Ubiquitin-specific protease 22 is critical to in vivo angiogenesis, growth and metastasis of non-small cell lung cancer.

BACKGROUND: Loss of monoubiquitination of histone H2B (H2Bub1) was found to be associated with poor differentiation, cancer stemness, and enhanced malignancy of non-small cell lung cancer (NSCLC). Herein, we investigated the biological significance and therapeutic implications of ubiquitin-specific protease 22 (USP22), an H2Bub1 deubiquitinase, in non-small cell lung cancer (NSCLC). METHODS: USP22 expression and its clinical relevance were assessed in NSCLC patients. The effects of USP22 knockout on sensitivity to cisplatin and irradiation, and growth, metastasis of NSCLC xenografts, and survival of cancer-bearing mice were investigated. The underlying mechanisms of targeting USP22 were explored. RESULTS: Overexpression of USP22 was observed in 49.0% (99/202) of NSCLC tissues; higher USP22 immunostaining was found to be associated with enhanced angiogenesis and recurrence of NSCLC. Notably, USP22 knockout dramatically suppressed in vitro proliferation, colony formation; and angiogenesis, growth, metastasis of A549 and H1299 in mouse xenograft model, and significantly prolonged survival of metastatic cancer-bearing mice. Furthermore, USP22 knockout significantly impaired non-homologous DNA damage repair capacity, enhanced cisplatin and irradiation-induced apoptosis in these cells. In terms of underlying mechanisms, RNA sequencing and gene ontology enrichment analysis demonstrated that USP22 knockout significantly suppressed angiogenesis, proliferation, EMT, RAS, c-Myc pathways, concurrently enhanced oxidative phosphorylation and tight junction pathways in A549 and H1299 NSCLC cells. Immunoblot analysis confirmed that USP22 knockout upregulated E-cadherin, p16; reduced ALDH1A3, Cyclin E1, c-Myc, and attenuated activation of AKT and ERK pathways in these cells. CONCLUSIONS: Our findings suggest USP22 plays critical roles in the malignancy and progression of NSCLC and provide rationales for targeting USP22, which induces broad anti-cancer activities, as a novel therapeutic strategy for NSCLC patient.

Targeting histone methyltransferase G9a inhibits growth and Wnt signaling pathway by epigenetically regulating HP1α and APC2 gene expression in non-small cell lung cancer.

BACKGROUND: Dysregulated histone methyltransferase G9a may represent a potential cancer therapeutic target. The roles of G9a in tumorigenesis and therapeutics are not well understood in non-small cell lung cancer (NSCLC). Here we investigated the impact of G9a on tumor growth and signaling pathways in NSCLC. METHODS: Immunohistochemistry analyzed G9a expression in NSCLC tissues. Both siRNA and selective inhibitor were used to target G9a. The impact of targeting G9a on key genes, signaling pathways and growth were investigated in NSCLC cells by RNA sequencing analysis, rescue experiments, and xenograft models. RESULTS: Overexpression of G9a (≥ 5% of cancer cells showing positive staining) was found in 43.2% of 213 NSCLC tissues. Multiple tumor-associated genes including HP1α, APC2 are differentially expressed; and signaling pathways involved in cellular growth, adhesion, angiogenesis, hypoxia, apoptosis, and canonical Wnt signaling pathways are significantly altered in A549, H1299, and H1975 cells upon G9a knockdown. Additionally, targeting G9a by siRNA-mediated knockdown or by a selective G9a inhibitor UNC0638 significantly inhibited tumor growth, and dramatically suppressed Wnt signaling pathway in vitro and in vivo. Furthermore, we showed that treatment with UNC0638 restores the expression of APC2 expression in these cells through promoter demethylation. Restoring HP1α and silencing APC2 respectively attenuated the inhibitory effects on cell proliferation and Wnt signaling pathway in cancer cells in which G9a was silenced or suppressed. CONCLUSIONS: These findings demonstrate that overexpressed G9a represents a promising therapeutic target, and targeting G9a potentially suppresses growth and Wnt signaling pathway partially through down-regulating HP1α and epigenetically restoring these tumor suppressors such as APC2 that are silenced in NSCLC.

