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Supplementary Figure 2A from ST14 Gene Variant and Decreased Matriptase Protein Expression Predict Poor Breast Cancer Survival

Supplementary Figure 3 from ST14 Gene Variant and Decreased Matriptase Protein Expression Predict Poor Breast Cancer Survival

Background: Matriptase plays a role in carcinogenesis, but the role of its genetic variation or that of the hepatocyte growth factor activator inhibitor-1 (HAI-1) has not been evaluated. This study aimed to examine the genetic variation of matriptase (ST14 gene) and HAI-1 (SPINT1 gene) in breast cancer risk and prognosis, to assess matriptase and HAI-1 gene and protein expression in breast tumors, and to identify their clinicopathologic correlations and prognostic significance.Methods: Five single nucleotide polymorphisms in ST14 and three in SPINT1 were genotyped in 470 invasive breast cancer cases and 446 healthy controls. Gene expression analysis was done for 40 breast cancer samples. Protein expression was assessed by immunohistochemical analyses in 377 invasive breast tumors. The statistical significance of the associations among genotypes, clinicopathologic variables, and prognosis was assessed.Results: The ST14 single nucleotide polymorphism rs704624 independently predicted breast cancer survival, a poor outcome associated with the minor allele (P = 0.001; risk ratio, 2.221; 95% confidence interval, 1.382-3.568). Moreover, ST14 gene expression levels were lower among the minor allele carriers (P = 0.009), and negative/low matriptase protein expression was independently predictive of poorer survival (P = 0.046; risk ratio, 1.554; 95% confidence interval, 1.008-2.396).Conclusions: The ST14 variant rs704624 and protein expression of matriptase have prognostic significance in breast cancer. This study adds to the evidence for the role of matriptase in breast cancer and has found new evidence for the genotypes having an impact in breast cancer.Impact: This is the first study showing that genetic variation in matriptase has clinical importance. The results encourage further study on the genetic variation affecting protein levels and function in type II transmembrane serine proteases. Cancer Epidemiol Biomarkers Prev; 19(9); 2133–42. ©2010 AACR.

Supplementary Figure Legends from ST14 Gene Variant and Decreased Matriptase Protein Expression Predict Poor Breast Cancer Survival

Supplementary figures S2-S6. Supplementary Figure S2: A diagram depicting the location of the studied SNPs and miR-200a binding site in KEAP1 gene. Supplementary Figure S3: Association of KEAP1 rs11085735 with breast cancer survival among the KBCP and OBCS breast cancer cases. Supplementary Figure S4: Association of KEAP1 rs11085735 with breast cancer survival among the ER positive cases. Supplementary Figure S5: Association of KEAP1 rs11085735 with breast cancer survival among KBCP cases with lower (

Supplementary tables S1-S10. S1. Significant associations of the KEAP1 protein expression and receptor statuses; S2. Association of the KEAP1 protein expression with NRF2 protein expression; S3. Analyzed polymorphisms in the KEAP1 gene; S4. Significant associations of the KEAP1 SNP genotypes with KEAP1 protein expression; S5. Associations of the KEAP1 SNP s11085735 genotypes with NRF2 protein expression; S6. Associations of the KEAP1 SNPs with breast cancer survival in multivariate analysis among invasive KBCP and OBCS breast cancer cases separately; S7. Variables significantly associated with breast cancer survival in multivariate analysis among invasive KBCP and OBCS breast cancer cases; S8. Significant associations with breast cancer survival in univariate analysis (Kaplan-Meier) according to KEAP1 SNP rs11085735 genotypes among KBCP and OBCS ER positive cases, KBCP ER positive cases and KBCP cases with low/negative ({less than or equal to}1.33) KEAP1 protein expression levelS9. Variables significantly associated with breast cancer survival in multivariate analysis among ER positive KBCP breast cancer cases; S10. Variables significantly associated with breast cancer survival in multivariate analysis among (KBCP) cases with lower KEAP1 protein expression

