Focused ultrasound-mediated blood-brain barrier opening is safe and feasible with moderately hypofractionated radiotherapy for brainstem diffuse midline glioma.

BACKGROUND: Diffuse midline glioma (DMG) is a pediatric tumor with dismal prognosis. Systemic strategies have been unsuccessful and radiotherapy (RT) remains the standard-of-care. A central impediment to treatment is the blood-brain barrier (BBB), which precludes drug delivery to the central nervous system (CNS). Focused ultrasound (FUS) with microbubbles can transiently and non-invasively disrupt the BBB to enhance drug delivery. This study aimed to determine the feasibility of brainstem FUS in combination with clinical doses of RT. We hypothesized that FUS-mediated BBB-opening (BBBO) is safe and feasible with 39 Gy RT. METHODS: To establish a safety timeline, we administered FUS to the brainstem of non-tumor bearing mice concurrent with or adjuvant to RT; our findings were validated in a syngeneic brainstem murine model of DMG receiving repeated sonication concurrent with RT. The brainstems of male B6 (Cg)-Tyrc-2J/J albino mice were intracranially injected with mouse DMG cells (PDGFB+, H3.3K27M, p53-/-). A clinical RT dose of 39 Gy in 13 fractions (39 Gy/13fx) was delivered using the Small Animal Radiation Research Platform (SARRP) or XRAD-320 irradiator. FUS was administered via a 0.5 MHz transducer, with BBBO and tumor volume monitored by magnetic resonance imaging (MRI). RESULTS: FUS-mediated BBBO did not affect cardiorespiratory rate, motor function, or tissue integrity in non-tumor bearing mice receiving RT. Tumor-bearing mice tolerated repeated brainstem BBBO concurrent with RT. 39 Gy/13fx offered local control, though disease progression occurred 3-4 weeks post-RT. CONCLUSION: Repeated FUS-mediated BBBO is safe and feasible concurrent with RT. In our syngeneic DMG murine model, progression occurs, serving as an ideal model for future combination testing with RT and FUS-mediated drug delivery.

Pemetrexed combined with dual immune checkpoint blockade enhances cytotoxic T lymphocytes against lung cancer.

Chemotherapy, in combination with immune checkpoint blockade (ICB) targeting to programmed death-1 (PD-1) or its ligand PD-L1, is one of the first-line treatments for patients with advanced non-small-cell lung cancer (NSCLC). However, a large proportion of patients, especially those with PD-L1 negative tumors, do not benefit from this treatment. This may be due to the existence of multiple immunosuppressive mechanisms other than the PD-1/PD-L1 axis. Human leukocyte antigen-G (HLA-G) has been identified as an immune checkpoint protein (ICP) and a neoexpressed tumor-associated antigen (TAA) in a large proportion of solid tumors. In this study, we evaluated the induction of HLA-G as well as PD-L1 using sublethal doses of chemotherapeutics including pemetrexed in different NSCLC cell lines. Except for gefitinib, most of the chemotherapeutic agents enhanced HLA-G and PD-L1 expression in a dose-dependent manner, whereas pemetrexed and carboplatin treatments showed the most consistent upregulation of PD-L1 and HLA-G in each cell line. In addition to protein levels, a novel finding of this study is that pemetrexed enhanced the glycosylation of HLA-G and PD-L1. Pemetrexed potentiated the cytotoxicity of cytotoxic T lymphocytes (CTLs) to treat NSCLC. Both in vitro and in vivo experiments revealed that CTL-mediated cytotoxicity was most pronounced when both anti-PD-L1 and anti-HLA-G ICBs were combined with pemetrexed treatment. In conclusion, anti-HLA-G could be an intervention strategy in addition to the anti-PD-1/PD-L1 pathway for NSCLC. Moreover, dual targeting of PD-L1 and HLA-G combined with pemetrexed might have a better extent of CTL-based immunotherapy.

Band-Selection of a Portal LED-Induced Autofluorescence Multispectral Imager to Improve Oral Cancer Detection.

