Jyh-Cherng Yu

Mechanisms of inactivation of E-cadherin in breast carcinoma: modification of the two-hit hypothesis of tumor suppressor gene.

Loss of heterozygosity (LOH) allows the expression of recessive mutation in tumor suppressor genes (TSG). Therefore, on the basis of Knudsons ‘two-hit’ hypothesis for TSG inactivation, the detection of a high LOH frequency in a chromosomal region is considered critical for TSG localization. One of these LOH regions in breast cancer is 16q22.1, which has been suggested to reflect the involvement of E-cadherin (E-cad), a cell–cell adhesion molecule. To confirm the tumorigenic role of E-cad, 81 sporadic invasive ductal carcinomas (IDCs) of the breast were tested for the ‘two hits’ required to inactivate this gene. A high frequency (37.3%) of LOH was detected in 67 informative tumors, but no mutation was found. To examine the possibility that transcriptional mechanisms serve as the second hit in tumors with LOH, specific pathways, including genetic variant and hypermethylation at the promoter region and abnormal expression of positive (WT1) and negative (Snail) transcription factors, were identified. Of these, promoter hypermethylation and increased expression of Snail were found to be common (>35%), and to be strongly associated with reduced/negative E-cad expression (P<0.05). However, unexpectedly, a significantly negative association was found between the existence of LOH and promoter hypermethylation (P<0.05), which contradicts the ‘two-hit’ model. Instead, since they coexisted in a high frequency of tumors, hypermethylation may work in concert with increased Snail to inactivate E-cad expression. Given that E-cad is involved in diverse mechanisms, loss of which is beneficial for tumors to invade but may also trigger apoptosis, this study suggests that maintaining a reversible mechanism, either by controlling the gene at the transcriptional level or by retaining an intact allele subsequent to LOH, might be important for E-cad in IDC and may also be common in TSGs possessing diverse functions. These findings provide clues to explain why certain TSGs identified by LOH cannot fulfil the two-hit hypothesis.

Small Hepatocellular Carcinoma—A Clinicopathological Study in Thirteen Patients

Abstract To investigate the diagnosis and clinicopathological features of hepatocellular carcinoma at its early stage, 13 patients with carcinoma ≤3.0 × 3.0 cm in size (small hepatocellular carcinoma) were studied with radionuclide scan, celiac arteriography, computed tomography, ultrasonography, and peritoneoscopy. Most of the patients were found from prospective studies in hepatoma high-risk subjects and were asymptomatic, with liver tests showing only mild abnormalities. Hepatitis B surface antigen was positive in all except one who was hepatitis B core antibody positive. Serum α-fetoprotein level was normal in 3 patients and in the remaining patients it increased to an extent far lower than in patients with advanced hepatocellular carcinoma. For detecting small tumors, ultrasonography, computed tomography, and celiac arteriography were superior to radionuclide scan or peritoneoscopy (detection rate: 95%, 94%, 89% vs. 16%, 12%, respectively). Small hepatocellular carcinoma was characterized as a hypoechoic shadow with weak internal echoes on ultrasonography, hypodense lesion on computed tomography, and hypervascular stain in angiography. None of these three examinations detected all the tumors in every patient and thus at least two of the three examinations should be done to avoid missing any tumor. Among them, the real-time ultrasonography was most practical for localizing small hepatocellular carcinoma and the diagnosis could be confirmed by ultrasound-guided biopsy. Ultrasonography was indispensable for identifying the small tumor during laparotomy in 3 patients. Ten patients had a single tumor. The small tumors were resected successfully in 10 patients. The tumor was frequently surrounded by a thin fibrous capsule and the most common cell type was of the trabecular pattern.

Glycan microarray of Globo H and related structures for quantitative analysis of breast cancer

Cancer-associated carbohydrate antigens are often found on the surface of cancer cells. Understanding their roles in cancer progression will lead to the development of new therapeutics and high-sensitivity diagnostics for cancers. Globo H is a member of this family, which is highly expressed on breast cancer cells. Here, we report the development of a glycan microarray of Globo H and its analogs for measurement of the dissociation constants on surface (KD,surf) with three different monoclonal antibodies (VK-9, Mbr1, and anti-SSEA-3), to deduce their binding specificity. The glycan microarray was also used to detect the amount of antibodies present in the plasma of breast cancer patients and normal blood donors. It was shown that the amount of antibodies against Globo H from breast cancer patients were significantly higher than normal blood donors, providing a new tool for possible breast cancer diagnosis. Compared with the traditional ELISA method, this array method required only atto-mole amounts of materials and is more effective and more sensitive (5 orders of magnitude). The glycan microarray thus provides a new platform for use to monitor the immune response to carbohydrate epitopes after vaccine therapy or during the course of cancer progression.

