Makio Shozu

The human CYP19 (aromatase P450) gene: update on physiologic roles and genomic organization of promoters ☆

The human CYP19 (P450arom) gene is located in the chromosome 15q21.2 region and is comprised of a 30 kb coding region and a 93 kb regulatory region. The Internet-based Human Genome Project data enabled us to elucidate its complex organization. The unusually large regulatory region contains 10 tissue-specific promoters that are alternatively used in various cell types. Each promoter is regulated by a distinct set of regulatory sequences in DNA and transcription factors that bind to these specific sequences. In most mammals, P450arom expression is under the control of gonad- and brain-specific promoters. In the human, however, there are at least eight additional promoters that seemed to have been recruited throughout the evolution possibly via alterations in DNA. One of the key mechanisms that permit the recruitment of such a large number of promoters seems to be the extremely promiscuous nature of the common splice acceptor site, since activation of each promoter gives rise splicing of an untranslated first exon onto this common junction immediately upstream of the translation start site in the coding region. These partially tissue-specific promoters are used in the gonads, bone, brain, vascular tissue, adipose tissue, skin, fetal liver and placenta for physiologic estrogen biosynthesis. The most recently characterized promoter (I.7) was cloned by analyzing P450arom mRNA in breast cancer tissue. This TATA-less promoter accounts for the transcription of 29-54% of P450arom mRNAs in breast cancer tissues and contains endothelial-type cis-acting elements that interact with endothelial-type transcription factors, e.g. GATA-2. We hypothesize that this promoter may upregulate aromatase expression in vascular endothelial cells. The in vivo cellular distribution and physiologic roles of promoter I.7 in healthy tissues, however, are not known. The gonads use the proximally located promoter II. The normal breast adipose tissue, on the other hand, maintains low levels of aromatase expression primarily via promoter I.4 that lies 73 kb upstream of the common coding region. Promoters I.3 and II are used only minimally in normal breast adipose tissue. Promoters II and I.3 activities in the breast cancer, however, are strikingly increased. Additionally, the endothelial-type promoter I.7 is also upregulated in breast cancer. Thus, it appears that the prototype estrogen-dependent malignancy breast cancer takes advantage of four promoters (II, I.3, I.7 and I.4) for aromatase expression. The sum of P450arom mRNA species arising from these four promoters markedly increase total P450arom mRNA levels in breast cancer compared with the normal breast.

High Aromatase Expression in Uterine Leiomyoma Tissues of African-American Women

CONTEXT Symptomatic uterine leiomyoma is associated with irregular uterine bleeding, anemia, and recurrent pregnancy loss. African-American women develop uterine leiomyomas at an earlier age and with higher frequency compared with Caucasian-American women or other races; however, the underlying mechanism for this discrepancy is unknown. OBJECTIVE Our objective was to determine whether gene targets of emerging leiomyoma therapeutics such as aromatase inhibitors and antiprogestins, which reduce tumor size and symptoms, are differentially expressed in tissues of African-American (n = 31), Caucasian-American (n = 34), and Japanese women (n = 36). RESULTS We found strikingly higher aromatase mRNA levels in leiomyoma compared with adjacent myometrium in African-American (83 fold), Caucasian-American (38 fold), and Japanese women (33 fold). Among the four major promoters that regulate aromatase expression in leiomyoma, the proximal promoter II accounted for higher aromatase mRNA levels in tissues from African-American women. Estrogen receptor subtype alpha mRNA levels were significantly, and 1.8- to 2.6-fold, higher in leiomyoma compared with adjacent myometrium in all groups, whereas leiomyoma estrogen receptor subtype beta mRNA levels were significantly elevated only in Japanese women. Leiomyoma progesterone receptor mRNA levels were significantly higher in Japanese women compared with African-American or Caucasian-American women. CONCLUSIONS Leiomyoma tissues from African-American women contained the highest level of aromatase expression, which may result in elevated tissue concentrations of estrogen, and account for the higher prevalence and earlier incidence. Analysis of leiomyoma tissue for biomarkers may predict the response to hormonal treatments such as aromatase inhibitors.

Successful treatment of a symptomatic uterine leiomyoma in a perimenopausal woman with a nonsteroidal aromatase inhibitor.

OBJECTIVE To assess the management of symptomatic leiomyomas using a nonsteroidal aromatase inhibitor in perimenopausal women. DESIGN Case report. SETTING Academic clinical practice. PATIENT(S) A 53-year-old woman suffering from recurrent urinary retention secondary to a uterine leiomyoma. INTERVENTION(S) Fadrozole, orally, 2 mg daily for 8 weeks and then 1 mg daily for 4 weeks. MAIN OUTCOME MEASURE(S) Measurements of leiomyoma volume, and levels of serum E(2), LH, and FSH. RESULT(S) Urinary retention resolved after 2 weeks of treatment and did not recur. Leiomyoma volume estimated by ultrasonography revealed a 71% reduction after 8 weeks of treatment. CONCLUSION(S) Fadrozole was useful for the management of a symptomatic leiomyoma without transient deterioration of symptoms. Clinical trials are warranted.

