| Gastric carcinoma is one of malignant neoplasms that has an serious influence on humanhealthy. The patients with gastric carcinoma had a poor improvement on the five-yearsurvival rate which was lower than30percent on the advanced stage. Chemotherapy ofcytotoxic drugs was major method except for surgical procedures and radiotherapy. Thecurative effects were limited because the cytotoxic effect was not specificity and not all of thegastric cancer cells were sensitive to chemotherapy.Sachs suggested differentiation therapy as he detected the phenotypic reversion of malignancyin mice myeloid leukaemia dysdifferentiation on the effects of the induced cell differentiatedand proliferation inhibition materials in1970s. Induced cell differentiated therapy couldinduced tumor cells differentiation and apoptosis because of immaturity anddysdifferentiation of tumor cells in which the controls of genetic regulation were abnormal.Induced cell differentiated therapy which is a new therapeutic method has been paid closeattention to oncotherapy.The clinic frequently used cell differentiating inducers are retinoids compounds whichproduced a marked biological effect by retinoic signal pathways to modulate geneticexpression after to combine with endonuclear retinoic acid receptors or retinoic acid Xreceptors, further to bind retinoic acid response element of gene promoter region, and toresult in cellular growth suppression, apoptosis and cellular differentiation. Now, it waspreferred therapeutic strategy to treat acute promyelocytic leukemia by induced differentiationtherapy to utilize all trans-retinoic acid (ATRA) which was also to be used on preventionand treatment in the solid tumor of mammary adenocarcinoma, thyroid cancer, non-small-celllung carcinoma, prostatic cancer and cervical cancer, however, there is no to confirm thetherapeutic effect on gastric carcinoma. ATRA has limited clinic applications because of theserous side effect of retinoic acid syndrome,inefficienc and resistance. Many kinds ofretinoic compounds were synthesized according to the lead compound of ATRA in the prophase study at the college of pharmacy Anhui medical university.4-anino-2-trifluoromethyl-phenyl retimate (ATPR) was showed to possess more potent oninducing cell differentiation in leukemic cells.This study was focus on the therapeutical effect of ATRA on gastric carcinoma and was tostudy the antitumous effect and partial mechanism of the derivative of ATPR in gastriccarcinoma in order to obtain experiment data on developing high performance andharmfulless derivative of ATPRObjective: To clarify the the expressions of RARs/RXRs and effects of the treatment byATRA on postsurgical prognosis in the patients with gastric carcinoma, and to investigatetheantitumous effect and partial mechanism of ATRA and the derivative of ATPR, and toprovide a new therapeutic intervention of inducing cell differentiation in gastric carcinoma.Methods:①The expressions of RARs/RXRs protein in the gastric cancer tissue weredeteced by immunohistochemisty, we analyzed the relation between the clinicopathologicfactors and expressions of RARs/RXRs, then, we detected the influence between the overallsurvival and expressions of RARs/RXRs by survival analysis in147gastric carcinomacases.②We performed a clinical trials to study the effect on overall survival of treatment by ATRAwith a oral dose at45mg/m2and6months course of treatment in80gastric cancer cases.③The relative expression of RARs/RXRs was quantified by Western-blotting analysis inMKN-45,MKN-74,NCI-N87cell line.④T heantitumous effects of ATPR on gastric cancer cell lines were analysed by MTT test.⑤ATPR combined with RAR or RXR selectively was analyzed by luciferase reporter gene.⑥C ell cycle, apoptosis and celldifferentiation were analyzed by flom cytometry (FCM) inMKN-45,MKN-74,NCI-N87cells with treatment by ATPR or ATRA on the time of72hrs.⑦T odetect the variance of expression of carcinoembryonic antigen (CEA) byWestern-blotting analysis in MKN-45,MKN-74,NCI-N87cells with treatment by5uM ATPR or ATRA on the time of72hrs.⑧The variance of relative expression of RARs/RXRs quantified by real-time fluorescentquantitative polymerase chain reaction (RT-FQ-PCR) and Western-blotting analysis in MKN-45,MKN-74,NCI-N87cells treated by5uM ATPR or ATRA on the time of72hrs.