Transcription factor ZNF488 accelerates cervical cancer progression through regulating the MEK/ERK signaling pathway.

Cervical cancer (CC) is one of the most common gynecological malignancies worldwide. Zinc Finger Protein 488 (ZNF488) has been identified as an oncogene in nasopharyngeal carcinoma. However, its biological role and potential mechanism in CC remain to be elucidated. In the present study, upregulation of ZNF488 expression in human CC tissues was found in clinical samples and analyzed in The Cancer Genome Atlas (TCGA) dataset, which was associated with clinical staging and lymph node metastasis. Quantitative real time polymerase chain reaction (PCR) and western blot assays indicated that the expression of ZNF488 was up-regulated in CC cells. Cell colony formation and cell cycle analysis assays suggested that ZNF488 promoted CC cell proliferation and cycle progression. Knockdown of ZNF488 inhibited tumor growth of xenograft tumor mice in vivo, in agreement with the levels of ZNF488 and Ki-67. Moreover, transwell and western assays demonstrated that ZNF488 enhanced CC cell migration and invasion. Additionally, knockdown of ZNF488 also inhibited lung metastasis of CC cells in vivo. Further mechanism analysis implied that ZNF488 promoted the MEK/ERK signaling pathway. ERK inhibitor PD98059 significantly weakened the proliferation and epithelial-mesenchymal transformation (EMT) promotion effect of ZNF488. Collectively, ZNF488 exerts its oncogene function partially through modulating MEK/ERK signaling pathway in CC, indicating that ZNF488 may provide a promising therapeutic target for the treatment of CC.

[Distribution characteristics and risk factors of colorectal adenomas].

OBJECTIVE: To determine the detection rate and distribution characteristics of colorectal adenomas in Ningbo area of China, and to identify the risk factors for colorectal adenoma, in order to provide reference for colorectal cancer screening. METHODS: A cross-sectional study was performed among 8660 subjects undergoing colonoscopy in the Ningbo No.2 Hospital between January and December 2016, using a questionnaire, including demographic data (age, gender, height and weight), history of diseases (diabetes, hypertension, hyperlipidemia, and family history of malignant neoplasm), lifestyle (smoking, alcohol, dietary bias on red meat, dietary bias on fruit and vegetables, dietary frequency of pickled food and physical activities), and intestinal early warning symptoms. All colonoscopically detected polyps were removed for histological examination. Polyps were histologically divided into non-adenomatous (hyperplastic polyps and inflammatory polyps) and adenomatous polyps (tubular, villous, tubulovillous and serrated adenomas). Pathologic features were analyzed according to anatomical site. Multivariate logistic regression analysis was used to identify the risk factors for colorectal adenoma. RESULTS: A total of 7077 subjects who received colonoscopic examination and completed the questionnaire survey were enrolled in this study. There were 3633 males and 3444 females with a median age of 53 (ranged 17 to 83) years. Adenoma detection rate was 15.6% (1103/7077) in all cases, 21.0%(762/3633) for males, and 9.9%(341/3444) for females(P=0.000). Detection rate of 6.2%(29/469) was recorded in individuals aged less than 30 years, 8.0%(87/1086) in those from 30 to 39 years, 12.1%(148/1222) in those from 40 to 49 years, 16.8%(272/1623) in those from 50 to 59 years, 20.4%(326/1601) in those from 60 to 69 years, and 22.4%(241/1076) in those ≥70 years. The detection rate increased according to age(P=0.000). A total of 1521 adenomas were detected in 1103 cases, including 1455 tubular adenomas, 33 tubulovillous adenomas, 9 villous adenomas and 24 serrated adenomas. Among 1521 adenomas, 44.1%(n=671) located in the right hemicolon, 39.0%(n=593) in the left hemicolon, and 16.9%(n=257) in the rectum. Significantly larger number of serrated adenomas and advanced adenomas (advanced adenoma was defined as any adenoma with high-grade intraepithelial neoplasia, diameter ≥10 mm or with villous component) was observed in the right hemicolon compared to left hemicolon and rectum [serrated adenomas: 2.5%(17/671) vs. 0.8% (5/593) and 0.8% (2/257), P=0.029; advanced adenoma: 9.2% (62/671) vs. 5.2% (31/953) and 6.6% (17/257), P=0.021]. Multivariate analysis showed that malely (P=0.003), elderly (P=0.000), obesity (P=0.014), smoking (P=0.001), alcohol (P=0.032), and family history of malignancy (P=0.000) were independent risk factors of colorectal adenoma. CONCLUSIONS: In view of a higher detection rate of colorectal adenoma in population aged 40 to 49 years especially in male individuals, the starting age of colonoscopy screening may be advanced to 40 years old. People with family history of malignancy, obesity, and habit of smoking or drinking should be regarded as important subjects for colonoscopy screening. During colonoscopy screening, special emphasis should be given to right hemicolon.