Overexpression of Flap Endonuclease 1 Correlates with Enhanced Proliferation and Poor Prognosis of Non-Small-Cell Lung Cancer.

Flap endonuclease 1 (FEN1) plays a crucial role in both DNA replication and damage repair. In this study, FEN1 expression and its clinical-pathologic significance in non-small-cell lung cancer (NSCLC) was investigated. Quantitative RT-PCR and immunohistochemistry analysis identified that both FEN1 mRNA and protein were highly overexpressed in about 36% of 136 cancer tissues compared to adjacent tissues, in which FEN1 was generally undetectable. Notably, patients with FEN1-overexpressed cancers were prone to have poor differentiation and poor prognosis. A strong positive correlation between the levels of FEN1 and Ki-67 staining was identified in these NSCLC tissues (r = 0.485), suggesting overexpressed FEN1 conferred a proliferative advantage to NSCLC. Furthermore, knockdown of FEN1 resulted in G1/S or G2/M phase cell cycle arrest and suppressed in vitro cellular proliferation in NSCLC cancer cells. Consistently, a selective FEN1 inhibitor was shown to effectively inhibit cellular proliferation of NSCLC cells in a dose-dependent manner. Additionally, knockdown of FEN1 significantly attenuated homologous DNA repair efficiency and enhanced cytotoxic effects of cisplatin in NSCLC cells. Taken together, these findings have indicated that overexpressed FEN1 represents a prognostic biomarker and potential therapeutic target for NSCLC treatment, which warrants further study.

Loss of H2B monoubiquitination is associated with poor-differentiation and enhanced malignancy of lung adenocarcinoma.

Deregulated monoubiquitination of histone H2B (H2Bub1), mainly catalyzed by E3 ubiquitin-protein ligase RNF20/RNF40 complex, may play an important role in cancer. Here we investigate potential roles of H2Bub1 and the underlying mechanisms through which it contributes to cancer development and progression in lung adenocarcinoma. We show that downregulation of H2Bub1 through RNF20 knockdown dramatically decreases H3K79 and H3K4 trimethylation in both normal and malignant lung epithelial cell lines. Concurrently, global transcriptional profiling analysis reveals that multiple tumor-associated genes such as CCND3, E2F1/2, HOXA1, Bcl2 modifying factor (BMF), Met, and Myc; and signaling pathways of cellular dedifferentiation, proliferation, adhesion, survival including p53, cadherin, Myc, and anti-apoptotic pathways are differentially expressed or significantly altered in these lung epithelial cells upon downregulation of H2Bub1. Moreover, RNF20 knockdown dramatically suppresses terminal squamous differentiation of cultured bronchial epithelial cells, and significantly enhances proliferation, migration, invasion, and cisplatin resistance of lung cancer cells. Furthermore, immunohistochemistry analysis shows that H2Bub1 is extremely low or undetectable in >70% of 170 lung adenocarcinoma samples. Notably, statistical analysis demonstrates that loss of H2Bub1 is significantly correlated with poor differentiation in lung adenocarcinoma (p = 0.0134). In addition, patients with H2Bub1-negative cancers had a trend towards shorter survival compared with patients with H2Bub1-positive cancers. Taken together, our findings suggest that loss of H2Bub1 may enhance malignancy and promote disease progression in lung adenocarcinoma probably through modulating multiple cancer signaling pathways.

Henoch-schonlein purpura-a case report and review of the literature.

