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Background The incidence of human papillomavirus (HPV) positive oropharyngeal cancer (OPC) is rising but HPV negative OPC is decreasing in Western countries. In Taiwan, the incidence of HPV negative OPC is common but the incidence of HPV positive OPC remains unknown. The objective of this study is to estimate the incidence trend and the survival of HPV positive OPC in Taiwan. Methods Between 1999 and 2014, primary tumor tissues from 425 incident OPCs were obtained from 5 medical centers in Taiwan. 408 OPCs were evaluated by the EasyChip HPV genotyping (King-Car, I-Lan, Taiwan) and 369 OPCs by p16 staining. The clinical data were retrospectively obtained from the medical records. Results In our study, 29% of OPCs were HPV positive. The percentage of HPV positive OPC was stable from 1999 to 2014 (25% (1999–2002), 30% (2003–2006), 30% (2007–2010), 29% (2011–2014)). The estimated crude incidence rate of HPV positive OPC increased significantly from 0.62 (1999–2002), 1.06 (2003–2006), 1.52 (2007–2010) to 1.74 (2011–2014) per 100,000 person-year. The sensitivity and specificity of p16 staining for positive HPV infection were 92% and 91%, respectively. The 5-year overall survival rates for patients with HPV positive OPC and with HPV negative OPC were 67.8% and 49.0%, respectively (HR = 0.52 (0.35–0.76), p = 0.0005). Patients with HPV positive OPC but no betel nut/cigarette exposure had the best overall survival (5-year: 88.2%, p p p Conclusion The incidence of HPV positive OPC is increasing along with HPV negative OPC, which leads to stably low percentage of HPV positive OPC in Taiwan. HPV positive OPC may become an important head and neck cancer when the incidence of HPV negative OPC declines in the near future. P16 is a useful surrogate marker for HPV infection in OPC and a good prognostic indicator for treatment outcome of OPC. Patients with HPV positive OPC but no betel nut/cigarette exposure has an excellent prognosis. Betel nut/cigarette exposure significantly worsens the prognosis of HPV positive OPC.

Extranodal extension (ENE) is a crucial prognostic factor of oral squamous cell carcinoma (OSCC). However, the role of ENE in regional recurrence (rENE) remains unclear. The purpose of our study is to assess the salvage outcome according to the presence of rENE in oral cancer patients with isolated nodal recurrence.Oral cancer patients diagnosed with isolated nodal recurrence at the National Taiwan University Hospital between January 2010 and December 2015 were reviewed. All patients were classified into two groups: with and without rENE. The treatment included salvage neck dissection (ND) ± metronomic chemotherapy, salvage ND and radiation (RT)/concurrent chemoradiation (CCRT), Salvage RT/CCRT alone, metronomic chemotherapy, or supportive care.We analyzed 198 patients, 156 with rENE and 42 without rENE. rENE presented more frequently in patients with initial ENE+ (OR = 3.17, p = 0.04), prior RT+ (OR = 2.96, p = 0.02), initial N2/N3 (OR = 2.76, p = 0.01), and recurrent LN size1.5 cm (OR = 2.33, p = 0.03). The extent of rENE were also significantly different in these patients. The 2-year disease-free survival for patients with and without rENE were 15.7% and 31.7%, respectively (p = 0.002). The 2-year overall survival for patients with and without rENE were 19.6% and 43.9%, respectively (p = 0.004). For patients without rENE, those received salvage ND had better survival outcome (p 0.001). By contrast, for patients with rENE, those received salvage RT/CCRT had better survival outcome (p 0.001).The rENE is frequently present (78.79%) in OSCC patients with isolated nodal recurrence. Individualized treatment modalities based on the presence of rENE should be recommended to achieve better salvage outcomes.

Besides being a predictive biomarker of response to immunotherapy in lung cancer in general, programmed death-ligand 1 (PD-L1) is not so well correlated with treatment outcomes of lung adenocarcinoma (ADC) harbouring epidermal growth factor receptor (EGFR) mutations, as reported studies are inconclusive and seldom addressed the issues of response to treatment and resistance. The primary objective is to evaluate the association of PD-L1 and EGFR tyrosine kinase inhibitor (TKI) efficacy, resistance, and relevant clinical outcomes. The secondary objective is to further explore the tumour microenvironments of EGFR mutant tumours with different PD-L1 expression.Using immunohistochemical (IHC) staining, we retrospectively tested PD-L1 expression (Dako 22C3) in the pre-treatment tumours from advanced EGFR mutant lung ADC patients, of whom all were treated with TKIs. Multiplex IHC assay was applied for exploring immune cells in tumour microenvironments.A total of 153 Taiwanese patients were enrolled in our study, of whom a majority of cases were female (58.9%) and non-smokers (75.8%). The objective response rate (ORR) to EGFR TKI and progression-free survival (PFS) were better in patients with PD-L1 expression50% (ORR/PFS in PD-L1 0% versus 1-49% versus ≥50%: 65.6%/12.5 months versus 56.4%/12.8 months versus 38.9%/5.9 months, P 0.05). The multivariate analysis showed that PD-L150% was an independent prognostic factor for longer PFS (hazard ratio (HR) 0.433, 95% confidence interval (CI) 0.250-0.751, P = 0.003). Furthermore, tumours with higher PD-L1 expression were less likely to develop a secondary T790M mutation (T790M+ in PD-L1 0% versus 1-49% versus ≥50%: 53.7% versus 35.7% versus 10%, P = 0.024). Multiplex IHC tests were applied in 15 cases and revealed a potential correlation between PD-L1, immune cells, and EGFR TKI responses.Lower pre-treatment PD-L1 is associated with better ORR, PFS, and higher frequency of T790M resistance in EGFR TKI-treated lung ADC patients.

