Effectiveness of prophylactic HPV vaccines against cervical abnormalities and HPV infection in Japan: The J-HERS 2021 multicenter study.

This study investigated the efficacy of the prophylactic human papillomavirus (HPV) vaccine, which was initiated between 2009 and 2013 in Japan. The study involved 1529 eligible women aged 16-39 years who visited 11 outpatient clinics in Japan for various reasons. These patients underwent HPV genotype analysis and a Pap test of cervical cell samples. A total of 299 women (19.6%) had received the prophylactic HPV vaccine (bivalent:quadrivalent vaccine ratio = 2:1). Of the 5062 participants in the Japanese Human Papillomavirus Disease Education and Research Survey (J-HERS 2011), which was conducted in the pre-vaccination era, 3236 eligible participants were included as controls. In this study (J-HERS 2021), the highest rate of HPV vaccination (53%) was observed in patients aged 22-27 years. Vaccinated individuals exhibited a 49% rate of protection against low-grade intraepithelial lesions (LSILs) and atypical squamous cells, not excluding high-grade squamous intraepithelial lesions (ASCH) or worse (LSIL/ASCH+), and a 100% rate of protection against high-grade squamous intraepithelial lesions (HSILs) or worse (HSIL+). Significant reductions in HPV16 (95%) and HPV18 (100%) infections were noted, but no differences were observed in HPV6 and HPV11 infections. The prevalences of HPV51 and HPV59 increased with vaccination, although these changes were not confirmed in the comparative study with J-HERS 2011. Comparing the prevaccination (J-HERS 2011) and postvaccination (J-HERS 2021) periods, 43%, 51%, 88%, and 62% reductions in HPV16, HPV18, HPV16/18, and HPV31/58 infection rates were observed, respectively. Similarly, 62% and 71% reductions in LSIL/ASCH+ and HSIL+ rates were noted, respectively. There were 88% and 87% reductions in LSIL/ASCH+ and HSIL+ rates in 16-21- and 28-33-year-old patients, respectively. Bivalent or quadrivalent vaccines provided 100% protection against high-grade squamous cell lesions (suggestive of CIN2 or CIN3) in young women aged <39 years at 9-12 years after initiation of Japan's first nationwide HPV vaccination program. Cross-protection against HPV31 and HPV58 is likely to occur, although some HPV-type replacements are inconsistent across vaccination regimens. This demonstrates the effectiveness of the HPV vaccine. However, continuous monitoring of cervical cancer and precancer is necessary in younger generations (born 1997-2007), who were rarely vaccinated due to the prolonged suspension of the vaccine recommendations in Japan.

Comparison of Aptima and hybrid capture-2 HPV tests and Pap test in the referral population in Japan.

The Aptima human papillomavirus (HPV) test (APTIMA) detects E6-E7 mRNA in abnormal cells in the uterine cervix. To investigate the accuracy of APTIMA for cervical cancer screening in Japan, 423 subjects, mostly referrals with abnormal cytology or being followed up for cervical intraepithelial neoplasia (CIN)1, were screened using two HPV tests, hybrid capture 2 (HC2) and APTIMA, and by the Pap test. Colposcopy was conducted in all subjects with a positive result in either test type. HPV genotyping was performed by Genosearch-31. A result of atypical squamous cells-undetermined significance (ASC-US) or worse on the HC2 test (ASC-US-HC2), and low-grade squamous intraepithelial lesion (LSIL) or worse (LSIL+) on the Pap test, was regarded as positive. APTIMA (97.5%) was more sensitive than LSIL+ (85.1%) for detecting CIN2 or worse (CIN2+) (McNemar test; p = .0003), and more sensitive (98.6%) than ASC-US-HC2 (92.7%) for detecting CIN3+. APTIMA and HC2 had similar sensitivities. HPV genotyping revealed that CIN2/3 with high-risk HPV (HR-HPV) was overlooked in five cases by ASC-US-HC2, and in four cases by HC2, while no such lesions were missed by APTIMA. Thus, APTIMA might be superior to HC2 for primary HPV screening in Japan. One cancer case positive for HPV67 (potentially high risk, [pHR]) was overlooked by Pap test and both HPV tests, suggesting a need for a new HPV test able to detect pHR-HPV types.

The prevalence of VAIN, CIN, and related HPV genotypes in Japanese women with abnormal cytology.

