Objective:To explore the effect of paclitaxel (PTX) and cisplatin (DDP) on the expression of NKG2D ligands of hu-man esophagus carcinoma cell EC9706 and on the cytotoxicity of cytokine-induced killer (CIK) cells, as well as to discuss its molecu-lar mechanisms. Methods: The half maximal inhibitory concentration (IC50) values of PTX and DDP against EC9706 cells for 24 h were measured by MTT assay. The expression levels of NKG2D ligands (MICA, MICB, ULBP1, ULBP2, and ULBP3) on the EC9706 cell surface before and after 24 h culture with 1/2 IC50 of PTX or DDP were assayed by flow cytometry. Cytotoxicity of CIK cells against EC9706 cells before and after 24 h culture with 1/2 IC50 PTX or DDP was analyzed by lactate dehydrogenase release assay at an effector to target cell ratio (E:T) of 20:1 and 30:1, respectively. The expression levels of DNA damage repair genes (ATM, ATR, CHK1, CHK2, and p53) of EC9706 cells before and after 24 h incubation with 1/2 IC50 PTX or DDP were detected by quantitative fluorescent PCR. Results:The IC50 values of PTX and DDP were 10 and 5μg/mL, respectively. MICB, ULBP2, and ULBP3 on EC9706 cells were upregulated after 24 h culture with 1/2 IC50 PTX (P<0.05), and the expression levels of MICA, MICB, ULBP2, and ULBP3 were higher after 24 h culture with 1/2 IC50 DDP (P<0.05). Cytotoxicity of CIK cells against EC9706 cells cultured with 1/2 IC50 of PTX or DDP at E:T of 20:1 and 30:1 was significantly enhanced compared with those untreated (P<0.05). The expression levels of DNA damage repair genes did not significantly increase after 24 h treatment with 1/2 IC50 PTX (P>0.05), whereas ATM, ATR, CHK1, and CHK2 were over-expressed after 24 h treatment with 1/2 IC50 DDP (P<0.05). Conclusion:PTX or DDP can enhance the susceptibility of EC9706 cells to CIK cell-mediated lysis by upregulating the expression of NKG2D ligands through activating DNA damage repair genes.

Objective: To analyze the expression of HLA class Ⅰ molecules and MHC classⅠ related chain A and B (MICA/MICB) in human nasopharyngeal carcinoma cell line (CNE2) and multi-drug resistant nasopharyngeal carcinoma cell line (CNE2/ DDP), and to assess their influence on NK cell-mediated lysis.Methods: Expression of HLA classⅠ molecules and MICA/MICB on the surface of CNE2 and CNE2/DDP cell lines was analyzed by flow cytometry. Cytotoxicity of NK cells (isolated from 3 healthy persons) against CNE2 and CNE2/DDP cells were detected by LDH releasing assay at different effect-to-target cell ratios (E∶T). In blocking experiments, anti-MHC class Ⅰ monoclonal antibody (mAb) (W6/32, a pan anti-HLA class Ⅰ antibody) and anti-MHC class I chain related molecules mAb (BAMO-1, specificly against MICA and MICB) were added to the target cells at a E∶T ratio of 10∶1. Results:It was found that the expression of HLA class Ⅰ molecules and MICA/MICB on CNE2 was higher than that on CNE2/DDP(P

Objective:To study the inhibitory effect of allogeneic natural killer(NK) cells on subcutaneously transplanted human multi-drug resistant nasopharyngeal carcinoma cells(CNE2/DDP) in BALB/c nude mice.Methods:Human leucocyte antigen(HLA) genotypes of CNE2/DDP cells and the genotypes of inhibitory killer cell immunoglobulin-like receptor(KIR) in NK cells(isolated from 3 healthy persons by immuno-magnetic microbead technique) were analyzed by PCR-SSP.Twelve BALB/c nude mice were evenly divided into 2 groups:the control group and the treatment group.Mice in the treatment group were injected subcutaneously with 1?106 CNE2/DDP cells together with 3?107 NK cells via the tail veins;mice in the control group were injected with 1?106 CNE2/DDP cells subcutaneously.The tumor formation time,tumor formation rate and changes of tumor size were observed.Three weeks after tumor formation,all the mice were killed and human NK cells in peripheral blood were analyzed by flow cytometry;the tumors were weighed and the tumor inhibitory rates were calculated.Results:The HLA genotypes of CNE2/DDPcells were A2,24,B18,35,Cw4,and 7;the KIR genotypes of the 3 healthy persons were KIR2DL1,KIR2DL3,KIR3DL1,and KIR3DL2.There were mismatches between the KIRs expressed in NK cells and HLA class Ⅰ molecules expressed in the CNE2/DDP cells.NK cells obviously inhibited the growth of CNE2/DDP xenograft in nude mice.The tumor formation periods of control group and NK cell group were(17.17?1.17) d and(24.83?1.47) d,respectively(P