This aim of this study was to find effective spectral bands for the early detection of oral cancer. The spectral images in different bands were acquired using a self-made portable light-emitting diode (LED)-induced autofluorescence multispectral imager equipped with 365 and 405 nm excitation LEDs, emission filters with center wavelengths of 470, 505, 525, 532, 550, 595, 632, 635, and 695 nm, and a color image sensor. The spectral images of 218 healthy points in 62 healthy participants and 218 tumor points in 62 patients were collected in the ex vivo trials at China Medical University Hospital. These ex vivo trials were similar to in vivo because the spectral images of anatomical specimens were immediately acquired after the on-site tumor resection. The spectral images associated with red, blue, and green filters correlated with and without nine emission filters were quantized by four computing method, including summated intensity, the highest number of the intensity level, entropy, and fractional dimension. The combination of four computing methods, two excitation light sources with two intensities, and 30 spectral bands in three experiments formed 264 classifiers. The quantized data in each classifier was divided into two groups: one was the training group optimizing the threshold of the quantized data, and the other was validating group tested under this optimized threshold. The sensitivity, specificity, and accuracy of each classifier were derived from these tests. To identify the influential spectral bands based on the area under the region and the testing results, a single-layer network learning process was used. This was compared to conventional rules-based approaches to show its superior and faster performance. Consequently, four emission filters with the center wavelengths of 470, 505, 532, and 550 nm were selected by an AI-based method and verified using a rule-based approach. The sensitivities of six classifiers using these emission filters were more significant than 90%. The average sensitivity of these was about 96.15%, the average specificity was approximately 69.55%, and the average accuracy was about 82.85%.

Long noncoding RNA HOTAIR promotes invasion of breast cancer cells through chondroitin sulfotransferase CHST15.

The long noncoding RNA HOTAIR plays significant roles in promoting cancer metastasis. However, how it conveys an invasive advantage in cancer cells is not clear. Here we identify the chondroitin sulfotransferase CHST15 (GalNAc4S-6ST) as a novel HOX transcript antisense intergenic RNA (HOTAIR) target gene using RNA profiling and show that CHST15 is required for HOTAIR-mediated invasiveness in breast cancer cells. CHST15 catalyzes sulfation of the C6 hydroxyl group of the N-acetyl galactosamine 4-sulfate moiety in chondroitin sulfate to form the 4,6-disulfated chondroitin sulfate variant known as the CS-E isoform. We show that HOTAIR is necessary and sufficient for CHST15 transcript expression. Inhibition of CHST15 by RNA interference abolished cell invasion promoted by HOTAIR but not on HOTAIR-mediated migratory activity. Conversely, reconstitution of CHST15 expression rescued the invasive activity of HOTAIR-depleted cells. In corroboration with this mechanism, blocking cell surface chondroitin sulfate using a pan-CS antibody or an antibody specifically recognizes the CS-E isoform significantly suppressed HOTAIR-induced invasion. Inhibition of CHST15 compromised tumorigenesis and metastasis in orthotopic breast cancer xenograft models. Furthermore, the expression of HOTAIR closely correlated with the level of CHST15 protein in primary as well as metastatic tumor lesions. Our results demonstrate a novel mechanism underlying the function of HOTAIR in tumor progression through programming the context of cell surface glycosaminoglycans. Our results further establish that the invasive and migratory activities downstream of HOTAIR are distinctly regulated, whereby CHST15 preferentially controls the arm of invasiveness. Thus, the HOTAIR-CHST15 axis may provide a new avenue toward novel therapeutic strategies and prognosis biomarkers for advanced breast cancer.

Interleukin-8/CXCR1 Signaling Contributes to the Progression of Pulmonary Adenocarcinoma Resulting in Malignant Pleural Effusion.