To determine the tumor genomic, immunologic expression, and risk factors of treatment outcomes for patients with double head and neck squamous cell carcinoma (HNSCC) and esophageal squamous cell carcinoma (ESCC).We reviewed patients with double HNSCC and ESCC between 1995 and 2014. The TP53 genomic mutation, CD8+ tumor infiltrating lymphocytes (TIL) and tumor programmed cell death ligand 1 (PD-L1) expression of paired HNSCC and ESCC were analyzed.A total of 116 patients (57 metachronous and 59 synchronous) were included. There were 88 (75.86%) patients with HNSCC and 80 (68.97%) with ESCC harboured TP53 disruptive mutation. Nearly 106 (91.38%) patients had different clonality of TP53 mutation in paired HNSCC and ESCC. The immunologic expression of synchronous and metachronous patients was significantly different. Compared to the metachronous patients, the synchronous patients had significantly higher HNSCC CD8+ TIL (p = 0.03), ESCC CD8+ TIL (p 0.001), HNSCC PD-L1+ tumor proportion score (TPS, p = 0.04), and ESCC PD-L1+ TPS (p = 0.04). Furthermore, among the synchronous patients, the immunologic expression between HNSCC and ESCC was significantly correlated. The CD8+ TIL and PD-L1 TPS had strongly (r = 0.63, p 0.0001) and moderately (r = 0.42, p = 0.001) positive correlations, respectively. Finally, advanced stage (III/IV) HNSCC was a significant factor for disease-free (p = 0.03) and overall survival (p = 0.005).In patients with double HNSCC and ESCC, nearly all HNSCC and ESCC were of multicentric origin. For the synchronous patients, there was more adaptive immune resistance in HNSCC and ESCC. The immunologic expression between paired HNSCC and ESCC was also significantly correlated.

Mixed squamous cell and glandular papilloma (mixed papilloma) is a very rare tumor, with fewer than 25 cases having been reported in the literature. Although a scattering of cases of p16Ink4a overexpression have been described to date, no human papillomavirus (HPV) DNA has been detected in these tumors, either by in situ hybridization (ISH) or polymerase chain reaction (PCR). This is the first case of mixed papilloma with PCR-confirmed HPV genotype 16, 35, 51 infections in an 18-year-old non-smoking male, coexisting with multiple atypical adenomatous hyperplasias (AAHs). Histologically, this tumor shows a predominant papillary architecture, covered by a mixture of stratified squamous cells, ciliated or non-ciliated cuboidal to columnar cells, mucous cells, and scattered goblet cells. Immunohistochemically, the squamous component was positive for p40, and the glandular cells were focally positive for TTF-1. Both components were diffusely immunoreactive to CK7. In addition, BRAF V600E mutation was also first demonstrated in mixed papilloma, but not in the AAHs. These findings suggest that HPV infection and the BRAF mutation may be important in the pathogenetic role in young non-smoking patients.

Background Programmed death-ligand 1 (PD-L1) expression is associated with clinical outcomes of epidermal growth factor receptor (EGFR) mutant lung adenocarcinoma (ADC) treated with tyrosine kinase inhibitors (TKIs). However, whether PD-L1 expression plays a role in anaplastic lymphoma kinase (ALK)-positive lung ADC is unknown. We aimed to evaluate the impact of PD-L1 in patients with ALK-positive lung ADC receiving crizotinib. Materials and Methods PD-L1 expression was identified by immunohistochemistry (IHC). Reverse transcriptase-polymerase chain reaction was used for ALK variant detection, and immunofluorescence-based multiplex staining was applied for exploring immune cells in tumor microenvironments. Results A total of 78 patients with ALK-positive advanced ADC were enrolled in our study, of whom 52 received crizotinib. Compared with EGFR/ALK wild-type tumors, PD-L1 expression was lower in ALK-positive ADC. ALK fusion variants were identified in 32 patients, and those with variant 3 and 5 (short variants) had higher PD-L1 expression than those with other variants. The crizotinib objective response rate (ORR) and progression-free survival (PFS) was better in tumors with negative PD-L1 expression (ORR/PFS in PD-L1 0% vs. 1%–49% vs. 50%–100%: 60.7%/11.8 months vs. 38.5%/6.5 months vs. 36.4%/4.0 months, p = .007/.022). The multivariate Cox proportional hazards model revealed that PD-L1 0% (vs. ≥1%) was an independent factor for longer PFS (adjusted hazard ratio 0.322, 95% confidence interval 0.160–0.650, p = .002). Multiplex IHC in three cases showed a varied extent of immune cell infiltrations in tumors with different PD-L1 expression. Conclusion Positive PD-L1 expression was associated with unfavorable clinical outcomes in patients with ALK-positive lung ADC receiving crizotinib. Implications for Practice Not all lung adenocarcinoma with sensitizing driver mutations experienced durable responses to small-molecule tyrosine kinase inhibitors (TKIs). Similar to the negative impact of programmed death-ligand 1 (PD-L1) in epidermal growth factor receptor mutant tumors treated with TKIs, this study demonstrated that positive PD-L1 expression was also associated with worse response rate and shorter progression-free survival of anaplastic lymphoma kinase (ALK)-positive adenocarcinoma treated with crizotinib. Among different ALK fusion partners, tumors with short variants (V3 and V5) had higher PD-L1 compared with long variants (V1, V2, and V6). Testing PD-L1 before initiating crizotinib for ALK-positive lung cancer could be a simple method to provide important prognostic information.