FGFR3b Extracellular Loop Mutation Lacks Tumorigenicity In Vivo but Collaborates with p53/pRB Deficiency to Induce High-grade Papillary Urothelial Carcinoma.

Missense mutations of fibroblast growth factor receptor 3 (FGFR3) occur in up to 80% of low-grade papillary urothelial carcinoma of the bladder (LGP-UCB) suggesting that these mutations are tumor drivers, although direct experimental evidence is lacking. Here we show that forced expression of FGFR3b-S249C, the most prevalent FGFR3 mutation in human LGP-UCB, in cultured urothelial cells resulted in slightly reduced surface translocation than wild-type FGFR3b, but nearly twice as much proliferation. When we expressed a mouse equivalent of this mutant (FGFR3b-S243C) in urothelia of adult transgenic mice in a tissue-specific and inducible manner, we observed significant activation of AKT and MAPK pathways. This was, however, not accompanied by urothelial proliferation or tumorigenesis over 12 months, due to compensatory tumor barriers in p16-pRB and p19-p53-p21 axes. Indeed, expressing FGFR3b-S249C in cultured human urothelial cells expressing SV40T, which functionally inactivates pRB/p53, markedly accelerated proliferation and cell-cycle progression. Furthermore, expressing FGFR3b-S243C in transgenic mouse urothelium expressing SV40T converted carcinoma-in-situ to high-grade papillary urothelial carcinoma. Together, our study provides new experimental evidence indicating that the FGFR3 mutations have very limited urothelial tumorigenicity and that these mutations must collaborate with other genetic events to drive urothelial tumorigenesis.

Role of isoenzyme M2 of pyruvate kinase in urothelial tumorigenesis.

The conversion of precancerous lesions to full-fledged cancers requires the affected cells to surpass certain rate-limiting steps. We recently showed that activation of HRAS proto-oncogene in urothelial cells of transgenic mice causes simple urothelial hyperplasia (SUH) which is persistent and whose transition to low-grade papillary urothelial carcinoma (UC) must undergo nodular urothelial hyperplasia (NUH). We hypothesized that NUH, which has acquired fibrovascular cores, plays critical roles in mesenchymal-to-epithelial signaling, breaching the barriers of urothelial tumor initiation. Using proteomics involving two-dimensional gel electrophoresis, immunoblotting with pan-phosphotyrosine antibody and MALDI-mass spectrometry, we identified isoform 2 of pyruvate kinase (PKM2) as the major tyrosine-phosphorylated protein switched on during NUH. We extended this finding using specimens from transgenic mice, human UC and UC cell lines, establishing that PKM2, but not its spliced variant PKM1, was over-expressed in low-grade and, more prominently, high-grade UC. In muscle-invasive UC, PKM2 was co-localized with cytokeratins 5 and 14, UC progenitor markers. Specific inhibition of PKM2 by siRNA or shRNA suppressed UC cell proliferation via increased apoptosis, autophagy and unfolded protein response. These results strongly suggest that PKM2 plays an important role in the genesis of low-grade non-invasive and high-grade invasive urothelial carcinomas.

Bilateral breast cancer following augmentation mammaplasty with polyacrylamide hydrogel injection: A case report.

Breast cancer occurring following injection with polyacrylamide hydrogel (PAMG) for augmentation mammaplasty is rare. The present study reports the case of a 43-year-old female presenting with bilateral breast cancer 10 years after augmentation mammaplasty with PAMG injection and no family history of breast cancer. A 5.5×6.0-cm mass in the right breast with multiple intumescent axillary lymph nodes was revealed and a palpable mass of ~1.0 cm was identified in the outer upper quadrant of the left breast. Multiple smaller nodules were observed in the pulmonary field. Pathological examination revealed invasive lobular grade II carcinoma in both breasts with ER(+++), PR(+++), C-erbB2(-), Top-2(+), in the right breast and ER(++), PR(++), c-erb-B2(-), Top-2(+) in the left. Preoperative chemotherapy, modified radical bilateral mastectomy with axillary clearance, postoperative chemotherapy, and an oophorectomy were conducted, followed by treatment with Arimedex® until the present date A number of valuable insights can be garnered from this case. First, close follow-up is required for female patients who receive an injection of PAMG for augmentation mammaplasty in order to achieve an early diagnosis and to intervene in any incidences of breast cancer. Second, the differential diagnosis of dual primary carcinoma versus metastatic breast cancer is important and may be aided by the use of molecular technology. Third, it remains difficult to determine gene expression values for the prediction of chemotherapy sensitivity. Thus, discrimination between primary and secondary carcinomas is the principle barrier for identifying an appropriate treatment strategy when a patient is diagnosed with bilateral breast cancer.

