Klebsiella pneumoniae liver abscess with metastatic complications is an emerging infectious disease in Taiwan. To identify genes associated with liver infection, we used a DNA microarray to compare the transcriptional profiles of three strains causing liver abscess and three strains not associated with liver infection. There were 13 clones that showed higher RNA expression levels in the three liver infection strains, and 3 of these 13 clones contained a region that was absent in MGH 78578. Sequencing of the clones revealed the replacement of 149 bp of MGH 78578 with a 21,745-bp fragment in a liver infection strain, NTUH-K2044. This 21,745-bp fragment contained 19 open reading frames, 14 of which were proven to be associated with allantoin metabolism. The K2044 (⌬allS) mutant showed a significant decrease of virulence in intragastric inoculation of BALB/c mice, and the prevalence of this chromosomal region was significantly higher in strains associated with liver abscess than in those that were not (19 or 32 versus 2 of 94; P ‫؍‬ 0.0001 [ 2 test]). Therefore, the 22-kb region may play a role in K. pneumoniae liver infection and serve as a marker for rapid identification.

Our results suggest that chromosomal heterogeneity is present in tissue-invasive K. pneumoniae strains. A genotype containing all 3 regions is strongly associated with PLA and metastatic infection. These regions may serve as convenient markers for the rapid diagnosis of emergent tissue-invasive strains.

Tamoxifen, a synthetic triphenyl-ethylene compound, is a member of a class of anticancer drugs known as selective estrogen receptor modulators. It may block tumor growth by mimicking estrogen and binding to the estrogen receptors, preventing cancerous growth. Clinical studies have demonstrated that a combination chemo/hormonal therapy regimen with tamoxifen and O 6 -alkylating drugs increased the tumor response rate in cancer patients. The mechanism of action of this combined regimen remains undefined. In this study, we demonstrated that treatment of human colorectal HT-29 carcinoma cells with tamoxifen decreased the repair activity and expression level of O 6 -methylguanine DNA methyltransferase (MGMT) protein in a concentration-and timedependent manner. This inhibition was also shown in other malignant human cells, regardless of their estrogen receptor status. Furthermore, MGMT inactivation by tamoxifen was associated with a significantly increased susceptibility of cells to 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). No alteration in MGMT mRNA levels was observed in tamoxifen-treated cells. The halflife of MGMT protein was markedly decreased in the presence of tamoxifen. Tamoxifen-induced MGMT degradation could be blocked by MG-132, a proteasome inhibitor. An increased level of ubiquitinated MGMT protein was found after tamoxifen treatment. We conclude that tamoxifen decreased the MGMT protein level by accelerating protein degradation through the ubiquitindependent proteasomal pathway. These findings provide a strong rationale for combined chemo/hormonal therapy with tamoxifen and BCNU in the treatment of human cancers. ' 2007 Wiley-Liss, Inc.Key words: tamoxifen; MGMT; ubiquitination; proteasomal pathway; BCNU Tamoxifen, a synthetic triphenyl-ethylene compound, is a member of a class of anticancer drugs known as selective estrogen receptor modulators (SERMs). It is used to treat patients with all stages of hormone-responsive breast cancer. 1 It has also been shown to prevent breast cancer in women who are at high risk for this disease, by mimicking estrogen and binding to the estrogen receptor (ER). 2 Although the primary mechanism of action of tamoxifen is believed to be through inhibition of the ER, research over the years has indicated that additional, non-ER-mediated mechanisms exist. These include modulation of signaling proteins, such as protein kinase C (PKC), 3,4 calmodulin, 3,5 transforming growth factor-b 3 and insulin-like growth factor I. 3,6 The antitumor effect of tamoxifen is thus believed to be a combination of ER-mediated and non-ER-mediated mechanisms. 5 In addition, tamoxifen has been used in the treatment of malignancies other than breast carcinomas, including hepatocellular, colorectal, ovarian, pancreatic, and renal cell carcinomas, malignant gliomas and melanomas.O 6 -methylguanine-DNA methyltransferase (MGMT), a ubiquitous DNA repair protein, is responsible for removal of alkyl adducts from the O 6 -position of guanine in a reaction that transfers the alkyl group from the DNA to an i...

O 6-Methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein that removes alkyl-adducts from the O 6 -guanine in DNA and is a crucial defense against O 6 -alkylating agentinduced cytotoxicity. We demonstrated here that two camptothecin (CPT)-resistant cell lines (CPT30 and KB100) were more sensitive to N,NЈ-bis(2-chloroethyl)-N-nitrosurea (BCNU) than their parental cells. Enhanced sensitivity to BCNU in these two CPT-resistant cells involved transcriptional repression of the MGMT gene. The mechanism of MGMT gene down-regulation in CPT-resistant cells was not through gene abnormality, mRNA stability, and CpG island hypermethylation. However, the high level of methyl-CpG-binding protein 2 (MeCP2) and dimethylation of H3K9 in the promoter region were found in CPT30 and KB100 cells. Furthermore, increased MeCP2 binding on MGMT promoter was also found to be correlated with MGMT genesilencing in short-term CPT treatment; thus, enhanced BCNU sensitivity was found in CPT-treated cells. Taken together, we suggest that CPT is able to suppress the transcription of the MGMT gene through recruiting of MeCP2 and H3K9 dimethylation, thus causing a synergistic interaction with BCNU. These findings provide a possible explanation regarding why the combination of CPT and BCNU results in a better objective response than single-use alone. In addition, this study supports a new indication for treating patients who are receiving refractory CPT derivatives with BCNU.