人类平衡型核苷转运蛋白1预测吉西他滨治疗胰腺癌患者预后的价值:一项Meta分析

目的:采用Meta分析探讨人类平衡型核苷转运蛋白1(human equilibrative nucleoside transporter 1,bENT1)预测接受吉西他滨治疗的胰腺癌患者预后的价值.方法:检索Cochrane Library、Medline和Embase等数据库,筛选有关hENT1表达与胰腺癌预后的相关文献,分析hENT1低表达和高表达与接受吉西他滨治疗的胰腺癌患者的总生存期和无病生存期的关系,采用Meta分析合并风险比(hazard ratio,HR).结果:共纳入9篇合格文献.在接受吉西他滨治疗的胰腺癌患者中,hENT1低表达组有较高的死亡风险(HR=2.61,95％可信区间:2.03～3.34)和复发风险(HR=2.62,95％可信区间:1.94～3.54).在接受吉西他滨术后辅助治疗的患者中,hENT1低表达组同样有着较高的死亡风险(HR=2.49,95％可信区间:1.86～3.33)和复发风险(HR=2.79,95％可信区间:1.98～3.94).hENT1蛋白或mRNA低表达均预示着较高的死亡和复发风险.结论:hENT1表达水平在接受吉西他滨治疗的胰腺癌患者中具有预测预后的价值.     

The absence of human equilibrative nucleoside transporter 1 is associated with reduced survival in patients with gemcitabine-treated pancreas adenocarcinoma.

PURPOSE: Gemcitabine monotherapy is the standard palliative chemotherapy for pancreatic adenocarcinoma. Gemcitabine requires plasma membrane nucleoside transporter proteins to efficiently enter cells and exert it cytotoxicity. In vitro studies have demonstrated that deficiency of human equilibrative nucleoside transporter 1 (hENT1), the most widely abundant and distributed nucleoside transporter in human cells, confers resistance to gemcitabine toxicity, but the distribution and abundance of nucleoside transporters in normal and malignant pancreatic tissue is unknown. EXPERIMENTAL DESIGN: We studied tumor blocks from normal pancreas and 21 Alberta patients with gemcitabine-treated pancreatic cancer. Immunohistochemistry on the formalin-fixed, paraffin-embedded tissues was performed with specific hENT1 and human Concentrative Nucleoside Transporter 3 monoclonal antibodies and scored by a pathologist blinded to clinical outcomes. RESULTS: hENT1 was detected in normal Langerhan cells and lymphocytes but not in normal glandular elements. Patients in whom all adenocarcinoma cells had detectable hENT1 had significantly longer median survivals from gemcitabine initiation than those for whom hENT1 was absent in a proportion (10 to 100%) of adenocarcinoma cells (median survival, 13 versus 4 months, P = 0.01). Immunohistochemistry for human Concentrative Nucleoside Transporter 3 revealed moderate to high-intensity staining in all adenocarcinoma tissue samples. CONCLUSIONS: Patients with pancreatic adenocarcinoma with uniformly detectable hENT1 immunostaining have a significantly longer survival after gemcitabine chemotherapy than tumors without detectable hENT1. Immunohistochemistry for hENT1 shows promise as a molecular predictive assay to appropriately select patients for palliative gemcitabine chemotherapy but requires formal validation in prospective, randomized trials.     

The absence of human equilibrative nucleoside transporter 1 expression predicts nonresponse to gemcitabine-containing chemotherapy in non-small cell lung cancer

We report here the development of a polyclonal antibody for human equilibrative nucleoside transporter 1 (hENT1) and assess the expression of hENT1 in non-small cell lung cancer (NSCLC) patients who were treated with gemcitabine-containing chemotherapy. hENT1 expression was analyzed by immunohistochemical staining in 24 NSCLC biopsy samples of formalin-fixed, paraffin-embedded tissues. The hENT1-positive staining in NSCLC samples was significantly associated with response to gemcitabine-containing chemotherapy. Responses to gemcitabine-containing chemotherapy were evident in none of the seven patients with no hENT1 expression. These results indicate that the absence of hENT1 expression may be useful to predict NSCLC patients who will not respond to gemcitabine-containing chemotherapy.     

