Immunohistochemical Demonstration of Dihydropyrimidine Dehydrogenase in Normal and Cancerous Tissues

Abstract

Dihydropyrimidine dehydrogenase (DPD) is a rate-limiting enzyme in the catabolism of 5-fluorouracil (5-FU). Previous studies have suggested that high enzyme activities and mRNA levels of DPD are involved in the resistance of cancers to 5-FU. For immunohistochemically demonstrating the tumor resistance to 5-FU, we determined the most suitable and reproducible condition of antigen retrieval for DPD localization in formalin-fixed, paraffin-embedded sections, and analyzed DPD expression in various types of normal and cancerous tissues. Pressure cooking in ethylenediaminetetraacetic acid, pH 8.0, was the best choice for retrieving the antigenicity of DPD. In normal tissues, DPD immunoreactivity was consistently detected in squamous epithelia, hepatocytes, Clara cells, ductal/glandular epithelia of various organs, myoepithelial cells, and non-epithelial cells such as macrophages, plasma cells, and part of the endothelial cells and fibroblasts. DPD was strongly expressed in adenocarcinomas of the lung, gallbladder and pancreas, hepatocellular carcinomas, and transitional cell carcinomas of the urinary bladder. Squamous cell carcinomas of the pharynx and lung were moderately reactive. Carcinomas of the breast and stomach revealed less consistent and less intense reactivity. Colorectal adenocarcinomas were not immunoreactive. The polyclonal antibody gave a broader and stronger reactivity than the monoclonal antibody KM1915, particularly in the normal epidermis and skin appendage as well as in squamous cell carcinomas and gastric cancers. The present results hopefully contribute to further investigations, in which the immunohistochemical demonstration of DPD is utilized as a tool of tailor-made chemotherapy for selecting of patients with cancer resistant to 5-FU and its derivatives.

Recurrence and 5-FU sensitivity of stage I/II node-negative breast, lung, or gastric cancer with occult neoplastic cells in lymph node sinuses

“…Then the primary tumors of the patients in the recurrence and non-recurrence groups were immunohistochemically stained for TS and DPD by the indirect immunoperoxidase method using polyclonal anti-TS and -DPD antibodies (Taiho Pharmachemical Co. Ltd., Japan) as the primary antibodies (25)(26)(27)(28)(29). To assess the extent of immunostaining, stained tumor cells were counted at a high magnification in the entire maximum cut surface of the lesion.…”

“…The mechanism by which 5-FU controls tumor growth involves FdUMP, a metabolite of 5-FU that forms a ternary complex with methylenetetrahydrofolate (CH 2 FH 4 ) and thymidylate synthase (TS), thereby inactivating TS and inhibiting DNA synthesis (24,25). It is also known that more than 80% of each dose of 5FU is directly metabolized by dihydropyrimidine dehydrogenase (DPD) in the liver (26,27). Therefore, TS and DPD are the target of 5-FU and are involved in its metabolism, respectively, making these enzymes important determinants of sensitivity to the anticancer effect of 5FU+LV therapy.…”

“…Therefore, TS and DPD are the target of 5-FU and are involved in its metabolism, respectively, making these enzymes important determinants of sensitivity to the anticancer effect of 5FU+LV therapy. Earlier studies have shown that tumors with high TS and low DPD expression are most sensitive to 5-FU+LV (24)(25)(26)(27). However, no previous study used immunohistochemistry to assess the sensitivity of the primary tumor to 5-FU in patients with stage I/II LNnegative breast, lung, or gastric cancer and ONCs in their regional LNs.…”

Advanced Application of the In Vivo Cryotechnique to Immunohistochemistry for Animal Organs

“…been frequently used to improve the immunoreactivity of antigens on paraffin-embedded sections [2,10,11,21,22,33]. They are particularly necessary for certain intranuclear antigens, due to masking or steric hindrance in compact chromatin components induced by the molecular crosslinking properties of common aldehyde fixatives [1,23].…”

Comparative Expression of mRNA and Protein of DPC4 in Relation to Ubiquitin Expression in Various Pancreatic Duct Lesions of Human Pancreatic Cancer Tissue

“…The sections were deparaffinized in xylene for 5 min 3 times, hydrated in 100% and 95% ethanol and finally immersed in phosphate-buffered saline (PBS). To detect Dpc4 protein, the sections were treated with 10 mM sodium citrate buffer (pH 6) at 95°C for 40 min in a water bath (Yamato BM400, Japan) and cooled for 20 min at room temperature [12]. Next, the sections were incubated with a 1:100 dilution of anti-Dpc4 protein monoclonal antibody (Ab) (clone B8, Santa Cruz Biotech, Santa Cruz, CA, U.S.A.) for 60 min at room temperature.…”