Pulmonary adenocarcinoma (PADC) treatment limited efficacy in preventing tumor progression, often resulting in malignant pleural effusion (MPE). MPE is filled with various mediators, especially interleukin-8 (IL-8). However, the role of IL-8 and its signaling mechanism within the fluid microenvironment (FME) implicated in tumor progression warrants further investigation. Primary cultured cells from samples of patients with MPE from PADC, along with a commonly utilized lung cancer cell line, were employed to examine the role of IL-8 and its receptor, CXCR1, through comparative analysis. Our study primarily assessed migration and invasion capabilities, epithelial-mesenchymal transition (EMT), and cancer stem cell (CSC) properties. Additionally, IL-8 levels in MPE fluid versus serum, along with immunohistochemical expression of IL-8/CXCR1 signaling in tumor tissue and cell blocks were analyzed. IL-8/CXCR1 overexpression enhanced EMT and CSC properties. Furthermore, the immunocytochemical examination of 17 cell blocks from patients with PADC and MPE corroborated the significant correlation between upregulated IL-8 and CXCR1 expression and the co-expression of IL-8 and CXCR1 in MPE with distant metastasis. In summary, the IL-8/ CXCR1 axis in FME is pivotal to tumor promotion via paracrine and autocrine signaling. Our study provides a therapeutic avenue for improving the prognosis of PADC patients with MPE.

CAR-T cells targeting HLA-G as potent therapeutic strategy for EGFR-mutated and overexpressed oral cancer.

Oral squamous cell carcinoma (OSCC) is a common malignancy in the world. Recently, scientists have focused on therapeutic strategies to determine the regulation of tumors and design molecules for specific targets. Some studies have demonstrated the clinical significance of human leukocyte antigen G (HLA-G) in malignancy and NLR family pyrin domain-containing 3 (NLRP3) inflammasome in promoting tumorigenesis in OSCC. This is the first study to investigate whether aberrant epidermal growth factor receptor (EGFR) induces HLA-G expression through NLRP3 inflammasome-mediated IL-1ß secretion in OSCC. Our results showed that the upregulation of NLRP3 inflammasome leads to abundant HLA-G in the cytoplasm and cell membrane of FaDu cells. In addition, we also generated anti-HLA-G chimeric antigen receptor (CAR)-T cells and provided evidence for their effects in EGFR-mutated and overexpressed oral cancer. Our results may be integrated with OSCC patient data to translate basic research into clinical significance and may lead to novel EGFR-aberrant OSCC treatment.

BiTE-Secreting CAR-γδT as a Dual Targeting Strategy for the Treatment of Solid Tumors.

HLA-G is considered as an immune checkpoint protein and a tumor-associated antigen. In the previous work, it is reported that CAR-NK targeting of HLA-G can be used to treat certain solid tumors. However, the frequent co-expression of PD-L1 and HLA-G) and up-regulation of PD-L1 after adoptive immunotherapy may decrease the effectiveness of HLA-G-CAR. Therefore, simultaneous targeting of HLA-G and PD-L1 by multi-specific CAR could represent an appropriate solution. Furthermore, gamma-delta T (γδT) cells exhibit MHC-independent cytotoxicity against tumor cells and possess allogeneic potential. The utilization of nanobodies offers flexibility for CAR engineering and the ability to recognize novel epitopes. In this study, Vδ2 γδT cells are used as effector cells and electroporated with an mRNA-driven, nanobody-based HLA-G-CAR with a secreted PD-L1/CD3ε Bispecific T-cell engager (BiTE) construct (Nb-CAR.BiTE). Both in vivo and in vitro experiments reveal that the Nb-CAR.BiTE-γδT cells could effectively eliminate PD-L1 and/or HLA-G-positive solid tumors. The secreted PD-L1/CD3ε Nb-BiTE can not only redirect Nb-CAR-γδT but also recruit un-transduced bystander T cells against tumor cells expressing PD-L1, thereby enhancing the activity of Nb-CAR-γδT therapy. Furthermore, evidence is provided that Nb-CAR.BiTE redirectes γδT into tumor-implanted tissues and that the secreted Nb-BiTE is restricted to the tumor site without apparent toxicity.

Tid1, CHIP and ErbB2 interactions and their prognostic implications for breast cancer patients.