Human equilibrative nucleoside transporter 1 is associated with the chemosensitivity of gemcitabine in human pancreatic adenocarcinoma and biliary tract carcinoma cells

Gemcitabine has been one of the most commonly used agents for pancreatic adenocarcinoma chemotherapy, but the determinants of the sensitivity of and resistance to this agent are not yet fully understood. In this study with pancreatic carcinoma and biliary tract carcinoma cell lines, we examined the gene expression levels of nucleotide transporters and others related to the metabolism of gemcitabine in the light of sensitivity to this agent. Quantitative RT-PCR demonstrated that one of the nucleotide transporter genes; human equilibrative nucleoside transporter 1 (hENT1) was associated with the sensitivity to gemcitabine as represented by IC50, while the other genes for nucleotide transporter and metabolism were not. We conclude that increased hENT1 expression is a most important determinant of gemcitabine sensitivity at least in an in vitro study.     

Concurrent analysis of human equilibrative nucleoside transporter 1 and ribonucleotide reductase subunit 1 expression increases predictive value for prognosis in cholangiocarcinoma patients treated with adjuvant gemcitabine-based chemotherapy

Background:

The aim of this study was to investigate the predictive and prognostic values of intratumoural human equilibrative nucleoside transporter 1 (hENT1) and ribonucleotide reductase subunit 1 (RRM1) expression in advanced cholangiocarcinoma patients treated with adjuvant gemcitabine-based chemotherapy (AGC).

Methods:

Intratumoural hENT1 and RRM1 expression levels were investigated immunohistochemically in 127 patients with advanced cholangiocarcinoma who underwent surgical resection (68 with AGC and 59 without AGC). The impacts of hENT1 and RRM1 expression on survival were evaluated.

Results:

High intratumoural hENT1 and RRM1 expression levels were observed in 86 (68%) and 67 (53%) patients, respectively. In a multivariate analysis of 68 patients who received AGC, high hENT1 (P=0.044) and low RRM1 expression (P=0.009) were independently associated with prolonged disease-free survival (DFS), whereas low RRM1 expression (P=0.024) was independently associated with prolonged overall survival (OS). Moreover, concurrent high hENT1 and low RRM1 expression was a powerful independent predictor of prolonged DFS (P<0.001) and OS (P=0.001) when the combined classification of hENT1 and RRM1 was introduced.

Conclusions:

Concurrent analysis of hENT1 and RRM1 expression may increase the predictive value of these biomarkers for survival of advanced cholangiocarcinoma patients treated with AGC.     

Human equilibrative nucleoside transporter‐1 expression is a predictor in patients with resected pancreatic cancer treated with adjuvant S‐1 chemotherapy

The high expression of human equilibrative nucleoside transporter‐1 (hENT1) and the low expression of dihydropyrimidine dehydrogenase (DPD) are reported to predict a favorable prognosis in patients treated with gemcitabine (GEM) and 5‐fluorouracil (5FU) as the adjuvant setting, respectively. The expression of hENT1 and DPD were analyzed in patients registered in the JASPAC 01 trial, which showed a better survival of S‐1 over GEM as adjuvant chemotherapy after resection for pancreatic cancer, and their possible roles for predicting treatment outcomes and selecting a chemotherapeutic agent were investigated. Intensity of hENT1 and DPD expression was categorized into no, weak, moderate or strong by immunohistochemistry staining, and the patients were classified into high (strong/moderate) and low (no/weak) groups. Specimens were available for 326 of 377 (86.5%) patients. High expression of hENT1 and DPD was detected in 100 (30.7%) and 63 (19.3%) of 326 patients, respectively. In the S‐1 arm, the median overall survival (OS) with low hENT1, 58.0 months, was significantly better than that with high hENT1, 30.9 months (hazard ratio 1.75, P = 0.007). In contrast, there were no significant differences in OS between DPD low and high groups in the S‐1 arm and neither the expression levels of hENT1 nor DPD revealed a relationship with treatment outcomes in the GEM arm. The present study did not show that the DPD and hENT1 are useful biomarkers for choosing S‐1 or GEM as adjuvant chemotherapy. However, hENT1 expression is a significant prognostic factor for survival in the S‐1 arm.     