Objective: To study the effects of IL-2 and IL-15 on the expression of NKG2D and the cytotoxicity of edited-NK cells against human nasopharyngeal carcinoma cell line CNE2.Methods: NK cells were purified by anti-CD56 MACS and were divided into four groups: non-edited-NK cells group(NK cells treated with 100 U/ml IL-2),edited-NK cells group(NK cells co-cultured with CNE2 cells at a ratio of 10∶1 and then treated with 100 U/ml IL-2),edited-NK cells retreated with 1 000 U/ml IL-2 group,and edited-NK cells retreated with 10 ng/ml IL-15 group.Expression of NKG2D in each group was determined by FACS 24 h later.Cytotoxicity of NK cells against CNE2 cells(NK∶CNE2 being 20∶1) was measured by LDH releasing assay.Results: The expression of NKG2D in non-edited-NK cells,edited-NK cells,edited-NK cells retreated with IL-2,and edited-NK cells retreated with IL-15 were(97.63?0.83)%,(53.50?1.25)%,(94.47?1.00)%,and(98.07?0.21)%,respectively.The expression of NKG2D on edited-NK cells retreated with IL-2 or IL-15 was significantly increased than that on edited-NK cells(P

Objective:To investigate the expression of MHC class Ⅰ chain-related gene A(MICA) on different human tumor cells(K562,MCF-7,HR-8348,CNE-2,HeLa) and their sensitivity to NK cytotoxicity.Methods:Cytotoxicity of NK cells(isolated from 3 healthy persons) were detected by LDH releasing assay at different effect-to-target cell ratios(E∶T). The expression of MICA was measured by FACS.Results:Different level of MICA was detected on surface of tumor cell lines, anti-MICAmAb could partially inhibit the cytotoxicity of NK cells.Conclusion:NKG2D-MICA interaction can trigger the cytotoxicity of NK cells against tumor,NK cells can be a kind of effector cell for the treatment of some certain tumors.

Objective: To inrestigate the association between thymidine phosphorylase (TYMP) polymorphisms and efficacy of postop-erative capecitabine-based adjuvant chemotherapy in colorectal cancer (CRC) patients. Methods: Two hundred and thirty-five patients with colorectal cancer who received surgical treatment and adjuvant chemotherapy between January 2010 and December 2016 from People's Hospital of Zhengzhou, were included in this study. Peripheral blood and postoperative tissue specimens of the CRC patients were collected for genotyping polymorphisms and measuring TYMP mRNA expression, respectively. The correlation between the poly-morphisms and efficacy of postoperative chemotherapy in CRC patients was analyzed. Results: The prevalence of 5633C>T in TYMP gene among the CRC patients was as follows: CC genotype, 149 cases (63.40%); CT genotype, 73 cases (31.06%); and TT genotype, 13 cases (5.54%); the minor allele frequency of 5633C>T was 0.21. Survival analysis of the patients revealed that the median overall sur-vival (OS) of patients with the CT/TT genotype and those with the CC genotype was 5.9 and 4.5 years, respectively; the result was sta-tistically significant (P=0.009). Following adjustment in multivariate Cox regression analysis, the CT/TT genotype was found to be an in-dependent favorable factor for OS (HR=0.67, P=0.015). Additionally, of the 87 postoperative tissue specimens, results show that the levels of TYMP mRNA in cancer tissues of patients with the CT/TT genotype were significantly higher than those with the CC genotype (P=0.019). Conclusions: TYMP mRNA expression may be influenced by the 5633C>T polymorphism, making CRC patients benefit from capecitabine treatment.

Objective To investigate the associations between CYP2A6 polymorphisms and treatment outcomes of adjuvant S-1 in postoperative gastric cancer patients.Methods 188 patients after D2 radical resection received S-1 based adjuvant chemotherapy.PBMC cell specimen were collected for the genotyping of genetic variation and CYP2A6 gene mRNA expression.Univariate analysis of genotypes and prognosis was carried out by Kaplan-Meier survival analysis,and multivariate analysis were adjusted by COX regression analysis.Results The polymorphisms included in this study were collected in the NCBI database with the minor allele frequency ＞ 10％ in Chinese population (rs8192725,rs8192720 and rs28399433),with rs8192725 only of clinical significance.The prevalence of rs8192725 in CYP2A6 were CC genotype 131 cases (69.7％),CT genotype 51 cases (27.1％),TT genotype 6 cases (3.2％),minor allele frequency of rs8192725 was 0.17.The 3 year disease-free survival (DFS) rate in patients with CT/TT genotype and CC genotype were 61.5％ and 72.5％,respectively (x2 =8.233,P =0.004).The 3 year overall survival rate of the two genotypes were 73.7％ and 79.4％ (x2 =4.863,P =0.021).CT/TT genotypes were an independent factor for DFS (OR =1.81,P =0.012).The expression of CYP2A6 in PBMC of the patients with CT/TT genotypes were significantly lower than those of the CC genotype patients (P ＜ 0.001).Conclusions After D2 gastric cancer patients treated by S-1,the polymorphism rs8192725 of CYP2A6 may effect the clinical outcomes of adjuvant chemotherapy S-1 treatment through influencing the mRNA expression of CYP2A6.