ErbB2 (HER2/neu) is overexpressed in about 25-30% of breast malignancies, and up-regulation of ErbB2 in breast cancer patients is associated with poor prognosis. It is known that the carboxyl terminus of heat shock cognate 70 interacting protein (CHIP) efficiently down-regulates ErbB2 in vitro. Human tumourous imaginal disc 1 (Tid1, DnaJa3), a co-chaperone of heat shock protein 70 (Hsp70), also suppresses ErbB2 expression in breast cancer cell lines. However, the intracellular interactions among Tid1, CHIP, and ErbB2 remain elusive, and the utilization of Tid1 and CHIP as breast cancer biomarkers has never been proposed. Herein, we analysed the expression and correlations among Tid1, CHIP, and ErbB2 in a total of 183 breast cancer histology sections, including 30 fresh tissue specimens, using immunohistochemistry (IHC) and immunoblotting assay. A computerized image analysis system was used for IHC scoring and determining relative immunoblot intensity. The immunohistochemical expression of Tid1 and CHIP were positively correlated with each other but were both inversely correlated to that of ErbB2. Odds ratio analyses showed that lower expression of Tid1 has a relatively higher risk of unfavourable tumour grade, later pathological stage, larger tumour size, and microscopic features of a more malignant histology including lymphovascular invasion, stromal inflammatory response, and tumour necrosis. Expression of CHIP displayed similar characteristics. Furthermore, expression of Tid1 and/or CHIP increases patients' 10-year overall and disease-free survival rate. Empirically, we also demonstrated that Tid1, CHIP, and ErbB2 interacted with each other through immunofluorescence or co-immunoprecipitation analyses. Functionally, Tid1 and CHIP acted synergistically to degrade ErbB2 in vitro. Conversely, Tid1 cannot compensate for the loss of proteolytic function noted in CHIP mutations for degradation of ErbB2. Overall, our data suggest that Tid1 and CHIP play pivotal roles in affecting the levels of ErbB2 protein, and that both are significant prognostic indicators of breast cancer patient survival.

Catamenial pneumothorax due to bilateral pulmonary endometriosis.

Co-existence of catamenial pneumothorax and hemoptysis is rare. We present a case of catamenial pneumothorax due to bilateral pulmonary endometriosis in a 45-year-old woman. The patient presented with a 3-year history of intermittent productive cough with blood-tinged sputum, chronic anemia, loss of appetite, and general weakness associated with menstruation. Three years prior to this presentation the patient had undergone a sigmoidectomy as treatment for endometriosis of the sigmoid colon with bleeding. Chest radiographs and computed tomography (CT) scan revealed multiple nodules in both lung parenchyma and recurrent pneumothorax. CT-guided biopsy revealed chronic inflammation of those pulmonary nodules, and laboratory studies disclosed elevated serum levels of carbohydrate antigen 19-9 (CA 19-9) and CA 125. Thoracoscopic wedge resection of the pulmonary nodules was performed, and histopathological examination of the resected nodules revealed endometriosis. At one-year follow-up there was no evidence of recurrence of gastrointestinal bleeding or pneumothorax.

Targeting human leukocyte antigen G with chimeric antigen receptors of natural killer cells convert immunosuppression to ablate solid tumors.

BACKGROUND: Immunotherapy against solid tumors has long been hampered by the development of immunosuppressive tumor microenvironment, and the lack of a specific tumor-associated antigen that could be targeted in different kinds of solid tumors. Human leukocyte antigen G (HLA-G) is an immune checkpoint protein (ICP) that is neoexpressed in most tumor cells as a way to evade immune attack and has been recently demonstrated as a useful target for chimeric antigen receptor (CAR)-T therapy of leukemia by in vitro studies. Here, we design and test for targeting HLA-G in solid tumors using a CAR strategy. METHODS: We developed a novel CAR strategy using natural killer (NK) cell as effector cells, featuring enhanced cytolytic effect via DAP12-based intracellular signal amplification. A single-chain variable fragment (scFv) against HLA-G is designed as the targeting moiety, and the construct is tested both in vitro and in vivo on four different solid tumor models. We also evaluated the synergy of this anti-HLA-G CAR-NK strategy with low-dose chemotherapy as combination therapy. RESULTS: HLA-G CAR-transduced NK cells present effective cytolysis of breast, brain, pancreatic, and ovarian cancer cells in vitro, as well as reduced xenograft tumor growth with extended median survival in orthotopic mouse models. In tumor coculture assays, the anti-HLA-G scFv moiety promotes Syk/Zap70 activation of NK cells, suggesting reversal of the HLA-G-mediated immunosuppression and hence restoration of native NK cytolytic functions. Tumor expression of HLA-G can be further induced using low-dose chemotherapy, which when combined with anti-HLA-G CAR-NK results in extensive tumor ablation both in vitro and in vivo. This upregulation of tumor HLA-G involves inhibition of DNMT1 and demethylation of transporter associated with antigen processing 1 promoter. CONCLUSIONS: Our novel CAR-NK strategy exploits the dual nature of HLA-G as both a tumor-associated neoantigen and an ICP to counteract tumor spread. Further ablation of tumors can be boosted when combined with administration of chemotherapeutic agents in clinical use. The readiness of this novel strategy envisions a wide applicability in treating solid tumors.