Background Our aim was to investigate the prognostic significance of tumor-infiltrating lymphocytes (TILs) in operable tongue cancer patients. Methods The presence of CD3+, CD4+, CD8+, and forkhead box protein P3-positive (FOXP3+) TILs in tumor tissues obtained from 93 patients during surgery was examined using immunohistochemistry. Results The 3-year overall survival (OS) of patients with a low CD8/FOXP3 ratio was significantly lower than that of patients with a high CD8/FOXP3 ratio (63.8% vs. 87.3%, p = 0.001). Patients with high FOXP3 had a significantly lower 3-year regional recurrence-free survival (RRFS) than did patients with low FOXP3 (49.3% vs. 87.3%, univariate log rank p = 0.000). A low CD4/FOXP3 ratio (68.4% vs. 93.7%, univariate log rank p = 0.002) was significantly unfavorable prognostic factors for 3-year distant metastasis-free survival (DMFS). Conclusions In addition to clinicopathological characteristics, TIL markers represent prognosticators for clinical outcomes.

Background/Purpose To investigate the M1/M2 polarity of macrophages in the endometrium among different menstrual cycles, normal and abnormal pregnancies, and unexplained recurrent spontaneous abortions (RSAs). Methods Endometrial tissue was obtained from 43 patients undergoing hysterectomy, either in the follicular phase (Group 1, n = 23) or in the luteal phase (Group 2, n = 20). In addition, decidual tissue was obtained from 53 pregnant women during the first trimester, either of normal pregnancies (Group 3, n = 12) or abnormal pregnancies (Group 4: spontaneous abortions, n = 20; Group 5: unexplained RSA, n = 21). Using immunofluorescence to examine the M1 and M2 macrophages in the endometrium and deciduae from cases with different menstrual phases and various pregnancy outcomes, respectively, we endeavored to learn the possible pathophysiology of abortions. Results M1 macrophages were abundant in the deciduae of spontaneous abortions and unexplained RSA, whereas the frequency of M2 macrophages was significantly higher in the endometrium of luteal phase and normal pregnancies. Conclusion M2 polarization is important for early successful pregnancies in humans.

This study aims to assess whether geranylgeranylacetone (GGA) could reduce ototoxicity induced by cisplatin through upregulation of not only heat shock protein(HSP)-70, but also HSP-27 and HSP-40, and to study if GGA would reduce cisplatin-induced increase in oxidative stress. 48 guinea pigs were used in this study and treated with the following regimen: 0.5% CMC (sodium carboxymethyl cellulose) control for 7days, GGA (600mg/kg/d) for 7days, a combination of GGA (600mg/kg) for 7days and then one dose of 10mg/kg cisplatin (GGA+Cis), and a combination of CMC for 7days and then 10mg/kg cisplatin (cisplatin group). Auditory brainstem response (ABR) measurement was performed in each animal at time before treatment and 7days after the last dose. Additionally, HSPs, nitric oxide (NO), and lipid peroxidation (LPO) levels in cochlear membranous tissues were assessed. The mean ABR thresholds in the cisplatin group were significantly (p

Lung cancer in East Asia is characterized by a high percentage of never-smokers, early onset and predominant EGFR mutations. To illuminate the molecular phenotype of this demographically distinct disease, we performed a deep comprehensive proteogenomic study on a prospectively collected cohort in Taiwan, representing early stage, predominantly female, non-smoking lung adenocarcinoma. Integrated genomic, proteomic, and phosphoproteomic analysis delineated the demographically distinct molecular attributes and hallmarks of tumor progression. Mutational signature analysis revealed age- and gender-related mutagenesis mechanisms, characterized by high prevalence of APOBEC mutational signature in younger females and over-representation of environmental carcinogen-like mutational signatures in older females. A proteomics-informed classification distinguished the clinical characteristics of early stage patients with EGFR mutations. Furthermore, integrated protein network analysis revealed the cellular remodeling underpinning clinical trajectories and nominated candidate biomarkers for patient stratification and therapeutic intervention. This multi-omic molecular architecture may help develop strategies for management of early stage never-smoker lung adenocarcinoma.

Background The Slug-E-cadherin axis plays a critical role in non-small-cell lung cancers (NSCLCs) where aberrant upregulation of Slug promotes cancer metastasis. Now, the post-translational modifications of Slug and their regulation mechanisms still remain unclear in lung cancer. Hence, exploring the protein linkage map of Slug is of great interest for investigating the scenario of how Slug protein is regulated in lung cancer metastasis. Methods The Slug associated proteins, Ubc9 and SUMO-1, were identified using yeast two-hybrid screening; and in vitro SUMOylation assays combined with immunoprecipitation and immunoblotting were performed to explore the detail events and regulations of Slug SUMOylation. The functional effects of SUMOylation on Slug proteins were examined by EMSA, reporter assay, ChIP assay, RT-PCR, migration and invasion assays in vitro, tail vein metastatic analysis in vivo, and also evaluated the association with clinical outcome of NSCLC patients. Results Slug protein could interact with Ubc9 and SUMO-1 and be SUMOylated in cells. Amino acids 130–212 and 33–129 of Slug are responsible for its binding to Ubc9 and protein inhibitor of activated STAT (PIAS)y, respectively. SUMOylation could enhance the transcriptional repression activity of Slug via recruiting more HDAC1, resulting in reduced expression of downstream Slug target genes and enhanced lung cancer metastasis. In addition, hypoxia could increase Slug SUMOylation through attenuating the interactions of Slug with SENP1 and SENP2. Finally, high expression Slug and Ubc9 levels were associated with poor overall survival among NSCLC patients. Conclusions Ubc9/PIASy-mediated Slug SUMOylation and subsequent HDAC1 recruitment may play a crucial role in hypoxia-induced lung cancer progression, and these processes may serve as therapeutic targets for NSCLC. Electronic supplementary material The online version of this article (10.1186/s13046-018-0996-8) contains supplementary material, which is available to authorized users.

