Abstract Nasopharyngeal carcinoma (NPC) is a common EBV-associated human malignancy in Southern China. NPC is highly radiosensitive and radiotherapy is the mainstay treatment for the patients. However, poor clinical outcome remains the major concern in NPC patients with advanced stage disease. Development of new clinical interventions to treat the disease are importance for control this cancer. PIN1 is a highly conserved enzyme that isomerizes specific phosphorylated Ser/Thr-Pro bonds in multiple proteins. It induces conformational changes and modulates functions of many proteins that promote oncogenesis. We have recently confirmed the high PIN1 expression in almost all NPC primary tumours. The aim of this study was to investigate the potential of PIN1 as a therapeutic target for the treatment of NPC. The oncogenic function of PIN1 was investigated in the EBV-positive NPC cell line C666-1 and an immortalized NP epithelial cells NP69. The antitumor effect of PIN1 inhibition was then studied in both in vitro and in vivo NPC models by using the PIN1 inhibitor juglone. In the stable PIN1-expressing NP cells, we found the upregulation of cyclin D1 and activation of MAPL/JNK pathway. Furthermore, we also demonstrated the interaction between PIN1 and EBNA1 in the NPC cells. PIN1 was also shown to interfere the transcription activity of EBNA1 through binding to the phosphorylation sites of EBNA1. Notably, knockdown of PIN1 by siRNA significantly inhibited cell proliferation, DNA synthesis and colony formation of NPC cells. Treatment with juglone demonstrated significant induction of apoptosis and suppression of cyclin D1 expression in C666-1 cells. The IC50 of C666-1 was approximately 6 μM while that of EBV-negative NPC cells HK-1was 10 μM. A dose-dependent inhibition of tumor growth was observed in mice treated with juglone. A significantly growth suppression and cyclin D1 downregulation was found in the tumours of the mice treated with 1-1.5 mg/kg juglone. In conclusion, our findings imply that targeting Pin1 may serve as a potential therapeutic approach for treating patients suffering from this EBV-associated cancer. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B238. Citation Format: Meng Xu, Chit Chow, Chartia Ching Mei Cheung, Grace Tin-Yun Chung, Kwok Wai Lo. PIN1 (Peptidyl-prolyl cis/trans-isomerase, NIMA-interacting 1) is a potential therapeutic target for EBV-associated nasopharyngeal carcinoma. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B238.
Abstract Nasopharyngeal carcinoma (NPC) is a distinctive type of squamous cell carcinoma of the head and neck region with remarkable ethnic and geographic distribution. It is consistently associated with Epstein–Barr virus (EBV) infection and serves as a fascinating model to understand the complex interaction among environmental, viral, and genetic factors in human tumourigenesis. The development of NPC involves clonal expansion of EBV‐infected cells and cumulative genetic and epigenetic changes. Both viral and cellular genes contribute to disease pathogenesis and their interplay creates distinct phenotypes, unique molecular features and tumour microenvironment. Recent findings on EBV‐encoded and cellular miRNAs demonstrate their importance in NPC progression by altering multiple cellular pathways. On the basis of current knowledge, we proposed a stepwise model describing the initiation events and the role of EBV latent infection and multiple genetic changes in NPC tumourigenesis. Key Concepts: Nonkeratinising NPC is consistently associated with EBV infection. The development of NPC involves EBV latent infection and cumulative genetic and epigenetic changes influenced by predisposing genetic factors and environmental carcinogens. Inactivation of tumour suppressor genes at chromosomes 3p and 9p and clonal expansion of EBV‐infected cells are critical initiation events in NPC tumourigenesis. EBV‐encoded and cellular miRNAs contribute to the NPC progression by altering multiple cellular pathways and modulating EBV latent gene expression. The interplay between viral and cellular genes contributes to the establishment of distinct phenotypes, unique molecular features and tumour microenvironment of NPC.