Vaginal intraepithelial neoplasia (VAIN) is often found by chance. We investigated the prevalence of VAIN and related human papillomavirus (HPV) types in comparison with cervical intraepithelial neoplasia (CIN). This study enrolled 648 women who were referred to the outpatient clinic of Kanazawa Medical University Hospital for abnormal cytology from January 2009 to January 2019. HPV genotypes were determined using Genosearch-31 + 4, which can detect 35 different HPV types. Colposcopy was performed at the first visit by an experienced gynecological oncologist. Among 611 subjects with squamous cell lesions, 107 (17.5%) VAIN cases were identified, and 67 (11.0%) women had both VAIN and CIN. Ultimately, 72 VAIN1, 15 VAIN2/3, 203 CIN1, 249 CIN2/3, 32 cervical squamous cell carcinomas (SCC), and one vaginal SCC (Vag-SCC) were identified. The prevalences of VAIN1, VAIN2/3, and Vag-SCC were 35.5%, 6.0%, and 3.1% of equivalent cervical lesions, respectively. The VAIN patients were older than the CIN patients (P = .002). About half of the VAIN cases were diagnosed during the follow-up. Multiple HPV infections were found in 42.9% of the VAIN and CIN patients. HPV52, 16, 51, 53, and 56 were the most common types in VAIN, whereas HPV16, 52, 58, 51, and 31 predominated in CIN. HPV18 was rare in VAIN, HPV58 was more common in CIN than in VAIN, and HPV53 and HPV73 were more common in VAIN. In conclusion, VAIN1 was identified more frequently than we expected. Various HPV types were identified in the vagina, which is likely a reservoir for HPV.

Detection of circulating tumor cells in cervical cancer using a conditionally replicative adenovirus targeting telomerase-positive cells.

Circulating tumor cells (CTC) are newly discovered biomarkers of cancers. Although many systems detect CTC, a gold standard has not yet been established. We analyzed CTC in uterine cervical cancer patients using an advanced version of conditionally replicative adenovirus targeting telomerase-positive cells, which was enabled to infect coxsackievirus-adenovirus receptor-negative cells and to reduce false-positive signals in myeloid cells. Blood samples from cervical cancer patients were hemolyzed and infected with the virus and then labeled with fluorescent anti-CD45 and anti-pan cytokeratin antibodies. GFP (+)/CD45 (-) cells were isolated and subjected to whole-genome amplification followed by polymerase chain reaction analysis of human papillomavirus (HPV) DNA. CTC were detected in 6 of 23 patients with cervical cancers (26.0%). Expression of CTC did not correlate with the stage of cancer or other clinicopathological factors. In 5 of the 6 CTC-positive cases, the same subtype of HPV DNA as that of the corresponding primary lesion was detected, indicating that the CTC originated from HPV-infected cancer cells. These CTC were all negative for cytokeratins. The CTC detected by our system were genetically confirmed. CTC derived from uterine cervical cancers had lost epithelial characteristics, indicating that epithelial marker-dependent systems do not have the capacity to detect these cells in cervical cancer patients.

Comparison of the digene hybrid capture 2 and Roche cobas 4800 HPV tests for detection of CIN2+ in a referral population in Japan.

To examine validity of the hybrid capture-2 and cobas 4800 HPV tests, 396 women including 188 women visiting for cancer screening, and 208 referral cases were examined with both HPV tests and the liquid-based cervical Pap test. Concordant results between the HPV assays were observed in 333 cases (coincident rates; 84.1%, kappa value; 0.682). The sensitivity for CIN2+ was 98.6% (69/70) and 82.9% (58/70) for HC2 and cobas 4800 (McNemar's test; P = 0.0026). The sensitivity for CIN3+ was 97.2% (35/36) and 83.3% (30/36) (Not significant, P = 0.0736). The specificities for CIN2+ or CIN3+ did not differ between the tests. The HPV16, 52, 18, 31, and 58 were the most common types in CIN2+ cases. Reasonable sensitivity for HPV52, and cross-hybridization with some probable high-risk HPV type such as HPV82 explain the higher sensitivity of HC2 than cobas 4800 in detection of CIN2+ in a referral population in Japan.

Synchronous endometrioid adenocarcinomas in the uterine cervix and corpus.