Purpose: Defective oxidative stress response may increase cancer susceptibility. In tumors, these rescue mechanisms may cause chemo- and radioresistance impacting patient outcome. We previously showed that genetic variation in the nuclear factor erythroid 2–related factor 2 (NFE2L2) is associated with breast cancer risk and prognosis. Here we further studied this pathway by investigating Kelch-like ECH-associated protein 1 (KEAP1).Experimental Design: Five tagging SNPs in the KEAP1 gene were genotyped in 996 breast cancer cases and 880 controls from two Finnish case–control sets. KEAP1 protein expression was studied in 373 invasive breast cancer tumors.Results: rs34197572 genotype TT was associated with increased risk of breast cancer in the KBCP samples [P = 1.8×10−4; OR, 7.314; confidence interval (CI), 2.185–24.478]. rs11085735 allele A was associated with lower KEAP1 protein expression (P = 0.040; OR,= 3.545) and high nuclear NRF2 expression (P = 0.009; OR, 2.445) and worse survival in all invasive cases (P = 0.023; HR, 1.634). When including treatment data, rs11085735 was associated with recurrence-free survival (RFS; P = 0.020; HR, 1.545) and breast cancer–specific survival (P = 0.016; HR, 1.683) and rs34197572 with overall survival (P = 0.045; HR, 1.304). rs11085735 associated with RFS also among tamoxifen-treated cases (P = 0.003; HR, 3.517). Among radiotherapy-treated cases, overall survival was associated with rs34197572 (P = 0.018; HR, 1.486) and rs8113472 (P = 0.025; HR, 1.455). RFS was associated with rs9676881 (P = 0.024; HR, 1.452) and rs1048290 (P = 0.020; HR, 1.468) among all invasive cases and among estrogen receptor (ER)-positive tamoxifen-treated cases (P = 0.018; HR, 2.407 and P = 0.015; HR, 2.476, respectively).Conclusions: The present findings suggest that the investigated SNPs have effects related to oxidative stress induced by cancer treatment, supporting involvement of the NRF2/KEAP1 pathway in breast cancer susceptibility and patient outcome. Clin Cancer Res; 21(7); 1591–601. ©2015 AACR.

Supplementary table and figure legends

PDF file - 40K, A diagram depicting the genetic location of the studied SNPs in the NRF2 and SRXN1 genes

PDF file - 96K, Breast cancer survival according to NRF2 and SRXN1 genotypes and NRF2 protein expression

PDF file - 43K, Clinicopathological characteristics of the patients

PDF file - 1.2MB, Survival curves for breast cancer survival according to NRF2 and SRXN1 genotypes and protein expression

PDF file - 272K

Supplementary figures S1-5, materials and methods. 1) Supplementary figures and associated legends: - Figure S1. ERK silencing decreases breast cancer cell migration and invasion but does not influence cell proliferation. - Figure S2. ERK2 does not interact with actin or keratin 8 and ERK2 phosphorylation is supported by vimentin but not actin. - Figure S3. Fluorescence recovery of wild-type Slug-GFP after photobleaching (FRAP). - Figure S4. Mass spectrometric identification of ERK1/2-dependent phosphorylation sites on Slug and validation using a newly generated Slug phospho-serine-87 antibody. - Figure S5. The phosphorylation status of Slug does not influence repression of E-Cadherin, Slug nuclear localisation or stability. 2) Supplementary materials and methods 3) Supplementary references