Focused Ultrasound-Mediated Blood-Brain Barrier Opening Increases Delivery and Efficacy of Etoposide for Glioblastoma Treatment.

PURPOSE: Glioblastoma (GBM) is a devastating disease. With the current treatment of surgery followed by chemoradiation, outcomes remain poor, with median survival of only 15 months and a 5-year survival rate of 6.8%. A challenge in treating GBM is the heterogeneous integrity of the blood-brain barrier (BBB), which limits the bioavailability of systemic therapies to the brain. There is a growing interest in enhancing drug delivery by opening the BBB with the use of focused ultrasound (FUS). We hypothesize that an FUS-mediated BBB opening can enhance the delivery of etoposide for a therapeutic benefit in GBM. METHODS AND MATERIALS: A murine glioma cell line (Pdgf+, Pten-/-, P53-/-) was orthotopically injected into B6(Cg)-Tyrc-2J/J mice to establish the syngeneic GBM model for this study. Animals were treated with FUS and microbubbles to open the BBB to enhance the delivery of systemic etoposide. Magnetic resonance (MR) imaging was used to evaluate the BBB opening and tumor progression. Liquid chromatography tandem mass spectrometry was used to measure etoposide concentrations in the intracranial tumors. RESULTS: The murine glioma cell line is sensitive to etoposide in vitro. MR imaging and passive cavitation detection demonstrate the safe and successful BBB opening with FUS. The combined treatment of an FUS-mediated BBB opening and etoposide decreased tumor growth by 45% and prolonged median overall survival by 6 days: an approximately 30% increase. The FUS-mediated BBB opening increased the brain tumor-to-serum ratio of etoposide by 3.5-fold and increased the etoposide concentration in brain tumor tissue by 8-fold compared with treatment without ultrasound. CONCLUSIONS: The current study demonstrates that BBB opening with FUS increases intratumoral delivery of etoposide in the brain, resulting in local control and overall survival benefits.

Focused ultrasound mediated blood-brain barrier opening is safe and feasible in a murine pontine glioma model.

Drug delivery in diffuse intrinsic pontine glioma is significantly limited by the blood-brain barrier (BBB). Focused ultrasound (FUS), when combined with the administration of microbubbles can effectively open the BBB permitting the entry of drugs across the cerebrovasculature into the brainstem. Given that the utility of FUS in brainstem malignancies remains unknown, the purpose of our study was to determine the safety and feasibility of this technique in a murine pontine glioma model. A syngeneic orthotopic model was developed by stereotactic injection of PDGF-B+PTEN-/-p53-/- murine glioma cells into the pons of B6 mice. A single-element, spherical-segment 1.5 MHz ultrasound transducer driven by a function generator through a power amplifier was used with concurrent intravenous microbubble injection for tumor sonication. Mice were randomly assigned to control, FUS and double-FUS groups. Pulse and respiratory rates were continuously monitored during treatment. BBB opening was confirmed with gadolinium-enhanced MRI and Evans blue. Kondziela inverted screen testing and sequential weight lifting measured motor function before and after sonication. A subset of animals were treated with etoposide following ultrasound. Mice were either sacrificed for tissue analysis or serially monitored for survival with daily weights. FUS successfully caused BBB opening while preserving normal cardiorespiratory and motor function. Furthermore, the degree of intra-tumoral hemorrhage and inflammation on H&E in control and treated mice was similar. There was also no difference in weight loss and survival between the groups (p > 0.05). Lastly, FUS increased intra-tumoral etoposide concentration by more than fivefold. FUS is a safe and feasible technique for repeated BBB opening and etoposide delivery in a preclinical pontine glioma model.