It is frequently difficult to distinguish multiple primary carcinomas from single primary carcinoma with metastasis. Here, we report a case of synchronous endometrioid adenocarcinomas that independently occurred in the uterine cervix and corpus. A 47-year-old woman complaining of genital bleeding was preoperatively diagnosed with cervical adenocarcinoma with an endometrial lesion. On surgical treatment, two separate malignant lesions bearing endometrioid adenocarcinoma were identified in the uterine cervix and cavity. Although both lesions expressed the same type of human papillomavirus (HPV) gene, type 16, microscopic continuity was not observed. Furthermore, we detected a critical difference in PTEN mutation between the tumors and finally diagnosed this case as multiple primary cancers. This is the first report to show multiple primary endometrioid adenocarcinomas simultaneously arising in the uterine cervix and corpus. Considering the rarity of this case, the coexistence of HPV suggests its possible involvement in the carcinogenesis of the endometrioid adenocarcinomas.

Risk factors of heavy uterine bleeding in patients with endometriosis and adenomyosis treated with dienogest.

OBJECTIVE: Dienogest (DNG), a fourth-generation progestin, reduces pain associated with endometriosis and uterine adenomyosis; however, it is associated with irregular uterine bleeding that can cause anemia and poor quality of life. We investigated risk factors for heavy bleeding following DNG administration. MATERIALS AND METHODS: We retrospectively investigated patients who received DNG for risk factors of heavy uterine bleeding, including clinical diagnosis, use of pretreatment gonadotropin-releasing hormone agonist, smoking, cancer antigen 125, and blood hormone levels. We additionally assessed the uterine area in patients with uterine adenomyosis, the major axis of the uterine body, the major axis of myometrial thickness, the site of tumor development, and the site of myoma development in patients with uterine fibroids. RESULTS: Eighty Japanese patients were administered DNG. The median age was 41 (range: 24-51) years. The odds ratio (OR) for moderate-to-severe bleeding according to clinical diagnosis were 0.33 (P = 0.011) for endometrioma and 9.00 (P = 0.049) for uterine adenomyosis. Receiver operating characteristic curve analysis of the uterine area associated with uterine adenomyosis showed an area under the curve (AUC) of 0.909 between those with major and minor bleeding, with an optimal cut-off value of 7388.2 mm2. The uterine body major axis had an AUC of 0.946, with an optimal cut-off value of 78.3 mm. The major axis of myometrial thickness had an AUC of 0.855, with an optimal cut-off value of 46.8 mm. CONCLUSION: Patients with endometrioma treated with DNG were less likely to experience heavy uterine bleeding. Uterine bleeding in patients with uterine adenomyosis and adenomyosis associated with uterine fibroids should be closely monitored while administering DNG.

Chemical Peeling Therapy Using Phenol for the Cervico-Vaginal Intraepithelial Neoplasia.

Objective: This study aimed to validate the use of liquid phenol-based chemical peeling therapy for cervical and vaginal intraepithelial neoplasia (CIN and VaIN, respectively), with the goal of circumventing obstetric complications associated with surgical treatment and to determine the factors associated with treatment resistance. Methods: A total of 483 eligible women diagnosed with CIN, VaIN, or both, participated in this study. Participants underwent phenol-based chemical peeling therapy every 4 weeks until disease clearance. Disease clearance was determined by negative Pap tests for four consecutive weeks or by colposcopy. HPV genotyping was conducted at the onset of the study and after disease clearance in select cases. Our preliminary analysis compared the recurrence and persistence rates between 294 individuals who received phenol-based chemical peeling therapy and 189 untreated patients. Results: At 2 years following diagnosis, persistent disease was observed in 18%, 60%, and 88% of untreated patients with CIN1-3, respectively, and <2% of patients with CIN who received phenol-based chemical peeling therapy. Among 483 participants, 10 immune-suppressed patients required multiple treatments to achieve disease clearance, and 7 were diagnosed with cervical cancer. Of the 466 participants, except those with cancer or immune suppression, the number of treatment sessions until CIN/VaIN clearance ranged from 2 to 42 (average: 9.2 sessions). In total, 43 participants (9.2%) underwent surgical treatment. Six patients (1.3%) experienced recurrence of CIN2 or worse, suggesting that treatment failed in 46 patients (9.9%). No obstetrical complications were noted among the 98 pregnancies following this therapy. Factors associated with resistance to this therapy include immune suppression, ages 35-39 years, higher-grade lesions, and multiple HPV-type infections. Conclusions: Phenol-based therapy is safe and effective for CINs and VaINs. Women aged < 35 years and with persistent CIN1 or CIN2 with a single HPV-type infection are suitable candidates for phenol-based chemical peeling therapy. However, this therapy requires multiple lengthy sessions.