PDF file - 69K, Analysed SNPs in the NRF2 and the SRXN1 genes

NRF2 activates several protective genes, such as sulfiredoxin (SRXN1), as a response to oxidative and xenobiotic stress. Defects in NRF2 pathway may increase cancer susceptibility. In tumor cells, activation of NRF2 may lead to chemo- and radioresistance and thus affect patient outcome. Nine single-nucleotide polymorphisms on NRF2 gene and eight on SRXN1 were genotyped in 452 patients with breast cancer and 370 controls. Protein expression of NRF2 and SRXN1 was studied in 373 breast carcinomas by immunohistochemistry. Statistical significance of the associations between genotypes, protein expression, clinicopathologic variables, and survival was assessed. A high level (>25%) of cytoplasmic NRF2 positivity was observed in 237 of 361 (66%) and SRXN1 positivity was observed in 82 of 363 (23%) cases. The NRF2 rs6721961 genotype TT was associated with increased risk of breast cancer [P = 0.008; OR, 4.656; confidence interval (CI), 1.350–16.063] and the T allele was associated with a low extent of NRF2 protein expression (P = 0.0003; OR, 2.420; CI, 1.491–3.926) and negative SRXN1 expression (P = 0.047; OR, 1.867; CI = 1.002–3.478). The NRF2 rs2886162 allele A was associated with low NRF2 expression (P = 0.011; OR, 1.988; CI, 1.162–3.400) and the AA genotype was associated with a worse survival (P = 0.032; HR, 1.687; CI, 1.047–2.748). The NRF2 rs1962142 T allele was associated with a low level of cytoplasmic NRF2 expression (P = 0.036) and negative sulfiredoxin expression (P = 0.042). The NRF2 rs2706110 AA genotype was associated with an increased risk of breast cancer, and the SRXN1 rs6053666 C allele was associated with a decrease in breast cancer risk (P = 0.011 and 0.017). NRF2 and SRXN1 genetic polymorphisms are associated with breast cancer risk and survival, implicating that mechanisms associated with reactive oxygen species and NRF2 pathway are involved in breast cancer initiation and progression. Cancer Res; 72(21); 5537–46. ©2012 AACR.

PDF file - 73K, Predicted functional effects of the studied SNPs in NRF2 and SRXN1 genes

PDF file - 68K, Association between NRF2 and SRXN1 protein expression

PDF file - 69K

Objectives: To investigate the protein expression of the epithelial-mesenchymal transition-inducing transcription factors (TFs) Twist, ZEB1 and Slug in peripheral T-cell lymphomas (PTCL) and their ...

BACKGROUND Oral lichen planus (OLP) is a chronic T-cell-mediated inflammatory disease, which is associated with increased risk of developing oral squamous cell carcinoma. Epithelial-to-mesenchymal transition is a physiological phenomenon occurring during growth and organogenesis, but it has also an important role in tumorigenesis. In the present work, we studied the expression of known epithelial-to-mesenchymal transition markers in oral lichen planus. METHODS In total, 54 oral lichen planus and 22 control samples were analyzed for epithelial-to-mesenchymal transition markers. Samples were immunohistochemically stained for claudin-1, claudin-4 and claudin-7, cadherin-1 (E-cadherin), Twist-related protein 1 (TWIST1) and zinc finger E-box-binding homeobox 1 (ZEB1). RESULTS The expression of claudin-1, claudin-4 and E-cadherin was significantly weaker in oral lichen planus epithelium compared to controls (P

Objectives: Protein levels of the transcription factor nuclear factor erythroid-derived 2-like 2 (Nrf2) and its inhibitor Kelch-like ECH-associated protein 1 (Keap1) have been proposed as prognostic factors in pancreatic ductal adenocarcinomas (PDACs). These cellular redox-state-regulating enzymes are targeted by several microRNAs, including miR-93 and miR-200a. Methods: We assessed mRNA levels of Nrf2 and Keap1 and tissue expression of miR-93 and miR-200a in 51 patients with surgically treated PDAC. Expression levels were separately measured in malignant cells and adjacent benign cells. Results: Keap1 and Nrf2 mRNA expression levels in cancer cells were lower than in adjacent benign tissue (Wilcoxon’s test; p = 0.0015 and p = 0.000032, respectively). Conversely, miR-93 expression was higher in cancer cells than in adjacent benign tissue (p = 0.00082). Low levels of miR-93 and miR-200a in cancer cells were associated with poorer differentiation (p = 0.004 and p = 0.002, respectively). In univariate survival analysis, benign-tissue levels of miR-200a above the median predicted better relapse-free survival (RFS) (p = 0.045). Conclusions: High miR-93 and miR-200a levels in cancer cells of PDAC were associated with better differentiation, and miR-200a expression in benign tissue with excellent RFS. Keap1 and Nrf2 mRNA levels showed prominent down-regulation in cancerous versus benign tissue, but they were not associated with disease aggressiveness or outcome.