We describe a case of an adolescent male with Henoch-Schonlein purpura (HSP), presenting with cutaneous and gastrointestinal manifestations. Endoscopy revealed diffuse ulcerations in the stomach, duodenum, and right colon. Biopsies revealed a leukocytoclastic vasculitis in the skin and gastrointestinal tract. Steroid therapy led to complete resolution of the symptoms. HSP is the most common childhood vasculitis, and is characterized by the classic tetrad of nonthrombocytopenic palpable purpura, arthritis or arthralgias, gastrointestinal and renal involvement. It is a systemic disease where antigen-antibody (IgA) complexes activate the alternate complement pathway, resulting in inflammation and small vessel vasculitis. Mild disease resolves spontaneously, and symptomatic treatment alone is sufficient. Systemic steroids are recommended for moderate to severe HSP. The prognosis depends upon the extent of renal involvement, which requires close followup. Early recognition of multiorgan involvement, especially outside of the typical age group, as in our adolescent patient, and appropriate intervention can mitigate the disease and limit organ damage.

Therapies for coronaviruses. Part I of II -- viral entry inhibitors.

BACKGROUND: Severe acute respiratory syndrome (SARS) coronavirus emerged fleetingly in the winter of 2002 and again in the winter of 2003, resulting in the infection of ~8,000 people and the death of ~800. The identification of the putative natural reservoir suggests that a re-emergence is possible. The functions of many coronaviral proteins have now been elucidated, resulting in many novel approaches to therapy. OBJECTIVE: To review anticoronaviral therapies based on inhibition of viral entry into the host cell and to cast light on promising approaches and future developments. METHOD: The published literature, in particular patent publications, is searched for relevant documents. The information is organized and critiqued. RESULTS/CONCLUSION: The approaches to combating coronaviral infections are built on the foundation of antivirals against other viruses and the fundamental insights gained by dissection of the coronaviral lifecycle. These approaches include the prevention of viral entry, reviewed here, and interference with the intracellular lifecycle of the virus in the infected cell, reviewed next. Of the viral-entry inhibitors, monoclonal antibodies have demonstrated efficacy, clinical application in other viral infections, and the potential to impact a future epidemic. Moreover, combinations of monoclonal antibodies have been shown to have a broader spectrum of antiviral activity.

Therapies for coronaviruses. Part 2: Inhibitors of intracellular life cycle.

BACKGROUND: Severe acute respiratory syndrome (SARS) coronavirus emerged from an animal reservoir in 2002 and has the potential to reemerge, as shown by the occurrence of non-laboratory-associated new cases in the winter of 2003. In the absence of a vaccine, broad spectrum anticoronaviral medications are needed. OBJECTIVE: Anticoronavirals targeting viral entry were reviewed in part I. Here we review anticoronaviral therapies directed against the intracellular life cycle, with an emphasis on allowed patents and pending patents. METHOD: The published literature, in particular, patent publications is searched for relevant documents. The information is organized and critiqued. RESULTS/CONCLUSION: Many promising anticoronaviral strategies are identified. Monoclonal antibodies, protease inhibitors, interferon-based drugs and nucleic-acid based antivirals are most advanced, each having its own advantages and disadvantages. A multi-pronged approach, keeping all venues open, is advocated.

Drug targets in severe acute respiratory syndrome (SARS) virus and other coronavirus infections.

Coronaviruses are important human and animal pathogens of the order Nidovirales. Several new members were discovered following the emergence of SARS-CoV in human populations, including two human coronaviruses and several animal coronaviruses. They cause respiratory and gastrointestinal illnesses and have been found in the brains of patients with multiple sclerosis. The high mortality of SARS, the identification of a natural reservoir, and the well-founded fear of provoking antibody-enhanced disease as a result of vaccination fueled the ongoing efforts in anti-coronavirus drug discovery. This review presents the results of current research.

Inter- and intra-observer concordance of cyberpathology in twenty-five cases.

To study the feasibility of anatomical pathology consultation in cyberspace (cyberpathology) and to determine inter- and intra-observer concordance. Twenty-five consecutive cytology and histopathology cases are photographed using a digital camera placed against the eyepiece, and uploaded to an image-server in the Internet. Participants view the images, rate their confidence, and provide a diagnosis. They then view the original glass slides and provide a final diagnosis. The diagnoses are compared for inter- and intra-observer concordance. Participants are confident of their diagnoses based on viewing images on the Internet. The intra-observer concordance exceeds 95% individually, and 96% overall. Inter-observer concordance was 100% in a subset of cases. Cyberpathology as described is both available and affordable and is a valid alternative to slide-based anatomic pathology consultation.