[Associations between Maternal Postpartum Depression and Psychosocial Factors Including Marital Relationship and Social Support].

OBJECTIVES: We aimed to investigate the psychosocial factors for postpartum depression as indicated by a high score of the Edinburgh Postnatal Depression Scale (EPDS), including marital relationship and social support. Relevant factors for antenatal depression were also analyzed. METHODS: Thirty-five wife-and-husband pairs who visited University Hospital A for the wife's antenatal health check-up participated in a questionnaire survey using the Japanese version of the EPDS. Social support from the wife's husband, kins, and others including friends at the third trimester of pregnancy and 1 month after birth was assessed. The Marital Love Scale (MLS) was also used, and two marital relationship questions were asked regarding the husband's and wife's considerate actions toward each other during pregnancy. Binary logistic regression analysis was conducted to determine adjusted associations between higher EPDS scores (≥5 for postpartum depression and ≥7 for antenatal depression) and indicators for social support and marital relationships. RESULTS: The most relevant factor for higher postpartum EPDS scores was a higher antenatal EPDS score, followed by the couple's poor communication skills (the wife did not feel any appreciation from her husband) during pregnancy and no support from the wife's husband during the postpartum period. The wife's poor marital communication skills and the husband's low MLS scores during pregnancy were associated (borderline significance) with the wife's higher antenatal EPDS scores. CONCLUSIONS: A good marital relationship before birth and support by the husband after birth may be important for preventing postpartum depression.

Massive hemothorax due to diaphragmatic endometriosis after a laparoscopic cystectomy of an ovarian endometrioma in a patient without a history of thoracic endometriosis.

BACKGROUND: Severe hemothorax is a rare complication after laparoscopic surgery for endometriosis, and the causes and proper management are not well understood. CASE: We report here the extremely rare case with massive hemothorax after laparoscopic surgery for ovarian endometrioma. A 40-year-old woman, gravida 1, para 1, underwent laparoscopic cystectomy of ovarian endometrioma. On postoperative day 2, she had progressive anemia (Hb 5.3) as well as dyspnea. A chest X-ray and computed tomography showed massive fluid collection in the right thoracic cavity, suggestive of intrapleural bleeding. TREATMENT: Thoracoscopic operation was performed and a total of 930 ml of blood retention in the right thoracic cavity was found. Scattered small endometriotic lesions were present on the pleural surface of the right diaphragm; pulsatile active bleeding was confirmed from one of these. Furthermore, two endometriotic lesions had perforated into the intraperitoneal cavity. The diaphragm containing bleeding spots was thoracoscopically resected and sutured. After thoracoscopic surgery, the dyspnea and anemia resolved. On postoperative day 5, the patient left the hospital. CONCLUSION: The present report reminds us of the importance of paying special attention to postoperative-thoracic complications caused by diaphragmatic endometriosis if the patient shows respiratory symptoms.

Clinical Landscape of PARP Inhibitors in Ovarian Cancer: Molecular Mechanisms and Clues to Overcome Resistance.

The survival of patients with advanced or recurrent ovarian cancer has improved tremendously in the past decade, mainly due to the establishment of maintenance therapy with poly (ADP-ribose) polymerase (PARP) inhibitors (PARPis) after conservative chemotherapies. Despite their superior efficacy, resistance to PARPis has been reported, and patients with resistance have a much worse prognosis. Therefore, the development of novel treatment strategies to overcome PARPi resistance is urgently needed. The present review article focuses on the molecular mechanisms of how PARPis exert cytotoxic effects on cancer cells through DNA repair processes, especially the genetic background and tumor microenvironment favored by PARPis. Furthermore, currently available information on PARPi resistance mechanisms is introduced and discussed to develop a novel therapeutic approach against them.

A retrospective study of immunotherapy using the cell wall skeleton of Mycobacterium bovis Bacillus Calmette-Guérin (BCG-CWS) for cervical cancer.