AIMS Oral tongue squamous cell carcinoma (OTSCC) has a relatively poor outcome, and there is a need to identify better prognostic factors. Recently, tumour-stroma ratio (TSR) has been associated with prognosis in several cancers. The aim of this multi-institutional study was to evaluate the prognostic value of TSR from original haematoxylin and eosin (HE)-stained tumour-resection slides in a series of early-stage (cT1-2N0) OTSCC patients. METHODS AND RESULTS A TSR cutoff value of 50% was used to divide the patients into stroma-rich (≥50%) and stroma-poor (

It is of great clinical importance to identify simple prognostic markers from preoperative biopsies that could guide treatment planning. Here, we compared tumor budding (B), depth of invasion (D), and the combined scores (i.e., budding and depth of invasion (BD) histopathologic model) in preoperative biopsies and the corresponding postoperative specimens of oral tongue squamous cell carcinoma (OTSCC). Tumor budding and depth of invasion were evaluated in the pre- and postoperative samples from 100 patients treated for OTSCC. Sensitivity and specificity statistics were used. Our results showed statistically significant (P < 0.001) relationship between pre- and postoperative BD scores. There was an agreement between the pre- and postoperative BD model scores in 83 cases (83%) with 57.1% sensitivity (95% CI 39.4 to 73.7%) and 96.9% specificity (95% CI 89.3 to 99.6%). Our findings suggest that the BD model, analyzed from representative biopsies, could be used for the treatment planning of OTSCC.

BACKGROUND/AIM The aim of the study was to investigate the prognostic role of androgen receptor (AR), mineralocorticoid receptor (MR) and glucocorticoid receptor β (GRβ) expression in HER-2 negative breast cancer patients. MATERIALS AND METHODS The study population (n=152) was enriched with triple-negative breast cancers (TNBC) (n=96; 63.2%). The median follow-up time was 100 months. AR, MR and GRβ immunocytochemical staining was compared with that of epithelial-mesenchymal transition (EMT) markers (vimentin, SIP1, ZEB1). RESULTS High expression of cytoplasmic MR was associated with dismal local relapse-free survival (RR=13.923; 95%CI=1.071-181.045; p=0.044) in tumours with non-TNBC phenotype. AR and GRβ were more frequently expressed in ER+/PR+/HER2- tumours, while cytoplasmic MR was more often expressed in TNBC tumours (for all, p

Background:Oral tongue squamous cell carcinoma (OTSCC) metastasises early, especially to regional lymph nodes. There is an ongoing debate on which early stage (T1-T2N0) patients should be treated with elective neck dissection. We need prognosticators for early stage tongue cancer. Methods: Mice immunisation with human mesenchymal stromal cells resulted in production of antibodies against tenascin-C (TNC) and fibronectin (FN), which were used to stain 178 (98 early stage), oral tongue squamous cell carcinoma samples. TenascinC and FN expression in the stroma (negative, moderate or abundant) and tumour cells (negative or positive) were assessed. Similar staining was obtained using corresponding commercial antibodies. Results: Expression of TNC and FN in the stroma, but not in the tumour cells, proved to be excellent prognosticators both in all stages and in early stage cases. Among early stages, when stromal TNC was negative, the 5-year survival rate was 88%. Correspondingly, when FN was negative, no cancer deaths were observed. Five-year survival rates for abundant expression of TNC and FN were 43% and 25%, respectively. Conclusions: Stromal TNC and, especially, FN expressions differentiate patients into low-and high-risk groups. Surgery alone of early stage primary tumours might be adequate when stromal FN is negative. Aggressive treatments should be considered when both TNC and FN are abundant.

Background: Epigenetic regulators, including Jumonji domain 2 (JMJD2/KDM4) proteins are involved in post-translational modification of histone demethylation and have a major role in carcinogenesis of many solid tumors. Materials and Methods: We assessed immunohistochemically the expression of lysine (K)-specific demethylase 4 (KDM4)A, KDM4B and KDM4D in tumors from 91 patients of adriamycin, bleomycin, vinblastine, darcabazine (ABVD)-treated classical Hodgkin lymphoma. Results: Strong cytoplasmic KDM4B expression in the reactive cellular infiltrate and also in Reed-Sternberg (RS) cells predicted poor relapse-free survival (RFS) (p=0.020 and p=0.022, respectively) in patients with limited-stage disease. Strong KDM4B expression in RS cells was also related to B-symptoms (p=0.007) and advanced stage (p=0.024). Strong KDM4D expression in the cytoplasm of RS cells was also associated with poor RFS in limited-stage patients RFS (p=0.043) and, most significantly, in patients receiving involved-field radiotherapy (p=0.007). Conclusion: KDM4B and KDM4D expression may associate with an aggressive subtype of classical Hodgkin lymphoma and be linked with radioresistance.