Figure S2. The activities of Slug mutants. (a) Mutation of individual lysine affects the SUMOylated level of Slug. HEK293T cells were cotransfected with plasmids encoding different 3xFlag-tagged Slug mutants and GFP-tagged SUMO-1. The lysates were used to examine the SUMOylation levels by immunoblotting with anti-Flag antibodies. (b) Different levels of SUMOylation between Slug mutants. HEK293T cells were transfected with expression vectors encoding GFP-tagged SUMO-1 and different 3xFlag-tagged Slug mutants (22 M, all lysines were replaced with arginines; 5 M: lysines at 239, 240, 244, 248, and 258 were replaced with arginines; 6 M: lysines at 188, 239, 240, 244, 248, and 258 were replaced with arginines). These lysates were also examined by immunoblotting with anti-Flag antibodies. The asterisk and arrowhead indicate Slug modified and not modified by SUMO-1, respectively. (c) The transcriptional repression activity of wild-type and mutant Slug proteins. HEK293T cells were cotransfected with the SBS–Gal4–luciferase reporter and Gal4–VP16 activator expression plasmids together with the wild-type or mutant Slug expression plasmid (8 M: lysines at 135, 145, 188, 239, 240, 244, 248, and 258 were replaced with arginines), and the luciferase assay was performed to determine the transcriptional repression activity of Slug. Immunoblotting results are presented alongside the luciferase assay results to demonstrate the expression of the Slug mutant proteins. (d) The DNA-binding activity of wild-type and mutant Slug proteins. The wild-type and mutant Slug proteins used in the EMSA were produced using an in vitro transcription/translation system. The protein expression levels were evaluated by immunoblotting with anti-Slug antibodies (top panel). Phosphor image analysis of the EMSA gel showing 32P-labeled E-box oligonucleotides incubated with in vitro-translated proteins (4 μl) or with Slug antibodies (Ab: antibody, 0.3 μg) (bottom panel). (PDF 152 kb)

Figure S11. Effects of Slug and Slug5M overexpression on cultured cell proliferation. HEK293 cells were driven to express wild-type or mutant Slug using a lentiviral system. Cells were counted at the indicated time points after plating. No significant difference in the cell proliferation rate was found between the different cell lines based on one-way ANOVA. Error bars indicate mean values Âą SEM. (PDF 69 kb)

Figure S8. HDAC1 stronger associated with the E-cadherin promoter in Hop62 cells overexpressing Ubc9 compare with the vector control cells. The normal mouse IgG and HDAC1 antibodies were used to pull down protein-DNA complexes in Hop62 cells with or without Ubc9 overexpression; and the E-cadherin promoter level in the samples was determined by PCR using a gene-specific primer set. Input, an aliquot of each sample was prepared and used as a template for PCR to examine the level of the E-cadherin promoter before immunoprecipitation (IP). (PDF 25 kb)

Figure S5. Structure of the Slug/PIASy/Ubc9/SUMO-1 complex. (a) Schematic showing the regions of Slug that interact with PIASy, Ubc9, and SUMO. Slug is 268 amino acids in length and contains a SNAG repression domain at its N-terminus and five zinc finger (ZnF) domains at its C-terminus. ND means no detection. (b) A 3D structure of Slug/PIASy/Ubc9/SUMO-1 complex was generated using prediction software (orange, Slug; purple, PIASy; green, Ubc9; gray, SUMO-1). A rotated view of this complex is shown in the lower panel. (PDF 127 kb)

Figure S7. Slug recruits corepressors more abundantly than Slug5M. The nuclear fractions of Slug- and Slug5M-overexpressing HEK293 cells were obtained by adding hypotonic buffer to the cells. Subsequently, the samples were subjected to immunoprecipitation using an anti-Slug antibody. The accompanying precipitates were analyzed by immunoblotting using the indicated antibodies (left panel). Lamin B was used as a nuclear marker. The relative densitometry results (the results for Slug were normalized to one) were calculated using two programs (ImageJ and GelPro3.1), and the average values are plotted in the right panel. (PDF 61 kb)

Figure S6. Characterization of Slug and Slug5M protein. (a) The DNA-binding ability of Slug is not altered by the inserted mutations. Equal amounts of in vitro-translated Slug and Slug5M were used in the EMSAs (left panel). Slug and Slug5M bound to the E-box C probes in a dose-dependent manner (+: 0.1 μl; ++: 0.3 μl; +++: 1 μl) (right panel). Anti-Slug antibodies were used to confirm that the shifted bands were formed specifically by Slug and Slug5M. (b) The protein stability of Slug is not altered by the inserted mutations. Protein stability was not significantly different between the wild-type and mutant forms of Slug. Slug- and Slug5M-overexpressing HEK293 cells were treated with cycloheximide (CHX) to prevent further protein synthesis for the indicated periods. The expression of Slug was analyzed by immunoblotting. β-actin was used as the internal control. Relative densitometry results are plotted in the bottom panel. (PDF 68 kb)

Figure S9. SUMOylation affects the expression of Slug-regulated downstream targets. (a) CL1–5 cells were induced to express the wild-type and mutant Slug, respectively, using a lentiviral system. The mRNA expression of the indicated genes was determined via RT-PCR. Gβ-like was used as the internal control. (b) The protein expression of the Slug downstream target, E-cadherin, in HEK293 (left) and CL1–2 (right) over-expressing Slug wild-type or Slug5M cells. The results were analyzed by immunoblotting with the indicated antibodies. β-actin was used as the internal control. (PDF 174 kb)