Temporary amniotic membrane patching for acute chemical burns

AbstractPurpose To describe the surgical technique, and its usefulness, of temporary amniotic membrane patching (AMP) in the acute phase of ocular chemical injury.Methods Temporary AMP with modification in suture placement was performed on five eyes of five consecutive patients inflicted with acute chemical injury having a greater than grade II injury by the Roper-Hall classification.Results All patients reported herein presented with a large epithelial defect on the cornea and conjunctiva. Case 3 was classified as grade III while the other four cases were classified as grade II. The causative chemical agents were anhydrous acetic acid in Case 1, calcium oxide in Case 2, sodium hydroxide in Case 3, sodium silicate in Case 4, and sulphuric acid in Case 5. All cases experienced rapid relief of pain after AMP. Epithelialization of the cornea with improvement of visual acuity was observed in all cases when the amniotic membrane was removed within 2 weeks after surgery. During the mean follow-up of 19.6 months, the ocular surface remained stable and no cicatricial complications were noted.Conclusions These results suggest that immediate AMP is quite useful for managing moderately severe acute ocular chemical injury by facilitating rapid epithelialization and pain relief, and securing ocular surface integrity.

Tumor suppressive microRNA-218 inhibits cancer cell migration and invasion by targeting focal adhesion pathways in cervical squamous cell carcinoma

Cervical cancer is one of the most common cancers in women. More than 275,100 women die from cervical cancer each year. Cervical squamous cell carcinoma (cervical SCC), one of the most frequent types of cervical cancers, is associated with high-risk human papilloma virus (HPV), although HPV infection alone may not be enough to induce malignant transformation. MicroRNAs (miRNAs), a class of small non-coding RNAs, regulate protein-coding gene expression by repressing translation or cleaving RNA transcripts in a sequence-specific manner. A growing body of evidence suggests that miRNAs contribute to cervical SCC progression, development and metastasis. miRNA expression signatures in SCC (hypopharyngeal SCC and esophageal SCC) revealed that miR-218 expression was significantly reduced in cancer tissues compared with adjacent non-cancerous epithelium, suggesting that miR-218 is a candidate tumor suppressor. The aim of this study was to investigate the functional significance of miR-218 in cervical SCC and to identify novel miR-218-mediated cancer pathways in cervical SCC. Restoration of miR-218 significantly inhibited cancer cell migration and invasion in both HPV-positive and HPV-negative cervical SCC cell lines. These data indicated that miR-218 acts as a tumor suppressor in cervical SCC. Our in silico analysis showed that miR-218 appeared to be an important modulator of tumor cell processes through suppression of many targets, particularly those involved in focal adhesion signaling pathways. Gene expression data indicated that LAMB3, a laminin protein known to influence cell differentiation, migration, adhesion, proliferation and survival, was upregulated in cervical SCC clinical specimens, and silencing studies demonstrated that LAMB3 functioned as an oncogene in cervical SCC. The identification of novel tumor-suppressive miR-218-mediated molecular pathways has provided new insights into cervical SCC oncogenesis and metastasis.

Effects of metformin on endometrial cancer cell growth in vivo: A preoperative prospective trial

Metformin, an antidiabetic drug, decreases the incidence of various cancers in diabetic patients. Metformin‐induced inhibition of cancer cell proliferation has been confirmed in vitro but not in humans. Because endometrial cancer is associated with insulin resistance, the authors investigated whether a diabetes‐therapeutic metformin dose inhibits cancer cell growth in patients with endometrial cancer.

Aromatase Expression in Stromal Cells of Endometrioid Endometrial Cancer Correlates with Poor Survival

Purpose and Experimental Design: To assess the prognostic significance of intratumoral aromatase in endometrioid endometrial cancer, sections from 55 patients with endometrial cancer were evaluated for expression of aromatase using immunohistochemistry, and the correlation between aromatase expression and clinicopathologic parameters were analyzed. Results: Immunohistochemical staining for aromatase was positive for 32 (58%), 20 (36%), and 19 (34%) patients in cancer epithelial cells, stromal cells, and myometrial cells around the flank invasion, respectively. In situ hybridization also detected aromatase mRNA in all three types of cells. RT-PCR analysis revealed that aromatase mRNA was 2.5 ± 1.0 amol/μg total RNA (mean ± SE; n = 7) in tumor tissue. Western blot analysis detected the expected aromatase protein size of 58 kDa in cancer tissues more abundantly than in cancer-free endometrium (n = 3). The immunoreactivity in stromal cells correlated positively with advanced surgical stage and poor survival. Survival analysis revealed that the immunoreactivity of stromal cells was a significant prognostic factor, independent of histologic grade, muscular invasion, and lymph node metastasis, but dependent on surgical stage. By contrast, the immunoreactivity of aromatase both in cancer epithelial cells and myometrial cells did not correlate with prognosis. Conclusions: To the best of our knowledge, this is the first evidence associating intratumoral aromatase expression in stromal cells and poor survival in endometrioid endometrial cancer. This positive linkage indicates that local expression of aromatase plays a role in tumor progression through the formation of in situ estrogens. In situ expression of aromatase may offer a potential target for management of endometrial cancers.