One of the main changes in the 8th edition of the American Joint Committee on Cancer (AJCC) for staging of oral cancer is the inclusion of depth of invasion (DOI) in the T category. However, cancers in different oral subsites have variable behavior, with oral tongue squamous cell carcinoma (OTSCC) being the most aggressive one even at early stage. Thus, it is necessary to evaluate the performance of this new T category in homogenous cohort of early OTSCC. Therefore, we analyzed a large cohort of patients with a small (ae4 cm) OTSCC to demonstrate the differences in T stage between the AJCC 7th and 8th editions. A total of 311 early-stage cases (AJCC 7th) of OTSCC were analyzed. We used 5 mm and 10 mm DOI for upstaging from T1 to T2 and from T2 to T3 respectively, as in the AJCC 8th. We further reclassified the cases according to our own proposal suggesting 2 mm to upstage to T2 and 4 mm to upstage to T3. According to AJCC 7th, there were no significant differences in the survival analysis. When we applied the 8th edition, many cases were upstaged to T3 and thus associated with worse disease-specific survival (HR 2.37, 95% CI 1.12-4.99) and disease-free survival (HR 2.12, 95% CI 1.09-4.08). Based on our proposal, T3 cases were associated with even worse disease-specific survival (HR 4.19, 95% CI 2.27-7.74). The 8th edition provides better survival prediction for OTSCC than the 7th and can be further optimized by lowering the DOI cutoffs.

Diffuse large B-cell lymphoma (DLBCL) is an aggressive lymphoma with diverse outcomes. Concurrent translocation of MYC and BCL-2 and/or BCL-6, and concurrent immunohistochemical (IHC) high expression of MYC and BCL-2, have been linked to unfavorable treatment responses. TP53-mutated DLBCL has also been linked to worse outcome. Our aim was to evaluate the aforementioned issues in a cohort of 155 patients uniformly treated with R-CHOP-like therapies. We performed direct sequencing of TP53 exons 5, 6, 7 and 8 as well as fluorescence in-situ hybridization (FISH) of MYC, BCL-2 and BCL-6, and IHC of MYC, BCL-2 and BCL-6. In multivariate analysis, TP53 mutations in L3 and loop-sheet helix (LSH) associated with a risk ratio (RR) of disease-specific survival (DSS) of 8.779 (p = 0.022) and a RR of disease-free survival (DFS) of 10.498 (p = 0.011). In IHC analysis BCL-2 overexpression was associated with inferior DFS (p = 0.002) and DSS (p = 0.002). DLBCL with BCL-2 and MYC overexpression conferred inferior survival in all patients (DSS, p = 0.038 and DFS, p = 0.011) and in patients with non-GC phenotype (DSS (p = 0.013) and DFS (p = 0.010). Our results imply that in DLBCL, the location of TP53 mutations and IHC analysis of BCL-2 and MYC might have a role in the assessment of prognosis.

Background/aim The role of histone demethylators, such as Jumonji domain 2 (JMJD2/KDM4) proteins, and histone deacetylases, such as sirtuins (SIRT) is poorly characterized in pancreatic carcinomas while they have a major role in the carcinogenesis of several other tumours. Materials and methods We assessed retrospectively with immunohistochemistry the expressions of KDM4A, KDM4B and KDM4D in 81 and SIRT1-4 in 102 pancreatic adenocarcinomas. Immunostaining was evaluated separately in benign pancreatic tissues and in malignant cells. Results High nuclear KDM4D expression in benign pancreatic tissue from resection margins associated with dismal disease-free survival (DFS) (OR=8.00; 95%CI=1.87-33.9; p=0.005), even more significantly than tumour size and lymph node involvement. High cytoplasmic SIRT2 expression in benign pancreatic tissues also associated with a shorter DFS, but only in univariate analysis (p=0.026). Conclusion Nuclear KDM4D and SIRT2 expression deviated from that of benign pancreatic tissue thus putatively influencing gene expression of tumor cells. Regardless, none of the enzymes studied had a decisive role in the spread of pancreatic cancer. A high nuclear expression of KDM4D in samples of pancreatic resection margins significantly and independently predicted an earlier recurrence and could thus be used in the assessment of risk of relapse in clinical practice.

Nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) is an indolent Hodgkin lymphoma (HL) entity counting for ∼5% of all HLs [1]. However, it seems to be a biologically distinct disease with a f...

Nuclear factor erythroid 2-related factor 2 (NRF2), DJ1 and sulfiredoxin 1 (SRXN1) are transcription factors which protect cells from the oxidative damage caused by reactive oxygen species and, on the other hand, are associated with resistance to cancer treatments. The immunohistochemical expression of NRF2, DJ1 and SRNX1 was assessed in human grade II-IV astrocytic gliomas. Their association to clinicopathologic and essential molecular factors was evaluated. The RNA expression levels and genetic alterations were analyzed from publicly available datasets. All studied molecules were commonly expressed. The cytoplasmic NRF2 expression was higher in tumors with a higher malignancy grade, whereas the nuclear and cytoplasmic DJ1 expression was associated with a lower grade. The presence of the isocitrate dehyrdogenase 1 mutation (IDH1) was associated with an increasing cytoplasmic and nuclear expression of NRF2 and a nuclear DJ1 expression. When primary grade IV astrocytomas were compared to secondary glioblastomas, nuclear DJ1 was associated with secondary tumors. In grade II-IV tumors, the cytoplasmic NRF2 expression was associated with a poor prognosis, whereas nuclear NRF2 and both cytoplasmic and nuclear DJ1 were associated with a better patient prognosis. Recurrent homozygous deletions of DJ1 were observed, especially in the IDH wild-type samples. When only the glioblastomas were evaluated, nuclear NRF2 and SRNX1 predicted better survival. As a conclusion, NRF2, DJ1 and SNXR1 can be used as prognosticators in gliomas.

Central nervous system (CNS) relapse occurs in around 5% of diffuse large B-cell lymphoma (DLBCL) cases. No biomarkers to identify high-risk patients have been discovered. We evaluated the expression of lymphocyte-guiding chemokine receptors in systemic and CNS lymphomas. Immunohistochemical staining for CXCR4, CXCR5, CCR7, CXCL12, and CXCL13 was performed on 89 tissue samples, including cases of primary central nervous system lymphoma (PCNSL), secondary CNS lymphoma (sCNSL), and systemic DLBCL. Also, 10 reactive lymph node samples were included. Immunoelectron microscopy was performed on two PCNSLs, one sCNSL, one systemic DLBCL, and one reactive lymph node samples, and staining was performed for CXCR4, CXCR5, CXCL12, and CXCL13. Chi-square test was used to determine correlations between clinical parameters, diagnostic groups, and chemokine receptor expression. Strong nuclear CXCR4 positivity correlated with systemic DLBCL, whereas strong cytoplasmic CXCR5 positivity correlated with CNS involvement (P = 0.003 and P = 0.039). Immunoelectron microscopy revealed a nuclear CXCR4 staining in reactive lymph node, compared with cytoplasmic and membranous localization seen in CNS lymphomas. We found that CNS lymphoma presented a chemokine receptor profile different from systemic disease. Our findings give new information on the CNS tropism of DLBCL and, if confirmed, may contribute to more effective targeting of CNS prophylaxis among patients with DLBCL.

Aims Diffuse large B-cell lymphoma (DLBCL) is an aggressive and potentially fatal disease. Prediction of risk of relapse is based on clinical markers. There is a need for more accurate biomarkers to select patients for more aggressive first-line treatments. Peroxiredoxins (Prxs) are a family of potent antioxidant proteins. Their prognostic role in DLBCL is unknown. Methods Altogether, 103 diagnostic biopsy samples from patients with DLBCL were immunohistochemically stained for Prxs I, II, III, V and VI. Results Strong Prx VI expression was associated with the presence of B-symptoms. There were no other significant associations with traditional risk factors. Five-year disease-specific survival was 68.6% in patients with high cytoplasmic Prx VI intensity vs 97.0% in those with low intensity. In multivariate analysis, high Prx VI expression (HR 12.846, 95% CI 1.722 to 95.807, p=0.013) was an independent risk factor of lymphoma-associated death not related to International Prognostic Index score (HR 2.514, 95% CI 1.040 to 6.073, p=0.041). Conclusions High intensity of cytoplasmic Prx VI expression in pretreatment DLBCL samples predicts worse outcome in patients with DLBCL. Whether Prx VI is associated with chemoresistance, and therefore a poorer outcome, needs to be evaluated. If Prx VI is a predictive marker and it proves causality, it would be crucial to study Prx VI ability to become a target enzyme for treatment.