Determinants of sensitivity and resistance to gemcitabine: the roles of human equilibrative nucleoside transporter 1 and deoxycytidine kinase in non-small cell lung cancer

Gemcitabine is one of the most commonly used agents for lung cancer chemotherapy, but the determinants of sensitivity and/or resistance to this agent are not yet fully understood. In this study we used quantitative RT-PCR to examine the expression levels of human equilibrative nucleoside transporter 1 (hENT1) and deoxycytidine kinase (dCK) genes in non-small cell lung cancer (NSCLC) cell lines in relation to sensitivity and resistance to gemcitabine. The basal expression levels of hENT1 were significantly correlated with the IC50 values for gemcitabine (r =-0.6769, P = 0.0005), whereas dCK expression levels were not. In a highly gemcitabine-sensitive cell line, NCI-H23, the sensitivity to gemcitabine was inhibited by nitrobenzylmercaptopurine ribonucleoside (NBMPR), an inhibitor of hENT1, without significant modulation of hENT1 expression. These data suggest that hENT1 is associated with gemcitabine sensitivity in lung cancer. We also continuously exposed NCI-H23 cells to gemcitabine and subsequently established the drug-resistant clone H23/GEM-R, which showed a significant decrease of dCK expression; however, hENT1 expression was not altered in the continuously exposed sublines or in the resistant clone. We conclude that increased hENT1 expression is a determinant of gemcitabine sensitivity, while decreased dCK expression is associated with acquired resistance to gemcitabine in NSCLC cells. Thus, hENT1 and dCK might be useful as predictive markers for efficacy of gemcitabine therapy in NSCLC.     

Human equilibrative nucleoside transporter 1 (hENT1) protein is associated with short survival in resected ampullary cancer.

BACKGROUND: Gemcitabine is an acceptable alternative to best supportive care in the treatment of advanced biliary tract cancers. The human equilibrative nucleoside transporter 1 (hENT1) is a ubiquitous protein and is the major means by which gemcitabine enters human cells. Moreover, recent reports indicate a significant correlation between immunohistochemical variations of hENT1 in tumor samples and survival after gemcitabine therapy in patients with solid tumors. Materials and methods: We used immunohistochemistry to assess the abundance and distribution of hENT1 in tumor samples from radically resected cancer of the ampulla, and sought correlations between immunohistochemical results and clinical parameters including disease outcomes. RESULTS: In the 41 individual tumors studied, 12 (29.3%) had uniformly high hENT1 immunostaining. Statistical analysis showed a significant correlation between hENT1 and Ki-67 (P = 0.04). No statistical significant differences were found between immunohistochemical findings and patient characteristics (sex, age, and tumor-node-metastasis). On univariate analysis, hENT1 and Ki-67 expression were associated with overall survival (OS). Specifically, those patients with overexpression of hENT1 showed a shorter OS (P = 0.022) and those with high Ki-67 staining showed a shorter survival (P = 0.05). CONCLUSIONS: hENT1 expression is a molecular prognostic marker for patients with resected ampullary cancer and holds promise as a predictive factor to assist in chemotherapy decisions.     