Breast cancer risk associated with genotype polymorphism of the catechol estrogen-metabolizing genes: a multigenic study on cancer susceptibility.

Estrogen has been suggested to trigger breast cancer development via an initiating mechanism involving its metabolite, catechol estrogen (CE). To examine this hypothesis, we carried out a multigenic case‐control study of 469 incident breast cancer patients and 740 healthy controls to define the role of important genes involved in the different metabolic steps that protect against the potentially harmful effects of CE metabolism. We studied the 3 genes involved in CE detoxification by conjugation reactions involving methylation (catechol‐O‐methyltransferase, COMT), sulfation (sulfotransferase 1A1, SULT1A1), or glucuronidation (UDP‐glucuronosyltransferase 1A1, UGT1A1), one (manganese superoxide dismutase, MnSOD) involved in protection against reactive oxidative species‐mediated oxidation during the conversion of CE‐semiquinone (CE‐SQ) to CE‐quinone (CE‐Q), and 2 of the glutathione S‐transferase superfamily, GSTM1 and GSTT1, involved in CE‐Q metabolism. Support for this hypothesis came from the observations that (i) there was a trend toward an increased risk of breast cancer in women harboring a greater number of putative high‐risk genotypes of these genes (p < 0.05); (ii) this association was stronger and more significant in those women who were more susceptible to estrogen [no history of pregnancy or older (≥26 years) at first full‐term pregnancy (FFTP)]; and (iii) the risks associated with having one or more high‐risk genotypes were not the same in women having experienced different menarche‐to‐FFTP intervals, being more significant in women having been exposed to estrogen for a longer period (≥12 years) before FFTP. Furthermore, because CE‐Q can attack DNA, leading to the formation of double‐strand breaks (DSB), we examined whether the relationship between cancer risk and the genotypic polymorphism of CE‐metabolizing genes was modified by the genotypes of DSB repair genes, and found that a joint effect of CE‐metabolizing genes and one of the two DSB repair pathways, the homologous recombination pathway, was significantly associated with breast cancer development. Based on comprehensive CE metabolizing gene profiles, our study provides support to the hypotheses that breast cancer can be initiated by estrogen exposure and that increased estrogen exposure confers a higher risk of breast cancer by causing DSB to DNA.

Anthropometric measures, plasma adiponectin, and breast cancer risk

Adiponectin is a peptide hormone secreted exclusively by adipocytes, and obesity is an established risk factor for breast cancer. We have, thus, evaluated the associations of anthropometric measures of adiposity and adiponectin with the development of breast cancer in a case-control study. Questionnaire information, anthropometric measures, and blood samples were taken before treatment from 244 incident cases with breast cancer, including 141 premenopausal and 103 postmenopausal cases, and 244 controls admitted for health examination at the Tri-Service General Hospital, Taipei between 2004 and 2005. Plasma levels of adiponectin were measured by RIA. The relationship between anthropometric measures of adiposity and breast cancer risk was modified by menopausal status, with a significant increase in risk observed in postmenopausal but not premenopausal women. Moreover, a fairly robust inverse association of adiponectin with the risk was observed only in postmenopausal women (adjusted odds ratio (OR), 0.55; 95% confidence interval (CI), 0.23-0.97), but not in premenopausal women. Additionally, the plasma adiponectin levels tended to be inversely associated with estrogen receptor (ER)-positive (adjusted OR, 0.53; 95% CI, 0.27-0.98) but not ER-negative breast tumors. Furthermore, the associations of adiponectin with breast cancer risk overall and by menopausal and ER status remained after adjustment for obesity indices. These results suggest that adiponectin may have an independent role in breast carcinogenesis, particularly in the postmenopausal and ER-positive breast cancer risk.

