Inhibition of epidermal growth factor receptor and vascular endothelial growth factor receptor phosphorylation on tumor-associated endothelial cells leads to treatment of orthotopic human colon cancer in nude mice.

The purpose of our study was to determine whether the dual inhibition of epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor (VEGFR) signaling pathways in tumor-associated endothelial cells can inhibit the progressive growth of human colon carcinoma in the cecum of nude mice. SW620CE2 human colon cancer cells growing in culture and orthotopically in the cecum of nude mice expressed a high level of transforming growth factor alpha (TGF-alpha) and vascular endothelial growth factor (VEGF) but were negative for EGFR, human epidermal growth factor receptor 2 (HER2), and VEGFR. Double immunofluorescence staining revealed that tumor-associated endothelial cells expressed EGFR, VEGFR2, phosphorylated EGFR (pEGFR), and phosphorylated VEGFR (pVEGFR). Treatment of mice with either 7H-pyrrolo [2,3-d]-pyrimidine lead scaffold (AEE788; an inhibitor of EGFR and VEGFR tyrosine kinase) or CPT-11 as single agents significantly inhibited the growth of cecal tumors (P < .01); this decrease was even more pronounced with AEE788 combined with CPT-11 (P < .001). AEE788 alone or combined with CPT-11 also inhibited the expression of pEGFR and pVEGFR on tumor-associated endothelial cells, significantly decreased vascularization and tumor cell proliferation, and increased the level of apoptosis in both tumor-associated endothelial cells and tumor cells. These data demonstrate that targeting EGFR and VEGFR signaling on tumor-associated endothelial cells provides a viable approach for the treatment of colon cancer.

Multiple organ failure due to 5-fluorouracil chemotherapy in a patient with a rare dihydropyrimidine dehydrogenase gene variant.

BACKGROUND: Dihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in the metabolism of the chemotherapeutic drug 5-fluorouracil (5-FU). Application of 5-FU is restricted by a narrow therapeutic index because of severe toxicity of WHO grades III-IV. The exon 14-skipping mutation (c.1905+1G>A) accounts for approximately a quarter of all severely toxic cases. However, numerous other polymorphisms have been identified within the DPYD gene in affected patients, and the pathophysiological significance of most of them is unclear. PATIENT AND METHODS: We report a patient with advanced caecum cancer who twice received 950 mg 5-FU and 45 mg folinic acid as adjuvant by bolus injection. 2 days after onset of chemotherapy, the patient developed a multiple organ dysfunction exhibiting a cardiogenic shock with severe left ventricular insufficiency, marked reduction of renal function, and beginning hepatic encephalopathy with somnolence, myoclonus, and a seizure. In order to investigate a possible defect within the DPYD gene direct sequencing of all 23 exons was carried out. RESULTS: Genotyping revealed a rare c.1601G>A polymorphism which causes a change in the protein sequence (S534N). Data regarding the clinical relevance are ambiguous. The polymorphism has been detected together with an intronic mutation and both polymorphisms have consistently been reported with reduced enzyme activity. CONCLUSION: The present case provides further evidence of an etiologic role of the c.1601G>A mutation for DPD deficiency and the occurrence of severe 5-FU-related toxicity and underlines the value of comprehensive pharmakogenetic diagnostics with respect to the dihydropyrimidine dehydrogenase.

Longitudinal variation in O(6)-alkylguanine DNA-alkyltransferase activity in the human colon and rectum.

In a systematic study of O(6)-alkylguanine DNA-alkyltransferase activity in the human colon and rectum, tumours were found to occur in regions of low activity. These results are consistent with the hypothesis that O(6)-alkylguanine DNA-alkyltransferase levels and alkylating agent exposure may be important determinants of large bowel tumorigenesis.

Enhanced expression of cyclooxygenase-2 and nuclear beta-catenin are related to mutations in the APC gene in human colorectal cancer.