Enhanced efficacy of gemcitabine by indole-3-carbinol in pancreatic cell lines: the role of human equilibrative nucleoside transporter 1

Pancreatic cancer patients treated with gemcitabine (2',2'-difluorodeoxycytidine) can eventually develop resistance. Recently, published data from our laboratory demonstrated enhanced efficacy of gemcitabine with the dietary agent, indole-3-carbinol (I3C). The current study examined the possible mechanism for this I3C-enhanced efficacy. Several pancreatic cell lines (BxPC-3, Mia Paca-2, PL-45, AsPC-1 and PANC-1) were examined for modulation of human equilibrative nucleoside transporter 1 (hENT1) expression, the major transporter for gemcitabine, by I3C alone and combined with gemcitabine. I3C significantly (p<0.01) up-regulated hENT1 expression in several cell lines. Gemcitabine alone showed no effect on hENT1 expression. However, combining gemcitabine with I3C further increased hENT1 expression. Cell viability assays revealed no effect of I3C on normal cells, hTERT-HPNE. hENT1-specific inhibitor, nitrobenzylthioinosine, significantly abrogated I3C-induced gemcitabine cytotoxicity, further demonstrating its specificity. This study demonstrates that up-regulation of hENT1 expression may be a novel mechanism involved in the additive effect of I3C and gemcitabine.     

cN-II expression predicts survival in patients receiving gemcitabine for advanced non-small cell lung cancer

Resistance to gemcitabine is likely to be multifactorial and could involve a number of mechanisms involved in drug penetration, metabolism and targeting. In vitro studies of resistant human cell lines have confirmed that human equilibrative nucleoside transporter 1 (hENT1)-deficient cells display resistance to gemcitabine. Overexpression of certain nucleotidases, such as cN-II, has also been frequently shown in gemcitabine-resistant models. In this study, we applied immunohistochemical methods to assess the protein abundance of cN-II, hENT1, human concentrative nucleoside transporter 3 (hCNT3) and deoxycitidine kinase (dCK) in malignant cells in from 43 patients with treatment-na?ve locally advanced or metastatic non-small cell lung cancer (NSCLC). All patients subsequently received gemcitabine-based chemotherapy. Response to chemotherapy, progression-free survival (PFS), and overall survival (OS) were correlated with abundance of these proteins. Among the 43 samples, only 7 (16%) expressed detectable hENT1, with a low percentage of positive cells, 18 expressed hCNT3 (42%), 36 (86%) expressed cN-II and 28 (66%) expressed dCK. In univariate analysis, only cN-II expression levels were correlated with overall survival. None of the parameters were correlated with freedom from progression survival nor with response. Patients with low levels of expression of cN-II (less than 40% positively stained cells) had worse overall survival than patients with higher levels of cN-II expression (6 months and 11 months, respectively). In a multivariate analysis taking into account age, sex, weight loss, stage and immunohistochemical results, cN-II was the only predictive factor associated with overall survival. This study suggests that cN-II nucleotidase expression levels identify subgroups of NSCLC patients with different outcomes under gemcitabine-based therapy. Larger prospective studies are warranted to confirm the predictive value of cN-II in these patients.     

Human equilibrative nucleoside transporter 1 (hENT1) expression is a potential predictive tool for response to gemcitabine in patients with advanced cholangiocarcinoma