Nasopharyngeal carcinoma (NPC) is a unique EBV-associated epithelial malignancy, showing highly invasive and metastatic phenotype. Despite increasing evidence demonstrating the critical role of cancer stem-like cells (CSCs) in the maintenance and progression of tumors in a variety of malignancies, the existence and properties of CSC in EBV-associated NPC are largely unknown. Our study aims to elucidate the presence and role of CSCs in the pathogenesis of this malignant disease. Sphere-forming cells were isolated from an EBV-positive NPC cell line C666-1 and its tumor-initiating properties were confirmed by in vitro and in vivo assays. In these spheroids, up-regulation of multiple stem cell markers were found. By flow cytometry, we demonstrated that both CD44 and SOX2 were overexpressed in a majority of sphere-forming C666-1 cells. The CD44+SOX2+ cells was detected in a minor population in EBV-positive xenografts and primary tumors and considered as potential CSC in NPC. Notably, the isolated CD44+ NPC cells were resistant to chemotherapeutic agents and with higher spheroid formation efficiency, showing CSC properties. On the other hand, microarray analysis has revealed a number of differentially expressed genes involved in transcription regulation (e.g. FOXN4, GLI1), immune response (CCR7, IL8) and transmembrane transport (e.g. ABCC3, ABCC11) in the spheroids. Among these genes, increased expression of CCR7 in CD44+ CSCs was confirmed in NPC xenografts and primary tumors. Importantly, blocking of CCR7 abolished the sphere-forming ability of C666-1 in vitro. Expression of CCR7 was associated with recurrent disease and distant metastasis. The current study defined the specific properties of a CSC subpopulation in EBV-associated NPC. Our findings provided new insights into developing effective therapies targeting on CSCs, thereby potentiating treatment efficacy for NPC patients.
Abstract The unique virus-cell interaction in Epstein-Barr virus (EBV)-associated malignancies implies targeting the viral latent-lytic switch is a promising therapeutic strategy. However, the lack of specific and efficient therapeutic agents to induce lytic cycle in these cancers is a major challenge facing clinical implementation. We develop a synthetic transcriptional activator that specifically activates endogenous BZLF1 and efficiently induces lytic reactivation in EBV-positive cancer cells. A lipid nanoparticle encapsulating nucleoside-modified mRNA which encodes a BZLF1 -specific transcriptional activator (mTZ3-LNP) is synthesized for EBV-targeted therapy. Compared with conventional chemical inducers, mTZ3-LNP more efficiently activates EBV lytic gene expression in EBV-associated epithelial cancers. Here we show the potency and safety of treatment with mTZ3-LNP to suppress tumor growth in EBV-positive cancer models. The combination of mTZ3-LNP and ganciclovir yields highly selective cytotoxic effects of mRNA-based lytic induction therapy against EBV-positive tumor cells, indicating the potential of mRNA nanomedicine in the treatment of EBV-associated epithelial cancers.
Abstract Circular RNAs (circRNAs) are abundantly expressed in cancer. Their resistance to exonucleases enables them to have potentially stable interactions with different types of biomolecules. Alternative splicing can create different circRNA isoforms that have different sequences and unequal interaction potentials. The study of circRNA function thus requires knowledge of complete circRNA sequences. Here we describe psirc, a method that can identify full-length circRNA isoforms and quantify their expression levels from RNA sequencing data. We confirm the effectiveness and computational efficiency of psirc using both simulated and actual experimental data. Applying psirc on transcriptome profiles from nasopharyngeal carcinoma and normal nasopharynx samples, we discover and validate circRNA isoforms differentially expressed between the two groups. Compared to the assumed circular isoforms derived from linear transcript annotations, some of the alternatively spliced circular isoforms have 100 times higher expression and contain substantially fewer microRNA response elements, demonstrating the importance of quantifying full-length circRNA isoforms.
Severe acute respiratory syndrome (SARS) is the first pandemic of the 21st century (1). Since its recognition, 8437 individuals have been affected and 813 have died (2). Approximately 20–30% of patients required intensive care admission (1). Although there was a slight predominance of female SARS patients, possibly because of the overrepresentation of female healthcare workers (1), male SARS patients were more likely to suffer poor outcomes (3). In a major hospital outbreak in Hong Kong (4), 32% of male and 15% of female SARS patients required intensive care or died. Remarkably, similar demographic data were seen among SARS patients in the greater Toronto area, Canada, where 32% of males and 14% of females with SARS required intensive care or died (5). Karlberg et al. (3) studied the case fatality rates among all confirmed SARS patients documented in the Hong Kong SARS epidemic in 2003. The authors concluded that the mortality rates differed significantly between males and females, being 21.9% and 13.2%, respectively. The relative risk for death in males was 1.62 after adjustment for age. It is thus an intriguing coincidence that ACE2 , the gene for the newly identified functional receptor for the SARS coronavirus, angiotensin-converting enzyme 2, maps to the X-chromosome (Xp22) (6).
