The nucleolar protein PinX1 has been proposed to be a putative tumor suppressor due to its binding to and inhibition of the catalytic activity of telomerase, an enzyme that is highly expressed in most human cancers in which it counteracts telomere shortening-induced senescence to confer cancer cell immortalization. However, the role of PinX1 in telomere regulation, as well as in cancer, is still poorly understood. In this study, we showed that the PinX1 protein is constitutively expressed in various human cells regardless of their telomerase activity and malignant status. Most interestingly, we found that silencing PinX1 expression by a potent short hairpin RNA construct led to a robust telomere length shortening and growth inhibition in telomerase-positive but not in telomerase-negative human cancer cells. We further showed that silencing PinX1 significantly reduced the endogenous association of telomerase with the Pot1-containing telomeric protein complex, and therefore, could account for the phenotypic telomere shortening in the affected telomerasepositive cancer cells. Our results thus reveal a novel positive role for PinX1 in telomerase/telomere regulations and suggest that the constitutive expression of PinX1 attributes to telomere maintenance by telomerase and tumorigenicity in cancer cells. [Cancer Res 2009;69(1):75-83]

Chronic rhinosinusitis (CRS) is a common chronic inflammatory disease of the upper airways involving nasal cavity and sinus. Deriving both from its clinical complexity with protean clinical manifestations as well its pathogenetic heterogeneity, the molecular mechanisms contributing to the pathogenesis of CRS remain unclear, and attract a wide interest in the field. Current evidences indicate that IL-17A is highly expressed in chronic rhinosinusitis with nasal polyps (CRSwNP). However, its pathogenetic role in regulation of tissue remodeling of CRSwNP remains unknown. The present study aimed to investigate the cellular origins and functions of IL-17A cytokine in CRSwNP, and further determined whether IL-17A could affect the expression of metalloproteinases (MMPs), the remodeling factors of CRSwNP. The results showed that the expression of IL-17A was upregulated in nasal tissues of patients with CRSwNP compared to those with chronic rhinosinusitis without nasal polyps (CRSsNP) and controls. CD8+ cytotoxic T lymphocytes (Tc) were major IL-17A producers in nasal tissues of CRSwNP. Interleukin (IL)-17-producing CD8+ T cells (Tc17) was significantly higher in nasal tissues of CRSwNP than CRSsNP and controls. Nonetheless, no difference was observed among the IL-17A in peripheral blood lymphocytes of these three groups. Moreover, in the same patients, IL-17A expression was negligible in lymphocytes of peripheral blood when compared with nasal tissues. Increased gene and protein expression of MMP-7 and MMP-9 in patients with CRSwNP compared with controls were observed. In CRSwNP samples, IL-17A receptor (IL-17AR) co-localized with MMP-9 and they were mainly expressed in the epithelial cells. MMP-9 expression was up-regulated both in Primary human nasal epithelial cells (PHNECs) and a nasal epithelial cell line (RPMI 2650) by IL-17A treatment, and diminished by anti-IL-17AR treatment. Furthermore, IL-17A promoted the expression of MMP-9 by activating the NF-κB signal pathway. Thus, our results have revealed a crucial role of IL-17A and Tc cells on pathogenesis and tissue remodeling of CRSwNP.

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Chemoradiotherapy (CRT) is a standard treatment for oesophageal squamous cell carcinoma (ESCC) in its advanced stages. The telomerase/telomere interacting protein PinX1 contributes to telomere maintenance, tumourigenicity, and influences the DNA damage agent-induced apoptotic response in telomerase-positive cancer cells. However, the clinical and biological significance of PinX1 in human ESCCs remains unclear. We examined the expression dynamics of PinX1 by immunohistochemistry in a learning cohort (n = 98) and a validation cohort (n = 59) of ESCC patients treated with definite chemoradiotherapy (CRT). A series of in vivo and in vitro assays were performed to elucidate the effect of PinX1 on ESCC cells' CRT response and underlying mechanisms. Knockdown of PinX1 did not affect ESCC cells' chemosensitivities to 5-fluorouracil and cisplatin, but substantially increased ESCC cells' therapeutic efficacy of radiation both in vitro and in vivo. Ectopic overexpression of PinX1 dramatically enhanced ESCC cells' resistance to radiotherapy. Furthermore, we demonstrated that PinX1 resistance to radiotherapy (RT) was attributed to PinX1 maintaining telomere stability, reducing ESCC cell death by RT-induced mitosis catastrophe (MC). High expression of Pinx1 correlated positively with ESCC's resistance to CRT, and was a strong and independent predictor for short disease-specific survival (DSS) of ESCC patients. Our data suggest that PinX1 could serve as a novel predictor for a CRT response to ESCC patients, and the pathway of PinX1-mediated telomere stability might represent a new target to improve the RT effect of ESCC.

Glioblastoma multiforme is the most aggressive form of human brain tumor, which has no effective cure. Previously, we have demonstrated that overexpression of the C-terminal fragment of the human telomerase reverse transcriptase (hTERTC27) inhibits the growth and tumorigenicity of human cervical cancer HeLa cells. In this study, the therapeutic effect and molecular mechanisms of hTERTC27-mediated cancer gene therapy were further explored in vivo in established human glioblastoma xenografts in nude mice. We showed that intratumoral injection of adeno-associated virus carrying hTERTC27 (rAAV-hTERTC27) is highly effective in reducing the growth of the subcutaneously transplanted glioblastoma tumors. Histological analyses showed that rAAV-hTERTC27 treatment leads to profound necrosis, apoptosis, infiltration of polymorphonuclear neutrophils and reduced microvessel density in the tumor samples. To study the molecular mechanism of rAAV-hTERTC27-mediated antitumor effects, we analyzed the global gene expression profiles of the rAAV-hTERTC27-treated tumor tissues and cell line as compared with that of the control rAAV-green fluorescent protein-treated samples by DNA microarray. Our results suggest that hTERTC27 exerts its effect through complex mechanisms, which involve genes regulating apoptosis, cell adhesion, cell cycle, immune responses, metabolism, signal transduction, transport, transcription and telomere maintenance.