Comparative proteomic analysis of the function and network mechanisms of MASPIN in human lung cells.

MASPIN, which is also known as Serpin B5, is a novel tumor suppressor. Emerging evidence suggests that MASPIN acts as a multifaceted protein in various types of cancer, including prostate, breast and pancreatic cancer. It interacts with diverse groups of intercellular and extracellular proteins, regulating cell adhesion, motility, apoptosis and angiogenesis, and is involved in mammary gland development. As MASPIN is a multifunctional factor in cancer pathways, its function remains poorly illuminated. In this study, we compared the protein profiles of LC5 cell lines with MASPIN overexpression and knockdown using comparative two-dimensional gel electrophoresis. The differences in protein expression, visualized as differences in spots, were identified by time-of-flight (TOF)/TOF mass spectometry. Significant differences were observed between overexpressing and knocked down cells, including eight spots that were unique and sixteen spots that were up- or down-regulated by more than 4-fold. Six genes, including Sdccag8, Ldoc1, SCAI, SDCCAG3, CT62 and NEDD9 were unique in MASPIN-expressing cell lines, but absent in knock-out cell lines, in which most of them play a significant role in the invasion of cancer cells. Moreover, the Brms1 and CAGE1 genes were identified as being uniquely expressed in knocked down cell lines, which were associated with the development and progression of tumors. The data from this study shed some light on the function, as well as the general network mechanisms of MASPIN in lung cancer.

Urothelial tumor initiation requires deregulation of multiple signaling pathways: implications in target-based therapies.

Although formation of urothelial carcinoma of the bladder (UCB) requires multiple steps and proceeds along divergent pathways, the underlying genetic and molecular determinants for each step and pathway remain undefined. By developing transgenic mice expressing single or combinatorial genetic alterations in urothelium, we demonstrated here that overcoming oncogene-induced compensatory tumor barriers was critical for urothelial tumor initiation. Constitutively active Ha-ras (Ras*) elicited urothelial hyperplasia that was persistent and did not progress to tumors over a 10 months period. This resistance to tumorigenesis coincided with increased expression of p53 and all pRb family proteins. Expression of a Simian virus 40 T antigen (SV40T), which disables p53 and pRb family proteins, in urothelial cells expressing Ras* triggered early-onset, rapidly-growing and high-grade papillary UCB that strongly resembled the human counterpart (pTaG3). Urothelial cells expressing both Ras* and SV40T had defective G(1)/S checkpoint, elevated Ras-GTPase and hyperactivated AKT-mTOR signaling. Inhibition of the AKT-mTOR pathway with rapamycin significantly reduced the size of high-grade papillary UCB but hyperactivated mitogen-activated protein kinase (MAPK). Inhibition of AKT-mTOR, MAPK and STAT3 altogether resulted in much greater tumor reduction and longer survival than did inhibition of AKT-mTOR pathway alone. Our studies provide the first experimental evidence delineating the combinatorial genetic events required for initiating high-grade papillary UCB, a poorly defined and highly challenging clinical entity. Furthermore, they suggest that targeted therapy using a single agent such as rapamycin may not be highly effective in controlling high-grade UCB and that combination therapy employing inhibitors against multiple targets are more likely to achieve desirable therapeutic outcomes.

Temporally and spatially controllable gene expression and knockout in mouse urothelium.

Urothelium that lines almost the entire urinary tract performs important functions and is prone to assaults by urinary microbials, metabolites, and carcinogens. To improve our understanding of urothelial physiology and disease pathogenesis, we sought to develop two novel transgenic systems, one that would allow inducible and urothelium-specific gene expression, and another that would allow inducible and urothelium-specific knockout. Toward this end, we combined the ability of the mouse uroplakin II promoter (mUPII) to drive urothelium-specific gene expression with a versatile tetracycline-mediated inducible system. We found that, when constructed under the control of mUPII, only a modified, reverse tetracycline trans-activator (rtTA-M2), but not its original version (rtTA), could efficiently trans-activate reporter gene expression in mouse urothelium on doxycycline (Dox) induction. The mUPII/rtTA-M2-inducible system retained its strict urothelial specificity, had no background activity in the absence of Dox, and responded rapidly to Dox administration. Using a reporter gene whose expression was secondarily controlled by histone remodeling, we were able to identify, colocalize with 5-bromo-2-deoxyuridine incorporation, and semiquantify newly divided urothelial cells. Finally, we established that, when combined with a Cre recombinase under the control of the tetracycline operon, the mUPII-driven rtTA-M2 could inducibly inactivate any gene of interest in mouse urothelium. The establishment of these two new transgenic mouse systems enables the manipulation of gene expression and/or inactivation in adult mouse urothelium at any given time, thus minimizing potential compensatory effects due to gene overexpression or loss and allowing more accurate modeling of urothelial diseases than previously reported constitutive systems.