Breast cancer risk associated with genotypic polymorphism of the mitosis-regulating gene Aurora-A/STK15/BTAK

Aneuploidy, an abnormal number of chromosomes, is relatively common and occurs early in breast cancer development. This observation supports a breast tumorigenic contribution of mechanisms responsible for maintaining chromosome number stability in which centrosomes play an essential role. We therefore speculated that the Aurora‐A/STK15/BTAK gene, implicated in the regulation of centrosome duplication, may be associated with breast tumorigenesis. To test this hypothesis, we conducted a case‐control study of 709 primary breast cancer patients and 1,972 healthy controls, examining single‐nucleotide polymorphisms (SNPs), including a suggested functional Phe31Ile SNP, in Aurora‐A. We were also interested in knowing whether any association between Aurora‐A and breast cancer was modified by reproductive risk factors reflecting susceptibility to estrogen exposure. Our hypothesis is that, since estrogen is known to promote breast cancer development via its mitogenic effect leading to malignant proliferation on breast epithelium and since Aurora‐A is involved in regulating mitosis, the discovery of a joint effect between the Aurora‐A genotype and reproductive risk factors on cancer risk might yield valuable clues to the association of breast tumorigenesis with estrogen. Support for this hypothesis came from the following observations. (i) Two SNPs in Aurora‐A were significantly associated with breast cancer risk (p < 0.05). (ii) Haplotype analyses, based on different combinations of multiple SNPs in Aurora‐A, revealed a strong association with breast cancer risk; interestingly, the genotypic distribution of the suggested functional Phe31Ile SNP was not significantly different between breast cancer patients and controls, but the specific haplotype containing the putative at‐risk Ile allele was more common in patients. (iii) This association between risk and putative high‐risk genotypes was stronger and more significant in women thought to be more susceptible to estrogen, i.e., those with a longer interval between menarche and first full‐term pregnancy. (iv) The protective effect conferred by a history of full‐term pregnancy was significant only in women with a putative low‐risk genotype of Aurora‐A. Our study provides new findings supporting the mutator role of Aurora‐A in breast cancer development, suggesting that breast cancer could be driven by genomic instability associated with variant Aurora‐A, the tumorigenic contribution of which could be enhanced as a result of increased mitosis due to estrogen exposure.

Breast Cancer Risk Is Associated with the Genes Encoding the DNA Double-Strand Break Repair Mre11/Rad50/Nbs1 Complex

The evolutionarily conserved Mre11-Rad50-Nbs1 (MRN) complex, consisting of proteins encoded by the genes Mre11, Rad50, and Nbs1, was recently shown to play a crucial role in DNA double-strand break (DSB) repair by recruiting the nuclear protein kinase ataxia telangiectasia mutated to DSB sites, leading to activation of this DNA repair network. Given the fact that carriers of defective mutation and polymorphic variants of ataxia telangiectasia mutated are at higher risk of developing breast cancer, we hypothesized a role of the MRN genes in determining breast cancer susceptibility. This hypothesis was examined both in a case control study of 559 breast cancer patients and 1,125 healthy women of single-nucleotide polymorphisms in Mre11, Rad50, and Nbs1 and by the in vivo detection of binding between Mre11 and BRCA1, encoded by the breast cancer susceptibility gene BRCA1. We were also interested in defining whether any association between MRN genes and breast cancer was modified by reproductive risk factors reflecting the level of estrogen exposure or susceptibility to estrogen exposure, as estrogen is known to initiate breast cancer development due to its metabolites causing DSB formation. Support for the hypothesis came from the observations that (a) one single-nucleotide polymorphism in Nbs1 was significantly associated with breast cancer risk, and a trend toward an increased risk of developing breast cancer was found in women harboring a greater number of putative high-risk genotypes of MRN genes (an adjusted odds ratio of 1.25 for each additional putative high-risk genotype; 95% confidence interval, 1.10-1.44); (b) this association between risk and the number of putative high-risk genotypes was stronger and more significant in women thought to be more susceptible to estrogen, i.e., those with no history of full-term pregnancy, those older (≥26 years of age) at first full-term pregnancy, or those having had fewer (<2) full-term pregnancies; the risk effect conferred by harboring a higher number of high-risk genotypes of MRN genes was more significant in women without a history of breast feeding; and (c) Mre11 and BRCA1 were shown to form a complex in vivo, and this interaction was increased by irradiation. This study supports the role of the MRN pathway in breast cancer development, further strengthening the suggestion that mechanisms regulating DSB repair may play a mutator role driving breast cancer pathogenesis. (Cancer Epidemiol Biomarkers Prev 2007;6(10):2024–32)

