PDAC remains a major therapeutic challenge. Recent randomized clinical trials showed a significant clinical benefit of gemcitabine-based chemotherapy in patients with both resected and unresectable PDAC [7,33
]. Therefore, gemcitabine-based chemotherapy remains the standard palliative chemotherapy for PDAC. However, there remains a substantial subset of cases in which gemcitabine-based chemotherapy is insufficient, suggesting the importance of introducing individualized chemotherapy into the clinical setting. Individualized chemotherapy, based on the expression of genes involved in cellular uptake and metabolism of gemcitabine, will be a potent strategy.
We and other investigators have demonstrated that several altered gene expression profiles, including those of hENT1, dCK, RRM1
, and RRM2
correlated with the sensitivity to gemcitabine in cancer cell lines [12,15,20,22
]. However, analysis of gene expression in two gemcitabine-resistant pancreatic cancer cell lines revealed that there were differences in gene expression between these cancer cell lines. These data suggest that there are different patterns of gene expression that can develop gemcitabine resistance, and combined evaluation of several genes may be required to predict gemcitabine sensitivity.
In the current study, univariate analyses showed that low hENT1
, low dCK
, high RRM1
, and high RRM2
correlated well with poor outcome in patients treated with gemcitabine-based AC, although these altered expression levels did not reach statistical significance in multivariate analysis. Recent clinical studies, including two prospective clinical trials, revealed that PDAC patients with high hENT1 immunoreactivity or high hENT1 expression gained significant benefit from gemcitabine-based AC [13,34,35
], and these data are consistent with our results. However, although Akita et al. [36
] and Zheng et al. [37
] revealed that high RRM1 and high excision repair cross-complementation group 1-expressing patients with PDAC or NSCLC had prolonged survival regardless of AC, Nakahira et al. [20
] and Akita et al. [36
] also demonstrated that only patients with low RRM1 derive significant benefit from gemcitabine on disease recurrence. Therefore, RRM1 expression may contribute to gemcitabine resistance in PDAC. Moreover, although Sebastiani et al. [38
] demonstrated that PDAC patients with high dCK expression had prolonged survival regardless of AC and concluded that genetic alterations of dCK
are not a common mechanism of resistance to gemcitabine, previous in vitro
] support our results showing that high dCK
and low RRM2
expressions are correlated with prolonged survival time in PDAC patients who received gemcitabine-based AC. Therefore, to introduce individualized AC into the clinical setting, based on gene expression profiles, the expression levels of several genes will need to be determined, and combined evaluation of these results may be needed. For this reason, we evaluated a simplified score, the GEM score, and found that a low GEM score was predictive for reduced DFS and prognostic for reduced survival in resected PDAC treated with gemcitabine-based AC. However, to evaluate the usefulness of this score, further studies, incorporating larger patient numbers, are required.
In contrast, we found that there was no evident correlation between CDA
expression levels and survival time. Recently, a single-nucleotide polymorphism in the CDA
gene, which was analyzed using the peripheral blood of cancer patients, was reported to influence the pharmacokinetics and toxicity of gemcitabine [39
]. Bengala et al. [40
] also demonstrated that high CDA
expression and CDA activity levels in peripheral blood mononuclear cells were correlated with shorter survival in gemcitabine-treated patients with advanced pancreatic carcinoma. These data suggest that simple quantification of CDA
mRNA in PDAC tissues is not helpful in predicting sensitivity of gemcitabine treatment.
Only 10% to 20% of patients with PDAC are candidates for curative resection [3
]; therefore, the remaining 80% to 90% of patients with unresectable advanced PDAC need cytopathologic assessment of EUS-FNA specimens, or pancreatic juice specimens, to predict their sensitivity to chemotherapeutic agents for individualized chemotherapy. The present analysis of mRNA is quantitative (even considering the small amount of specimen available, including cytologic specimens). In addition, the present results revealed that quantification of mRNA in neoplastic cells microdissected from cytologic samples was more useful to distinguish between samples with higher and lower gene expression levels compared with the analysis of WCP samples. The reliability of tests based on tissue or cell extracts is often crucially dependent on the relative abundance of the target cell population, and sampling errors or a large number of “contaminating cells” can lead to false-negative results [26
mRNA were reported to be expressed in human T lymphocytes and monocytes [41,42
], and RRM1 and RRM2 are essential for DNA synthesis in somatic cells. For these reasons, quantification of gene expression in cells microdissected from EUS-FNA cytologic specimens is likely to be useful for predicting gemcitabine sensitivity in patients with PDAC, although further investigations are needed before this approach can be introduced into the clinical setting.
In conclusion, we demonstrated that quantitative analysis of hENT1, dCK, RRM1, and RRM2 mRNA using FFPE tissue samples and evaluation of a combined GEM score were useful in predicting the sensitivity to gemcitabine-based AC in patients with PDAC. In addition, quantitative analysis of these genes in neoplastic cells microdissected from EUS-FNA specimens was useful in determining the treatment for patients with PDAC even when the tumor is unresectable. Quantitative analyses of genes related to cellular uptake and metabolism of cytotoxic agents can be a potent tool to perform individualized chemotherapy.