Serum YKL-40 as a marker for cervical adenocarcinoma

BACKGROUND The current study examined the clinical usefulness of YKL-40 in detection and prognosis of uterine cervical cancer. PATIENTS AND METHODS Serum levels of YKL-40, cancer antigen 125 (CA 125), carbohydrate antigen 19-9 (CA19-9), and squamous cell carcinoma (SCC) antigen were determined by enzyme-linked immunosorbent assay in women with benign gynecologic disease (n=24), cervical malignancy (SCC, n=104; adenocarcinoma, n=37), and age-matched healthy controls (n=45). Immunohistochemical analysis for local YKL-40 expression was carried out on 28 adenocarcinomas. RESULTS Receiver operating characteristic curve analysis showed that YKL-40 [area under the curve (AUC)=0.882] was significantly better at discriminating adenocarcinoma from healthy control than SCC antigen, CA 125, and CA19-9. For SCC, YKL-40 (AUC=0.898) carried out similarly to SCC antigen and was better than CA 125 and CA19-9. Using a cut-off YKL-40 value of 92.2 ng/ml, sensitivity of YKL-40 in stage I adenocarcinoma (68%) was higher than that of the other three markers (11%-21%). Tumor-associated macrophages showed immunoreactivity for YKL-40 in 2 of 28 adenocarcinoma tissue samples, but adenocarcinoma cells themselves were nonimmunoreactive in all samples. Multivariate Cox regression analysis revealed that elevated pretreatment YKL-40 levels predicted unfavorable prognosis, independent of International Federation of Gynecology and Obstetrics stage and age at diagnosis. CONCLUSIONS Pretreatment serum YKL-40 level is a possible prognosticator of cervical adenocarcinoma.

Early diagnosis and risk factors for lymphedema following lymph node dissection for gynecologic cancer.

Background: Although early diagnosis is important for selecting an effective surgical treatment for secondary lymphedema, an efficient screening test for detecting early-stage lymphedema has not yet been established. Serial changes of lymphatic function before and after lymph node dissection and risk factors for secondary lymphedema are important indicators. Methods: A prospective cohort observational study was conducted with 100 consecutive gynecologic cancer patients who underwent pelvic lymph node dissection. Lymphatic function was assessed by noninvasive lymphography using indocyanine green fluorescence imaging on a routine schedule. Earliest findings after lymphadenectomy and risk factors for lower leg lymphedema were investigated. Results: Atypical transient dermal backflow patterns were observed in an early postoperative period in 50 cases, all of which disappeared within 3 months. Of these patterns, the splash pattern was observed in 31 patients, of which five improved to normal following a natural course. In contrast, the stardust pattern was observed in 27 patients, and none had improved with conservative therapy. Postoperative radiotherapy was a significant risk factor for the stardust pattern. Conclusions: All patients who undergo lymphadenectomy for gynecologic malignancies should be examined for secondary lower extremity lymphedema by qualitative evaluation methods on a routine schedule to determine the earliest possible diagnosis. Because the splash pattern on indocyanine green lymphography is a reversible lymphatic disorder following a natural course, surgical treatments are not recommended. The decision regarding surgical treatment can be made after observing the stardust pattern. CLINICAL QUESTION/LEVEL OF EVIDENCE: Diagnostic, IV.

The risk of post-molar gestational trophoblastic neoplasia is higher in heterozygous than in homozygous complete hydatidiform moles

BACKGROUND Complete hydatidiform mole (CHM) is a high-risk pregnancy for gestational trophoblastic neoplasia (GTN). Patients with CHM have a 10-30% chance of trophoblastic sequelae. CHM includes androgenic homozygous (monospermic) and androgenic heterozygous (dispermic) moles. It is controversial whether the risk of GTN is higher with heterozygous than with homozygous CHM. A prospective cohort study was conducted to assess risk of GTN in homozygous and heterozygous CHM using short tandem repeat (STR) polymorphisms, and a meta-analysis of previous reports. METHODS Twenty-eight consecutive molar pregnancies were evacuated and followed by regular hCG measurements to detect GTN. Persistent GTN was diagnosed according to the International Federation of Gynecology and Obstetrics 2000 system. Cytogenesis of the mole was determined by STR polymorphisms of molar tissue and parental blood. A meta-analysis of the GTN rate from previous reports was conducted using Mantel-Haenszel methods. RESULTS Of 28 molar pregnancies, 24 were homozygous and three were heterozygous CHM. The remaining mole was diandric triploidy (a partial hydatidiform mole). Of the 24 homozygous CHMs, six (25%) cases developed GTN and received chemotherapy. Meanwhile, all three cases (100%) of heterozygous mole developed GTN and needed chemotherapy. The GTN risk was higher in heterozygous (P = 0.029, Fishers exact test) than homozygous moles. A systematic review revealed only five previous reports (with more than 15 cytogenetically diagnosed cases), and the pooled relative risk of persistent GTN for heterozygous mole was not significant (odds ratio, 2.0; 95% confidence interval, 0.98-4.07). CONCLUSIONS Heterozygous CHM had a higher risk for GTN than homozygous CHM.