⑨To detect the variance of expression of alkaline phosphatase (ALP) and lactatedehydrogenase (LDH) by spectrophotography in MKN-45,MKN-74,NCI-N87cells withtreatment by5uM ATPR or ATRA on the time of72hrs.Results:(1) RARs/RXRs existed in all of gastric normal epithelium, gastric carcinomas,There was lower expression of RXRβ not only in gastric normal epithelium, but also ingastric carcinomas and cell lines, and no strongly positive by immunohistochemisty. Positivecorrelation was present between RARs and RXRs.(2) The more advanced on AJCC TNM staging system, the lower expression of RARβ, RARγ,RXRa(p<0.05). The worse on cell differentiation,the lower expression of RARa and RARβ(p<0.05).(3) RARa and AJCC TNM staging system were independent prognosis factors, the patientwith positive expression of RARa had a longer overall survival time (hazard rate,HR=0.42),and prolonged the overall survival time moreover if treated by ATRA with a oraldose at45mg/m2and6months course of treatment in gastric cancer cases(HR=0.41).(4) The antitumous effect of ATPR was obvious in the gastric cancer cell lines: MKN45,MKN74,NCI-N87which was dose dependence and time dependence (p<0.05). IC50of theantitumous effect of ATPR was3.0uM～6.2uM, however,that of ATRA was6.6uM～8.5uM.The growth suppression effect of ATPR on the normal SVHUC cell was lower than ATRA onthe gastric cancer cell lines with the concentration of10-4mol/L.(5) Luminescence value of (RARE-TK-LUC+RARa+ATPR)was lager than Luminescencevalue o(fRARE-TK-LUC+RARa),Luminescence value o(fRARE-TK-LUC+RXRa) andLuminescence value of(RARE-TK-LUC+RXRa+ATPR)(P<0.05)；Luminescence value of(RARE-TK-LUC+RXRa+RARa+ATPR)was lager thanLuminescence value of(RARE-TK-LUC+RARa+ATPR)(P<0.05).(6) Flow cytometric analysis indicated that the percentage of cells in subG1phase wasimproved obviously in MNK-45and MKN-74cells by only10uM not5uM ATPR on thetime of72hrs than the control group(P<0.05),on the other hand,the percentage of cells insubG1phase was increased significantly by all of5uM and10uM ATPR on the time of72 hrs (P<0.05). The percentage of cells in G0/G1phase was increased up to75.11%inNCI-N87cells with treatment by5uM ATPR on the time of72hrs than the control group.(7) The rate of apoptosis cells in MKN-45,MKN-74,NCI-N87was36.8%,41.3%and46.5%respectively by only5uM ATPR on the time of72hrs, and that of ATRA was36.0.2%,36.1%and35.7.3%.(8) The expression of CEA was decreased significantly in MKN-45,MKN-74cells exceptNCI-N87cells with treatment by5uM ATPR on the time of72hrs(P<0.05).(9) the expression of RARa mRNA was increased obviously in the MKN-45, MKN-74,NCI-N87with treatment by5uM ATPR on the time of72hrs(P<0.05).(10) The expression of RARa protein was increased significantly in MKN45,MKN74,NCI-N87with treatment by5uM ATPR on the time of72hrs(P<0.05).(11) The vitality of LDH and ALP was decreased significantly in MKN45,MKN74,NCI-N87by5uM ATPR on the time of72hrs, and ATPR had more potent than ATRA indecreasing the vitality of LDH and ALP(P<0.05).Conclution:①RARa is a positive prognosis factor, and a marker of cell differentiateddegree. ATRA has an ability of antitumous effect, it could prolonged the postoperativeoverall survival time if treated with ATRA.②ATPR could inhibit gastric cancer cellular proliferation by inducing cancer cells todifferentiate and apoptosis and cell cycle arresting. When the gastric cancer cells are on themore poorly differentiated state,ATPR could induce cancer cells to differentiate easily. Whenthe gastric cancer cells are on the better differentiated state,ATPR could induce apoptosiseasily.③The antitumous effect of ATPR could depend on retinoic signal transduction pathway:ATPR is a agonist of RAR, RAR forms a heterodimers with RXR, ATPR combines withRAR to form a compound of ATPR/RAR/RXR, then the ATPR/RAR/RXR binds the RARE ofthe gene which controls cell differentiation and apoptosis④ATPR has more potent on the the antitumous effect than ATRA, which is preferable toinduce cell differentiation on more poorly differentiated gastric cancer by3.0～6.2uM. Thenew retinoid of ATPR shall become a potent therapeutic drug on inducing gastric cancer celldifferentiation... |
PDF Full Text Request