Despite early diagnosis and treatment, almost 20 % of patients with early-stage (cT1-cT2N0) oral tongue squamous cell carcinoma (OTSCC) still die of their disease. The prognosis of OTSCC patients is influenced by several demographic, clinical, and histopathologic factors. The aim of this multicenter international study was to find which of the factors age, gender, stage, grade, lymphocytic host re- sponse, perineural invasion, worst pattern of invasion, or depth of invasion has the strongest prognostic power in early-stage OTSCC. Patient data of 479 patients with early-stage (cT1-2N0) OTSCC in Finland, Brazil, and the USA were retrieved and analyzed using Cox proportional hazards regression models. Our results indicate that depth of invasion (DOI) and worst pattern of invasion (WPOI) are the strongest pathological predictors for locoregional recurrence, with a hazard ratio (HR) for 4 mm DOI of 1.67 (95 % confidence interval (CI) 1.07-2.60) and HR for WPOI of 1.46 (95 %C I 0.95-2.25). In addition, mor- tality from early OTSCC was also predicted by DOI (HR 2.44, 95 % CI 1.34-4.47) and by WPOI (HR 2.34, 95 % CI 1.26-4.32). We suggest that clinically early-stage oral tongue carcinomas 4 mm or deeper, or with a growth pat- tern of small cell islands or satellites, should be considered as high-risk tumors which require multimodality treatment.

Aims To investigate redox-regulating enzymes that may have a special role in melanoma pathogenesis due to continuous exposure to microenvironment-produced and ultraviolet radiation-induced oxidative stress. Methods and results We assessed immunohistochemically the expression of antioxidant enzymes peroxiredoxins (Prxs) I–IV, sulfiredoxin (Srx) and redox-regulated proto-oncogene DJ-1 in material consisting of 30 benign naevi, 14 lentigo malignas and 67 malignant melanomas. Evaluation of immunostaining was performed with special attention paid to protein expression in different tumour compartments. In particular, the expression patterns of nuclear Prx I and Prx II and cytoplasmic DJ-1 were decreased significantly in melanomas compared with dysplastic and benign naevi. In multivariate analysis, several prognostic factors were identified: Prx III expression in the cytoplasm of stromal fibroblasts was associated with shortened melanoma-specific survival [hazard ratio (HR) 6.730; 95% confidence interval (CI) 1.579–28.689], while cytoplasmic Prx IV expression in endothelial cells (HR 6.563; 95% CI 1.750–24.620) and Srx expression in the cytoplasm of keratinocytes (HR 6.988; 95% CI 1.559–31.324) were associated with better prognosis independently of ulceration, thickness of melanoma or its diagnostic type. Conclusions Redox-regulating enzymes have the potential to serve as novel prognostic factors and targeting them may offer new therapeutic options in malignant melanoma.

The prognostication of patient outcome is one of the greatest challenges in the management of early stage oral tongue squamous cell carcinoma (OTSCC). This study introduces a simple histopathological model for the prognostication of survival in patients with early OTSCC. A total of 311 cases (from Finland and Brazil) with clinically evaluated early stage OTSCC (cT1-T2cN0cM0) were included in this multicentre retrospective study. Tumour budding (B) and depth of invasion (D) were scored on haematoxylin-eosin-stained cancer slides. The cut-off point for tumour budding was set at 5 buds (low