Thyroidectomy for Hashimoto's Thyroiditis: Complications and Associated Cancers

BACKGROUND Hashimotos thyroiditis is usually treated medically; however, thyroidectomy is sometimes indicated. Thyroiditis can make thyroid dissection more difficult and possibly increase the risk of surgical complications. The aim of this study was to determine the rate of complications and associated cancer in patients with Hashimotos thyroiditis. METHODS Retrospective series of 474 patients treated surgically at the University of California, San Francisco, between January 1985 and June 2005 with final pathology demonstrating Hashimotos thyroiditis, chronic lymphocytic thyroiditis, or chronic thyroiditis. Parameters evaluated included demographics, surgical indications, and postoperative complications. RESULTS Among the 474 patients, 133 had thyroidectomy because of preoperative diagnosis of thyroid cancers (median age 39 years; 116 females and 17 males), 316 had thyroidectomy because of benign thyroid nodules or goiter (median age 47.5 years; 292 females and 24 males), and 25 had thyroidectomy to relieve local symptoms caused by thyroiditis but did not have thyroid nodules (median age 42 years; 25 females). No death or permanent surgical complications occurred. One hundred and fifty-two patients (32.1%) had transient postoperative hypocalcemia, 2 (0.4%) had transient recurrent nerve palsy, and 4 (0.8%) had a postoperative neck hematoma. Fifty-three percent had thyroid cancer at final histological examination. CONCLUSIONS Thyroidectomy can be performed in patients with Hashimotos thyroiditis with a low risk of permanent surgical complications. Cancer is common in patients who have a thyroidectomy for Hashimotos thyroiditis even when not suspected preoperatively.

Allelic loss of the BRCA1 and BRCA2 genes and other regions on 17q and 13q in breast cancer among women from Taiwan (area of low incidence but early onset)

We have examined the role of the breast cancer susceptibility genes BRCA1 and BRCA2 and other loci in the vicinity of these 2 genes on the long arms of chromosomes 17 and 13 (17q and 13q) for the presence of genomic deletions in breast cancer among Taiwanese women. Breast cancer in Taiwan is particularly characterized by its low incidence rate and its early age of tumor onset. Twelve microsatellite markers spanning the region 17q12– 21 and 8 microsatellite markers spanning the region 13q12– 14 were analyzed for allelic loss or loss of heterozygosity (LOH) in 90 patients with primary infiltrating ductal carcinoma. Compared with the background LOH level (10– 12%) estimated by LOH at 4 unrelated loci, 17 markers (11 at 17q and 6 at 13q) demonstrated a significantly increased frequency (21– 42%) of allelic loss (p < 0.05). Subsequent construction of deletion maps based on LOH at these significant loci localized the 6 smallest regions of overlap, including those harboring BRCA1, BRCA2, the retinoblastoma gene and 3 novel regions (the 1st located approximately 0.5 to 1 cM telomeric to BRCA1, the 2nd centromeric to BRCA1 flanked by D17S857 /D17S846 and the 3rd closely adjacent to BRCA2 ), suggesting sites of susceptibility genes. Allelic loss at BRCA1 and BRCA2 was specifically associated with poorly differentiated tumors. Int. J. Cancer (Pred. Oncol.) 79:580–587, 1998.

Genetic variants of BLM interact with RAD51 to increase breast cancer susceptibility

The role of the familial breast cancer susceptibility genes, BRCA1 and BRCA2, in the homologous recombination (HR) pathway for DNA double-strand break (DSB) repair suggests that the mechanisms involved in HR and DNA DSB repair are of etiological importance during breast tumorigenesis. Bloom (BLM) helicase directly interacts with RAD51 recombinase, which is involved in regulating HR, and it is thus of particular interest to examine whether this interaction is associated with breast cancer susceptibility. This single-nucleotide polymorphism (SNP)-based case-control study was performed to examine this hypothesis using specimens from 933 patients with breast cancer and 1539 healthy controls. The results showed that one SNP (rs2380165) in BLM and two (rs2412546 and rs4417527) in RAD51 were associated with breast cancer risk. Furthermore, haplotype and diplotype analyses based on combinations of five SNPs in RAD51 revealed a strong association between RAD51 polymorphisms and breast cancer risk (P < 0.05). Support for the interaction between BLM and RAD51 in determining breast cancer risk came from the finding that the association between cancer risk and at-risk genotypes/haplotype pairs of RAD51 was stronger and more significant in women harboring homozygous variant alleles of BLM (P for interaction < 0.05). Interestingly, not only the intronic SNP located within the region encoding the helicase domain of BLM but also those within the RAD51-interaction domain-encoding region showed an interaction with RAD51 polymorphisms in determining breast cancer susceptibility. Our results suggest a contribution of BLM and RAD51 to breast cancer development and provide support for the tumorigenic significance of the functional interaction between these two HR proteins.