Elimination of head and neck cancer initiating cells through targeting glucose regulated protein78 signaling.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is a highly lethal cancer that contains cellular and functional heterogeneity. Previously, we enriched a subpopulation of highly tumorigenic head and neck cancer initiating cells (HN-CICs) from HNSCC. However, the molecular mechanisms by which to govern the characteristics of HN-CICs remain unclear. GRP78, a stress-inducible endoplasmic reticulum chaperone, has been reported to play a crucial role in the maintenance of embryonic stem cells, but the role of GRP78 in CICs has not been elucidated. RESULTS: Initially, we recognized GRP78 as a putative candidate on mediating the stemness and tumorigenic properties of HN-CICs by differential systemic analyses. Subsequently, cells with GRP78 anchored at the plasma membrane (memGRP78+) exerted cancer stemness properties of self-renewal, differentiation and radioresistance. Of note, xenotransplantation assay indicated merely 100 memGRP78+ HNSCCs resulted in tumor growth. Moreover, knockdown of GRP78 significantly reduced the self-renewal ability, side population cells and expression of stemness genes, but inversely promoted cell differentiation and apoptosis in HN-CICs. Targeting GRP78 also lessened tumorigenicity of HN-CICs both in vitro and in vivo. Clinically, co-expression of GRP78 and Nanog predicted the worse survival prognosis of HNSCC patients by immunohistochemical analyses. Finally, depletion of GRP78 in HN-CICs induced the expression of Bax, Caspase 3, and PTEN. CONCLUSIONS: In summary, memGRP78 should be a novel surface marker for isolation of HN-CICs, and targeting GRP78 signaling might be a potential therapeutic strategy for HNSCC through eliminating HN-CICs.

Central obesity and atherogenic dyslipidemia in metabolic syndrome are associated with increased risk for colorectal adenoma in a Chinese population.

BACKGROUND: Metabolic syndrome (MetS) is composed of cardiovascular risk factors including insulin resistance, obesity, dyslipidemia, and hypertension. Most of the components of MetS have been linked to the development of neoplasm. The purpose of this study was to evaluate the relationship between individual components of MetS and colorectal adenoma. METHODS: The study subjects were recruited from a pool of 4872 individuals who underwent a health check-up examination during the period January 2006 to May 2008. Each participant fulfilled a structured questionnaire. MetS was defined based on the America Heart Association and National Heart Lung Blood Institute criteria. Subjects with history of colon cancer, colon polyps, colitis, or prior colonic surgery were excluded. RESULTS: A total of 4122 subjects were included for final analysis (2367 men and 1755 women; mean age, 49.6 +/- 11.7 years). Of them, MetS was diagnosed in 708 men (29.9%) and in 367 women (20.9%). Among the patients with MetS, 34.6% had adenoma, 31.7% had hyperplastic polyps and 23.3% were polyp-free (p < 0.0001, Chi-square test). The adjusted OR for colorectal adenoma was significantly higher in the subjects with MetS (OR, 1.31, CI: 1.09-1.57). A stronger association between MetS and colorectal adenoma was found in men (OR:1.44, CI:1.16-1.80) than in women (OR:1.04, CI:0.74-1.46). The adjusted OR for adenoma increased as the number of MetS components increased (p for trend = 0.0001 ). When the individual components of MetS were analyzed separately, only central obesity (OR:1.36, CI:1.14-1.63), low HDL cholesterol levels (OR:1.30, CI:1.10-1.54) and high triglyceride levels (OR:1.26, CI:1.04-1.53) were independently associated with colorectal adenoma. CONCLUSIONS: Of the components of MetS analyzed in this study, central obesity and dyslipidemia are independent risk factors for colorectal adenoma. With regard to the prevention of colorectal neoplasm, life-style modification such as weight reduction is worthwhile.

Tid1 functions as a tumour suppressor in head and neck squamous cell carcinoma.