Identification of HPV-16 in Borderline Mucinous Cystic Neoplasm of Pancreas.

Pancreatic mucinous cystic neoplasms (PMCN) predominantly affect women in the reproductive age, are located in the body and tail of the pancreas, and share morphological features with similar tumors of the ovary. We report the detection of human papillomavirus (HPV) using several different PCR protocols in a borderline PMCN from a female patient. Type-specific PCR demonstrated the HPV to be type 16. If confirmed by others, this group of neoplasms might become preventable by HPV vaccination.

Cervical Cancer Screening for the Reluctant - HPV Testing of Air-Dried Vaginal Discharge.

Despite the availability of the PAP test, cervical cancer continues to cause considerable morbidity and mortality. Many women default cervical cytology for a variety of reasons. This demands the development of alternative screening strategies, such as HPV testing on self-procured cervical-vaginal specimens in order to capture this group of women. We investigated the self-procured air-dried vaginal discharge for HPV testing. We recruited 82 patients with HPV-associated cervical lesions and 36 patients with normal cervical pathology. Participants were briefed and informed consents obtained. Each was then given a kit containing written instructions, a slim napkin, an empty zip-lock plastic bag for soiled napkin specimen, and a return envelope. After wearing the napkin for the day, the patient removes it, dries it, and returns the specimen by mail. Specimens were batched and a 0.5 cm area of each stained napkin was tested for HPV by PCR. Specimens from all 26 patients with high-grade (CIN 2 or above) HPV-induced cervical lesions and 4 of 36 normal subjects tested positive for HPV, giving a sensitivity and specificity of 100% and 88.9%, respectively. We propose offering to women who refuse cervical cytology the alternative screening strategy of testing of self-procured air-dried vaginal discharge for HPV. This method of cervical cancer screening is also suitable for people living in remote regions of the world.

Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV).

Severe acute respiratory syndrome (SARS) is caused by a coronavirus (CoV), SARSCoV. SARS-CoV belongs to the family Coronaviridae, which are enveloped RNA viruses in the order Nidovirales. Global research efforts are continuing to increase the understanding of the virus, the pathogenesis of the disease it causes (SARS), and the "heterogeneity of individual infectiousness" as well as shedding light on how to prepare for other emerging viral diseases. Promising drugs and vaccines have been identified. The milestones achieved have resulted from a truly international effort. Molecular studies dissected the adaptation of this virus as it jumped from an intermediary animal, the civet, to humans, thus providing valuable insights into processes of molecular emergence.

SARS coronavirus anti-infectives.

Severe acute respiratory syndrome (SARS) emerged in late 2002 and was controlled in July 2003 by public health measures. Its causative agent, SARS coronavirus (SARS-CoV) jumped from an animal reservoir to humans and has the potential to re-emerge. Following the sequencing of the genetic code and the deciphering of some of the functions of its proteins, including the cellular receptors and host proteins that participate in the life cycle of the virus, promising lead drugs and new uses of old drugs have been discovered. Patent applications for cathepsin L inhibitors have taken new relevance because of the role of cathepsin L in the entry of SARS-CoV into host cells. Likewise, patent applications for SARS-CoV protease inhibitors and interferon and mismatched dsRNA also need to be watched for potential application in treatment and prevention of SARS-CoV. Here, we review the recent advances and inventions that target SARS-CoV infection in humans.

Detection of human papillomavirus in sanitary napkins: a new paradigm in cervical cancer screening.

Human papillomavirus was successfully detected by polymerase chain reaction (PCR) in menstrual blood or vaginal discharge collected in sanitary napkins in 100% of 17 women having koilocytosis, cervical intraepithelial neoplasia, or squamous carcinoma. We advocate this form of cervical cancer screening because of its high sensitivity and acceptance by patients.