Figure S4. Direct interaction of Slug with PIAS family members. A pull-down assay was used to determine the physical interaction between Slug and PIAS family members. Recombinant GST and GSTâ Slug proteins were produced from bacteria, and the translated products of HA-tagged PIAS family member genes were obtained using an in vitro transcription/translation system. The production of these proteins was demonstrated by immunoblotting using anti-GST and anti-HA antibodies, respectively. GSTâ Slug was used in the pull-down assay for in vitro interaction with HA-tagged PIAS family members. The GST protein alone was used as a negative control. (PDF 24 kb)

Pulmonary peripheral-type squamous cell carcinoma (p-SqCC) has been increasing in incidence. However, little is known about the clinicopathologic features of p-SqCC. This study aimed to investigate the clinicopathologic characteristics and clinical outcomes of p-SqCC compared with central-type SqCC (c-SqCC) in a large cohort of surgically resected lung SqCC patients with long-term follow-up results. The study included 268 patients with SqCC who underwent surgical resection at the authors’ institute from January 1990 to September 2013. The mean follow-up period was 67.1 months. The clinicopathologic and genetic characteristics were investigated in relation to their association with progression-free survival (PFS) and overall survival (OS) based on tumor location. The study cohort included 120 patients with p-SqCC and 148 patients with c-SqCC. Compared with c-SqCC, p-SqCC was correlated with older age (p = 0.002), female sex (p = 0.033), better performance status (p

Mixed squamous cell and glandular papilloma (mixed papilloma) is a very rare tumor, with fewer than 25 cases having been reported in the literature. Although a scattering of cases of p16

Summary Objective The treatment strategies for advanced oral squamous cell carcinoma (OSCC), especially with necrotic changes, are not effective. The programmed death ligand 1 (PD-L1) immune escape may be one of the underlying sources of resistance. Furthermore, anti-PD-L1 directed immunotherapy may be another choice for adjuvant therapy. Therefore, the expression of PD-L1 in advanced OSCC with necrotic changes is very important. Materials and methods A total of 218 eligible patients with advanced stage (stage III/IV) OSCC and neck metastasis were enrolled. The presence of necrosis was reviewed by pretreatment magnetic resonance imaging. Paired paraffin-embedded primary tumor and metastatic lymph nodes (LN) sections were stained with antibodies against hypoxia-inducible factor-1α (HIF-1α) and PD-L1. Moderate-to strong HIF-1α nuclear staining in >10% and cell surface PD-L1 expression in >5% of OSCC cells were recorded as a positive result. Results For advanced OSCC with necrotic changes, there was substantial agreement in primary tumor (kappa value 0.54) and almost perfect agreement in metastatic LN (kappa value 0.86) between HIF-1α and PD-L1 expression. The patients with both necrosis and positive PD-L1 expression in OSCC surrounding necrosis had worse disease control and survival outcomes. After multivariate analysis, metastatic LN necrosis and positive PD-L1 expression were found to be significant independent adverse factors. Conclusion Advanced OSCC patients with both necrosis and positive PD-L1 expression in OSCC surrounding necrosis had worse outcome. The aggressive behavior of advanced OSCC could be partially related to PD-L1 immune escape. These patients may be good candidates for anti-PD-L1 immunotherapy.

Melanoma differentiation-associated gene-9 (MDA-9)/Syntenin is a novel therapeutic target because it plays critical roles in cancer progression and exosome biogenesis. Here we show that Slug, a key epithelial-mesenchymal-transition (EMT) regulator, is a MDA-9/Syntenin downstream target. Mitogen EGF stimulation increases Slug expression and MDA-9/Syntenin nuclear translocation. MDA-9/Syntenin uses its PDZ1 domain to bind with Slug, and this interaction further leads to HDAC1 recruitment, up-regulation of Slug transcriptional repressor activity, enhanced Slug-mediated EMT, and promotion of cancer invasion and metastasis. The PDZ domains and nuclear localization of MDA-9/Syntenin are both required for promoting Slug-mediated cancer invasion. Clinically, patients with high MDA-9/Syntenin and high Slug expressions were associated with poor overall survival compared to those with low expression in lung adenocarcinomas. Our findings provide evidence that MDA-9/Syntenin acts as a pivotal adaptor of Slug and it transcriptionally enhances Slug-mediated EMT to promote cancer invasion and metastasis.

Phosphaturic mesenchymal tumours (PMTs) are uncommon soft tissue and bone tumours that typically cause hypophosphataemia and tumour-induced osteomalacia (TIO) through secretion of phosphatonins including fibroblast growth factor 23 (FGF23). PMT has recently been accepted by the World Health Organization as a formal tumour entity. The genetic basis and oncogenic pathways underlying its tumourigenesis remain obscure. In this study, we identified a novel FN1–FGFR1 fusion gene in three out of four PMTs by next-generation RNA sequencing. The fusion transcripts and proteins were subsequently confirmed with RT-PCR and western blotting. Fluorescence in situ hybridization analysis showed six cases with FN1–FGFR1 fusion out of an additional 11 PMTs. Overall, nine out of 15 PMTs (60%) harboured this fusion. The FN1 gene possibly provides its constitutively active promoter and the encoded protein's oligomerization domains to overexpress and facilitate the activation of the FGFR1 kinase domain. Interestingly, unlike the prototypical leukaemia-inducing FGFR1 fusion genes, which are ligand-independent, the FN1–FGFR1 chimeric protein was predicted to preserve its ligand-binding domains, suggesting an advantage of the presence of its ligands (such as FGF23 secreted at high levels by the tumour) in the activation of the chimeric receptor tyrosine kinase, thus effecting an autocrine or a paracrine mechanism of tumourigenesis. Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Background Globo H is a tumor-associated carbohydrate antigen exclusively expressed in cancer cells rather than normal tissue. Globo H has been found on many cancers of epithelial origins, and become an attractive target for cancer vaccine. Objectives We aimed to study the expression of Globo H in non-small cell lung cancer (NSCLC) patients, and correlated its expression with common driver mutations, clinical outcomes, and status of immune checkpoint, programmed death-ligand 1 (PD-L1). Methods The study enrolled 228 patients with surgically resected stage I NSCLC, including 139 patients with adenocarcinoma (ADC) and 89 patients with squamous cell carcinoma (SqCC). Using immunohistochemistry, tumors with moderate to strong membranous staining in ⩾ 1% tumor cells per section were scored as positive Globo H expression. Driver mutations including EGFR, KRAS, BRAF were detected by direct sequencing, while ALK, PI3KCA, FGFR1 and PD-L1 expression was detected by immunohistochemical (IHC) staining. Results Positive Globo H expression was detected in 88 of the 228 (38.6%) patients. These included 51 of 139 (36.7%) patients with ADC and 37 of 89 (41.6%) patients with SqCC. Positive Globo H expression was significantly associated with EGFR mutation and PD-L1 expression in the ADC group, and PI3KCA overexpression in the SqCC group. The survival analysis showed that Globo H expression was not an independent prognostic factor in stage I NSCLC. Conclusions Globo H expression was correlated with specific driver mutations in ADC and SqCC NSCLC tumors, as well as PD-L1 status. Immunotherapy targeting Globo H may have potential application in lung cancer treatment.