Mycobacterium bovis Bacillus Calmette-Guérin (BCG) has the potential to promote adaptive immunity. We sought to examine the synergistic effect of BCG-CWS vaccination on cervical cancer patients undergoing standard treatments including surgery, chemotherapy, and/or radiation. We retrospectively analyzed 103 patients (13 cases administered with BCG-CWS vaccine and 90 controls without BCG-CWS) who underwent a standard treatment for cervical cancer from 2005 to 2021. The BCG-CWS group underwent repeated intradermal injections of the BCG-CWS vaccine before or immediately after the standard therapy start from 2011 to 2018. The vaccination was repeated weekly for 1 month, and then every 4 weeks thereafter. The effectiveness of the BCG-CWS vaccination on cervical cancer treatment was evaluated by determining the hazard ratios of overall survival between the BCG-CWS group and the control group with multivariate analysis using the Cox model. Hazard ratios between 2 groups were determined after adjustment by clinical parameters including surgery, chemotherapy, radiation, age, clinical stage, presence of human papillomavirus, and pathology. Long-term follow-up revealed a significantly better prognosis (hazard ratio: 0.2108, P = .008 by the Cox model) for patients with cervical cancer in the BCG-CWS group compared to patients in the control group. Among patients with advanced cancer worse than stage IB2, some completely cleared the disease, whereas the others showed long-term survival with recurrence. BCG-CWS therapy appears to be an effective immune adjuvant therapy for cervical cancer, although randomized control studies are needed to confirm this. We also need to clarify the underlying mechanisms slowing the progression of cervical cancer in those receiving this vaccination. This study sheds light on the potential of immunostimulatory drugs such as BCG-CWS and suggests the important role of immunity for cancer elimination in combination therapy.

Determination of human papillomavirus type in archival tissue specimens of invasive cervical cancer using molecular mapping and E6/E7-based polymerase chain reaction.

In our previous study, an L1-based human papillomavirus (HPV) test using liquid-based cytology revealed that some invasive cervical cancers (ICC) exhibited multiple HPV types or harbored no HPV DNA. Here, molecular mapping of formalin-fixed paraffin-embedded cancer tissue specimens from the same patients were conducted to confirm these observations. Among 377 ICC cases, 73 eligible specimens (9 positive for multiple HPV types, 16 negative for HPV, and 48 positive for a single HPV type from the previous study) were reexamined by manual microdissection of cancer lesions, then subjected to HPV genotyping using the uniplex E6/E7 polymerase-chain-reaction method to detect all high-risk and potentially high-risk HPV types. The HPV typing results were confirmed in 52 of 73 cancer cases; among the 21 remaining cases, 15 were discordant and 6 were partially concordant. In total, 8 of 16 (50%) HPV-negative samples became positive; 6 were positive for HPV16 and 2 were positive for HPV67. Moreover, two samples previously positive for HPV6 and HPV53 were negative for HPV. All nine cancers with multiple HPV types were found to harbor only a single HPV type. In total, 63 cancer tissues exhibited a single HPV type. HPV16 and HPV18 were detected in squamous cell carcinoma (SCC) and adenocarcinoma (ADC). Alpha-5 (HPV82), -6 (HPV56), and -9 (HPV31/52/67) HPV types were detected in SCC, whereas Alpha-7 (HPV59/68) types were detected in ADC and adenosquamous carcinoma (ADSCC). These findings suggested that the different HPV types induced different histological cancers. Furthermore, all SCCs and 10 of 11 usual-type ADCs were positive for high-risk HPV types, supporting the use of HPV screening for the detection of these cancers and associated premalignant lesions. HPV16 is likely to remain undetected in some cervical cancer tissues because of low viral-copy-numbers. Putative high-risk HPV types (e.g., HPV67 and HPV82) might be high risk in Japan.

A Novel Liquid Biopsy Strategy to Detect Small Amounts of Cancer Cells Using Cancer-Specific Replication Adenoviruses.