Background We studied the expression of some major proteins involved in cell-cycle regulation and DNA repair, the roles of which are not well known in pancreatic ductal adenocarcinoma (PDAC), but which have a significant impact on carcinogenesis of many other cancers. Methods We immunohistochemically assessed expression levels of the cell-cycle regulators Rb1, p16 and cyclin-dependent kinase 4 (CDK4), and the DNA repair enzymes O6-methylguanine-DNA-alkyltransferase (MGMT) and flap endonuclease-1 (FEN1) separately in malignant tissue and benign tissue from resection margins in 102 cases of PDAC. Nearly all (95.1%) patients had undergone pancreaticoduodenectomy. Results The studied proteins showed wide but somewhat variable expression in both benign and malignant pancreatic tissues. Strong CDK4 expression in islets of Langerhans predicted poor relapse-free survival (RFS) (HR 2.874; 95% CI 1.261–6.550; p = .012) and within T3–4 tumors CDK4 expression in adenocarcinoma cells also predicted poor disease-free survival (DFS) (RR 2.148; 95% CI 1.081–4.272; p = .029). Strong MGMT expression was associated in N1 patients with weak local relapse-free survival (RFS), DFS and overall survival; all significantly in Cox regression analysis. FEN1 was also an independent predictor of decreased DFS (in the whole study population) and worse RFS (in the patients with T3–4 tumors). Conclusions Major cell-cycle regulator also have predictive significance, but further studies are required to evaluate this.

Background: The subtype of claudin-low breast cancer can be reliably determined only by gene-expression profiling. Attempts have been made to develop immunohistochemical surrogates, which nearly always focus on membranous claudin expression. Methods: We assessed the immunohistochemical expression of both membranous and cytoplasmic claudins 3, 4 and 7 in a series of 197 non-metastatic breast cancers, enriched with triple-negative breast cancers (TNBCs; 60%). The expression of epithelial-to-mesenchymal transition-regulating transcription factors Sip1, Zeb1 and vimentin had previously been determined in the same material. Results: In multivariate analysis, strong cytoplasmic claudin 3 expression was associated with poor relapse-free survival (RFS), disease-free survival, distant disease-free survival, breast cancer-specific survival and overall survival among TNBC patients (for RFS, RR 5.202, 95% CI 1.210–22.369, p = 0.027, vs. T-class, RR 0.663, 95% CI 0.168–2.623, p = 0.558, and N-class, RR 3.940, 95% CI 0.933–16.631, p = 0.062). Cytoplasmic claudin 3 expression was also associated with strong nuclear Sip1 expression (p = 0.000053), TNBC phenotype (p = 0.012) and within them, non-basal-like phenotype (p = 0.026). Cytoplasmic claudin 7 was associated with dismal RFS (RR 6.328, 95% CI 1.401–28.593, p = 0.016, vs. T-class, RR 0.692, 95% CI 0.242–1.982, p = 0.493, and N-class, RR 2.981, 95% CI 1.1016–8.749, p = 0.047). Low cytoplasmic expression of claudins 3, 4 and 7 together also predicted poor RFS (RR 6.070, 95% CI 1.347–27.363, p = 0.019, vs. T-class, RR 0.677, 95% CI 0.237–1.934, p = 0.467, and N-class, RR 3.167, 95% CI 1.079–9.290, p = 0.036). Conclusions: Immunohistochemical expression levels of cytoplasmic claudins 3 and 7 appear to be novel prognostic factors in TNBC.

Background Hypoxia-inducible factors (HIFs) and prolyl hydroxylase domain (PHD) proteins control cellular oxygen homeostasis and a wide range of other processes. Materials and methods We immunohistochemically assessed the expression of HIF1α, HIF2α, PHD1, PHD2 and PHD3 in 115 cases of classical Hodgkin's lymphoma, all treated in the first line with doxorubicin, bleomycin, vinblastine and darcabazine (ABVD) chemotherapy. Results In advanced-stage patients treated with involved-field radiotherapy (IFRT), nuclear HIF1α expression in reactive cellular infiltrate predicted prolonged relapse-free survival (RFS) (p=0.026). Strong cytoplasmic PHD1 expression in Reed-Sternberg cells was associated with poor RFS among patients treated with IFRT and advanced-stage patients treated with ABVD and IFRT (p=0.0028 and p=0.0058, respectively). In Cox regression analysis, PHD1 was a more significant predictor of relapse (risk ratio=18.383; 95% confidence interval(CI)=1.521-222.246; p=0.022) than the International Prognostic Score. Conclusion HIF and PHD expression appear to be novel prognostic biomarkers in classical Hodgkin's lymphoma.