BackgroundCholangiocarcinoma (CC) is a rare cancer of the liver. Surgery offers the only chance for cure. When surgery is unfeasible, chemotherapy is the backbone of treatment. The combined administration of cisplatin and gemcitabine is considered standard of care. Human equilibrative nucleoside transporter 1 (hENT1) is the major transporter responsible for gemcitabine uptake into cells. hENT1 expression is associated with an increased survival for patients receiving gemcitabine after pancreatic cancer surgery, suggesting that hENT1 is predictive of response to gemcitabine.AimTo determine whether there is a correlation between the expression of hENT1 and disease outcome in CC.MethodsA retrospective study on 43 patients treated at our centre with a locally advanced or metastatic CC, who received first line treatment with gemcitabine, was performed.ResultsFor the whole population, median Progression Free Survival (PFS) and overall survival (OS) were 4.0 (95% Confidence Interval 2.7–5.3 months) and 10.0 months (95% CI 6.8–13.2 months), respectively. From the 26 samples available for hENT1 staining, 18 (69%) and 8 (31%) patients had high and low hENT1 immunostaining, respectively. The median PFS were 2.0 versus 6.0 months for low versus high staining respectively (p = 0.012). The median OS were 5.0 versus 11.0 months for low versus high staining, respectively (p = 0.036). On multivariate analysis, hENT1 expression was the single independent predictive factor associated with prolonged PFS (HR 0.35, p = 0.023) and OS (HR 0.41, p = 0.046).ConclusionIn this study we show the potential of hENT1 expression as a predictor of outcome in CC treated with gemcitabine. Larger studies are necessary to confirm these promising results.     

Prognostic significance of RASSF1A promoter methylation on survival of non-small cell lung cancer patients treated with gemcitabine

The epigenetic inactivation of genes plays an important role in lung cancer. We have investigated the methylation status of the promoter region of seven genes (APC1A, DAPK, FHIT, p14(ARF), p16(INK4a), RARbeta, RASSF1A) in serum DNA of NSCLC patients. The objective of our study was to reveal the influence of such alterations on overall survival. Blood samples were drawn pretherapeutically. Genomic DNA was purified from serum, treated with sodium bisulfite and hypermethylation was detected by a nested methylation-specific PCR in a group of 92 patients with histologically confirmed stage IIIB and IV NSCLC. All patients received gemcitabine first-line alone or in combination with other drugs. The vast majority (n=87) showed at least one epigenetic alteration. The methylation frequencies of individual genes varied between 25.9 and 47.3%. The hypermethylation status of none of the genes had a significant influence on median overall survival of the total population. In contrast, patients with a methylated RASSF1A gene who showed a partial response survived significantly longer (33.6+/-10.4 month) compared to those with a wild-type allele (12.9+/-4.7 month, P=0.0045). This effect became even more pronounced in combination with p14(ARF) (P=0.0004). This difference was not seen in patients with stable or progressive disease. A multivariate analysis confirmed that RASSF1A methylation was an independent prognostic factor. Our results show that the hypermethylation frequency of single genes and the accumulation of epigenetic alterations in individual samples of NSCLC patients may vary considerably. Molecular parameters such as hypermethylation of RASSF1A or p14(ARF) may be useful prognostic markers in subpopulations.     

Decrease of CA 19-9 during chemotherapy with gemcitabine predicts survival time in patients with advanced pancreatic cancer

Chemotherapy with gemcitabine has been shown to be an effective regimen in advanced or metastatic pancreatic cancer with improvement of both quality of life and survival time. The response of the tumour marker CA 19-9 to chemotherapy with gemcitabine was studied in order to find out whether it is related to survival time of patients. Forty-three consecutive patients (median age 61 years, range 39-76 years; 20 males, 23 females) suffering from histologically proven locally advanced or metastatic pancreatic adenocarcinoma and a baseline Karnofsky-index > or = 60 were treated with gemcitabine in a dose of 1,000 mg/m(-2) weekly x 7 followed by 1 week of rest during the first cycle and thereafter 1,000 mg/m(-2) weekly x 3 followed by 1 week of rest until progression. In 36 of 43 patients serial measurements of CA 19-9 could be performed. Patients with a decrease of > 20% of the baseline CA 19-9 level after 8 weeks of treatment (n = 25) had a significantly better median survival than patients with a rise or a decrease < or = 20% (n = 11) (268 vs 110 days; P < 0.001). The response of CA 19-9 was the strongest independent predictor of survival (P < 0.001) in the multivariate analysis. In conclusion, a decrease of CA 19-9 > 20% during the first weeks of chemotherapy with gemcitabine is associated with a better survival of patients with locally advanced or metastatic pancreatic cancer. Serial measurements of CA 19-9 are useful to decide whether further chemotherapy after the first weeks of treatment is indicated.     