Objective:Oral fluoropyrimidine S-1 contains tegafur,gimeracil,and oteracil;among them,tegafur is the major active pre-cursor,which is metabolized to 5-fluorouracil by cytochrome P4502A6(CYP2A6).We examined the associations between CYP2A6 poly-morphisms and the treatment outcomes of adjuvant S-1 in patients with gastric cancer.Methods:Two hundred patients diagnosed with pathological stageⅡ-Ⅲgastric cancer were included in this study,and they received adjuvant S-1(40 mg/m2,bid,days 1-28,ev-ery 6 weeks for eight cycles)after curative surgery.Additionally,we analyzed the wild-type allele(W)(CYP2A6*1)and four variant al-leles(V)(CYP2A6*4,*7,*9,and*10).Results:Two hundred patients were enrolled in this study between November 2007 and July 2013.With a median follow-up of 46.4 months(range:12.5-80.1),the 3-year relapse-free survival(RFS)and overall survival(OS)rates were 83.1%(95% confidence interval(CI),77.7%-88.5%)and 94.8%(95% CI,91.6%-98.0%),respectively.However,RFS differed signifi cantly according to the CYP2A6 genotype.The 3-year RFS rates were 95.9% for W/W,83.1% for W/V,and 72.5% for V/V(P=0.032)gen-otypes.Grades 3 and 4 overall toxicity did not differ according to genotype for any grade(P=0.628 and P=0.227,respectively).Conclu-sions:CYP2A6 genotypes correlate with the outcome of S-1 chemotherapy,wherein patients with the variant genotypes show worse prognosis.Additionally,polymorphism detection may be used as a biomarker to guide clinical chemotherapy choices for adjuvant ad-ministration of gastric cancer therapy.

Objective:To investigate the efficacy, safety and common adverse reactions of Apatinib monotherapy in elderly patients with advanced colorectal cancer(CRC)who failed to respond to standard regimens.Methods:This was a retrospective study.A total of 106 elderly patients with advanced CRC who had failed standard regimens from January 2015 to December 2019 were included.Patients enrolled in this study received Apatinib with an initial dosage of 500 mg or 250 mg.The objective response rate(ORR)and disease control rate(DCR)were assessed after treatment with apatinib.The progression-free survival(PFS)and overall survival(OS)were evaluated during the follow-up period.Additionally, adverse reactions during treatment with apatinib were recorded.Results:The efficacy was assessed by using the best overall response during apatinib treatment.Of 106 patients, there were 9 patients with partial response(PR), 68 patients with stable disease(SD)and 29 patients with progressive disease(PD). The ORR was 8.5% and the DCR was 72.6%.The median PFS was 3.6 months and the median OS was 10.1 months.Relatively common adverse reactions in these patients were hypertension(63 patients, 59.4%)and hand-foot syndrome(HFS)(51 patients, 48.1%)during apatinib treatment.The median PFS of patients with hypertension and of patients without hypertension were 5.0 months and 3.0 months, respectively( P=0.008). The median PFS of patients with and without HFS were 5.4 months and 3.0 months, respectively( P=0.013). Conclusions:Preliminary evidence suggests that Apatinib monotherapy has good efficacy and safety in elderly patients with advanced CRC who have failed standard regimens, and patients with adverse reactions such as hypertension and HSF still have a good prognosis.

Objective:To investigate the effects of Fructus Corni extract on the B7-H6 expression in primary liver cancer cells of rats. Methods:Sixty SD rats were randomly divided into three groups, namely, model, matrine, and Cornus officinalis. The rat model bear-ing the primary liver cancer was induced by diethylnitrosamine, except for the rats in the control group. The rats in both the matrine and Cornus officinalis groups were fed with matrine and Cornus officinalis. The rats in model groups were fed with 0.9％sodium chlo-ride solution. The number of hepatocellular carcinoma nodules was calculated, and the tumor growth inhibition rate was also calculat-ed. The pathological changes of hepatic tissues in rats of each group were observed by hematoxylin and eosin staining. The expression levels of B7-H6 in these three groups were determined by immunohistochemistry and Western blot. Results:The number of liver nod-ules of the matrine and Fructus Corni group rats was lower than that of the model group (P<0.05). The tumor inhibition rate of the Cor-nus group was significantly higher than that of the matrine group (P<0.05). The tumor growth inhibition rate of the Cornus officinalis group was significantly higher than that of the matrine group (P<0.05). Immunohistochemistry showed that the positive expression of B7-H6 in the Cornus officinalis group and the matrine group was significantly higher than that in the model group (P<0.05), and the positive expression of B7-H6 in the Cornus officinalis group was significantly higher than that in the matrine group (P<0.05). Similarly, the protein expression of B7-H6 in the Cornus officinalis and matrine groups was significantly higher than that in the model group (P<0.05) by Western blot, while the protein expression of B7-H6 in the Cornus officinalis group was significantly higher than that in the matrine group (P<0.05). Conclusion:Fructus Corni extract may inhibit the growth of hepatocellular carcinoma through upregulating the B7-H6 expression.