ACE2 was first identified as a homolog of angiotensin-converting enzyme with zinc metalloproteinase activity (7). Many of its activities differ from those of angiotensin-converting enzyme (8). ACE2 has been found …
Abstract Nasopharyngeal carcinoma (NPC) is an aggressive head and neck cancer characterized by Epstein-Barr virus (EBV) infection and dense lymphocyte infiltration. The scarcity of NPC genomic data hinders the understanding of NPC biology, disease progression and rational therapy design. Here we performed whole-exome sequencing (WES) on 111 micro-dissected EBV-positive NPCs, with 15 cases subjected to further whole-genome sequencing (WGS), to determine its mutational landscape. We identified enrichment for genomic aberrations of multiple negative regulators of the NF-κB pathway, including CYLD , TRAF 3, NFKBIA and NLRC5, in a total of 41% of cases. Functional analysis confirmed inactivating CYLD mutations as drivers for NPC cell growth. The EBV oncoprotein latent membrane protein 1 (LMP1) functions to constitutively activate NF-κB signalling, and we observed mutual exclusivity among tumours with somatic NF-κB pathway aberrations and LMP1-overexpression, suggesting that NF-κB activation is selected for by both somatic and viral events during NPC pathogenesis.
Abstract Introduction: EBV-associated nasopharyngeal carcinoma (NPC) is a distinct type of head and neck cancer that is prevalence in southern China and Southeast Asia. While radiotherapy or concurrent chemo-radiotherapy are efficient for the patients with early disease, more than 60% of NPC patients are diagnosed at an advanced stage and frequently develop local failure and distant metastases. The lack of druggable targets is a challenge for controlling this common cancer. By whole-genome sequencing and FISH analysis, we have defined the frequent homozygous deletion of 9p21.3 in NPC. The loss of MTAP that encodes methylthioadenosine phosphorylase on this region was found in 32% and 34% of primary and recurrent NPC respectively, indicating vulnerability to targeted-therapies for MAT2A/PRMT5 axis. Methods: The preclinical efficacy of MAT2A inhibitors, FIDAS-5 and BT115386 was elucidated in a panel of MTAP-deleted EBV-positive NPC cell lines and patient-derived xenografts. In vitro and in vivo growth inhibition of NPC cells by these compounds were determined. In addition to the suppression of PRMT5 activity, the effects of MAT2A inhibitor treatment on activating p53 pathway, inducing EBV lytic genes and modulating expression of proteins involved in cellular differentiation, apoptosis and lipogenesis in NPC were accessed. Results: In vitro study has revealed that treatment of the MAT2A inhibitors markedly reduced PRMT5 activity and cell growth in all MTAP-deleted NPCs. Compared to the MTAP-wild type NPC cells, the MTAP-deleted cells, either with wild type or mutated TP53, showed heightened sensitivity (>9 fold more sensitive) to the MAT2A inhibition by FIDAS-5 and BT115386. The potent in vivo growth inhibitory effect of BT115386 were observed in the MTAP-deleted xenografts, but not in that with wild type MTAP. In the MTAP-deleted NPC treated with MAT2A inhibitors, activation of p53 pathway and induction of BAX apoptotic protein were consistently found. The treatment also promoted differentiation, suppressed stemness transcription factors and inhibited lipogenesis. Notably, the MAT2A inhibitors also disrupted EBV latency in the MTAP-deleted NPC, induction of multiple EBV lytic genes including BZLF1 and BMRF1 were demonstrated. Conclusion: Selective sensitivity of MTAP-deleted NPC to the treatment of MAT2A inhibitors including FIDAS-5 and BT115386 was demonstrated. To facility its growth inhibitory effects, the compounds targeted PRMT5 activity, subsequently modulated multiple cellular mechanisms and induced EBV lytic reactivation. The potent antitumor effect of BT115386 was demonstrated in the in vivo MTAP-deleted NPC mouse models. The findings warrant future assessments of MAT2A inhibitors in clinical studies for NPC, which might well impact a large subset of patients. Acknowledgment: Acknowledgment: The work was supported by Hong Kong RGC funding (GRF:14101721 and AoE/M-401/20) and HMRF grant (#09203176) Citation Format: YUK-Yu Chan, Chi-Man Tsang, Grace T.Y. Chung, Ka-Fai To, Yi Zhang, Xiaodong Zhang, Jun Tang, Kwok Wai Lo. Therapeutic vulnerability of MTAP-deleted nasopharyngeal carcinoma by MAT2A inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6226.
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