Given salvage treatment for recurrent nasopharyngeal carcinoma (NPC) remains a clinical dilemma, immunotherapy targeting NPC-specific immunosuppression may bring new hope. We analyzed the expression of CD8, CD4, Foxp3 and Tim-3 in lymphocytes, and of Galectin-9 in tumour cells between paired primary and recurrent NPC from 95 patients and we noted that there was significant increase in the expression of Galectin-9+ tumour cells (p

Mycobacterium avium complex-induced lung disease (MAC-LD) becomes important due to its increasing prevalence. Attenuated cellular immunity associated with programmed cell death (PD)–1 may play a pathophysiological role in MAC-LD but lacks of investigation. We enrolled 80 participants in this prospective study, including 50 with MAC-LD and 30 healthy controls. Peripheral blood mononuclear cells (PBMCs), lymphocytes and monocyte-derived macrophages were used for MAC antigen stimulation. Patients with MAC-LD had lower tumor necrosis factor-α and interferon-γ responses compared to the healthy controls in PBMC stimulation assays with MAC bacilli. These responses improved after MAC treatment. The PD-1 and PD ligand expressions and apoptosis were higher in the lymphocytes of the patients with MAC-LD compared to the controls. Both PD-1 and apoptosis on T lymphocytes were significantly increased in the patients with MAC-LD, either by direct MAC stimulation or by MAC-primed macrophage activation. Partially blocking PD-1 and the PD ligand with antagonizing antibodies in the stimulation assay significantly increased the cytokine production of IFN-γ and decreased the apoptosis on T lymphocytes. In conclusion, the patients with MAC-LD have attenuated lymphocyte immunity, which might be associated with increasing activation of PD-1 and PD-1 ligand. Regulating such activation might improve the lymphocytic secretion of IFN-γ and reduce apoptosis.

Aims Dedifferentiated liposarcoma (DDLPS) is traditionally defined as a non-lipogenic high-grade sarcoma arising from a well-differentiated liposarcoma that confers metastatic potential. Recently, DDLPSs with lipoblastic differentiation, i.e. morphologically lipogenic DDLPSs, were reported. Because of the lipoblastic differentiation, these tumours caused confusion, and were reported under different names. However, cytogenetic and molecular studies have revealed their DDLPS nature. So far, the cases reported have been high-grade pleomorphic liposarcoma-like tumours. In this study we have collected another series that contains low-grade tumours, and expand the histological spectrum. Methods and results Eighteen cases of DDLPS with lipoblastic differentiation from various anatomical locations were analysed by routine histology, immunohistochemistry, and MDM2 fluorescence in-situ hybridization. Two main histological patterns were seen: one featured a spindle cell sarcoma containing lipoblasts with variable nuclear pleomorphism, and the other a pleomorphic liposarcoma-like tumour including the epithelioid variant. Two cases showed low nuclear grade and lipogenic activity in the metastatic foci. CDK4, MDM2 and p16INK4a overexpression was seen in all except one case. MDM2 amplification was found in all 16 cases tested. Conclusions We have expanded the spectrum of this variant of DDLPS to include low-grade tumours, in which a careful search for increased mitotic activity is essential. Like conventional DDLPS, these tumours are capable of metastasis.

This study aimed to investigate the clinicopathological factors that influence recurrence and survival in patients who undergo operations for T3–4 hypopharyngeal squamous cell carcinomas (SCCs). One hundred and five patients who underwent surgery between 2001 and 2008 for advanced hypopharyngeal SCCs were consecutively enrolled and reviewed. The pretreatment neutrophil-to-lymphocyte ratio (NLR; median 3.22, range 0.62–46.50) was associated with disease recurrence and patient survival. A difference in the 5-year cumulative disease recurrence rate between patients with high (≥3.22) and low (

This study aimed to investigate the clinicopathologic prognostic predictors of stage 4 hypopharyngeal cancer and to extend the traditional tumor-node-metastasis classification system to advance its predictive ability.The study enrolled 120 patients with pathologically stage 4 hypopharyngeal cancer treated with pharyngolaryngectomy and neck dissection between 2001 and 2007.The study showed a 5-year overall survival (OS) of 44.6%, a disease-specific survival (DSS) of 51.6%, and a disease-free survival (DFS) of 48% for all the patients. In the multivariate analysis, a lymph node (LN) ratio of 0.113 or higher was a significant poor prognostic factor for OS (hazard ratio [HR] 1.89; 95% confidence interval [CI] 1.17-3.05; p = 0.009), DSS (HR 2.17; 95% CI 1.29-3.64; p = 0.003), and DFS (HR, 2.24; 95% CI 1.12-4.52; p = 0.024) in stage 4 hypopharyngeal cancer. In addition, pretreatment neutrophil-lymphocyte ratio, lymphovascular invasion, and margin status also were predictors of survival outcomes. Furthermore, the study found that disease recurrence differed significantly between the patients with a LN ratio of 0.113 or higher (68.2%) and those with a LN ratio lower than 0.113 (39.5%) (p = 0.002).A LN ratio of 0.113 or higher is a strong predictor of disease recurrence and survival for patients with stage 4 hypopharyngeal cancer.

