2386 Factor miR-142-5p pN status R factor TABLE 3 Multivariate survival analysis (Cox regression model) of clinical prognostic factors and miR-142-5p expression in patients with gemcitabine treatment Relative risk 3.109 2.055 2.197 95% CI 1.087–10.01 0.646–9.219 0.823–5.904 P value 0.0340 0.2395 0.1146 CI conﬁdence interval status (P = 0.24) or R factor (P = 0.11). In the gemcita- bine group, miR-142-5p expression was an independent prognostic marker for pancreatic cancer patients, with a relative risk of 3.109 (Table 3). Univariate analysis of the nongemcitabine group showed that the only signiﬁcant clinical factor was pN status. Therefore, multivariate survival analysis was per- formed by using miR-34a expression and pN status. The results show that the overall survival time was signiﬁcantly dependent on both miR-34a expression and pN status (Table 4, P = 0.0103 for miR-34a; P = 0.0035 for pN status). In the nongemcitabine group, miR-34a expression was an independent prognostic marker for pancreatic can- cer patients with a relative risk of 2.920 (Table 4). DISCUSSION The present miRNA proﬁling study using two gemcit- abine-resistant pancreatic cancer cell lines and their parent cell lines identiﬁed 24 miRNAs candidates, which were up- or downregulated in gemcitabine resistant cells. The pres- ent results also showed that high miR-142-5p and miR-204 patients had statistically signiﬁcantly longer survival times than the low miR-142-5p and miR-204 patients in the gemcitabine group, but not in nongemcitabine group, although further examination is needed because the number of patients in the nongemcitabine group is too small to conclude that there were no differences in survival between high and low expression patients in the nongemcitabine group. Liu et al. reported that miR-142-5p was repressed in human lung cancer, and the transfection of miR-142-5p signiﬁcantly repressed lung cancer cell growth.27 miR-204 has been also reported to be downregulated in intrahepatic cholangiocarcinoma, and the level of miR-204 expression TABLE 4 Multivariate survival analysis (Cox regression model) of clinical prognostic factors and miR-34a expression in patients without gemcitabine treatment Factor miR-34a pN status CI conﬁdence interval Relative risk 2.920 2.957 95% CI 1.303–6.295 1.410–6.812 P value 0.0103 0.0035 K. Ohuchida et al. was inversely correlated with that of Bcl-2 expression, possibly leading to chemotherapeutic drug-triggered apoptosis.28 Taken together, these data suggest that miR- 142-5p and miR-204 are promising predictive markers for chemotherapeutic responses in patients with resected pan- creatic cancer. In neuroblastomas, miR-34a was generally expressed at lower levels in unfavorable primary tumors, and the rein- troduction of miR-34a results in a dramatic reduction in cell proliferation.29 miR-34a expression was decreased in 9 of 25 (36%) colon cancers, and transient introduction of miR-34a suppressed the in vitro and in vivo growth of colon cancer.30 In non-small-cell lung cancer, the miR-34 family was downregulated in tumors compared with normal tissues, and low levels of miR-34a expression correlated with a high probability of relapse.31 These data suggest that miR-34a functions as a potential tumor sup- pressor. Our results show that miR-34a is a favorable prognostic marker in patients without gemcitabine treat- ment after resection. We also found no correlation between miR-34a expression and survival time in patients treated with gemcitabine, possibly suggesting that pancreatic cancers with low levels of miR-34a are more sensitive to gemcitabine treatment than those with high levels of miR- 34a, although larger studies are needed to conﬁrm this. In conclusion, miR-142-5p expression is correlated with survival time in patients treated with gemcitabine after sur- gical resection of pancreatic cancer, but not in patients without gemcitabine treatment. miR-142-5p would be a promising predictive marker for gemcitabine treatment in patients with resected pancreatic cancer, although further examination are needed to analyze the functional role of these microRNAs. In the present study, we used FFPE samples to measure the indicated miRNAs. Measurement of such miRNAs may be possible using plasma, serum, and pancreatic ﬂuids, suggesting that these miRNAs may be useful in predicting the effects of chemotherapy for unre- sectable pancreatic cancer and of neoadjuvant chemotherapy for resectable pancreatic cancer, where it is difﬁcult to obtain tissues samples without the use of invasive procedures. ACKNOWLEDGMENT Supported in part by a Grant-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan, and a grant from the Kato Memorial Bioscience Foundation. We are grateful to Emiko Manabe, and Miyuki Omori (Department of Surgery and Oncology, Kyushu University) for skillful technical assistance. CONFLICT OF INTEREST None. OPEN ACCESS This article is distributed under the terms of the Creative Commons Attribution Noncommercial License which permits any noncommercial use, distribution, and reproduction in any medium, provided the original author(s) and source are credited.