Clinical trial and post-licensure safety profile of a prophylactic human papillomavirus (types 6, 11, 16, and 18) l1 virus-like particle vaccine.

BACKGROUND: We describe the safety of the human papillomavirus (HPV)-6/11/16/18 vaccine using updated clinical trial data (median follow-up time of 3.6 years) and summarize up to 3 years of post-licensure surveillance. METHODS: In 5 clinical trials, 21,480 girls/women aged 9 to 26 years and boys aged 9 to 16 years received >or=1 dose of HPV-6/11/16/18 vaccine or placebo. All serious and non-serious adverse experiences (AEs) and new medical conditions were recorded for the entire study period(s). As of June 2009, >25 million doses of HPV-6/11/16/18 vaccine had been distributed in the United States with >50 million doses globally. Post-licensure safety as summarized by the Centers for Disease Control and Prevention using the United States Vaccine Adverse Event Reporting System database is also reported. RESULTS: Eight subjects experienced a treatment-related serious AE (0.05% vaccine; 0.02% placebo). Of 18 deaths (0.1% vaccine; 0.1% placebo), all were considered unrelated to study treatment. New medical conditions which were potentially consistent with autoimmune phenomena were reported in 2.4% of both vaccine and placebo recipients. Pain, the most common injection-site AE, occurred more frequently with vaccine (81% vaccine; 75% placeboaluminum; 45% placebo-saline). No differences were seen in the incidence of the most common non-serious AEs-headache and pyrexia. The Vaccine Adverse Event Reporting System has received 14,072 reports for the HPV-6/11/16/18 vaccine since licensure, with only 7% being serious AEs, about half the average reported for licensed vaccines in general. CONCLUSIONS: HPV-6/11/16/18 vaccination was associated with more injection-site pain than placebo but similar incidences of systemic and serious AEs and new medical conditions potentially consistent with autoimmune phenomena. Based on review of post-licensure safety information, the benefits of vaccination to prevent the majority of genital tract precancers and cancers continue to far outweigh its risks.

Pregnancy and infant outcomes in the clinical trials of a human papillomavirus type 6/11/16/18 vaccine: a combined analysis of five randomized controlled trials.

OBJECTIVE: To present a combined analysis of the pregnancy outcomes for women aged up to 45 years enrolled in five phase III clinical studies of the prophylactic quadrivalent human papillomavirus 6/11/16/18 vaccine. METHODS: Twenty thousand five hundred fifty-one women aged 15-45 years received quadrivalent HPV vaccine or placebo at day 1 and months 2 and 6. Urine pregnancy tests were performed immediately before each injection; participants testing positive were not vaccinated. Women who became pregnant after enrollment were discontinued from further vaccination until resolution of pregnancy. All pregnancies were followed for outcomes. RESULTS: During the studies, 1,796 vaccine and 1,824 placebo recipients became pregnant, resulting in 2,008 and 2,029 pregnancies with known outcomes. No significant differences were noted overall for the proportions of pregnancies resulting in live birth, fetal loss, or spontaneous abortion. A total of 40 neonates born to vaccinated women and 30 neonates born to women given placebo had one or more congenital anomalies (P=.20). The anomalies were diverse and consistent with those most commonly observed in the general population. The vaccine was well tolerated among women who became pregnant. CONCLUSION: Administration of quadrivalent human papillomavirus vaccine to women who became pregnant during the phase III clinical trials did not appear to negatively affect pregnancy outcomes. The vaccine is a U.S. Food and Drug Administration pregnancy category B medication (animal studies revealed no evidence of fetal harm, but there are no adequate and well-controlled studies in pregnant women); however, vaccination is not recommended during pregnancy. Postlicensure surveillance is ongoing. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, www.clinicaltrials.gov, NCT00092521, NCT00092534, NCT00092495, NCT00092547 and NCT00090220. LEVEL OF EVIDENCE: II.