We report a case of giant cell tumor of the left distal femur with simultaneous bilateral pulmonary metastases. Pathologic examination of the left lung nodule showed a metastatic giant cell tumor with a noncontinuous ossified rim, and the right lung nodules were totally hyalinized or ossified without residual giant cell tumor components. Hyalinization was a consistent finding in both the primary bone lesion and the pulmonary metastases. Because the patient did not receive chemotherapy or radiotherapy before her surgical procedure, we believe that these changes represent spontaneous tumor regression.

Introduction Tumor tissue is often not obtainable or suitable for molecular-based epidermal growth factor receptor ( EGFR ) mutational analysis in advanced non–small-cell lung cancer (NSCLC). This retrospective and single-institution study was conducted to evaluate the role of effusion immunocytochemistry using two EGFR mutant-specific antibodies for the detection of relevant EGFR mutations in NSCLC, along with the selection of candidates for first-line therapy with EGFR tyrosine kinase inhibitors (TKIs). Methods Immunocytochemistry using two antibodies binding specifically to the major forms of mutant EGFR, L858R, and E746-A750 deletion (delE746-A750), was performed on cell blocks of malignant pleural effusion (MPE) from 78 patients with lung adenocarcinoma, who received first-line EGFR TKIs. The yield of EGFR -mutation detection and prediction of response rate and progression-free survival to TKI treatment by immunocytochemistry were compared with those by clinical characteristics and EGFR sequencing using cell-derived RNA from MPEs. Results Of the 78 MPE samples, direct sequencing using cell-derived RNA identified L858R mutation in 42 cases, deletions in exon 19 in 12 cases (delE746-A750 in eight cases), other types of mutations in three cases, and wild-type EGFR in 21 cases. Effusion immunocytochemistry with these two mutant-specific antibodies exhibited a sensitivity of 71% and 88% and a specificity of 86% and 96% for identifying predefined L858R and delE746-A750 mutations, respectively. Effusion immunocytochemistry provided a superior prediction of tumor response and progression-free survival to first-line EGFR TKIs than did clinical characteristics like sex and smoking status. Patients whose effusion immunocytochemistry showed a reaction to either of the two antibodies had a comparable TKI response rate (67% versus 72%) to those with EGFR mutations assessed by direct sequencing from cell-derived RNA. Conclusions Effusion immunocytochemistry could be introduced into clinical practice to identify more NSCLC patients likely to have benefit from first-line TKI treatment, especially for those without adequate tissue for molecular-based EGFR analysis.

In order to improve prognostic applications and treatment decisions, we report our experiences of visceral pleural surface invasion (VPSI) in non-small cell lung cancers (NSCLCs) with pleural retraction. A total of 321 NSCLCs with pleural retraction were identified by carefully inspecting surgically resected specimens. The extent of pleural invasion, including the use of elastic stain, was evaluated. Patients with and without VPSI were compared for clinicopathologic parameters and survival. VPSI was identified in 170 (53.0 %) of the stage I–III cases and 98 (43.4 %) of the patients with stage I disease. VPSI was associated with a higher frequency of tumor size greater than 3 cm, moderate/poor differentiation, vascular invasion, mediastinal lymph node metastasis, extranodal involvement, and higher TNM stages. Multivariate analysis revealed VPSI to be a significant independent predictor of unfavorable prognosis. The 5-year survival of patients with and without VPSI was 57.9 and 83.0 %, respectively (P = 0.001), and was 74.3 and 88.5 % (P = 0.005) in stages I–III and stage I disease, respectively. VPSI is an independent factor for poor prognosis in NSCLCs, regardless of lymph node status. Stage IB NSCLCs with PL1 pleural invasion are associated with a survival rate similar to that of stage IA NSCLCs and could be classified as T1 lesions. While surgical treatment is adequate in these patients, stage IB NSCLCs with VPSI have poor prognosis, and these patients should be considered for adjuvant chemotherapy.

Trop-2, a cell surface glycoprotein, contains both extracellular epidermal growth factor-like and thyroglobulin type-1 repeat domains. Low TROP2 expression was observed in lung adenocarcinoma tissues as compared with their normal counterparts. The lack of expression could be due to either the loss of heterozygosity (LOH) or hypermethylation of the CpG island DNA of TROP2 upstream promoter region as confirmed by bisulphite sequencing and methylation-specific (MS) polymerase chain reaction (PCR). 5-Aza-2′-deoxycytidine treatment on lung cancer cell (CL) lines, CL1-5 and A549, reversed the hypermethylation status and elevated both TROP2 mRNA and protein expression levels. Enforced expression of TROP2 in the lung CL line H1299 reduced AKT as well as ERK activation and suppressed cell proliferation and colony formation. Conversely, silencing TROP2 with shRNA transfection in the less efficiently tumour-forming cell line H322M enhanced AKT activation and increased tumour growth. Trop-2 could attenuate IGF-1R signalling-mediated AKT/β-catenin and ERK activation through a direct binding of IGF1. In conclusion, inactivation of TROP2 due to LOH or by DNA methylation may play an important role in lung cancer tumourigenicity through losing its suppressive effect on IGF-1R signalling and tumour growth.