Human tumourous imaginal disc (Tid1), a human homologue of the Drosophila tumour suppressor protein Tid56, is involved in multiple intracellular signalling pathways such as apoptosis, cell proliferation, and cell survival. Here, we investigated the anti-tumourigenic activity of Tid1 in head and neck squamous cell carcinoma (HNSCC) in vitro and in vivo. Firstly, the clinical association between Tid1 expression and progression of HNSCC was explored. It was found that expression of Tid1 was negatively associated with tumour status, recurrence, and survival prognosis using immunohistochemical analysis of primary HNSCC patient tumour tissue. Secondly, ectopic expression of Tid1 in HNSCC cells was shown to significantly inhibit cell proliferation, migration, invasion, anchorage-independent growth, and xenotransplantation tumourigenicity. Thirdly, we showed that overexpression of Tid1 attenuated EGFR activity and blocked the activation of AKT in HNSCC cells, which are known to be involved in the regulation of survival in HNSCC cells. On the other hand, ectopic expression of constitutively active AKT greatly reduced apoptosis induced by Tid1 overexpression. Together, these findings suggest that Tid1 functions as a tumour suppressor in HNSCC tumourigenesis.

MicroRNA-7 targets T-Box 2 to inhibit epithelial-mesenchymal transition and invasiveness in glioblastoma multiforme.

The dysregulation of microRNA expression in cancer has been associated with the epithelial-mesenchymal transition (EMT) that triggers invasive ability and increases therapeutic resistance. Here, we determined the microRNA expression profile of seven tumor tissues from patients with glioblastoma multiforme (GBM) by use of microRNA array analysis. We discovered that microRNA-7 (miR-7) is consistently downregulated in all tumor samples. Using the microRNA.org algorithm, the T-box 2 gene (TBX2) was identified as a candidate gene targeted by miR-7. In contrast to miR-7, TBX2 had an increased expression in GBM tumors and was linked to poor prognosis. We confirmed that TBX2 mRNA and protein production are significantly repressed by overexpressing miR-7 in GBM cells in vitro. The reporter assay showed that miR-7 significantly represses the signal from luciferase with the 3' UTR of TBX2. Furthermore, TBX2 overexpression decreased E-cadherin expression and increased Vimentin expression, causing an increasing number of invaded cells in the invasion assay, as well as pulmonary metastasis in vivo. Our findings demonstrated that overexpression of TBX2 in GBM tumors via the downregulation of miR-7 leads to EMT induction and increased cell invasion.

Distinct population of highly malignant cells in a head and neck squamous cell carcinoma cell line established by xenograft model.

The progression and metastasis of solid tumors, including head and neck squamous cell carcinoma (HNSCC), have been related to the behavior of a small subpopulation of cancer stem cells. Here, we have established a highly malignant HNSCC cell line, SASVO3, from primary tumors using three sequential rounds of xenotransplantation. SASVO3 possesses enhanced tumorigenic ability both in vitro and in vivo. Moreover, SASVO3 exhibits properties of cancer stem cells, including that increased the abilities of sphere-forming, the number of side population cells, the potential of transplanted tumor growth and elevated expression of the stem cell marker Bmi1. Injection of SASVO3 into the tail vein of nude mice resulted in lung metastases. These results are consistent with the postulate that the malignant and/or metastasis potential of HNSCC cells may reside in a stem-like subpopulation.

Novel K6-K14 keratin fusion enhances cancer stemness and aggressiveness in oral squamous cell carcinoma.

Keratin intermediate filament (IF) is one component of cellular architectures, which provides necessary mechanical support to conquer environmental stresses. Recent findings reveal its involvement in mechano-transduction and the associated stem cell reprogramming, suggesting the possible roles in cancer development. Here, we report t(12;17)(q13.13;q21.2) chromosomal rearrangement as the most common fusion event in OSCC, resulting in a variety of inter-keratin fusions. Junction site mapping verified 9 in-frame K6-K14 variants, three of which were correlated with lymph node invasion, late tumor stages (T3/T4) and shorter disease-free survival times. When expressed in OSCC cells, those fusion variants disturbed wild-type K14 organization through direct interaction or aggregate formation, leading to perinuclear structure loss and nuclear deformation. Protein array analyses showed the ability of K6-K14 variant 7 (K6-K14/V7) to upregulate TGF-ß and G-CSF signaling, which contributed to cell stemness, drug tolerance, and cell aggressiveness. Notably, K6-K14/V7-expressing cells easily adapted to a soft 3-D culture condition in vitro and formed larger, less differentiated tumors in vivo. In addition to the anti-mechanical-stress activity, our data uncover oncogenic functionality of novel keratin filaments caused by gene fusions during OSCC development.

