated human cancer antigen in esophageal lesion tissues may serve as a potential new biomarker for early identification of esophageal cancer.

The abnormal expression of retinoic acid receptor-beta, p 53 and Ki67 protein in normal, premalignant and malignant esophageal tissues.

AIM: Esophageal cancer remains a significant health problem worldwide. It is important to investigate alterations in expression of retinoic acid receptor-beta, p 53 and Ki67 proteins in esophageal carcinogenesis. METHODS: To find biomarkers for early identification of esophageal cancer, we analyzed the retinoic acid receptor-beta, p 53 protein and the proliferation marker Ki67 in surgical specimens of normal, mildly, and severely dysplastic and malignant esophageal tissues by in situ hybridization of RNA and immunohistochemistry. RESULTS: RAR-beta was expressed in 94.3%(33/35) of normal mucosae, 67.8%(19/28) of the mild, 58.1% (18/31) of the severe lesions and 53.2%(116/218) of tumor samples. RAR-beta mRNA was expressed in 62.7%(42/67), 55.1%(43/78) and 29.2%(7/24) of well, moderated and poorly differentiated SSCs. The p 53 and Ki67 proteins were 5.9%(2/34) of the normal mucosa. P53 and Ki67 stained positively in 10.7%(3/28) and 21.4% (6/28) of mild dysplasia, and 51.6%(16/31) and 58.1% (18/31) of severely dysplasia respectively. Samples from esophageal cancer showed no higher levers of p 53 and Ki67 expression than seen in severely dysplastic lesions. There was significant difference of RAR-beta p 53 and Ki67 expression between normal mucosa and dysplastic tissue or esophageal cancer. CONCLUSION: Loss of RAR-beta expression and accumulation of p 53 and Ki67 proteins may serve as biomarkers for early identification of esophageal cancer in the high-risk populations.