Laser Microdissection in Pharmacological Study of Pancreatic Cancer
Funel N1, Giovanetti E2, Campani D1, Pollina LE1, Mey V2, Nannizzi S2,Danesi R2, Del Chiaro M3, Boggi U3, Del Tacca M2, Bevilacqua G1, Mosca F3
1Division of Surgical, Molecular and Ultrastructural Pathology, 2Division of Pharmacology and Chemotherapy, 3Division of General Surgery and Transplantation and Regional Referral Center for Pancreatic Diseases Treatment. Department of Oncology, University and Hospital of Pisa. Pisa, Italy
ABSTRACT
Background Despite the knowledge of molecular pathology of pancreas cancer (PC), the systemic chemotherapy still relies on few drugs (gemcitabine and 5-fluorouracil). The rate-limiting step of gemcitabine activation is catalyzed by deoxycytidine kinase (dCK), whereas 5’-nucleotidase (5’-NT) and cytidine deaminase (CDA) inactivate the drug. Similarly, thymidylate synthase (TS) is the target enzyme of 5-fluorouracil, while polyglutamylation by folylpolyglutamate synthetase (FPGS) of intracellular 5,10-methylenetetrahydrofolate may enhance the cytotoxicity of 5-fluorouracil by allowing more efficient formation and stabilization of the inhibitory ternary complex involving thymidylate synthase and the 5-fluorouracil metabolite.
Aim In this work we compare, on samples microdissected and no-microdissected, RNA expression of key genes involved in pharmacological treatment of PC.
Methods One hundred and six cases of PC were collected in Pisa between December 2001 and June 2004 and RNA extraction was performed in all samples after laser micro-dissection (Leica ASLMD). In 17 cases RNA extraction was also obtained from the whole tumor (comprising stroma, vessels, inflammatory cells). Quantitative PCR analysis was carried out for dCK, 5’-NT, CDA, TS and FPGS genes in both microdissected and no-microdissected samples.
Results Data obtained by no-microdissected were not so informative compared to microdissected specimens. In particular, in microdissected samples the quantitative analysis of the results showed an evident difference of mRNA expression of dCK (1.031±0.171), 5’-NT (0.932±0.166), CDA (0.959±0.154) and TS (1.088±0.189), and particularly for FPGS (0.609±0.399).
Conclusions As expected, laser microdissection is a valid tool for molecular analysis and this technique is much more valid in pancreatic carcinoma where the desmoplastic reaction can underestimate the molecular expression of epithelial component, the true target of chemotherapy. This suggests a possible stratification of patients on the basis of their genotype to create homogeneous groups with different likelihood to respond to gemcitabine or fluorouracil treatment.
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