Mutational inactivation of the human tumour suppressor gene adenomatous polyposis coli (APC) results in constitutive activation of beta-catenin/T cell factor-4 (Tcf-4) mediated transcription of target genes. Up-regulation of cyclooxygenase-2 (COX-2) protein is frequently found in human colorectal cancer (CRC). We analysed 38 CRC for mutations in APC and beta-catenin and found an association between APC mutations and elevated COX-2 levels. Furthermore, APC mutations were predominantly observed in tumour tissues from the rectum compared to tumours of colonic origin. Western blot analysis revealed that nuclear beta-catenin levels were generally higher in tumours with APC mutations compared to tumours with wild type APC. However, there was also a higher level of nuclear beta-catenin in tumour compared to normal tissue, but nuclear Tcf-4 protein was constitutively expressed in tumour and normal tissue and showed no differences. An identified putative Tcf-4 binding element in the COX-2 promoter may partly explain the enhanced level of COX-2 and support the idea that COX-2 may be a downstream target of the APC/beta-catenin/Tcf-4 pathway.

Flaxseed and lignans increase cecal beta-glucuronidase activity in rats.

Flaxseed has been shown in previous studies to decrease some early markers of colon cancer risk in part because of its lignans. This study determined whether the intake of flaxseed and lignans is related to the activity of bacterial beta-glucuronidase, an enzyme suggested to increase colon cancer risk. Seven groups of six female rats each were fed, for four weeks, a basal high-fat (20%) diet (BD), BD supplemented with 2.5%, 5.0%, or 10.0% flaxseed, or BD with daily gavage of 0.75, 1.5, or 3.0 mg of secoisolariciresinol diglycoside (SDG), the major mammalian lignan precursor. The specific and total activities of beta-glucuronidase in the cecum were significantly related to the levels of flaxseed (r = 0.539, p < 0.008 and r = 0.599, p < 0.002, respectively) and SDG (r = 0.567, p < 0.007 and r = 0.435, p < 0.04, respectively). The urinary mammalian lignan excretion also increased with increasing flaxseed or SDG levels and thus was significantly related to the specific activity (r = 0.38, p < 0.017) and total activity (r = 0.429, p < 0.007) of beta-glucuronidase. Because flaxseed and lignans are colon cancer protective, it is concluded that, in contrast to other studies, beta-glucuronidase activity may play a beneficial role in their presence by increasing mammalian lignan absorption and enterohepatic circulation.

Histochemical demonstration of alkaline phosphatase in human large intestine, normal and diseased.

Alkaline phosphatase activity has been investigated by histochemical methods in normal and diseased human large intestine. The tissues were constantly maintained at 4 degrees C or below. Specimens were either frozen in liquid nitrogen, freeze-dried and embedded in glycol methacrylate for sectioning at 2 mu, or, fixed in ice-cold formol-calcium for frozen sectioning at 10 mu. The simultaneous coupling azo dye method using the substrates sodium alpha-naphthyl phosphate and Naphthol AS-BI phosphate, resulted in the demonstration of alkaline phosphatase activity in the surface epithelial cells, and the middle and upper crypts, of normal and transitional mucosa.

Nuclear DNA polymerases of human carcinomas.

Comparisons have been made of the DNA polymerases of normal human lung and cecum, primary carcinomas of human lung, breast, and cecum, and resting and regeneration rat liver. The picture for the normal human tissues is similar to the one for unstimulated rat liver, that for the human carcinomas resembles regenerating rat liver. The human tissues contain two polymerases with sedimentation coefficients of about 3 and 7 S, the enzymes are restricted to the nucleus, and the specific activities of the 7 S polymerase, but not of the 3 S enzyme, are elevated in the cancers. Just as with the regenerating rat liver polymerases, the 3 S activity of a bronchogenic carcinoma is unaffected by cytosine arabinoside 5'-triphosphate and only little reduced by novobiocin, whereas DNA synthesis by the 7 S enzyme is abolished by both compounds. A variety of other inhibitory agents have similar effects on the 7 S polymerases of the human carcinomas and regenerating rat liver.