Circulating tumor cells (CTCs) are a promising source of clinical and biological cancer information and can be a material for liquid biopsy. However, detecting and capturing these cells remains a challenge. Various biological factors (e.g., cell surface proteins, cell size, deformability, or dielectrophoresis) have been applied to detect CTCs. Cancer cells dramatically change their characteristics during tumorigenesis and metastasis. Hence, defining a cell as malignant using such a parameter is difficult. Moreover, immortality is an essential characteristic of cancer cells. Telomerase elongates telomeres and plays a critical role in cellular immortality and is specifically activated in cancer cells. Thus, the activation of telomerase can be a good fingerprint for cancer cells. Telomerase cannot be recognized by antibodies in living cells because it is a nuclear enzyme. Therefore, telomerase-specific replication adenovirus, which expresses the green fluorescent protein, has been applied to detect CTCs. This review explores the overview of this novel technology and its application in gynecological cancers.

Diagnostic potential and limitation of imaging cancer cells in cytological samples using telomerase-specific replicative adenovirus.

Cytological cancer screening that targets genetic or epigenetic abnormalities may be a viable alternative to morphological screening. Detecting cancer cells by specific genetic markers helps their easy detection in cytological samples. We recently established the telomerase-specific replication-selective adenovirus OBP-401, in which the human telomerase reverse transcriptase (hTERT) gene promoter has been inserted upstream of the E1 genes, and in which the green fluorescent protein (GFP) gene is driven by the CMV promoter. This virus selectively replicates only in telomerase-positive cells, expressing GFP, and therefore may be a tool for cancer screening. In the present study, we first confirmed that cytological samples can easily be infected with OBP-401, allowing visualization of GFP-positive cells under fluorescent microscopy 24 h after infection. After 32 cytological samples from patients with cervical, endometrial or ovarian cancers were infected with OBP-401, GFP signals were detected in 31 (96%) of the samples. However, some normal endometrial scrapings exhibited GFP-signals, possibly due to endometrial glandular cells with constitutive telomerase activity. The ability of OBP-401 to enrich cancer cells was then tested. Cytological samples containing cervical or endometrial cancer cells were infected with OBP-401, and GFP-positive cells were sorted by flow cytometry; DNA was extracted from the GFP-positive cells. Direct DNA sequencing or methylation-specific PCR identified cancer-derived mutations or hypermethylations of tumor suppressor genes more efficiently than analyses using crude cytological samples. Thus, OBP-401-based sorting of GFP-positive cells successfully enriched cancer cells, allowing efficient detection of genetic or epigenetic abnormalities in cytological samples.

Bone marrow-derived cells from male donors can compose endometrial glands in female transplant recipients.

OBJECTIVE: For continuous regeneration of human endometrium in menstrual cycles, endometrial stem cells are assumed to supply differentiating endometrial glandular cells. To elucidate the origin of endometrial stem cells, we examined the presence of donor-derived cells in endometria from patients who received bone marrow transplantation from male donors. STUDY DESIGN: Endometrial specimens biopsied after hormone replacement therapy were obtained and examined using fluorescent in situ hybridization analysis targeting X or Y chromosomes. RESULTS: All recipients had donor-derived Y chromosome-positive endometrial cells, accounting for 0.6-8.4% of glandular epithelial cells and 8.2-9.8% of stromal cells. Most of the endometrial glands were chimeric, consisting of both donor-derived and recipient cells. CONCLUSION: Donor-derived cells are capable of composing endometrium in recipients, even those of the opposite sex. These results suggest unexpected plasticity of bone marrow stem cells as well as a potential origin of endometrial stem cells.

Experience and efficacy of a bipolar vessel sealing system for radical abdominal hysterectomy.

The present study evaluated the efficacy of an electrosurgical bipolar vessel sealing system for radical abdominal hysterectomy (RAH). A total of 85 consecutive patients who underwent RAH with pelvic lymphadenectomy for cervical cancer or endometrial cancer were analyzed; in 18 patients, the LigaSure system (Valleylab, Boulder, Colo) was used to secure the vascular pedicles, and in the remaining 67 patients, clamps were used, and the pedicles suture ligated. Compared with the conventional suture group, the patients in the LigaSure group had a significantly shorter operation time (mean, LigaSure 242.8 +/- 36.1 minutes vs conventional, 349.1 +/- 82.6) minutes; P < 0.001) and lower blood loss (mean, 583.1 +/- 287.6 mL vs 999.0 +/- 524.2 mL; P < 0.005). Only 1 (0.06%) of the 18 patients in the LigaSure group was transfused, whereas 27 (40.2%) of the 67 patients in the conventional suture group were transfused. Because transfusion requirements may be affected by the surgeon's bias, we compared blood loss among untransfused patients and found that the LigaSure group still had significantly lower blood loss than the conventional suture group (mean, 550.9 +/- 233.1 mL vs 745.49.0 +/- 230.4 mL; P < 0.01). Hemoglobin level reduction after surgery in untransfused patients was significantly lower in the LigaSure group than in the conventional suture group (mean, 2.31 +/- 2.22 mg/dL vs 3.22 +/- 1.11 mg/dL; P < 0.05). These findings indicate that the LigaSure vessel sealing system is useful to reduce blood loss and shorten operating time at RAH.