ERCC1 predicts outcome in patients with gastric cancer treated with adjuvant cisplatin-based chemotherapy

Background

Adjuvant chemotherapy is gaining an increasing role in resectable gastric cancer. Customizing chemotherapy on the basis of chemosensitivity may improve outcome, and putative predictive molecular markers have been mostly evaluated in Asian patients. We profiled key DNA and damage signaling factors and correlated them with outcome, in a European cohort.

Methods

Formalin-fixed tumor samples obtained from surgical specimens of patients treated with adjuvant cisplatin-based chemotherapy for gastric cancer were analyzed. Immunohistochemistry (IHC) was performed to analyze excision repair cross-complementing gene 1 (ERCC1) and thymidylate synthase (TS) expression, and p53 mutations were detected with direct sequencing.

Results

Among the 68 patient recruited, the median age was 69 (range 30–74), and UICC stage was III in 44 patients (65 %). With a median follow-up of 40.5 months, disease-free and overall survival were 18.0 (95 % CI 13.4–22.76) and 56 months (95 % CI 44.87–67.13), respectively. ERCC1 score was 0 in 14 out 67 (21 %) cases, 1 in 19 (28 %), 2 in 20 (30 %) and 3 in 14 cases (21 %). Longer overall survival (p = 0.04) was found in patients categorized as ERCC1 negative by IHC according to median score. TS score was 0 in 16 out 67 (24 %) cases, 1 in 27 (40 %), 2 in 16 (24 %) and 3 in 8 cases (12 %). Mutations of p53 were found in 21 out 66 (32 %) cases. Neither TS nor p53 were found to correlate with outcome.

Conclusion

Excision repair cross-complementing gene 1 by IHC might predict patients more likely to benefit from adjuvant cisplatin-based chemotherapy in curatively resected gastric cancer. In patients exhibiting ERCC1 positive tumors, alternative regimens should be evaluated.     

Immunohistochemical variation of human equilibrative nucleoside transporter 1 protein in primary breast cancers.

Gemcitabine and capecitabine are nucleoside analogues used in chemotherapy strategies for the treatment of breast cancer. We previously demonstrated that deficiency in hENT1, the most abundant and widely distributed plasma membrane nucleoside transporter in human cells, confers high-level resistance to gemcitabine toxicity in vitro, whereas the relationship between hENT1 activity and capecitabine toxicity is unknown. To determine the relationship between capecitabine cytotoxicity and hENT1 abundance, cultured MDA-MB-435s human mammary carcinoma cells were exposed to graded concentrations of the capecitabine metabolites, 5'-deoxy-5-fluorouridine or 5-fluorouracil, in the presence and absence of nitrobenzylmercaptopurine ribonucleoside (NBMPR), a tight-binding inhibitor of hENT1. The presence of NBMPR reduced the cytotoxic effects of 5'-deoxy-5-fluorouridine, indicating that hENT1 also enabled cellular uptake of this capecitabine metabolite by breast cancer cells. We report here the development of an immunohistochemical method to assess the hENT1 abundance of malignant cells in solid tumors. Frozen sections of 33 primary breast cancers were stained with monoclonal antibodies raised against a synthetic peptide derived from the large intracellular loop of hENT1, and staining intensity was scored on a 0-4+ scale. hENT1 staining intensity varied markedly among breast samples (4 with score 0, 5 with score 1+, 7 with score 2+, 14 with score 3+, 3 with score 4+), suggesting that at least 9 of the tumors were hENT1 deficient. We conclude that because hENT1 deficiency has previously been associated with nucleoside drug resistance, immunohistochemical staining of hENT1 warrants further study as a predictive tool for guiding the appropriate use of gemcitabine and capecitabine in the treatment of breast cancer.