Predictors of Response to Autologous Dendritic Cell Therapy in Glioblastoma Multiforme.

Background: Glioblastoma (GBM) is the most common and lethal primary malignant glioma in adults. Dendritic cell (DC) vaccines have demonstrated promising results in GBM clinical trials. However, some patients do not respond well to DC therapy, with survival rates similar to those of conventional therapy. We retrospectively analyzed clinical and laboratory data to evaluate the factors affecting vaccine treatment. Methods: Forty-seven patients with de novo GBM were enrolled at China Medical University Hospital between 2005 and 2010 and divided into two subgroups. One subgroup of 27 patients received postsurgical adjuvant immunotherapy with autologous dendritic cell/tumor antigen vaccine (ADCTA) in conjunction with conventional treatment of concomitant chemoradiotherapy (CCRT) with temozolomide. The other 20 patients received only postsurgical conventional treatment without immunotherapy. Immunohistochemistry for CD45, CD4, CD8, programed death ligand 1 (PD-L1), and programed death 1 (PD-1) was performed on sections of surgical tumor specimens and peripheral blood mononuclear cells (PBMCs). Pearson's correlation, Cox proportional hazard model, and Kaplan-Meier analyses were performed to examine the correlations between the prognostic factors and survival rates. Results: Younger age (<57 years), gross total resection, and CCRT and PD-1+ lymphocyte counts were significant prognostic factors of overall survival (OS) and progression-free survival (PFS) in the ADCTA group. Sex, CD45+ lymphocyte count, CD4+ or CD8+ lymphocyte count, tumor PD-L1 expression, isocitrate dehydrogenase 1 mutation, and O6 methylguanine-DNA methyltransferase promoter methylation status were not significant factors in both groups. In the ADCTA group, patients with tumor-infiltrating lymphocytes (TILs) with a lower PD-1+/CD8+ ratio (≤0.21) had longer OS and PFS (median OS 60.97 months, P < 0.001 and PFS 11.2 months, P < 0.008) compared to those with higher PD-1+/CD8+ ratio (>0.21) (median OS 20.07 months, P < 0.001 and PFS 4.43 months, P < 0.008). Similar results were observed in patients' PBMCs; lymphocyte counts with lower PD-1+/CD8+ ratio (≤0.197) had longer OS and PFS. There was a significant correlation of PD-1+/CD8+ ratio between TILs and PBMCs (Pearson's correlation R2 = 0.6002, P < 0.001). By contrast, CD4-, CD8-, but PD-1+, CD45+ tumor-infiltrating lymphocytes have no impact on OS and PFS (P = 0.073 and P = 0.249, respectively). Conclusion: For patients receiving DC vaccine adjuvant therapy, better outcomes are predicted in patients with younger age, with TILs or PBMCs with lower PD-1+/CD8+ ratio, with gross tumor resection, and receiving CCRT.

Primary Sjögren's syndrome initially presenting as submandibular mucosa-associated lymphoid tissue lymphoma: A case report.

The present study reports the case of a 24-year-old female affected with primary Sjögren's syndrome (pSS), who presented with mucosa-associated lymphoid tissue (MALT) lymphoma of the submandibular gland. Reports of such cases, particularly in young patients, are very rare. The patient, who presented no oral or ocular symptoms prior to the development of the mass, underwent surgical ablation of the gland, and MALT lymphoma was diagnosed by histopathology. Since MALT lymphoma in the submandibular gland is rarely observed in otherwise healthy young females, a rheumatologist and an oncologist were consulted. Following a number of immunological tests, the results of the Schirmer's and Saxon tests were negative. However, the antinuclear antibody test revealed a speckled appearance, and there was also strong positivity for the serological markers of Sjögren's syndrome. Consequently, pSS was diagnosed, despite the fact that the patient did not fulfill all the diagnostic criteria for the disease. Therefore, MALT lymphoma in a single salivary gland should be used as a differential diagnosis for Sjögren's syndrome in young asymptomatic patients. Additionally, a multidisciplinary team is required for the treatment and management of these patients.