Understanding and exploiting hTERT promoter regulation for diagnosis and treatment of human cancers.

Telomerase activation is a critical step for human carcinogenesis through the maintenance of telomeres, but the activation mechanism during carcinogenesis remains unclear. Transcriptional regulation of the human telomerase reverse transcriptase (hTERT) gene is the major mechanism for cancer-specific activation of telomerase, and a number of factors have been identified to directly or indirectly regulate the hTERT promoter, including cellular transcriptional activators (c-Myc, Sp1, HIF-1, AP2, ER, Ets, etc.) as well as the repressors, most of which comprise tumor suppressor gene products, such as p53, WT1, and Menin. Nevertheless, none of them can clearly account for the cancer specificity of hTERT expression. The chromatin structure via the DNA methylation or modulation of nucleosome histones has recently been suggested to be important for regulation of the hTERT promoter. DNA unmethylation or histone methylation around the transcription start site of the hTERT promoter triggers the recruitment of histone acetyltransferase (HAT) activity, allowing hTERT transcription. These facts prompted us to apply these regulatory mechanisms to cancer diagnostics and therapeutics. Telomerase-specific replicative adenovirus (Telomelysin, OBP-301), in which E1A and E1B genes are driven by the hTERT promoter, has been developed as an oncolytic virus that replicates specifically in cancer cells and causes cell death via viral toxicity. Direct administration of Telomelysin was proved to effectively eradicate solid tumors in vivo, without apparent adverse effects. Clinical trials using Telomelysin for cancer patients with progressive stages are currently ongoing. Furthermore, we incorporated green fluorescent protein gene (GFP) into Telomelysin (TelomeScan, OBP-401). Administration of TelomeScan into the primary tumor enabled the visualization of cancer cells under the cooled charged-coupled device (CCD) camera, not only in primary tumors but also the metastatic foci. This technology can be applied to intraoperative imaging of metastatic lymphnodes. Thus, we found novel tools for cancer diagnostics and therapeutics by utilizing the hTERT promoter.

Role of menin in the regulation of telomerase activity in normal and cancer cells.

Transcriptional activation of human telomerase reverse transcriptase (hTERT) is critical for telomerase expression, a major step for cellular immortality and carcinogenesis. Although several transcriptional activators have been identified, factors responsible for repressing the hTERT promoter are largely unknown. Gene screening that employed enhanced retroviral mutagenesis has identified potential hTERT repressors. Among these, menin, which is a tumor suppressor and a gene product of MEN1, has been reported to play a critical role. In the present study, we further analyzed menin's role in the transcriptional regulation of hTERT in normal and cancer cells. Luciferase reporter assays that use the hTERT promoter have demonstrated that an overexpression of menin decreases the transcriptional activity of the hTERT gene in a cell-type specific manner. Mutation and deletion analyses of the hTERT promoter demonstrated that there was no specific site on the promoter that was responsible for the menin-mediated transcriptional inhibition. An electrophoretic mobility shift assay using recombinant menin protein generated the binding complexes with the hTERT promoter, which was completely diminished by the addition of poly-dI-dC. This indicates that there is a sequence-independent binding of menin. RT-PCR assays have revealed that overexpression of menin inhibits hTERT mRNA expression in some cell types, although this inhibition does not lead to a significant down-regulation of telomerase activity. In cancer cell lines and in normal cells, the siRNA-based inhibition of MEN1 does not lead to the up-regulation of hTERT mRNA expression. No significant correlation has been found between menin and hTERT mRNA expressions in a variety of cancer cell lines and clinical tissue samples. Thus, while menin appears to have some inhibitory effects on the hTERT promoter, possibly via the sequence-independent binding to the promoter, the present study does not support the hypothesis that menin has a crucial role in the determination of telomerase activity in normal and cancer cells.