Objective. Tongue cancer is well known to have a high poten- tial for locoregional metastasis. However, controversy about electively treating the neck in early-stage tongue cancer re- mains. Although many risk factors related to cervical occult nodal metastasis (ONM) have been investigated, the ability of the tumor to spread, a phenomenon that results from the intrinsic property of the tumor and its interaction with the surrounding environment, has seldom been addressed. Study Design. Retrospective case series with chart review. Setting. Tertiary referral hospital of university. Subjects. Patients with early-stage squamous cell carcinoma of the oral tongue. Results. In 71 eligible enrolled patients, ONM was detected in 19 (27%) patients, while the results were negative (ONM(−)) in 52 (73%) patients. The average tumor satellite distance (TSD) in the ONM(+) group was 4.1 ± 4.3 mm, in contrast to that in the ONM(−) group (1.0 ± 1.5 mm; P < .001). When stratified by increased TSD values, the significance of the dif- ference between the 2 groups increased. For clinical applica- tions, the optimal TSD threshold for determining the ONM probability was 3.5 mm. Multivariate analyses demonstrated that TSD was an independent prognosticator. Conclusions. The results indicate that TSD is a feasible pathologi- cal parameter that is useful for determining the status of cervical nodal metastasis. It can be used as an indicator of potential cervi- cal subclinical disease and as a guideline for deciding the necessity and modality of neck treatment.

Pleomorphic carcinomas of the lung are uncommon malignant tumors composed of carcinomatous and sarcomatous components and are distinguished from other non-small-cell lung carcinomas by a more aggressive clinical course with early distant metastases and far worse survival. Epidermal growth factor receptor (EGFR) and p53 are common genes involved in the pathogenesis of non-small-cell lung carcinomas, but their roles in pleomorphic carcinomas are unclear. The potential clinical activity of EGFR-targeted therapy is also unknown. A total of 42 pleomorphic carcinomas were identified to investigate somatic mutations of EGFR and p53. Genomic DNA was extracted from microdissected cells of paraffin-embedded tumor tissues. Somatic mutations in EGFR (exons 18–21) and p53 (exons 5–8) were examined. EGFR mutations were detected in 10 of 42 cases. Five of these patients had point mutations in exon 21 majorly with L858R; this mutation was found in both adenocarcinomatous and sarcomatous components in 1 case. The other 5 cases harbored 4 deletions and 1 mutation in exon 19. p53 mutations were found in 12 patients. Notably, identical mutation was observed in carcinomatous and sarcomatous components in 3 patients, and this finding strongly supported the theory of monoclonal histogenesis. The occurrence (23.8%) of EGFR mutations, including the exons 19 and 21 mutations observed frequently in our series, suggests that the patients with inoperable pleomorphic carcinomas are likely to benefit from treatment with EGFR tyrosine kinase inhibitors.

Mediastinal seminomas are uncommon primary mediastinal neoplasms, and most are solid in appearance. Cystic mediastinal seminoma is an unusual type of extragonadal germ cell tumor that has rarely been reported in the literature. Here we describe a 36-year-old man with a 7.5-cm cystic mediastinal seminoma. The tumor was excised successfully by video-assisted thoracoscopic surgery. No recurrence was noted during 28 months of follow-up.

p53 and epidermal growth factor receptor (EGFR) are common genes involved in the pathogenesis of lung cancer, but their roles in lymphoepithelioma-like carcinomas (LELC) are unclear. In this study, we investigate the roles of p53 and EGFR in LELC carcinogenesis. Forty-six pulmonary LELCs were identified to evaluate p53 and EGFR aberrations. p53 mutations were identified in three patients, which all occurred in exon 8. EGFR mutations were detected in 8 of 46 cases with a majority of exon 21 mutations but without L858R. The other cases harbored mutations in exons 20 and 18. Only one case gained a deletion in exon 19. Notably, EGFR mutation was more commonly observed in patients with tumor size ≤ 3 cm (P = 0.014). In addition, there was a trend of more common EGFR overexpression in female (22/30) than in male patients (7/16, P = 0.061). However, there was no correlation between p53/EGFR mutations and protein expressions, suggesting the presence of complex mechanisms. p53 and EGFR mutations are uncommon in LELCs, indicating that these genes are not the important events in carcinogenesis for this tumor subtype. The EGFR mutation in 35% patients with LELC tumors

Autogenic bone graft is the first choice for managing bone defects. However, donor site-associated morbidity and limited bone volume are constraints in clinical applications. Allografts can provide sufficient amounts for bone defects but have a high risk of infection. Bone substitute composed of hydroxyapatite (HA) is an alternative material for avoiding the aforementioned risks. Sintered bovine bone is a naturally occurring HA that has been proved to have excellent bioactivity for inducing osteoblastic expression and new bone formation in animal studies. The objective of this study was to evaluate the interactions between the tissue and the bone substitute composed of HA (sintered from bovine bone) in the human body.From 2003 to 2005, a total of 33 patients were enrolled to receive the sintered bovine HA as a bone substitute. Inclusion criteria were fractures with bony defects, benign bone tumors with a cavity, and spinal fusions. Bone healing was monitored by a series of radiographs, and bone microstructure was checked by scanning electron microscopy (SEM) and von Kossa staining.In 81.8% (27/33) of cases, significant fusion mass formation was visible in the radiographs after 6-12 months. New bone formation on the surface of the sintered bovine HA was seen under microscopic observation. Tight bonding between the interface of the bone and the sintered bovine HA was shown with SEM/energy-dispersive spectroscopy and von Kossa staining.Sintered bovine HA is a suitable material as a bone substitute to provide bone growth and promote bone healing.